Exhibit Hall B
Session 1: Cardiovascular, Chair: Lisa Haynie
8:00 Opening Remarks
8:02 MATRIX METALLOPROTEASE ACTIVITY IS INCREASED IN OBESE DOG KIDNEYS
Katrice Martin* and Jeffrey R. Henegar, Tougaloo College, Tougaloo, MS 39174 and University of Mississippi Medical Center, Jackson, MS 39216
Obesity is associated with changes in the extracellular matrix in the kidney including increases in collagen IV, glomerulosclerosis, and interstitial fibrosis. The extracellular matrix is maintained by a balance of production of matrix components by mesangial cells and fibroblasts and degradation of matrix components by matrix metalloproteases. It is unknown if the increase in extracellular matrix in the kidneys in obesity is due to an increase in production of or a decrease in degradation. Thus, the purpose of this study was to determine if matrix metalloprotease levels and activity are altered in obesity. We hypothesized that matrix metalloprotease activity would decrease thus allowing accumulation of extracellular matrix. Matrix metallopreotease activity was measured by zymography. Briefly, extracts of renal tissue were run on polyacrylamide gels containing either gelatin or casein. Gels were incubated in development buffer and lytic bands were visualized after staining with Coomassie Blue. The results show that both the 62kD collagenase and the 92kD gelatinase have increased activity in obese kidneys. There were no differences in the other metalloproteases present. These data suggest that metalloprotease activity is not responsible for the accumulation of extracellular matrix in obesity and, in fact, is working to decrease the amount of matrix. Thus, we conclude that extracellular matrix production and not decreased matrix metalloprotease activity is responsible for matrix accumulation in obese kidneys. This project was supported by the Mississippi Functional Genomic Network.
8:15 ROLE OF ALDOSTERONE IN MEDIATING HYPERTENSION AND TARGET ORGAN INJURY DURING OBESITY
Paul Brown* and Elizabeth Brandon, Tougaloo College, Tougaloo, MS 39174 and University of Mississippi Medical Center, Jackson, MS 39216
Obesity is a major risk factor for cardiovascular disease particularly hypertension. With the mechanisms involved still unclear, two studies were performed to examine the pathological changes associated with obesity. First, we tested whether obesity causes collagen accumulation in coronary arteries, leading to fibrosis and coronary artery dysfunction. Cardiac tissue from lean dogs and obese dogs fed a high fat diet for 6 weeks was stained with picrosirus red labeling collagen. An area of interest was defined using light microscopy and included the border of collagen around each coronary artery. To obtain the border containing collagen, the cross-sectional area of the artery was subtracted from the total artery area. In obese dogs, perivascular collagen was 17.5% greater than in lean dogs, indicating early fibrosis in coronary arteries after only 6 weeks of high fat diet. The second study tested the hypothesis that down-regulation of 11fO-hydroxysteroid dehydrogenase 2 (11 [pounds sterling]]-HSD2), an enzyme that inactivates cortisol in the kidney, may contribute to obesity and hypertension by permitting renal mineralocorticoid receptor (MR) activation by cortisol. Western blots for 11 [pounds sterling]]-HSD2 were performed using kidneys from 4 lean and 4 obese dogs. While two obese dogs had high levels of 11 [pounds sterling]]-HSD2, two had levels similar to the lean dogs. Preliminary results do not permit firm conclusions about the changes in 11 [pounds sterling]]-HSD2 during obesity.
8:30 EXPRESSION LEVELS OF ALLOGRAFT INFLAMMATORY FACTOR-1 (AIF-1) AND INTERLEUKIN-18 (IL-18) MIGHT PREDICT ALLOGRAFT REJECTION AFTER CARDIAC TRANSPLANTATION
Laura Piazza (1*), Charles Moore (2), Tammy Thomas (2), Sebron Harrison (1), Andy Barker (1), and D. Olga McDaniel (2), (1) University of Mississippi School of Medicine, Jackson, MS 39216 and (2) University of Mississippi Medical Center, Jackson, MS 39216
Background: Coronary vasculopathy (CV) is a major factor in long-term survival of heart transplantation. Inflammation might be a common background for both allograft rejection and CV. Cytokine gene arrays showed allograft inflammatory factor-1 (AIF-1) and interleukin-18 (IL-18) to be over-expressed in patients with severe rejection episodes. The specific objective was to determine the association of pro-inflammatory cytokines, specifically AIF-1 and IL-18, with allograft rejection scores to investigate their usefulness as prognostic markers. Hypothesis: Abnormal increases in pro-inflammatory cytokines may promote allograft rejection and vasculopathy following cardiac transplantation. Methods: Allograft recipients with varied rejection episodes were classified by histopathologic rejection assessment (grade 1A-3A/3B) and coronary vasculopathy. PBMCs were tested for AIF-1 and IL-18 expression using Reverse Transcriptase-PCR. Amplified DNA fragments were visualized using 2.0% agarose gel, and mRNA transcript levels quantified using UVP-8000 gel-base program. Results: mRNA transcript levels for IL-18 and AIF-1 expression were higher in samples from patients experiencing grade 3A allograft rejection compared to the same patient's expression in grade 0-1A and grade 2 rejections. A 1.1-fold increase in IL-18 and AIF-1 mRNA transcripts was observed in grade 3A rejection compared to grade 2 rejection, and 1.15-fold increase compared to grades 0-1A rejection. Conclusions: A direct correlation in the expression of AIF-1 and IL-18 and the clinical transplant results could provide a useful prognostic marker for the management of allograft rejection following heart transplantation.
8:45 THE IMPACT OF OBESITY ON VENOUS THROMBOSIS WHEN ASSOCIATED WITH TRAVEL
Kimberly C. Palmer, University of Mississippi Medical Center, Jackson, MS 39216
Background: There are estimated 200,000 new cases of venous thrombosis each year. Obesity and extended travel are two major risk factors. With the high prevalence of overweight/obesity in Mississippi, this study explored the link between extended travel, obesity, and the risk for venous thrombosis. Objectives: This study investigated the risk of developing VTE due to extended travel and obesity, and associated risk factors. Methods: The qualitative technique of ethnography was utilized. Two field observations were conducted at 1) the airport in Jackson, MS; and 2) truck stop in Brookhaven, MS. Both locations were assessed based on gender, race, estimated age, health status, and estimated travel. Results: Estimated 50% of the passengers at the airport were overweight/obese. Smoking status could not be determined. It was observed at the truck stop, that 65% of truckers were overweight while 30% were obese. Ninety percent of the truckers had the risk factor of being smokers. Most of the truckers were extended travelers from other states. In comparing the observations, at least half of all participants were overweight/obese. The truck drivers appear to be more at risk due to the known overweight/smoking combination. Conclusion: There is a need to make the public, especially extended travel truckers, aware of their risk for venous thrombosis due to existing risk factors. Many cases of venous thrombosis could be avoided if the public were more educated on the importance of mobility during extended travel. More research is needed to explore the risk among truckers.
9:00 ADENOSINE A1 RECEPTORS IN THE METABOLIC SYNDROME AND CORONARY ARTERY DISEASE
Alexis Hand (1*), Michael Sturek (2), Pamela Llyod (2), and Joseph A. Cameron (1), (1) Jackson State University, Jackson, MS 39217 and (2) Indiana University, School of Medicine, Indianapolis, IN 46202
In atherosclerotic cardiovascular disease (CVD), arteries are narrowed or completely blocked by atherosclerotic plaque. Metabolic syndrome, in humans, increases CVD risk. Previous studies from our lab have shown that activation of the adenosine A1 receptor (A1R) induces coronary smooth muscle cell proliferation; however, the association of A1R to CVD is unknown. Thus, the purposes of this study were to: 1) compare the effects of a high fat diet on development of metabolic syndrome and CVD in swine models and 2) determine whether A1R expression is altered by a high-fat diet. Male Yucatan and Ossabaw swine were treated for approximately 45 weeks with a normal low fat control diet (C) or a high fat/cholesterol (HF; 2% cholesterol) diet. Blood glucose and insulin during glucose tolerance test were obtained. Neointima formation was quantified using Verhoff-Van Gieson stain of the left main coronary artery as an index of CVD. A1R receptor mRNA levels were measured in the left anterior descending coronary artery (LAD) using real-time RT-PCR. Insulin to glucose ratios and neointima thickening were greater in the Ossabaw compared to the Yucatan swine, although no significant difference in A1R mRNA expression was found between C and HF swine. These results suggest that Ossabaw swine are a better model for metabolic syndrome and CVD, but the role of A1R is not clear. Supported in part by Am. Diab. Assoc., NIH RR013223, HL062552, R25 GM067592 and P01 AI56097-02S1.
9:30 Poster Session 1, Chair: Lisa Haynie
SURVEILLANCE OF GRAM NEGATIVE BACTERIA IN NON ICU SETTINGS: ANTIMICROBIAL SUSCEPTIBILITIES AND PREVALENCE
Erin C. Wiggers*, Mallory Phillips, and Donna C. Sullivan, University of Mississippi Medical Center, Jackson, MS 39216
Infections due to multidrug resistant gram negative bacteria are very common in the ICU but have been generally less common in the non ICU setting. This report presents results from the first 2 years of a 5 year study to determine the pattern of infection and antimicrobial resistance in non ICU settings. We collected 100 gram negative bacterial isolates from hospitalized, non ICU patients. Isolates were identified, speciated, and resistance patterns were determined. Bacteria were grown in Mueller Hinton broth and stored in glycerol at -80[degrees]C. Standardized microtiter minimal inhibitory concentration (MIC) panels for 13 antibiotics were used. Most isolates were cultured from urine (43% in 2004; 37% in 2005), miscellaneous wound sites (29% in 2004; 26% in 2005), respiratory sites (22% in 2004; 25% in 2005), and blood (9% in 2004; 13% in 2005). The distribution of sites of isolation, particularly urinary tract and wound infections, reflects the type of patient in non-ICU settings. In 2004, the most common isolates were Escherichia coli (39%), Pseudomonas aeruginosa (22%), and Enterobacter cloacae (12%). In 2005, the most common isolates were E. coli (33%), P. aeruginosa (16%) and Klebsiella pneumoniae (15%). Rates of resistance to various antibiotics increased across the board in all species tested in 2005 compared to 2004, following national trends of increased resistance in both hospital and community acquired pathogens.
EVALUATION OF THREE POLYPHENOLS AS POTENTIAL CHEMOTHERAPEUTIC AGENTS
Katie Womack*, Maxine Anderson, Michelle Tucci, and Hamed A. Benghuzzi, University of Mississippi Medical Center, Jackson, MS 39216
Polyphenols are compounds that are natural plant antioxidants. Antioxidants have been shown to prevent damage caused by free radicals to DNA and other molecules. Polyphenols have demonstrated several cancer preventive properties. In addition to antioxidant activity, these compounds may reduce abnormal cell growth and inflammation; help the body get rid of cancer-causing agents; and restore communication between different cells in the body. The objective of this study is to compare nontoxic phenols as cancer chemotherapeutic, agents and to apply these agents in treating human cancers. In this study the effects of a single dose of 10 [micro]M Green Tea Extract (ECGC), 5 [micro]M Tannic Acid (TA), or 5 [micro]M Thymoquinone (TQ) on Hep-2 cells were determined. Compounds were evaluated in their effectiveness to reduce cell number. The cells were also evaluated for membrane damage, and alterations in cellular morphology after 24, 48, and 72 hours in culture. The results showed a 50% reduction in Hep-2 cell numbers after 24 hours in TQ treated cells. After 48 hours, the cells treated with TQ and TA exhibited a four-fold decrease in total cell number compared to the control and ECGC treated cells. Cell numbers were similar in all treatment groups by 72 hours. At 48 hours the only significant increase in cell damage was seen in cells treated with ECGC. The results indicated that naturally occurring polyphenols given in a bolus dose could alter cellular viability. It appears the drug is either utilized by the cells or chemically degraded in the media leading to increased cell numbers back toward control values with time in culture. In order for the compounds to be effective chemotherapeutic agents, the data also suggests a need for continuous administration over bolus dosing.
DIFFERENTIAL EFFECTS OF CORTISOL ON NORMAL MRC-5 FIBROBLASTS AND HYPERTROPHIC LL-29 FIBROBLASTS
Casey McLendon*, Hamed A. Benghuzzi and Michelle Tucci, University of Mississippi Medical Center, Jackson, MS 39216
Cortisol is a glucocorticoid secreted by the adrenal cortex that helps facilitate the body's response to stress and regulate the immune system. Glucocorticoid receptors can be found on most cell types and as a consequence, cortisol hormone plays an essential role on the bodies physiologic systems. Corticosteroids are commonly employed in the treatment of inflammatory and fibrotic pulmonary disorders. It is unclear as to whether cortisol elicits differing responses from normal fibroblasts in comparison to hypertrophic fibroblasts. The purpose of this experiment is to analyze the differential effects of cortisol on normal MRC-5 lung fibroblasts and hypertrophic LL-29 lung fibroblasts from a patient with idiopathic pulmonary fibrosis. The objectives of the experiment were to obtain and culture normal and hypertrophic lung fibroblasts, to treat cells with hypophysiological, physiological, and supraphysiological doses of cortisol for 24, 48, and 72 hour incubation periods and to analyze cellular activity using the methods of cell count, protein assay, MDA, and morphological evaluation. In LL-29 cells, supraphysiological levels of cortisol stimulated cell growth only in the 24 hour incubation period without showing any changes in protein levels or cell damage. In contrast, MRC-5 cells showed increased growth at 48 hours with a dose specific increase and also had significantly increased protein levels. The largest increase in cell number was seen at the physiological concentration of cortisol in the MRC-5 cells. In conclusion, the two cell lines differ in their response to cortisol concentration in a dose and time dependent manner. Cortisol concentrations did not induce cellular damage throughout the experiment, and may be reflective of cortisol's role in membrane stabilization.
AQUEOUS V. AMYGDALINA EXTRACTS ALTER MCF-7 CELL MEMBRANE PERMEABILITY AND EFFLUX
Michael Opata*, and Ernest Izevbigie, Jackson State University, Jackson, MS 39217
Breast cancer is the second leading cause of cancer related deaths of women in the United States. Several treatment strategies have been developed over the past decade to reduce cancer morbidity and mortality rates. While mortality rates have declined in some ethnic populations, overall cancer incidence continues to grow. It was recently reported for the first time that, cancer has surpassed heart disease as the leading cause of deaths in the U.S. Therefore, novel chemotherapeutic agents are needed to improve cancer treatment outcome. Previous studies show that low concentration (microgram/ml) of water soluble leaf extracts of a Nigerian edible plant, V. amygdalina (VA), potently retarded the proliferative activity of human breast cancerous cells in vitro in a concentration-dependent fashion. Other reports show that the anti-proliferative activity of VA may be mitogen-activated protein kinase (MAPK)-dependent. The purpose of this study was to evaluate other mechanisms that could contribute to VA inhibitory actions. We hypothesized that VA exposure may compromise cell membrane resulting in alteration of efflux. To test this hypothesis, cells were incubated with 1[micro]Ci/ml [3H] thymidine for six hours at 37[degrees]C. After incubation, cells were washed with PBS, medium was replaced with a fresh medium, and [3H] thymidine release into the supernatant was determined. Our results showed that exposure of cells to VA decreased [3H] thymidine uptake in a concentration-dependent manner (P<0.03) but increased [3H] thymidine release into the supernatants (P<0.05). Thus suggesting that the membranes in the VA treated cells have been compromised. Studies to access effects of VA on apoptosis are underway in our Laboratory. Acknowledgement: This work was supported in part by Research Centers in Minority Institution (RCMI)/NIH grant # G122RR13459-07S1; National Center for Minority Health Disparities (NCMHD)/NIH grant # P20MD000534-01; and JSU Center for University Scholars Program.
ACTIVATION OF [NA.sup.+]/ [H.sup.+] EXCHANGER AND ALKALIZATION AS A MEDIATOR FOR ALDOSTERONE-INDUCED VASCULAR FIBROSIS
Tracie Perkins* and Jiankang Liu, Tougaloo College, Tougaloo, MS 39174 and University of Mississippi Medical Center, Jackson, MS 39216
Background: Increased aldosterone is believed to stimulate vascular fibrosis but the mechanisms involved are not clear. One mechanism that may link aldosterone with vascular fibrosis is activation of the [Na.sup.+]/[H.sup.+] exchanger (NHE). The NHE transports [Na.sup.+] out of the cell in exchange for [H.sup.+] and is believed to regulate cell proliferation. In vivo studies have demonstrated that aldosterone can increase NHE activity and intracellular pH (pHi) in vascular smooth muscle cells (VSMC) and treatment with the mineralocorticoid inhibitor, spironolactone attenuates these effects. Whether these effects of aldosterone are mediated via hemodynamic effects, such as increased blood pressure, by sodium retention, or by direct effects on VSMC are uncertain, and whether aldosterone-induced vascular fibrosis is mediated via the activation of the NHE is also poorly understood. Objectives: To determine whether aldosterone directly stimulates NHE and increases pHi in VSMC. Methods: Vascular smooth muscle cells (VSMC) from rat aorta were isolated grown to confluence in cell culture media. Once confluent, the cells were made quiescent for 24 hrs and then treated with [10.sup.-9] M aldosterone or [10.sup.-9] M aldosterone plus [10.sup.-6] spironolactone for 24-hrs. Untreated control cells were studied for comparison with the treated cells. After 24-hrs, pHi was measured using a spectroflourometer. To determine pHi, a non-carbonate solution and BCECF, an intracellular fluorescent dye, were used and the light density was found using a spectroflourometer. Nigericin equilibrates pHi and pHe, and a calibration curve with known pH of 6.8, 7.2, and 7.4 was constructed in order to determine pHi. Student's t-test was used to analyze the data. Results: Aldosterone significantly increased pHi (7.8 [+ or -] 0.12) compared to the control (7.5 [+ or -] 0.08) group, and the aldosterone and spironolactone group (7.4 [+ or -] 0.08) had a significantly lower pHi compared to the aldosterone group. There were no significant differences between the control and aldosterone and spironolactone group. Conclusion: Aldosterone directly increases intracellular pH in VSMC and this effect was prevented by spironolactone.
SEXUAL DIMORPHISM IN RENAL NADPH OXIDASE ACTIVITY IN DAHL SS
Rachel Lockhart, Licy L. Yanes, and Jane F. Reckelhoff, University of Mississippi Medical Center, Jackson, MS 39216
Salt sensitivity (SS) of blood pressure is a common occurrence in African American and aging individuals. SS Hypertension (SSHT) has been observed in the Dahl salt-sensitive rat (DS), and male DS develop higher blood pressure on high salt diet (HSD) than do females. Although HT is associated with increased oxidative stress (OxSt), its role in the sex differences in SSHT is unknown. Therefore, the hypothesis was tested that superoxide and NADPH oxidase activity are higher in kidneys from male DS. Methods: DS males and females were placed on HSD (8%) for 4 weeks. Rats were anesthetized, kidneys perfused and dissected into cortex and medulla, and homogenized. OxSt was measured by incubating homogenates with lucigenin (5 [micro]M) for chemiluminescent detection of superoxide. NADPH oxidase activity was measured by adding NADPH (100 [micro]M). Results: After 4 weeks of HSD, basal superoxide was not different in cortex or medulla of male vs. female DS (cortex: 91.91[+ or -]1.42 vs. 94.40[+ or -]6.22 RLU/mg protein; medulla: 110.9[+ or -]25.31 vs. 117.4[+ or -]12.80 RLU/mg protein). NADPH-stimulated activity was also similar in cortex of males and females (792.25[+ or -]13.4 vs. 931.89[+ or -]79.93 RLU/mg protein), but was significantly higher in medulla of females than males (977.82[+ or -]64.92 vs. 1640.53[+ or -]155.12 RLU/mg protein, males vs. females (p<0.05)). Conclusion: These data suggest that OxSt may play a role in SSHT in DS independent of the sex, but that the sex differences in NADPH oxidase activity cannot account for the sexual dimorphism of SSHT in DS rats.
EFFECTS OF SYNTHETIC FIRE ANT VENOM ALKALOIDS ON HUMAN CELLS
LaRue Sutton, Brenda Chapman, Heather White, Donna C. Sullivan, Robin R. Rockhold, Richard Deshazo, H.M.T. Bandara Herath and N.P.D. Nanayakkara, University Medical Center, Jackson, MS, 39216 and University of Mississippi, University, MS, 38677
The antibacterial activity of synthesized solenopsins, components of red imported fire ant venom, have been studied in our laboratory. These compounds were evaluated on the human monocyte line U937 to determine whether they are toxic for human cells at concentrations toxic for microorganisms. Viability of U937 cells following exposure to varying concentrations of solenopsin B and isosolenopsin B was determined by staining with trypan blue. Briefly, 1 X [10.sup.6] U937 cells were plated in triplicate for six different concentrations (3, 10, 13, 20, 23, and 30 [micro]M) of solenopsin tested. U937 cells in media alone and in 3 and 10 [micro]M concentrations of solenopsin B continued to replicate, increasing half a [log.sub.10] over 48 hours. Between 4 and 24 hours, cell counts and viability remained constant at 13[micro]M concentrations and decreased at all other concentrations of solenopsin B. At 48 hours, cell viability was decreased by more than one and a half [log.sub.10] at 30 [micro]M concentrations. Isosolenopsin B was less toxic with half a [log.sub.10] decrease observed at 23 and 30[micro]M after 24 hours when compared to media control. Thus, solenopsin B has a more profound and earlier detrimental effect on U937 cells than isosolenopsin B.
NERVE GROWTH FACTOR REGULATION OF ACIDSENSING ION CHANNEL 3 IN VASCULAR SMOOTH MUSCLE CELLS
Gina Hamilton (1*), Nikki L. Jernigan (2), and Heather A. Drummond (2), (1) Murrah High School, Jackson, MS 39202 and (2) University of Mississippi Medical Center, Jackson, MS 39216
In the peripheral nervous system, the Acid-Sensing Ion Channels (ASICs), members of the Degenerin/Epithelial [Na.sup.+] Channel protein family (DEG/ENaC), are putative mechanosensitive ion channel proteins found in some sensory neurons of the Dorsal Root Ganglia (DRG). Although the physiological role of ASIC proteins remains uncertain, they may play a significant role in pain perception following tissue acidosis since ASIC channels are activated by decreases in pH. Dr. Drummond's lab has recently identified ASIC message and proteins (1, 2, and 3) are expressed in vascular smooth muscle cells (VSMC). In DRG neurons, Nerve Growth Factor (NGF) stimulates the expression of ASIC3. We want to determine if NGF also stimulates ASIC3 expression in VSMCs since the effect of NGF remains unknown. We performed reverse transcriptase-polymerase chain reaction (RT-PCR) to amplify ASIC3 message expression in cultured VSMCs treated with or without NGF overnight. The VSMCs were treated with NGF concentrations at 10 ng/ml and 50 ng/ml to compare any possible increases due to higher application of NGF. PCR products were separated by gel electrophoresis. We could not detect an increase in ASIC3 expression following treatment with NGF. This result suggests NGF may not regulate ASIC3 expression in cultured VSMCs. Future experiments will determine if cultured VSMCs express functional receptors for NGF by a method of Real-Time PCR, which will provide quantitative assessments.
EFFECTS OF CORTISOL ON RHESUS MONKEY KIDNEY CELL VIABILITY IN CULTURE
DeAna Davis (1,2), Erica McClendon (1,2), Clement C. Yedjou (2), Michelle Tucci (3), Hamed A. Benghuzzi (3) and Joseph A. Cameron (2), (1) Hinds Community College, (2) Jackson State University, Jackson, MS 39217, and (3) University of Mississippi Medical Center, Jackson, MS 39216
Glucocorticoids are known to affect a variety of physiological functions e.g. stress response, and metabolic processes, e.g gluconeogenesis, proteolysis and lipolysis, in specific tissues, particularly liver, skeletal muscle and adipose. However, the direct effects of glucocorticoids in kidney cell cultures have not been studied extensively. The purpose of this investigation was to further investigate the action of glucocorticosteroids at the cellular level and provide knowledge regarding the potential use of kidney cells to counteract the effects of kidney failure that often results from alteration or dysfunction of kidney tissues. Specifically the present study examined the effects of cortisol on the viability of Rhesus Monkey Kidney cells (RMKC) cells in culture. Rhesus monkey kidney cells were obtained from Diagnostic Hybrids, Inc. in Athens, OH. Cells were cultured for 24 and 72 hours in the absence and presence of cortisol (.1 ug/dl and 2 ug/dl) in Diagnostic Hybrids, Inc. pre-prepared media at 37[degrees]C in a 5% C[O.sub.2] incubator. At termination, cells were analyzed with the MTT assay for cell viability using a microplate reader at a wavelength of 550 nm. The results showed that cortisol slightly enhanced or maintained cell viability at 24 hours and 72 hours compared to control. These data suggests that the effects of cortisol on cultured kidney cell survival and metabolic functions may vary according to dose and duration of culture. Supported in part by NIGMS R25 GM50117.
THYMOQUINONE PROTECTS A549 CELLS EXPOSED TO LOW OXYGEN CONCENTRATION FROM CELLULAR DAMAGE
Phatia Wells (1,2), Cherece Rigsby (1,2), Latasha Thurman (2,3,4), Corina Lewis (2,3,4), Hamed A. Benghuzzi (3), Michelle Tucci (3), and Joseph A. Cameron (2), (1) Hinds Community College, Raymond, MS 39154, (2) Jackson State University, Jackson, MS 39217, (3) University of Mississippi Medical Center, Jackson, MS 39216, and (4) Bailey Magnet High School, Jackson, MS 39217
Lung represents a unique tissue as it is exposed to various oxygen tensions and oxidants depending upon disease status and environment. Reactive oxygen species that are produced as byproducts of cellular metabolism, are toxic, and at low concentrations they are known to function as messengers in the signal transduction processes. Thymoquinone (TQ) is considered a potent antioxidant that has been used in traditional medicine to help alleviate symptoms and induce healing in respiratory disease processes. The goal of the project was to mimic lung disease process by reducing the cellular oxygen levels to 3% from normal 21% and evaluate the cell for cell viability as well as for cellular membrane integrity by MDA analysis and cellular morphology in the presence of TQ. The result of the investigation clearly show an increase in cellular damage in the low oxygen treated cells by 72 hours, which was reversed with the addition of TQ back to ward control values. It is possible that TQ works to scavenge ROS induced by normal metabolic processes of the cells when held at low oxygen tension. If ROS is produced it can result in significant cellular peroxidation as seen in the low oxygen treated group as evidenced by increasing MDA levels. Low oxygen treatment of cells in the presence of TQ reduced these levels back toward control values. The data clearly supports the use of TQ as adjuvant in the treatment of respiratory diseases. Supported in part by NIGMS R25 GM50117 and REAP, NAS.
HEME OXYGENASE-1 DECREASES ANGIOTENSIN II DEPENDENT REACTIVE OXYGEN SPECIES PRODUCTION IN MOUSE THICK ASCENDING LOOP OF HENLE CELLS
B.J. Patel, T. Vera, Heather A. Drummond, and David Stec, University of Mississippi Medical Center, Jackson, MS 39216
Heme oxygenase-1 (HO-1) induction can attenuate the development of angiotensin II (Ang II)-dependent hypertension. However, the mechanism by which HO-1 can lower blood pressure in this model is not clear. The goal of this study was to test the hypothesis that induction of HO-1 in the kidney can reduce the increase in oxidative stress in the thick ascending loop of Henle (TALH) during Ang II-dependent hypertension. Studies were performed on an immortalized cell line of mouse T ALH cells. Cells were exposed to Ang II (10-12M) for 1 hour and the production of reactive oxygen species (ROS) were measured by the increase in dihydroethidium (DHE) fluorescence using confocal microscopy. HO-1 was induced in cultured mTALH cells by treatment with cobalt protoporphyrin (CoPP), hemin (50 [micro]M) or by transfection with a plasmid containing the human HO-1 isoform. Treatment of mTALH cells with 10-12M Ang II increased DHE fluorescence from 35.5[+ or -]5 to 136[+ or -]18 RFU/[[micro]m.sup.2]. Induction of HO-1 via CoPP, hemin, or overexpression of the human HO-1 isoform significantly reduced Ang II induced DHE fluorescence to 64[+ or -]5, 64[+ or -]8, and 41[+ or -]4 RFU/[[micro]m.sup.2], respectively. In order to determine which metabolites ofHO-1 may be responsible for reducing Ang II mediated increases in ROS production in mTALH cells, cells were preincubated with bilirubin or the carbon monoxide (CO) releasing molecule, CORMA1 (100 [micro]m each) prior to exposure to Ang II. DHE fluorescence averaged 80[+ or -]7 RFU/[[micro]m.sup.2] after incubation with Ang II and was significantly decreased to 55[+ or -]7 and 53[+ or -]4 RFU/[[micro]m.sup.2] after pretreatment with bilirubin and CORMA1. These results demonstrate that induction ofHO-1 in mTALH cells reduces the levels of Ang II mediated ROS production through both the production of bilirubin and CO.
SURVEILLANCE OF GRAM NEGATIVE BACTERIA IN THE ICU: ANTIMICROBIAL SUSCEPTIBILITIES AND PREVALENCE
Mallory Phillips*, Erin C. Wiggers, and Donna C. Sullivan, University of Mississippi Medical Center, Jackson, MS 39216
Infections due to gram negative bacteria are very common in the ICU. Increasing antimicrobial resistance has made such infections very difficult to treat. This report presents results from the first 3 years of a 5 year study to determine the pattern of infection and antimicrobial resistance in the ICU. We collected 100 gram negative bacterial isolates from the ICU. Isolates were identified, speciated, and resistance patterns were determined. Bacteria were grown in Mueller Hinton broth and stored in glycerol at -80[degrees]C. Standardized microtiter minimal inhibitory concentration (MIC) panels for 13 antibiotics were used. Most isolates were cultured from sputum (43% in 2003; 33% in 2004; 51% in 2005), respiratory sites (33% in 2003; 34% in 2004; 22% in 2005) and blood (10% in 2003; 18% in 2004; 9%). In 2003, the most common isolates were Enterobacter cloacae (20%), Pseudomonas aeruginosa (16%) and Klebsiella pneumoniae (14%). In 2004, E. coli accounted for 20%, K. pneumoniae 14%, and P. aeruginosa 12% of the isolates compared to K. pneumoniae (22%), E. coli (18%), and P. aeruginosa (17%) for 2005. In 2003, susceptibility of the most common isolates to ciprofloxacin was 91.4% but dropped to 81% in 2004 and 72% in 2005, following the national trend of 76%. Additional changes have been noted and analysis of prescribing patterns in patients from whom isolates were obtained in ongoing.
ASSESSMENT OF SURVIVAL RESPONSES OF A549 CELLS TO MODULATION OF INTRACELLULAR ENERGETICS
Jeremie Brown (1,2), Ashley Whipps (1), Ashley Blackwell (1), Clement C. Yedjou (1), Ibrahim Farah (1), and Joseph A. Cameron (1), (1) Jackson State University Jackson, MS 39217 and (2) Hinds Community College, Raymond, MS 30154
Studies showed that tumor growth and abnormal cell survival was associated with a number of metabolic abnormalities. Glucose metabolism deranged, lipoprotein-lipase activity depressed and protein metabolism showed abnormality as revealed by changes in plasma amino acid profile and evidenced by increased plasma free tryptophan levels in patients with breast, lung, colon, stomach, and other cancers from various origins. Glucose was established to be the main energy source in cancer cells. However, cancer cells were not able to utilize aerobic respiration due to either defective mitochondria or hypoxia within the tumor microenvironments. The role of energy modulation and the use of glycolytic inhibitors on cancer cell survival are not clearly established in the literature. Therefore, the purpose of this study was to evaluate five glycolytic inhibitors namely, sodium ascorbate, oxalic acid, oxaloacetic acid, sodium citrate and fructose diphosphate (FDP) on A549 cell line exposed to 10, 100, 400 and 800ul of a 1% solution of each for 48h and tested with phase contrast microscanning to determine their influence on cell survival. Oxalic acid and oxalacetic acid both influenced the PH of the medium and resulted in massive cell debris within the exposure period and at 400 and 800 ul levels. Sodium ascorbate, Sodium citrate and FDP did not cause PH changes or massive cell debris even at the highest concentration. However, they caused detectable cell disfigurement and the cells generally remained attached. Future studies will focus on the mechanisms of survival/death in response to modulation of cellular energetics in cancer. Supported by NIGMS R25 GM50117 and RISE NIH
ASSOCIATION OF SERUM TOTAL BILE ACID CONCENTRATIONS WITH INCIDENCE OF FATTY LIVER HEMORRHAGIC SYNDROME AND F-STRAIN MYCOPLASMA GALLICEPTICUM INFECTION IN COMMERCIAL LAYING HENS
Shelia B. Cagle (1), Thomas A. Lenarduzzi (1*), E. David Peebles (1), Robert W. Keirs (1), Scott L. Branton (2), and Patrick D. Gerard (1), (1) Mississippi State University, Mississippi State, MS 39762 and (2) USDA-ARS-Poultry Research Unit, Mississippi State, MS 39762
Fatty Liver Hemorrhagic Syndrome (FLHS) incidence in chickens may be increased by F-strain Mycoplasma gallisepticum (FMG) infection and may also result in decreased egg production. The plasma enzymes aspartate aminotransferase, lactate dehydrogenase, and glutamate dehydrogenase have increased profiles in birds with liver damage and could possibly be used to diagnosis FLHS. In this study, we investigated whether or not total bile acid levels in the serum of egg-laying chickens change with FMG inoculation and incidence of FLHS. Day-old Single Comb White Leghorn pullets of a single genetic strain (Hy-line W-36) were obtained from a commercial source that was monitored certified free of MG and Mycoplasma synoviae (MS). At 22 wk of age, 120 sham-(control) and 120 FMG-inoculated (treated) birds were randomly assigned to individual cages in one of two enclosed and isolated ends of a caged layer facility. At 58 wk, all bled birds were euthanized, and livers from both groups were examined for FLHS. Serum total bile acid concentration was determined in serum from birds in control and FMG-vaccinated groups with and without FLHS. There were no significant main effects or interactions due to FLHS incidence or FMG treatment on serum total bile acid concentration.
COMPARISON OF HEP-2 CELLULAR FUNCTION FOLLOWING EITHER A BOLUS ADMINISTRATION OR CONTINUOUS SUSTAINED RELEASE OF IP-6
Zahra Norozy*, Hamed A. Benghuzzi and Michelle Tucci, University of Mississippi Medical Center, Jackson, MS 39216
Inositol 6-phosphate (IP-6) has demonstrated novel anti-cancer activity using several different tumor models. IP-6, or phytic acid, has antioxidant properties that directly act to inhibit cancer cell growth. In previous experiments cells treated with a single treatment dose of IP-6 (1 mM) showed a decrease in number only at 72 hours. The goal of this experiment was to deliver IP-6 in a sustained continuous manner for periods of 24, 48 and 72 hours to Hep-2 cancer cells and compare the changes in cell growth and morphology with a single bolus dose of IP-6. Our results indicated that IP-6 administered as a bolus dose or in a sustained continuous manner was unable to reduce Hep-2 cell numbers after 24 hours. By 48 and 72 hours an approximate 50% increase in cell numbers were seen in both IP-6 groups. The dose of IP-6 given did not induce changes in cellular protein concentrations nor increase cellular membrane change. Morphologically, the cells appeared small, round to cuboidal with dark eccentric nuclei and scant cytoplasm, and when given IP-6, the cells showed degeneration with irregular cell borders and hyperchromatic nuclei. Overall, IP-6 induced Hep-2 cell proliferation; however, further evaluation of the Hep-2 cytology revealed significant cellular degenerative changes.
DEVELOPMENT OF A FISH MODEL TO STUDY ALCOHOLIC LIVER DISEASE
Bennie Jacobs (1*), Chris Purser (2), Naila M. Mamoon (2), Rodney Baker, (1) Murrah High School, Jackson, MS 39202 and (2) Mississippi Medical Center, Jackson, MS 39216
Alcoholic liver injury is generally divided into four stages, hepatic steatosis (fatty liver) early fibrosis, alcoholic hepatitis and cirrhosis. The prevalence of cirrhosis in chronic alcoholics is relatively low, about 18% of individuals that drink heavily over extended periods of time develop cirrhosis. Since many individuals that drink heavily do not develop alcoholic cirrhosis other genetic or environmental factors must contribute to cirrhosis in addition to the amount and period of alcohol use. Identification of genetic or environmental factors that contribute to alcoholic liver disease has been hampered by the lack of alcoholic disease models that can be both manipulated genetically and controlled environmentally. Rodent models have proven ineffective as rodents do not readily consume alcohol and under most conditions do not develop end stage alcoholic liver disease. Models based on primates are expensive and are not suitable for genetic studies. Small fish such as the zebrafish offer a convenient model. The fish can be treated with ethanol simply by adding it to the aquarium water, and since the fish are inexpensive, large numbers of the fish can be housed and treated. Extensive information is available relating to genetics of zebrafish, and it has been proven efficient as a subject for gene manipulation techniques. Preliminary studies have demonstrated that zebrafish can be treated with 50 mM ethanol for up to 5 months without morbidity. The liver increases in size dramatically, and undergoes significant morphological changes. Fatty acids isolated from livers of ethanol treated fish are more saturated and the chain length is longer as compared to the untreated livers. The ethanol treated fish consumed food slower than controls, but no significant difference in body weight was measured.
MERCURY-INDUCED EXTERNALIZATION OF PHOSPHATIDYLSERINE IN HUMAN LIVER CARCINOMA CELLS
Vantrako Crockett (1*), Dwayne J. Sutton (2), Paul B. Tchounwou (2), and Joseph A. Cameron (2), (1) Hinds Community College, Raymond, MS 39154 and (2) Jackson State University, Jackson, MS 39217
Programmed cell death (apoptosis) is a normal physiologic process, which occurs during embryonic development as well as in the maintenance of tissue homeostasis. It functions as a mechanism to eliminate unwanted or irreparably damaged cells. However, inappropriate induction of apoptosis by environmental agents has broad ranging pathologic implications and has been associated with several diseases including cancer. The toxicity of several heavy metals such as mercury has been attributed to their high affinity to sulfhydryl groups of proteins and enzymes, and their ability to disrupt cell cycle progression and/or apoptosis in various tissues. The aim of this study was to assess the potential for mercury to induce early-stage apoptosis in human liver carcinoma (Hep[G.sub.2]) cells. Annexin-V assay was performed by flow cytometry to determine the extent of phosphatidylserine externalization in mercury-treated Hep[G.sub.2] cells. Cells were exposed to mercury for 10 hours at doses of 0, 1, 2, and 3 [micro]g/mL based on previous cytotoxicity results in our laboratory indicating an [LD.sub.50] of 3.5 [+ or -] 0.6 [micro]g/mL for mercury in Hep[G.sub.2] cells. The study data indicated a dose response relationship between mercury exposure and the degree of early apoptosis in Hep[G.sub.2] cells. The percentages of cells undergoing early apoptosis were 0.03 [+ or -] 0.03%, 5.19 [+ or -] 0.04%, 6.36 [+ or -] 0.04%, and 8.84 [+ or -] 0.02% for 0, 1, 2, and 3 [micro]g/mL of mercury respectively, indicating a gradual increase in apoptotic cells with increasing doses of mercury. Supported in part by NIH 12RR13459, U.S.DA DACA-42-02-C-0057 and NIGMS R25 GM50117.
RESEARCH TO ASSIST SINGING RIVER HOSPITAL'S CANCER CENTER MEDICAL STAFF IN DEVELOPMENT OF A CLINICAL RESEARCH TRIAL INVOLVING RADIATION THERAPY FOR PATIENTS WITH BONE METASTASES
Tammy Goff (1*) and Marguerite Clarkson (2), (1) Mississippi Gulf Coast Community College, Gautier, MS 39533, National Aeronautics and Space Administration, Stennis Space Center, MS 39522 and (2) Singing River Hospital Cancer Research Center, Pascagoula, MS 39581
The Singing River Hospital System (SRHS) Cancer Grants Program developed a clinical research trial testing a radiation therapy treatment dose schedule of 550 cGy for each of three treatments as palliative therapy for cancer patients with bone metastases. The student was involved in researching and writing the background section supporting the proposed treatment dose schedule. In addition, the student assisted the investigators in writing and formatting the study document for final review and approval by the Scientific Review Committee at the SRHS. The final aspect of this project involved working with the regulatory staff in the cancer clinical research office to write the informed consent form for the study, complete the Institutional Review Board (IRB) application and submit the study with application to the SRHS IRB for approval and activation.
DEVELOPMENT OF THE PHARMACEUICAL POLICIES, PROCEDURES, AND TRACKING FORMS FOR STUDY-RELATED AND SUPPLIED DRUGS OR DEVICES FOR THE CANCER CENTER CLINICAL RESEARCH OFFICE STAFF OF SINGING RIVER HOSPITAL
Victoria Olson (1*) and Marguerite Clarkson (2), (1) Mississippi Gulf Coast Community College, Gautier, MS 39533, National Aeronautics and Space Administration, Stennis Space Center, MS 39522, and (2) Singing River Hospital Cancer Research Center, Pascagoula, MS 39581
The Singing River Hospital (SRHS) Cancer Center Office of Clinical Research (OCR) needed to establish policies, procedures and tracking systems for the pharmaceuticals and devices supplied by the study sponsors for clinical trials activated for patients in the cancer center. The internship involved research of the NHI, FDA, and study sponsor regulations relating to study drug procurement, monitoring, dispensing and auditing. Using the regulations, guidelines were given to the OCR staff and SRHS pharmacy staff for revising and updating existing policies for study drugs. The project allowed the pharmacy and OCR staff to set up a monitoring and tracking system (paper as well as computer).
THE EFFECTS OF INCREASING [H.sub.2][O.sub.2] CONCENTRATIONS ON THE VIABILITY AND MORPHOLOGY OF A549 CELLS
Phatia Wells (1,2), Cherece Rigsby (1,2), Latasha Thurman (2,3,4) Corina Lewis (2), (3,4) Hamed A. Benghuzzi (2), Michelle Tucci (2), and Joseph A. Cameron (3), (1) Hinds Community College, Raymond, MS 30154, (2) University of Mississippi, Medical Center, Jackson, MS 39216, (3) Jackson State University, Jackson, MS 39217, and (4) Bailey Magnet High School, Jackson, MS 39217
Before the discovery of antibiotics in the 1920's, physicians were infusing hydrogen peroxide to help fight infections. Hydrogen peroxide is a natural substance manufactured in all our bodies. It is a product of what is called intermediary metabolism in the body and it participates in a significant number of biochemical reactions in the body. The body causes a rapid reduction of hydrogen peroxide to water and by an enzyme, called catalase, which catalyzes (breaks down) the reaction of hydrogen peroxide plus water to oxygen plus two water molecules. However, the practice remains controversial. The aim of this increase in oxidative reactions in the body is to help regulate tissue repair, cellular respiration, immune functions, the energy system, most hormone systems and the production of cytokines. Therefore, the objective of this study was to determine the effects of low (44.1 [micro]M), medium (88.2 [micro]M) and high (220 [micro]M) concentrations of hydrogen peroxide on cellular viability and morphology after periods of 24, 48 and 72 hours. The results of the experiment show that low and high levels of hydrogen peroxide affected initial cellular numbers. By 48 hours the low-level treatment had numbers similar to control, and by 72 hours the low-level treatment resulted in higher number of cells. This suggest that low level treatment may actually stimulate cellular repair mechanisms. Cellular damage was evident in the medium and high treatment doses by 48 and 72 hours. Cellular morphology showed the greatest changes in the high and medium dose treatments by 48 and more substantial degradation of the cell by 72 hours. Low dose treatment by 72 hours showed increased number of cells, which appeared similar to control in cellular detail. The data suggests that low level hydrogen peroxide treatment does not cause significant cellular damage, and may also help stimulate cellular repair mechanisms as indicated by increased cell numbers by 72 hours. Supported in part by NIGMS R25 GM50117 and REAP.
METHODS USED FOR DETECTING BACTERIAL CONTAMINATION IN PLATELET PRODUCTS BY BLOOD SUPPLIERS TO SOUTH MISSISSIPPI AND LOUISIANA
Lori Mosley, University of Southern Mississippi, Hattiesburg, MS 39406
Bacterial Contamination of platelet products has become the number one risk associated with transfusion-transmitted infections in the United States. This heightened risk is due to the storage requirements of platelet products. These products must be stored at room temperature, which provides an excellent environment for bacterial growth. To date, there has been no common methodology determined in which to test platelet products for bacterial contamination. However, in March 2004, the American Association of Blood Banks (AABB) implemented a policy which stated that all transfusion services must have a method to determine bacterial contamination in their platelet products. Several of the large blood suppliers in South Mississippi and Louisiana were contacted to see which methods are utilized in these facilities. Results showed the methods were based on the type of platelet products the facility produces, either random donor platelets or single donor platelets. In each case, the methodology chosen proved to be efficient in detecting bacterial contamination, and in most cases, positive units were found to be from improper donor arm disinfection and handling of the platelet products during preparation.
Session II: Microbiology, Chair: Tina Martin
11:15 ANTIBIOTIC PROPERTIES OF SPICE EXTRACTS
Sarah Ali* and Sabine Heinhorst, Oak Grove High School, Hattiesburg, MS 39402 and University of Southern Mississippi, Hattiesburg, MS 39406
Because of the existing controversy about natural antibiotic properties of spices, this project attempted to determine how compounds extracted from ground cloves, ginger, garlic, pepper, cinnamon, and from chili and curry powder affect the growth of Staphylococcus aureus and Escherichia coli as representatives of Gram-positive and Gram-negative bacteria, respectively. Organic and aqueous extracts, as well as dried and fresh spices were compared. Several spice extracts stimulated bacterial growth. Of all spices tested, only garlic had a growth inhibitory effect on both bacteria. The active compound(s) in garlic appeared to be sensitive to heat.
11:30 ANALYSIS OF HUMAN COMPLEMENT FACTOR H BINDING TO STREPTOCOCCUS PNEUMONIAE CLINICAL ISOLATES
Chinwendu Onwubiko*, Lisa R. Quin, Stephanie Carmicle, and Larry S. McDaniel, University of Mississippi Medical Center, Jackson, MS 39216
Streptococcus pneumoniae is responsible for most cases of community acquired pneumonia. Pneumococcal surface protein C (PspC) plays a role in allowing the pneumococcus to evade innate host immune responses. PspC binds factor H (FH), a protein involved in the regulation of complement activation. This binding inhibits the alternative pathway of complement thus increasing the survival of the pneumococcus within the host. We previously demonstrated variation in binding of FH to PspC by different pneumococcal isolates. This variability in binding occurred across different PspC groups and was independent of capsular serotype. The goal of this study was to determine if the variability in FH binding was based on differences in the anatomical source of the pneumococcal isolate. Using flow cytometry, we examined 89 clinical isolates, divided into 3 groups based on source type (respiratory, systemic, or carriage). The mean fluorescence intensity of respiratory strains was 66.3, of systemic strains was 100.2, and of carriage strains was 165.2. The difference in binding by respiratory versus carriage strains was statistically significant (p<0.05). These results suggest that strains isolated from asymptomatic carriage individuals bind more FH than those pneumococci causing respiratory disease.
11:45 ANALYSIS OF STREPTOCOCCUS PNEUMONIAE FROM MIDDLE EAR EFFUSIONS OF CHILDREN IN MISSISSIPPI USING PSPA FAMILY TYPING AND BOX PCR BASED DNA FINGERPRINTING
Courtney Shires*, Lisa R. Quin, and Larry S. McDaniel, University of Mississippi Medical Center, Jackson, MS 39216
Because Streptococcus pneumoniae is the leading bacterial cause of otitis media which is frequently seen in young children, a vaccine that will be both affordable and protective is in great demand. Pneumococcal surface protein A (PspA) has potential to be included in a vaccine that would be broadly protective. While the gene encoding PspA is variable among isolates, 95% of previously examined clinical isolates fall into two family groupings. We determined the PspA family type of pneumococcal isolates from middle ear effusions of children in Mississippi. Isolates from middle ear effusions of 31 patients (30 were under the age of 5 years) were analyzed. Seventy one percent of the isolates belonged to PspA family type 1, 13% belonged to family 2, and 13% belonged to family 1 and 2. We also performed BOX PCR characterization of the isolates using the BOX A1R primer. The isolates were placed into 24 different groups based on banding patterns, allowing a 2% difference in the fragment base pair length. Some fragments were more common than others with one fragment present in 77.4% of strains, one in 32.3%, and another in 29%. We conclude that a vaccine targeting PspA families 1 and 2 would cover 97% of the pneumococci in the middle ear effusions of children in our study.
Exhibit Hall B
Symposium 1: Nursing, Chair: Michelle Tucci
1:00 HISTORY OF NURSING: PAST, PRESENT, AND FUTURE
Mary Tan, University of Mississippi Medical Center, Jackson, MS, 39216
The purpose of this lecture is to discuss nursing past, present, and future. Nursing has undergone dramatic changes in response to societal needs and influences. From the beginning of time, women have cared for infants and children; thus, nursing could be said to have its roots in "the home." Religion also had a significant role in the development of nursing. During the third and fourth centuries, wealthy matrons of the Roman Empire converted to Christianity and used their homes to provide for the sick. Throughout history, inadequacy of care during the Crimean, American Civil and World Wars have accentuated the need for nurses. Nursing's image before the 1800's is reflected in Charles Dicken's (1896) writings of Sairy Gamp as "neglecting, stealing, and physically abusing the sick." Florence Nightingale's Angel of Mercy established respectability for nursing in the latter part of the 19th century. To understand nursing as it is practiced today and how it will be practiced tomorrow requires understanding social forces affecting the entire health system. Nurses are challenged to care for more acute illnesses, decreased length of stays and less contact time with clients than ever because of shifts from inpatient to outpatient care. With the establishment of health care reform, computerized charting, and telenursing we are confronted with the new nursing era. Registered nurses make up the largest group of health care providers. Yet for the future the supply of nurses is inadequate to meet the demand.
1:15 EDUCATION PREPARATION IN NURSING
Tina Mitchell Martin, University of Mississippi Medical Center, Jackson, MS 39216
Nursing is the nation's largest health care field with 2.7 million registered nurses nationwide. There are various educational routes for becoming a nurse. The purpose of this lecture is to discuss multiple entry points to nursing practice. Educational programs prepare nursing assistants and licensed practical nurses (LPNs) for roles in the health care delivery system. Nursing assistants are certified after completing a specific course of study. LPNs practice under the supervision of a registered nurse after completing a program of study (usually 12 months) and successfully passing a licensing examination. There are three major educational paths that prepare graduates to take the licensing examination for registered nursing: diploma, associate degree (AD), and baccalaureate (BSN). These basic programs of nursing vary in specific courses offered, program duration and cost. A brief description of each approach will be discussed as well as the various career opportunities open to nurses as they acquire experience and continue their education. Master's and doctoral degrees offer potential leadership positions in the profession. Nurses with advanced degrees may be researchers, nurse practitioners, nurse anesthetists, clinical specialists, educators or administrators. Trends in nursing education, including emergence of the practice doctorate, will be discussed. In addition, the various nontraditional approaches to nursing education that have evolved will be mentioned.
1:30 NURSING: "SO MANY OPTIONS, WHICH ONE TO CHOOSE?"
Lisa A. Haynie, University of Mississippi Medical Center, Jackson, MS 39216
The profession of nursing is the most diverse of all healthcare professions. It is a recognizable role in some form in every culture. Nurses have never been more vital to the health care industry than they are today. They must be well-educated, adaptable, and ready for change at all times. Fortunately, there is a strong demand for nurses and the career prospects are plentiful. In the United States there are a multitude of specialty areas within nursing. These opportunities encompass care throughout the lifespan and are based on patient needs. There are over 200 nursing specialties and sub-specialties; examples include: surgery, emergency, pediatric, psychiatric, school, public health, OB-GYN, neonatal, nurse midwifery, and others. Nurses who choose a specialty for which they focus can become certified in that area, signifying that they possess expert knowledge of the specialty. Additionally, certified nurses often earn a salary differential over their non-certified co-workers, and research has shown that these nurses have higher rates of patient satisfaction, as well as lower rates of work-related errors in patient care.
1:45 RESPONSIBLILITIES OF AN ADVANCED CLINICAL PRACTICE NURSE
Anne Afeman Norwood, University of Mississippi Medical Center, Jackson, MS 39216
Advance practice nurses have been an integral part of the health care system for years, and today, there are more and more nurses who are returning to school to complete a master of science in nursing to attain one of these highly sought opportunities within this diverse profession. Several advance practice nursing opportunities include: nurse educator, nurse anesthetist, and nurse practitioner. These three areas of nursing have grown to be more and more popular, primarily due to floor nurses becoming overworked and fatigued on the job due to the increased acuity level of patients in our hospitals. Furthermore, men and women are attracted to these types of positions because it allows more flexibility, more control, and of course, more financial security. Nurse practitioners are practicing in a variety of settings and perform similar duties as the physician. These areas include the emergency departments, after-hour clinics, and industries, as well as school clinics and primary care settings. The experiences nurse practitioners come upon in these areas are anywhere from routine physical assessments and decision making processes to unbelievable, incredible, exciting, and challenging incidents that create life long impressions. Whatever the experience, nurse practitioners are truly the epitome of holistic care!
2:00 Symposium I--Panel Discussion
3:00 Poster Session II, Chair: Ibrahim Farah & Olga McDaniel
INHIBITORY EFFECT OF CORTISOL IN HUMAN LUKEMIA CELLS
DeAna Davis (1,2), Erica McClendon (1,2), Clement C. Yedjou (2), Michelle Tucci (3), Hamed A. Benghuzzi (3) and Joseph A. Cameron (2), (1) Hinds Community College, Raymond, MS 39154, (2) Jackson State University, Jackson, MS 39217, and (3) University of Mississippi Medical Center, Jackson, MS 39216
Cortisol is an adrenal corticosteroid hormone that is released in the body during stressed or agitated states. It increases blood pressure and stimulates protein catabolism to release amino acids for use in repair, enzyme synthesis, and energy production. However, the mechanisms of action underlying its effects, particularly at low doses, on some mammalian cell lines including Jurkat cells are still largely unknown. To address this issue, the MTT assay was performed to assess the Jurkat cell viability upon exposure to cortisol. Cells were cultured for 24 and 48 hours in the absence and presence of cortisol (0.1 YMBOL32\f"Symbol"\s12 [micro]g/dL and 2 [micro]g/dL) in media at 37[degrees]C in a 5% C[O.sub.2] incubator. At termination, cells were analyzed with the MTT assay for cell viability using a microtiterplate reader at a wavelength of 550 nm. Study results indicated that cortisol significantly (p< 0.05) reduced the viability of Jurkat cells. Upon 24 hours of exposure, the percentages of cell viability were recorded to be 100%, 22%, and 37%, for 0, 0.1, and 2 ug/dL of cortisol, respectively. In addition, the percentages of cell viability computed for 48 hours of exposure were 100%, 28%, and 65% for 0, 0.1, and 2 ug/dL of cortisol, respectively, suggesting a time and dose dependent response. Interestingly, the data showed significant reduction of cell viability in cortisol-treated Jurkat cells as compared to the control upon 24 hrs, but demonstrated a slight increase in cortisol-treated Jurkat cells upon 48 hrs of exposure as compared to 24 hours. These results suggest an inhibitory effect of low doses of cortisol on Jurkat cell viability that declines in a time dependent manner. Supported in part, by the National Institutes of Health Grant No. 1G12RR13459through the Center for Environmental Health and in part by NIGMS R25 GM50117.
THE EFFECTS OF FRUCTOSE-1,6-BISPHOSPHATE ON LUNG CELLS AT REDUCED OXYGEN LEVELS
Derrick Huang (1*), Ameze Adah (1), Michelle Tucci (2), and Hamed A. Benghuzzi (2), (1) University of Mississippi, Oxford, MS 38762 and (2) University of Mississippi Medical Center, Jackson, MS 39216
Background and significance: MRC-5 cells are lung fibroblast cells. Two well-known diseases involving the lung are cystic fibrosis and asthma. Both conditions can result in altered oxygen delivery to the lung tissue. Leaving either of these conditions untreated for an extended period of time could result in the loss of lung function. Fructose-1,6-bisphosphate (FDP) is a glycolytic intermediate that has been used to protect tissues in various hypoxic and ischemic conditions. Under ischemic conditions where ATP levels are much lower than normal, bypassing the initial steps of glycolysis offers protection by allowing the cell to produce ATP without expending energy. Hypothesis: Exogenously added fructose-1,6-bisphosphate to MRC-5 fibroblast cells under ischemic conditions will allow the cells to grow similarly to untreated cells maintained at ambient air. Objectives: (1) To provide increasing concentrations of fructose-1,6-bisphosphate for 24, 48, and 72 hours under ischemic and ambient conditions and compare growth characteristics of cells under the similar conditions, and (2) To evaluate the cell viability and damage. Results: Under ambient conditions, FDP increased cell umber in a dose dependent fashion at 24 hours. Cell number was similar at 48 and 72 hours. Cellular glutathione levels were decreased in all treatment groups as early as 24 hours, and MDA was increased in the medium and high dose treatment group for the duration of the study. Under ischemic conditions, FDP reduced cell numbers by 50% in the medium and high dose group at 24 hours. Cell numbers were not different at 48 and 72 hours of treatment. Cellular glutathione levels were not significantly different from control. Cellular MDA levels were increased in the medium and high dose levels. Conclusions: Overall FDP was able to protect cellular glutathione levels under ischemic conditions. Increasing concentrations of FDP regardless of oxygen concentration resulted in increased evidence of cellular damage.
ASSESSING THE VARYING CAUSES AND RISK FACTORS ASSOCIATED WITH PEDIATRIC HYPERTENSION
Joshua Champion* and Jimmy Stewart, Murrah High School, Jackson, MS 39202 and University of Mississippi Medical Center, Jackson, MS 39216
Background. Pediatric hypertension is making a significant increase among the youth of Mississippi. Rising numbers of adolescents are being diagnosed with hypertension at younger ages each year. In many areas of Mississippi these young people are being exposed to unhealthy lifestyles and are not concerning themselves with the importance of cardiovascular health. The major objective of this study is finding the associations of a child's environment, demographics, education, religion, daily diet, physical activity, and heredity on his or her heart health. The results of this survey were expected to reveal the associations of unhealthy life practices and hypertension at early age and hopefully influence individual and community efforts in reducing childhood hypertension. Methods. One hundred children were selected to take part in this study. Fifty hypertensive and fifty normotensive patients ages 10-18 were given questionnaires. All of them were normal routine patients of the University Medical Center Hypertension Clinic and North Clinic, in Jackson, MS. Each of these adolescents was asked to complete a short telephone survey of 44 questions. The information used in the survey was compiled from sources such as the Behavioral Risk Factor Surveillance Survey (BRFSS), and The Jackson Heart Study questionnaires. Results. The study did not give a full proof result in all areas. The survey showed that the habits of young people are improved after they have the disease. The youth represented in this survey lead very healthy lives now that they have confirmed hypertension. Conclusions. In the areas of religious practice and education we are still in need of more research. These areas will be studied more closely this year in an effort to conclude more insightfully, their affects on hypertension.
THE EFFECTS OF PMMA PARTICLE NUMBER ON MG-63 OSTEOBLAST CELL FUNCTION
Heather DeLaSalle*, Hamed A. Benghuzzi, Robert Deville, and Michelle Tucci, University of Mississippi Medical Center, Jackson, MS 39216
Implants used for joint replacement are often cemented into place to increase stability. As the person ambulates the implanted materials slide against each other producing small wear debris particles. There is increasing evidence that wear debris particles that are present in periprosthetic tissues have direct effects on osteoblasts. Particles resulting from polymethylmethacrylate (PMMA) cements used for fixation may also be involved directly in aseptic loosening of implants. However, it is not known if these particles have a direct or indirect affect bone formation. The objective of this study was to determine the effect of PMMA particle number on osteoblast cells. MG63 osteoblast-like cells were challenged with 200 [micro]g/mL, 20 [micro]g/mL and 2 [micro]g/mL PMMA diluted in culture medium. Cells were incubated with the particles for 24, 48 and 72 hours. MG-63 cellular proliferation was unaffected at 24 and 48 hours regardless of the concentration. However, at 72 hours the low and high dose treatments resulted in a 50% reduction in cell numbers. Interestingly, at 72 hours cellular protein levels were increased in all treated groups by at least 50%. Cellular damage was evident as early as 24 hours in all treatment groups and continued for the duration of the study. Morphological observations showed significant vacuole formation in all treatments. Also, there was an overall increase in cell size that coincided with increasing levels of PMMA with noted macronucleoli at the highest dosage. The increase in macronucleoli may be reflective of increased cellular protein concentration. Overall, a PMMA particle challenge resulted in significant membrane perturbations, and the overall effects on cell number are PMMA dose and time dependent.
EBSTEIN-BARR VIRUS: EPSTEIN-BARR VIRUS: CLONING, EXPRESSION, AND PURIFICATION OF GP350
Jonathan Batty (1*), Joyce Fingeroth (2), and Shawn Clark (2), (1) Tougaloo College, Tougaloo, MS 39174 and (2) Harvard Institute of Medicine, Boston, MA 02215
The general objective of this research is to prevent the Epstein-Barr virus from attaching to human cells. By studying protein structure through crystallization it is hoped a drug can be produced to prevent cellular attachment of EBV and thereby prevent or reduce infection. Hence, the cloning, expression, and purification of gp350 were performed in this particular research. A truncated form of glycoprotein 350 was cloned and expressed in Cho-Lec and 293T cell lines. The cellular supernatants of the cell cultures were purified using Concanavalin A matrix. A Sodium Dodecyl Sulphate-Polyacrylamide gel electrophoresis analysis was executed on the purified eluate to separate proteins and to verify the presence of the purified glycoprotein in the supernatant. The glycoprotein was cloned and expressed as a fusion protein with RFP in Cho-Lec and 293T cells. During the purification process, the protein was retained on the columns used for purification. One part of the SDS-Page analysis was the western blot technique, which made it possible to develop film that demonstrated the molecular weight of the purified protein alone. In addition, the Coomassie Blue staining added tint to each band on the polyacrylamide gel to make each band visible for molecular weight determination by another method that showed whether contaminating proteins were present or not. To date, single cell cloning has been performed and there is high expression of glycoprotein in cells. Partial purification of the protein has been achieved.
NEURON SPECIFIC ENOLASE (NSE) AND FERRITIN COMPARED WITH ELEVEN OTHER TUMOR ANTIGENS FOR THE SERODIAGNOSIS OF PANCREATIC AND GASTRIC CANCER
Rasheeda Crowell (1*), Tammy Sims-Davis (1), Sharae Johnson (1), Mary Guo (1), Wileen Cooksey (1), Slobodanka D. Manceva (1), Sabrina Bryant (1), Margaret Jackson (1), James T. Johnson (1), Harold Schultze (1), Shawn Clinton (1), Kevin Beason (1), Cynthia Wilson (2), Debbie Fortenberry (1), Cynthia Bright (1), Helen Hua (1), Jiarong Ying (1), Paul Sykes (1), Kay Hollifield (3), Charlton Vincent (3), and Margot Hall (1), (1) University of Southern Mississippi, Hattiesburg, MS 39406, (2) University Medical Center, Jackson, MS 39216, and (3) Laurel Clinic for Women, Laurel, MS 39442
With 32,180 (pancreatic) and 21,860 (gastric) new cases and 31,800 (pancreatic) and 11,550 (gastric) deaths estimated during 2005, pancreatic and gastric cancers are important pathologies in the USA. Tumor antigens have been used in combination with other methods for diagnosis. The objective of this study was the comparison of NSE and ferritin with eleven other tumor antigens for diagnostic efficacy in pancreatic and gastric cancer. Sera from 554 patients (16 pancreatic cancer, 12 gastric cancer, 331 other cancers, and 195 non-cancer) were assayed for the presence of tumor antigens and the results correlated with diagnoses established pathologically. Immunoassay test kits from Diagnostic Automation (NSE, Ferritin, CA242), Hybritech (CEA, CA195), Centocor/Fugirebio Diagnostics (CA125, CA19-9, CA72-4, CA15-3, CA27.29, Cyfra21-1), CIS Biointernational (CA50), and Abbott (AFP) were used to test for the concentration of these antigens. Using the manufacturers' decision values the following diagnostic sensitivities were obtained: Pancreatic cancer: NSE 0.0%, Ferritin 50.0%, CEA 37.5%, CA19-9 66.7%, CA195 100.0%, CA50 66.7%, CA242 66.7%, CA72-4 31.3%, CA125 40.0%, CA 15-3 26.7%, CA27.29 40.0%, AFP 18.2%, Cyfra21-1 26.7%; Gastric CA NSE 0.0%, Ferritin 11.1%, CEA 50.0%, CA19-9 63.6%, CA195 58.3%, CA50 70.0%, CA242 70.0%, CA72-4 27.3%, CA125 40.0%, CA 15-3 45.5%, CA27.29 30.0%, AFP 22.2%, Cyfra21-1 9.0%. Diagnostic specificities were >75%. From these data we conclude that ferritin was a possible marker for pancreatic cancer but not for gastric cancer and that NSE was not a useful marker.
DEVELOPMENT OF A DNA IDENTIFICATION PROGRAM FOR ELEMENTARY STUDENTS BASED ON PARTICIPATION IN THE BASE PAIR SUMMER RESEARCH INSTITUTE
Erin C. Wiggers*, Susan Bender, Cindy Cook, Robin R. Rockhold, and Donna C. Sullivan, University of Mississippi Medical Center, Jackson, MS 39216
Senior Projects have been instituted in many Mississippi Public Schools as teaching and training tools. Senior high school students identify mentors who will help them select topics for research that will result in a product that can provide a community service. Students prepare a research paper, develop personal portfolios, and present PowerPoint-based talks to describe their projects to community, business and school leaders. The Howard Hughes Medical Institute-supported Base Pair Summer Research Institute at the University of Mississippi Medical Center provides training for middle and high school science teachers as well as hands-on activities for high school students. The program is responsible for the development of the Crime Scene Investigation teaching module adopted by many science teachers throughout the state. Following participation in this program, an extension of such a teaching module was developed for use in an elementary school setting. A lesson plan for elementary students concerning the use of DNA as a method of identification was developed. This program has been incorporated into the agenda of the Northshore Elementary School as partial fulfillment of the Senior Project. The lesson plan includes both a classroom presentation and the use of DNA collection kits for each elementary student in the class. (Supported by the Howard Hughes Medical Institute)
SYNTHESIS AND CHARACTERIZATION OF COMPLEXES OF CU(II), ZN(II), NI(II) AND SN(IV) WITH ACETONE N(4)-PHENYLTHIOSEMICARBAZONE (HAPTSC)
Derrick D. McNutt (1,2), Mantrako F. Crockett (1,2), Ramaiyer Venkatraman (1) and Joseph A. Cameron (1), (1) Jackson State University, Jackson, MS 39217 and (2) Hinds Junior College, Raymond, MS 39154
Thiosemicarbazones exhibit a broad spectrum of pharmacological properties, including antibacterial, antiviral, antifungal, antimalarial, and antineoplastic activities. In solution, thiosemicarbazone molecules can exist in thione-thiol tautomeric form. The unique property of thiosemicarbazones is related not only to the presence of many electron donors centers in the structure but also to the bonding characteristics. As a ligand, thiosemicarbazones are well known to behave as chelating agent towards a wide range of metallic ions forming structurally different complexes. In many instances, thiosemicarbazones act as a bidentate ligand and bind to the metals through the sulfur and hydrazinic nitrogen atom. Further, metal complexes often display enhanced activities when compared with the parent compound. Thiosemicarbazones can easily be modified by variation of parent aldehyde or ketone used for their synthesis. For this reason, there has been a steady growth in the synthesis, structure and reactivity studies of heterocyclic thiosemicarbazones and their metal complexes. However, relatively little is known about the complexing behavior of aliphatic thiosemicarbazones with metal ions. In this research, we have synthesized and characterized copper, zinc, nickel and tin metal complexes of acetone phenylthiosemicarbazone using it as a ligand. The synthesized compounds were characterized using thermal, spectral and molar conductivity methods. The molar conductivity studies indicate that the complexes exist in the nonionic form. Solubility studies indicate the non-polar nature of the complexes. Further, structural characterizations were carried out using infrared, UV-Visible and proton NMR techniques. The results indicate the formation of five coordinate chloro-complex of the formula, [M[(aptsc).sub.2]x]x (where M = Cu, Ni, Zn, Sn and x = [Cl.sup.-]] with the participation of thione sulfur and hydrazinic nitrogen atoms from the ligand molecule. The metal complexes exhibit trigonal bipyramid geometry, which is typical for nonaromatic thiosemicarbazones. Supported in part by NIGMS R25 GM50117.
THE EFFECT OF GREEN TEA EXTRACT ADMINISTERED BOTH PREVENTATIVELY AND UPON INFECTION IN TRYPANOSOMA LEWISI INFECTED RATS
Alan Jarrett and Laura Herrington*, Belhaven College, Jackson, MS 39202
Trypanosoma lewisi, a blood parasite found in rats, has many similarities to the trypanosome species causing human trypanosomiasis. For this reason, experimental data from studies involving T. lewisi could be valuable in the search for new treatments for human trypanosomiasis. Green tea has been used in many recent cancer studies because it contains natural antioxidants which are known to target rapidly dividing cells. Because blood parasites establish and maintain parasitemia in the host by rapid division, green tea extract was administered to rats: 1) fifteen days prior to infection (preventative group) and 2) upon establishment of infection (treatment group) to determine its effects on parasitemia. Both the treatment and preventative groups were divided into subgroups receiving different dosages of tea extract. The treatment group experienced a significant decrease in parasitemia, the highest dosage showing the greatest difference compared with the control, followed by the moderate and low dosages respectively. The preventative group showed no significant decrease in parasitemia.
EFFECTS OF POLYSTYRENE PARTICLE NUMBER ON TYPE II PNEUMOCYTES
Constance R. Gassion*, Sommer Welsh-Guedon, Michelle Tucci, and Hamed A. Benghuzzi, University of Mississippi Medical Center, Jackson, MS 39216
Studies into the effects of ultrafine particles in the lung have shown adverse effects and are considered to be due in part to the particle size. Inhalation of small particles is associated with exacerbation of respiratory diseases in epidemiological studies. Ultrafine particles have been hypothesized to play an important role, but it is unclear as to whether a dose response type of relationship exists. The aim of the present study was to investigate the role of ultrafine particle number on lung cells and to describe the affects due to phagocytosis of particles by the cells. A549 cells are a transformed cell line similar to type II lung pneumocytes. These 5 X [10.sup.4] cells were treated with 1000, 5000, and 10,000 polystyrene particles and incubated at 37[degrees]C for periods of 24, 48, and 72 hours. Cell number, protein, and MDA levels were determined on the treated cells and compared with untreated controls. The lung cells were also observed microscopically to assess cell damage. Lung cells treated for 24 hours with 1000 and 5000 particles showed the greatest increase in cell number. Cellular protein levels were similar for all groups (p> 0.05) for the duration of the study. MDA levels for treated groups at 24 and 72 hours were not statistically different from the control groups. Cells treated at 48 hours with 5000 and 10,000 particles showed slight increases in the MDA levels above low particle number and control treated groups. Morphological evaluation of the cells revealed increased inclusions with increasing dose. The results from this study indicate the ability of A549 cells to respond to a challenge with ultrafine particles. The concentrations tested caused an initial stimulation cell proliferation at 24 hours followed by increased damage at 48 hours. Future studies will focus on the inflammatory products formed by ingestion of the ultrafine particles.
THE EFFECT OF CURCUMIN & BET A-GLUCAN INDIVIDUALLY & INCOMBINATION ON TRYPANOSOMA LEWISI INFECTED RATS
Gabrielle Pickle*, Brian Kirby*, and John Shapiro*, Belhaven College, Jackson MS, 39202
Trypanosomiasis is the infection of vertebrates by hemoflagellated parasites known astrypanosomes. Trypanosoma brucei, the causative agent for African sleeping sickness, isone of the leading causes of death in Africa, andcurrent treatments are either costly ortoxic to the host. Two herbal supplements, beta-glucan and curcumin, were used in thisexperiment because of their availability and low cost. Recent literature has shown the effectiveness of beta-glucan, a derivative of yeast cells, in immune system enhancement. Curcumin, a substance found in curry, has shown success in inhibiting blood parasites. Inthis experiment, a rat-specific trypanosome, Trypanosoma lewisi, was used to inoculatelaboratory rats. Beta glucan and curcumin injections were administered to test groups, both separately and together, to determine inhibitory effects on the infection and possiblesynergistic effects of the drugs. The results were extremely variable. Unfiltered beta-glucan resulted in lesions in several rats, however these rats showed decreasedtrypanosome counts. Neither beta-glucan nor curcumin appeared to have any effect on the parasitemia. However, rats that developed an infection following the first inoculationshowed decreased trypanosome counts after the second inoculation. Further studies mightprovide evidence on correlations between decreased trypanosome counts and beta-glucanor curcumin.
METABOLIC EFFECTS OF FRUCTOSE 1,6-BIDSPHOSPHATE IN NORMOXIC AND HYPOXIC STATES OF MG63 OSTEOSARCOMA CELLS
Ameze Adah (1), Hamed A. Benghuzzi (2), Michelle Tucci (2), (1) University of Mississippi, University, MS 38677 and (2) University of Mississippi Medical Center, Jackson MS 39216
Glycolysis is a very important process that contains very intricate steps that play a role in cellular performance and viability. Fructose 1,6-bisphosphate (FBP) is a glycolytic intermediate that has proven to be replete in literature in improving cellular conditions under hypoxic and ischemic conditions. Osteoblasts are key regulators of skeletal matrix synthesis and degradation. Thus, considering FBP's positive effects on amelioratig hypoxia-induced injuries, the objective of this study was to determine its effects and comparative effects on osteoblast cells under normoxic and hypoxic states. MG63 osteoblast-like cells were cultured in 24-well culture plates and treated with high, medium and low dosages of FBP at 24, 48, and 72 hours. At the end of each time period, cellular number, damage (MDA), and glutathione levels were evaluated. There was a significant increase in cell number for the low level of FBP in normoxia at 48 hours (p <0.05). For the cells in hypoxia, there was a significant decrease in cell number for the medium level at 48 hours (p <0.05). At 48 hours there was a significant decrease in cell damage for the cells in normoxia and hypoxia when compared to the control. Cellular damage was not evident in the supernatant in either oxygen condition for the duration of the study. A significant decrease in glutathione levels was also noted for the cells in hypoxia. Cellular morphology included multiple nucleoli, vacuolated cytoplasm, abnormal cells, and web-like cytoplasm. The results indicate that FBP does protect bone cells exposed to hypoxic injuries, and while doing so, ameliorating the states of the cells in shock.
NEURON SPECIFIC ENOLASE (NSE) AND FERRITIN COMPARED WITH TWELVE OTHER TUMOR ANTIGENS FOR THE SERODIAGNOSIS OF BREAST CANCER
Sharae Johnson (1*), Rasheeda Crowell (1), Tammy Sims-Davis (1), Mary Guo (1), Wileen Cooksey (1), Slobodanka D. Manceva (1), Sabrina Bryant (1), Margaret Jackson (1), James T. Johnson (1), Harold Schultze (1), Shawn Clinton (1), Kevin Beason (1), Cynthia Wilson (2), Debbie Fortenberry (1), Cynthia Bright (1), Helen Hua (1), Jiarong Ying (1), Paul Sykes (1), Rafat AlKurd (1), Kay Hollifield (3), Charlton Vincent (3), and Margot Hall (1), (1) University of Southern Mississippi, Hattiesburg, MS 39406, (2) University Medical Center, Jackson, MS 39216, and (3) Laurel Clinic for Women, Laurel, MS 39442
With 212,930 new cases and 40,870 deaths estimated during 2005, breast cancer is the major cancer in females for the USA. Tumor antigens have been used in combination with other methods for diagnosis. The objective of this study was the comparison of NSE and ferritin with twelve other tumor antigens for diagnostic efficacy in breast cancer. Sera from 554 patients (87 breast cancer, 272 other cancers, and 195 non-cancer) were assayed for the presence of tumor antigens and the results correlated with diagnoses established pathologically. Immunoassay test kits from Diagnostic Automation (NSE, Ferritin, CA242), Hybritech (CEA, CA195, CA549), Centocor/Fugirebio Diagnostics (CA125, CA19-9, CA72-4, CA15-3, CA27.29, Cyfra21-1), CIS Biointernational (CA50), and Abbott (AFP) were used to test for the concentration of these antigens. Using the manufacturers' decision values the following diagnostic sensitivities were obtained: NSE 0.0%, Ferritin 40.0%, CA 15-3 63.4%, CA27.29 39.3%, CA549 40.3%, CEA 22.4%, CA195 31.8%, CA19-9 12.2%, CA50 22.2%, CA72-4 12.9%, CA125 12.1%, Cyfra21-1 12.2%, AFP 21.8%, CA242 29.3%. Diagnostic specificities were >75%. From these data we conclude that ferritin was inferior to CA15-3, equal to CA27.29 and CA549 (traditional breast cancer markers) and superior to all other markers for the diagnosis of breast cancer.
NEURON SPECIFIC ENOLASE (NSE) AND FERRITIN COMPARED WITH ELEVEN OTHER TUMOR ANTIGENS FOR THE SERODIAGNOSIS OF LEUKEMIA AND LYMPHOMA
Tammy Sims-Davis (1*), Sharae Johnson (1), Rasheeda Crowell (1), Mary Guo (1), Wileen Cooksey (1), Slobodanka D. Manceva (1), Sabrina Bryant (1), Margaret Jackson (1), James T. Johnson (1), Harold Schultze (1), Shawn Clinton (1), Kevin Beason (1), Cynthia Wilson (2), Debbie Fortenberry (1), Cynthia Bright (1), Helen Hua (1), Jiarong Ying (1), Paul Sykes (1), Kay Hollifield (3), Charlton Vincent (3), and Margot Hall (1), (1) University of Southern Mississippi, Hattiesburg, MS 39406, (2) University Medical Center, Jackson, MS 39216, and (3) Laurel Clinic for Women, Laurel, MS 39442
Leukemia and lymphoma are serious pathologies. The American Cancer Society estimates that there will be 34,810 and 63,740 new cases respectively and 22,570 and 20610 deaths respectively from these cancers in the USA during 2005. Classical tumor antigens have been used for therapeutic monitoring but not for diagnosis of the blood cancers. The objective of this study was the comparison of NSE and ferritin with eleven other tumor antigens for diagnostic efficacy in these cancers. Sera from 554 patients (13 leukemia, 16 lymphoma, 330 other cancers, and 195 non-cancer) were assayed for the presence of tumor antigens and the results correlated with diagnoses established pathologically. Immunoassay test kits from Diagnostic Automation (NSE, Ferritin, CA242), Hybritech (CEA, CA195), Centocor/Fugirebio Diagnostics (CA125, CA19-9, CA72-4, CA15-3, CA27.29, Cyfra21-1) CIS Biointernational (CA50), and Abbott (AFP) were used to test for the concentration of these antigens. Using the manufacturers' decision values the following diagnostic sensitivities were obtained: Leukemia: NSE 0.0%, Ferritin 58.3%, CA27.29 7.7%, CA195 7.7%, CA19-9 0.0%, CA50 0.0%, CA72-4 7.7%, CA125 0.0%, Cyfra21-1 0.0%, CEA 15.4%, CA15-3 15.4%, AFP 27.3%, CA242 30.8%; Lymphoma: NSE 0.0%, Ferritin 66.7%, CA27.29 25.0%, CA195 12.5%, CA19-9 12.5%, CA50 12.5%, CA72-4 0.0%, CA125 12.5%, Cyfra21-1 0.0%, CEA 12.5%, CA15-3 43.8%, AFP 6.7%, CA242 6.3%. Diagnostic specificities were >75%. From these data we conclude that ferritin was useful for the diagnosis of leukemia and lymphoma and that all other markers were not.
PROJECT F.I.R.M. (FAMILY-BASED INSULIN RESISTANCE MANAGEMENT)
Erin Reynolds (1*), Scott McIntosh (2), LaKeyah Quinn (2), and Cammie Hilliard (2), (1) Tougaloo College, Tougaloo, MS 39174 and (2) Rochester, New York 14642
The objective of this pilot study is to assess the needs of the overweight residents of the underserved areas in Rochester, New York and gain qualitative details on the best procedures for prevention of type 2 diabetes mellitus in these populations. Ten families were referred by participating doctors based on children being overweight or displaying risk factors for becoming overweight. The families were then followed to study for the necessary methods of intervention, or prevention of overweight resulting in type 2 diabetes mellitus. In order to expand on ways to improve the outcome of the study and/or the health of its participants, contributing doctors were interviewed with qualitative questions about Project FIRM. The responses to the questions were analyzed to determine the merit of the study. These consultations were effective ways to evaluate the study as a whole and identify the necessary improvements for future research. Overall, participants are aware that Project FIRM is an invaluable tool in decreasing the prevalence of overweight and obesity in the underserved areas of Rochester, New York. Along with other health issues, children who are overweight, obese, or have the risk factors for becoming overweight or obese are at risk for developing type 2 diabetes mellitus. If this health concern can be mended in these underserved areas in Rochester, it can serve as a model for cities nationwide in an effort to decrease the frequency of pediatric obesity resulting in the development of type 2 diabetes mellitus.
6:00 Poster Session III
THE ROLE OF (-) EPIGALLOCATECHINE GALLATE (EGCG) AND THYMOQUINONE ON THE PROLIFERATION OF PANC-1 CELL LINE IN CULTURE
Mary Tan*, Hamed A. Benghuzzi, Michelle Tucci, and Laura Franklin, University of Mississippi Medical Center, Jackson, MS
The limited ability of current treatments to control pancreatic cancer prompted us to examine the effect of antioxidants (EGCG and thymoquinone) on pancreatic cell proliferation in vitro. Cultured human pancreatic cancer cells (PANC-1) were exposed to EGCG or thymoquinoe alone or in combination for 24, 48, and 72 hours. Selected dosages for EGCG and thymoquinone (5, 25, 50 ug/dL) were delivered both by direct administration and a drug delivery system using TCPL ceramic capsules for 24, 48, and 72 hrs. Ceramic capsules were gas sterilized and prepared using standard procedures. Data collected from this study indicated a dose dependent relationship with direct administration of EGCG alone or in combination with thymoquinone. Conversely data collected from sustained delivery of EGCG alone or in combination with thymoquinone was dose independent. All selected dosages did affect the proliferation of PANC-1 cells during the direct administration and ceramic drug delivery. However a rebound effect was observed after 48 and 72 hours exposure to both EGCG and thymoquinone using direct delivery method. These results indicate that direct administration or ceramic drug delivery of thymoquinone (P<0.05) or EGCG (P<0.05) alone or in combination can limit PANC-1 cell proliferation. These findings suggest that specific dosages of (-) epigallocatechin gallate and thymoquinone tested significantly reduces the growth of PANC-1 cells and therefore has a strong potential as a useful therapeutic regimen for inhibiting pancreatic cell proliferation.
DIFFERENTIAL EFFECTS OF CYCLOSPORINE A ON RHESUS MONKEY KIDNEY EPITHELIAL CELLS IN CULTURE
Stacy Hull Vance*, Michelle Tucci, and Hamed A. Benghuzzi, University of Mississippi Medical Center, Jackson, MS 39216
Cyclosporine A (CsA) is extracted from Tolypocladium inflatum Gams, which is metabolized through the superfamily of hepatic isoenzymes P-450. CsA has a mean life of 6.4-8.7 h, although this varies among different individuals. Ninety percent of the drug is withdrawn through biliary excretion and only 6% appears unchanged in the urine. The exact mechanism of action of CsA is unknown; however, CsA has the ability to act on the immune system by blocking the biosynthesis of some lymphokines produced by T lymphocytes and interleukne-2 synthesis at the transcriptional level. It has been suggested that CsA acts by interacting with cytoplasmic membrane and activates the intracellular calcium pathway, or binds to cytoplasmic proteins (Cid, 2003). The specific objective of this study was to investigate the effects of various concentrations of CsA on the proliferation and viability of Rhesus Monkey Kidney epithelial cells (RMKEC) in culture. Thirty-five wells were plated with RMKEC and sub-divided into five equal groups. Group 2 was treated with 10mL of ETOH (vehicle for CsA). Groups 3-5, were treated with 5, 25, and 0.50 [micro]M of CsA, respectively. Data obtained suggested the following: (1) The 24 hour phase, showed no significance difference in cell number in the CsA 10 [micro]M and CsA 25 [micro]M in comparison to the control. (2) The use of 50 [micro]M CsA suppressed cell proliferation as early as 24 hours in comparison to the control and caused an increase in all biochemical markers) MDA, protein, and glutathione). (3) Interestingly, the vehicle resulted in an increase in cell proliferation and a decrease in the biochemical markers MDA and Glutathione at 24 hours. The data suggested the need for further studies need to be conducted to determine the full impact of CsA on Kidney Epithelial function.
MECHANICAL STRENGTH REPERCUSSIONS OF VARIOUS FIXATIVE STORAGE METHODS ON BONE
Graham Calvert, Scott Wingerter*, Michelle Tucci, Hamed A. Benghuzzi, and Aaron Puckett, University of Mississippi Medical Center, Jackson, MS 39216
This study compensates for the lack in literature on the actual effects that various fixative storage methods have on the mechanical strength characteristics of bone. Researchers usually operate under the assumption that fixation of bone in formalin has no effect on such material properties. Such assumptions could introduce error into a great number of bone fracture studies if a disparity in the mechanical properties of fixed bone versus in vivo does actually exist. Furthermore, such assumptions could go on to pose clinical risks for patients. This study focuses on the mechanical strength testing of four different groups of rat femurs that were extracted at various times and subject to differing storage procedures. The first, Group N, are fresh, new femurs extracted just days before testing. The second, Group F, are femurs that have been fixed in a 10% formalin bath for just over a year prior to testing. The third, Group W, are femurs that have also been fixed in 10% formalin for just over a year but were washed out just prior to testing. The fourth, Group P, are femurs that have been taken from rats that were perfused with formalin immediately following euthanasia. Mechanical strength tests on the four groups revealed that fixing bone in a 10% formalin bath significantly reduces the mechanical fracture strength properties of the bone regardless of whether the formalin is washed out prior to testing. The tests also revealed that bone from perfused animals behave mechanically like fresh bones from non-perfused animals suggesting that the formalin did not entirely infiltrate the bone and permanently fix the material. These results could have profound implications on how studies equate the behavior of in vitro bone to in vivo bone which could manifest as clinical complications for patients.
PATHOPHYSIOLOGICAL RESPONSES OF MRC-5 CELLS EXPOSED TO VARIOUS DOSES OF X-RAY RADIATION
Pamala Jones*, La'Toya Ross Richards, Hamed A. Benghuzzi, and Michelle Tucci, University of Mississippi Medical Center, Jackson, MS 39216
The numbers of x-ray technology procedures used as diagnostic tools is on the rise, the radiation effect from these procedures are said to be minimal, however there is a significant increase in developing cellular damage, that may lead to cellular aberrations. The objective of this study was to expose MRC-5 cells to various doses of x-ray radiation and evaluate its effects on the proliferation and morphology of the cells. The doses were 2, 6 and 10 (Gy), evaluated after 24, 48 and 72 hours of incubation. The results indicated a slight decrease in the cell number after 48 and 72 hours (p <0.05), in all groups exposed to either dose of radiation. After 24 hours, there was not a statistically significant difference between the groups exposed to either dose of radiation. Morphologically, the cells exhibited significant changes when exposed to either, 2, 6 or 10 Gy of radiation. The cells showed multiple nucleoli with all doses, and mitotic figures were observed with a dose of 2 Gy. After a dose of 6 Gy, hydropic swelling was noted as well as multiple nucleoli within the nucleus of the cells. After 48 and 72 hours, severe vacuolization with various degree of pleomorphism was noted with a dose of 10 Gy. Further analysis revealed that the higher the radiation dose, morphologically the more severe the damage after 48 and 72 hours, however, significant damage was evident with all doses, even as low as 2 Gy. The role of free radicals as a major factor in cell membrane damage is still being uncovered by current research.
ROLE OF NITRIC OXIDE IN DOMOIC ACID INDUCED HIPPOCAMAPL DEGENERATION
Manju Pande (1*), Joseph Wahome (1), David Desouza (2), and PJS Vig (2), (1) Mississippi Valley State University, Itta Bena, MS 38941 and (2) University of Mississippi Medical Center; University of Mississippi Medical Center, Jackson, MS 39216
Nitric oxide (NO) acts as a transynaptic retrograde messenger in the nervous system. The initial observation from our earlier studies showed that NO inhibitors (LNAME and 7NI) potentiated Domoic Acid (DA) induced stereotypic seizures indicating that NO may possess anticonvulsant properties. The immunohistochemical studies from CA 1/2 regions of hippocampus immunostained for glial fibrillary acidic protein (GFAP) showed large number of reactive glial cells in DA treated animals and in groups treated with NO Inhibitors compared to controls. Loss of calbindin D28k positive neurons in CA1/2 regions were also observed in 7NI and LNAME treated animals indicating neuronal damage in the hippocampus. In the present study, to quantify the earlier data we ran western blots to assess the specific biochemical changes in the hippocampus. Cytosolic fraction was prepared form brain tissue obtained from animals treated with DA, DA+LNAME, DA+ 7NI. Frozen hippocampal tissue samples were homogenized in homogenization bufferand centrifuged. The cytoplasmic protein fractions were prepared using cytosol-nuclear frationation kit. The cytosolic frations with equal amount of protein were ran on gels for immunoblot analysis for Beta-III-tubulin, GFAP and calbindin (CaB), GAPDH was used as house-keeping protein. No significant changes were observed from the immunoblot analysis for beta-111-tubulin. The western blot analysis for GFAP also showed no significant changes in treated samples as compared to control, the reason could be use of the total hippocampus and not specific regions. Sampling of specific area, where we observed immunohistochemical changes could have demonstrated some changes. There was a marked decrease in the expression of CaB in animals treated with NO inhibitors LNAME and 7NI compared to control and DA alone treated animals. The loss of calbindin positive neurons and further reduced expression of CaB on Western blots in animals treated with nitric oxide inhibitors suggest a positive role of NO in DA induced toxicity.
STEM CELL COLLECTION METHODS AND THE STEM CELL RESOURCES IN MISSISSIPPI
Anna Stevens, University of Southern Mississippi, Hattiesburg, MS 39406
I performed a study of the various types of stem cells in the human body and the methods used to collect those stem cells. I also researched the institutions in Mississippi that collect stem cells and the type of stem cells they utilize. The three main sources of stem cells in use today are bone marrow, peripheral blood, and umbilical cord blood. Bone marrow is the traditional source of stem cells, but peripheral blood is being utilized more often. Stem cells are collected from the peripheral blood via apheresis. The newest source of stem cells is umbilical cord blood. Mississippi has only one facility that collects and transplants stem cells. The University of Mississippi Medical Center in Jackson, MS is the location of the UMC Hematopoietic Cell Transplant Program. UMC\'s facility only collects bone marrow and peripheral blood stem cells; however they do utilize cord blood for pediatric cases. The UMC Hematopoietic Cell Transplant program is a member of the National Marrow Donor Program, which allows them to send out stem cells and also bring in stem cells that they need. UMC is providing Mississippi with a much needed service.
CHRONIC BLOCKADE OF VEGF RECEPTOR 2 CAUSES PROLONGED INCREASES IN VEGE EXPRESSION IN SKELETAL MUSCLES OF TREADMILL-EXERCISED MICE
Janelle Pryor*, A.J. Thibodeaux, P.B. McDonnell, and T.H. Adair, University of Mississippi Medical Center, Jackson MS 39216
Long-term exercise causes a VEGF-mediated increase in angiogenesis in skeletal muscle. The VEGF mRNA response consists of an initial large increase in expression (days 1-7), which returns to nearly normal levels after 14-28 days when the muscle capillarity has adapted to the exercise. This temporal relation between muscle capillarity and VEGF expression supports the contention that VEGF production may be regulated by a negative feedback mechanism. Presumably, the increase in capillarity induced by VEGF returns tissue oxygenation to normal, and VEGF expression, in turn, returns to nearly normal levels. To test the hypothesis that VEGF is subject to negative feedback regulation, we attempted to "open" the feedback loop using a VEGF receptor inhibitor, PTK787 (Novartis) to prevent angiogenesis in skeletal muscle during exercise conditioning. Male C57BL/6J mice were treated with PTK787 (50 mg/kg/day, oral), or an equivalent volume of vehicle, and exercised on a motorized rodent treadmill for 1 hr/day at 18 m/min with a 10 degree incline. After 14 days, VEGF mRNA expression in gastrocnemius muscle was 2-fold higher in exercised mice treated with PTK787 (n=8) compared to exercised mice treated with vehicle (n=4). These results suggest that inhibition of VEGF receptor function in the face of chronically increased metabolic demand leads to chronically elevated expression of VEGF. NHLBI (HL-51971)
TIME COURSE OF PROANGIOGENIC GROWTH FACTOR EXPRESSION DURING EXERCISE CONDITIONING IN MICE
Janelle Pryor*, P.B. McDonnell, and T.H. Adair, University of Mississippi Medical Center, Jackson, MS 39216
Male C57BL/6 mice (6-7 weeks old) were exercised on a rodent treadmill (18 m/min, 10 degree incline, 1 h/day). Gastrocnemius muscles were collected 1-2 h after exercise; agematched sedentary mice of the same strain served as controls. Relative mRNA expression of angiogenic growth factors was assessed by real-time RT-PCR. Vascular endothelial growth factor A (VEGF-A) mRNA expression was significantly increased in exercised mice compared to control on days 1 and 4, but not significantly different from control levels on day 14. Similar temporal changes in mRNA expression patterns were observed for angiopoietin-2, VEGF receptor-1, and the angiopoietin receptor, Tie-2. Angiopoietin-1 and VEGF receptor-2 mRNA expression in gastrocnemius muscle did not change significantly during the 14 days of the experiment. These results suggest that VEGF, as well as several other growth factors and their receptors, may be subject to negative feedback regulation during exercise conditioning, and that significant adaptation of skeletal muscle to exercise conditioning occurs by day 14 in C57BL/6 mice. NHLBI (HL-51971).
THE EFFECTS OF ESTROGEN ON THE VIABILITY AND PROLIFERATION OF CERVICAL TUMOR CELL LINES, SW 756 AND HELA, AND NORMAL CERVICAL CELLS, ECT 1/ E6E7 IN CULTURE.
Melissa P. Daniel* and Hamed A. Benghuzzi, University of Mississippi Medical Center, Jackson, MS 39216.
Cervical cancer remains a major health threat and the role of steroid hormones in cervical cancer is not clearly defined. This study investigated the effects of the steroid hormone, estrogen, on the viability and proliferation of two cervical tumor cell lines, SW 756 and HeLa, and normal cervical cells, Ect 1/ E6E7 cells, that were HPV transformed. In this study, all cell lines were treated with physiological and supraphysiological doses of estrogen (EP and EH) and examined biochemically and morphologically at 24, 48, and 72 hours. Biochemical assays performed at all time periods, included the MTS Assay to determine actual cell counts, the MDA and LDH Assays to evaluate cellular damage, and the Pierce BCA Protein Assay to determine total protein. In addition, morphological characteristics were evaluated using Papanicolaou and H & E staining. All values were compared to an appropriate control. Highlights of the results from this study indicated the following: Proliferation rates (MTS) were higher than the controls at all time periods in the HeLa cell line following EP (P<0.05, Dunnett's Test, at 24 and 72 hours) while the SW 756 cell line matched the control values. MTS values were lower than the controls at all time periods in SW 756 cells following EH (P<0.05) while the reverse was true for HeLa cells. MDA and LDH levels were higher in SW 756 cells following treatment with EH at 24, 48, and 72 hours (P<0.05): however, MDA levels did not differ from the controls in HeLa cells following EH at any time period. Cellular protein levels (Pierce BCA Protein Assay) were unremarkable at both doses for all time periods. Papanicolaou and H & E stains provided information on the cellular morphology of these cell lines. Conclusions were that the HPV-transformed tumor cell line (HeLa) was more resistant/ robust than the HPV-negative tumor cell line (SW 756).
THE EFFECTS OF GROWTH FACTORS ON THE PRODUCTION OF BONE MATRIX PROTEINS
Joel Davis*, Michelle Tucci, Laura Franklin, George Russell, and Hamed A. Benghuzzi, University of Mississippi Medical Center, Jackson, MS 39216
The bone morphogenetic proteins (BMPs) and transforming growth factor-betas (TGF-beta), are a group of structurally related proteins which have been shown to stimulate bone formation in vivo. Since these proteins are concentrated in the organic matrix of bone and would be released during bone resorption, they are likely to have a profound effect on the remodeling bone and may provide a link between bone resorption and bone formation. Osteopontin is a phosphorylated glycoprotein that is abundant in bone mineral matrix and accelerates bone regeneration and remodeling of bone. Osteocalcin, or bone gla protein, is a noncollagenous protein in mature bone. It is synthesized by osteoblasts and incorporated into bone matrix, although a fraction of the newly synthesized osteocalcin is released into the circulation. Osteocalcin is metabolized mainly in the kidney and to a lesser extent in the liver, the half-life in the circulation is about five minutes. Osteocalcin production is dependent on 1,25 dihydroxy vitamin D; vitamin K; and vitamin C. Serum osteocalcin correlates with bone formation. Osteoblast like cells were challenged with either OP-1, demineralized bone matrix proteins or IGF-1 for 24, 48 and 72 hours. After each incubation period, ELISA assay determined the levels of Osteopontin and Osteocalcin production and collagen type-I were determined by immunocytochemistry. The results clearly demonstrated an increase in Ostepontin and osteocalcin production by DBX as early as 24. All three treatments resulted in increased collagen type-I. This information is important for understanding the signaling pathways that may be innervated in the osteoblast following stimulation with growth factors.
THE EFFECTS OF GLUCOSAMINE, CHONDROITIN, AND THYMOQUINONE ON HTB-93 SYNOVIAL CELLS
Marilyn May*, Hamed A. Benghuzzi, and Michelle Tucci, University of Mississippi Medical Center, Jackson, MS 39216
Glucosamine, chondroitin, and thymoquinone (TMQ) are the complementary medicines under investigation in this study. Glucosamine is a naturally occurring glycoaminoglycan that contributes to the development of proteoglycans needed for the development of cartilage development and regeneration. Chondroitin is also a naturally occurring glycoaminoglycan that seems to support the efforts of glucosamine as well as provides chondroprotection while serving as a 'water magnet' within the joint matrix. Thymoquinone is derived naturally from the black seed plant that is extremely popular within Middle Eastern countries. Its benefits are multiple, including both antioxidant and anti-inflammatory properties. These products were administered to HTB-93 synovial cells and cell viability, damage and alterations in morphology were analyzed after 72 hours. Preliminary results revealed that chondroitin increased cell number in the high treatment group with increased nitric oxide production and decreased glutathione content compared to control, glucosamine and TMQ. Decreased glutathione levels were seen in the medium and high doses of both glucosamine and chondroitin. Increased levels of glutathione were seen with increasing TMQ, with out changes in cell numbers or nitric oxide. The data indicates that medium and high doses of glucosamine and chondroitin may be cytotoxic to HTB-93 synovial cells.
PREVALENCE OF ANTIBODIES TO HEPATITIS C VIRUS IN A UNIVERSITY SETTING
Carolyn Beck*, Virginia M. Crawford, Kim R. Henson, Annette High, and Sonja Hollingshead, University of Southern Mississippi, Hattiesburg, MS 39406
The purpose of this study was to determine the prevalence of antibodies to the Hepatitis C Virus(HCV) in a volunteer sample of university students and then compare the rate of infection to that of the general public of the United States (1.8%). The occurrence of HCV in this age population has been not been widely reported. Such studies are of value to health care providers, health educators and to blood donor centers. A sample of convenience utilized students visiting the on campus health care service. Volunteers donated a blood sample and completed a researcher developed survey. Two hundred seventy six students participated. Blood samples were tested at a major blood center for antibodies to HCV using Abbott Laboratories EIA 2.0 methodology. Positive samples were repeated in duplicate. Students testing positive were contacted and received counselling regarding further testing. The mean age of the participants was 23 +/- 5 years. Characteristics of the group included: 63% female, 57% Caucasian, 38% African American, 29% with tattoos, 67% had pierced ears, and 19% had other body piercings. Participants also reported having other sexually transmitted diseases (17%) and 53% had multiple sex partners. Despite reported risks, only 2 of the 276 students tested EIA positive. On further testing by RIBA both were shown to be negative. The prevalence of anti-HCV in this population was 0%. Therefore, the incidence of HCV in this sample of university students was much lower than the prevalence in the general population.
THE PHYSIOLOGICAL EFFECT OF CONVENTIONAL TREATMENT WITH EPIGALLOCATECHIN-3-GALLATE, THYMOQUINONE, AND TANNIC ACID ON THE LNCaP CELL LINE
La'Toya Ross Richards*, Pamala Jones, Hamed A. Benghuzzi, and Michelle Tucci, University of Mississippi Medical Center, Jackson, MS 39216
Several antioxidants have been discussed for use in prevention and treatment of prostate carcinoma. Epidemiological evidence has indicated that these antioxidants may reduce the risk of prostate cancer by underlying mechanisms that remain unclear. The aim of this study was to use the androgen-dependent LNCaP human prostate cancer cell line as a cell model to evaluate the physiological effects to conventional treatments with both low (LD) and high doses (HD) of epigallocatechin-3-gallate (EGCG), thymoquinone (TQ), and tannic acid (TA). Following treatment, cells were incubated and the various groups were evaluated at 24, 48, and 72 hours. After 24, 48, and 72 hours of incubation, all groups suppressed the cells, but the TQHD treated group seemed to be the most potent. The TQHD group also demonstrated the greatest decrease in total protein levels in comparison to the control. According to one-way analysis of variance (ANOVA), significant differences were observed (P<0.001). Upon observation of the PSA values, all groups showed decreased levels; however, the TQHD treated group showed an initial suppression after 24 hours and then finally portrayed adaptation after 48 and 72 hours. Maliondialdehyde (MDA) values were also assessed at the same three time periods (24, 48, and 72 hours) as an indicator of membrane integrity. After 24 hours of incubation, the TAHD group demonstrated the greatest increase in MDA levels. Morphologically, the cells demonstrated significant changes upon antioxidant exposure. These findings reveal that antioxidants may serve as agents for prostate cancer prevention; however, further experiments are needed to understand the interactions involved.
IMMUNOSTAINING TO DETERMINE OSTEOCYTE APOPTOSIS BY CASPASE-3
J. Belcher (1,2*), H. Follet (2), and D. Burr (2), (1) Jackson State University, Jackson, MS 39217 and (2) Indiana University, School of Medicine, Indianapolis, IN 46202
Caspases are cysteine proteases that play a critical role in apoptosis. Activated caspase-3 is a downstream cell death signal. Thus, immunostaining by caspase-3 can be used as an effective tool for the assessment of osteocyte apoptosis. The specific intent of this work was to optimize caspase-3 immunostaining protocol. Working antibody dilution, method of epitope retrieval, peroxidase inactivation, and blocks were examined through various series of antibody runs. 1:300 appears to be a good working dilution for the caspase-3 antibody. It produced a high signal to noise ratio with minimal nonspecific expression. Of the retrieval techniques analyzed, DeCal retrieval appeared to limit nonspecific staining the most. We have yet to determine the significance of peroxidase inactivation with 3% [H.sub.2][O.sub.2] in methanol. Casein appears to be a better blocking agent than serum. Caspase-3 immunostaining is a vital tool in the evaluation of apoptosis. By considering the previously mentioned variables, we hope to have a protocol for a more accurate measure of osteocyte apoptosis. Supported in part by NIH R25 GM067592-02.
EFFECTS OF AMBULATION WITH STANDARD WALKER AND/OR ROLLING WALKER WITH PLATFORMS ON CARDIOPULMONARY FUNCTIONS AND BIOCHEMICAL STRESS MARKERS IN NON-WEIGHT BEARING INDIVIDUALS
Veronica Taylor, Jim Garrett, Jason Reeves, Leann Staton, Hunter Stark, Ameze Adah, and Derrick Huang, Hamed A. Benghuzzi, and Felix Adah*, University of Mississippi Medical Center, Jackson MS 39216
Research shows an increase in cardiopulmonary demands with exercise. Patients with traumatic injuries referred for Physical Therapy may be placed on non-weight bearing (NWB) on the affected limb for healing to occur. Few studies have explored the physiological impact of various walkers in a non-weight bearing individuals. The purpose of this study was to examine the effects of ambulation with a standard walker (SW) and a platform walker with wheels (RW) using cardiopulmonary parameters and biochemical stress markers in one legged ambulation. The study consisted of 2 phases using male and female students (n=14), age 23 to 27 years (mean [+ or -] SE of 24.21 [+ or -] 1.24). Phase I consisted of the participants ambulating with a SW and RW for 6 minutes. The heart rate (HR), respiratory rate (RR), systolic blood pressure (SBP), and diastolic blood pressure (DBP) before and after were recorded. Phase II consisted of the same participants and measurements, but instead of ambulating for 6 minutes, they ambulated for 12 minutes. In addition, urine and saliva samples were collected before and after ambulation in Phase II. Results indicated a statistically significant difference between before and after measurements of HR, SBP, and RR for SW & RW for the 6 minute-walk and 12 minute-walk levels and DBP for the RW for 12 minutes walk (p<0.05). The cardiovascular demands were not statistically different using each type of walker and walking for 6 or 12 minutes (p>0.05). There was no significant difference in cortisol levels in Phase II (12 minutes walk) in both the saliva and urine. Our study suggests that ambulation using the types of walker used in this study for short and long period of walk resulted in increased cardiovascular demands.
POXVIRUS PHOSPHATASE VH1 ALTERS IFN-GAMMA REGULATED GENE EXPRESSION IN MACROPHAGES
Andre W. Hite, Jr. (1,2*) and Michael J. Klemsz (1), (1) Indiana University, School of Medicine, Indianapolis, IN 46202 and (2) Jackson State University, Jackson, MS 39217
Poxviruses including Variola (small pox), and Vaccinia, which is used to vaccinate against small pox have developed numerous methods for altering and/or evading an immune response. One such way is by the ability of the viral VH1 phosphatase to alter signaling cascades, which is vital for viral replication and may play a key role in the invasion of host defense during infection. Our studies focus on understanding if either the Vaccinia or Variola VH1 protein blocks interferon-gamma induced STAT1 regulated gene expression in macrophages. It is our hypothesis that the Variola protein will function more efficiently than Vaccinia in blocking STAT1 function and may contribute to the pathogenicity of the smallpox virus. We first wanted to show that VH1 is present in the virus prior to infection. Results using RT-PCR showed that VH1 was expressed in Vaccinia virus that was used to infect cells. Next, a comparison of transfection reagents showed that Gene Porter 2 was optimal for our experiments in the macrophage cell P388D1. A series of co-transfection experiments showed that transfection of a VH1 expression plasmid reduced the STAT1 regulated reporter gene expression of the TAP-1 promoter. These data suggest that VH1 in the virus may prevent the induction of interferon-gamma regulated genes in macrophages during viral infection. Supported in part by NIH R25 GM067592-02 and P01 AI56097-02S1.
THE EFFECTS OF GROWTH FACTORS ON CELLULAR PHOSPHOLIPIDS
Laura Franklin*, ShaDonna Jefferson, Michelle Tucci, Hamed A. Benghuzzi, and Rodney Baker, University of Mississippi Medical Center, Jackson, MS 39216
Signals are transduced across the cell membrane through a series of events that are triggered by the activation of receptors or opening of ion channels. It is evident that the stimulation of cells can elicit a series of catabolic cascades, which cause degradation of several membrane phospholipids. Several breakdown products participate directly in the intracellular signaling cascade. The objective of this study was to establish the changes in phospholipid membrane fractions of MG-63 cells following stimulation with growth factors (DBX, OP-1, and IGF-1) for 15 and 30 minutes. After collection of the cells, methods were performed to isolate cellular phospholipids and final analysis of each phsopholipid fraction by thin-layer chromatography and liquid scintillation counting. The results showed OP-1 and IGF-1 treatments at 15 and 30 minutes caused a 50% decrease in phosphatidylcholine fraction and increased phosphatidylserine and phosphatidylethanolamine fractions compared to DBX and control. DBX had 25% increase in phosphatidylinositol fraction when compared to control, OP-1 and IGF-1. Levels of phosphatidic acid and phosphatidylglycerol remained unchanged. Understanding the dynamics of the membrane may help in establishing the intracellular signaling pathways which are activated by various growth factors.
EFFECTS OF DRUGS ON THE DIFFERENTIATION OF PC12 CELLS
Ujjwal K. Rout*, Laura Vick, Dirk M. Dhossche, and John R. Gosche, University of Mississippi Medical Center, Jackson, MS 39216
Epidemiological studies show that autism may result from exposure to valproic acid, thalidomide or alcohol during gestation. This commonality in the behavioral outcomes resulting from the exposure of different teratogens suggests that autism may result from disturbances in developmental process common to these teratogens. Normal brain development requires intricate balance amongst proliferation, differentiation, migration and apoptosis of different cell types in the developing tissues that are regulated by changes in the expression pattern of genes. Therefore autism-like symptoms resulting from these teratogens may derive from changes in the expression and function of genes regulated by common genetic elements. To test this possibility, we examined effects of these teratogens on the activity of transcription factor EGR that is known to regulate differentiation of several cell types. PC12 cells were cultured on laminin-coated plates. At 1h of incubation, cells were induced to differentiate into neuronal phenotype by the addition of nerve growth factor (NGF) in the culture medium. At the same time, cells were treated with alcohol, thalidomide or valproate and incubated for additional 3h. EGR activity was determined in the nuclear extracts of untreated and treated cells by a fluorescent based method using Luminex-100 instrument. Effects of low and higher concentrations of teratogens were examined on the activity of EGR. NGF increased EGR activity in all experimental conditions, and higher concentrations of teratogens elevated NGF-induced EGR activity. Results suggest that exposure to these teratogens may cause excessive and premature differentiation of neurons during brain development resulting in abnormal neural positioning or connection. Our data indicate that EGR may be a common target of these teratogens in the developing brain.
THE NEWEST MEMBER OF THE INHIBITORS OF APOPTOSIS FAMILY, IAP LIKE PROTEIN 2 DEMONSTRATED GROWTH FACTOR REGULATION IN CD34+ AND MO7E CELLS
Leshundra Young (1,2*), Seiji Fukuda (2), Janardhan Sampath (2), and Louis M. Pelus (2), (1) Jackson State University, Jackson, MS 39204 and (2) Indiana University Purdue University at Indianapolis, Indianapolis, IN 46202
Inhibitors of apoptosis proteins are a family of natural inhibitors of cysteine proteases that regulate apoptosis through caspase dependent and independent mechanisms. ILP2 (IAP-like protein-2) is the newest member of the IAP family. ILP2 regulates apoptosis by restricted specificity for caspase-9 and neutralization of the Smac/Diablo complex. It is expressed in the testis and in lymphoblastoid and other transformed cell lines. Of the eight members of the IAP family, only Survivin is growth factor regulated in normal human CD[34.sup.+] stem and progenitor cells and the human hematopoietic cell line, MO7e. In order to determine if ILP2 is similarly expressed and regulated, we examined mRNA and protein expression of ILP2 before and after incubation with growth factors in CD[34.sup.+] and MO7e cells and examined if ILP2 expression was associated with cell cycle progression and apoptosis. Western analysis and quantitative-RT-PCR revealed that ILP2 was expressed in MO7e and CD[34.sup.+] cells and that growth factor starvation reduced ILP2 protein and mRNA. Addition of stem cell factor (SCF) or granulocyte-macrophage colony-stimulating factor (GM-CSF) blocked downregulation of ILP2 protein induced by growth factor deprivation and enhanced cell survival and cell cycle progression in MO7e cells. ILP2 mRNA was significantly up-regulated by the early acting cytokine SCF but not by the late acting cytokine GM-CSF. ILP2 mRNA was down-regulated by the combination GM-CSF + SCF, suggesting that ILP2 may be regulated with differentiation. These results suggest that ILP2 may be involved in the growth factor mediated proliferation, survival or differentiation of primitive hematopoietic cells. Supported in part by NIH R25 GM067592-02 and P01 AI56097-02S1.
Exhibit Hall A1
Session III: Biology, Chairs: Joseph A. Cameron & Ham Benghuzzi
9:00 Opening Remarks
9:02 MARKERS OF INFLAMMATION AND OXIDATIVE STRESS IN ALZHEIMER'S DISEASE
Christopher Bennett* and March Ard, Alcorn State University, Lorman, MS 39096 and University of Mississippi Medical Center, Jackson, MS 39216
Alzheimer's disease (AD) is a neurodegenerative disease that affects millions of people throughout the world. The risk of obtaining AD usually increases after the age of 70 due to increased levels of oxidative stress and inflammatory processes. Oxidative stress and inflammation may cause AD. Amyloid beta-protein plaque accumulation is believed to initiate the main events that are believed to cause AD. The purpose of this study is to investigate links between oxidative stress and inflammatory processes in AD. Blood samples were collected to measure oxidative stress through lipid peroxidation measurements. The blood samples were from the age groups of 20-40 and 60-80. Brain tissue samples were also used in this study. The brain tissues were collected from AD patients and from controls, which did not have AD. The brain sections were immunostained for light microscopy to determine if inflammatory and oxidative stress related molecules were in the Alzheimer brain tissue. Electron microscopy (EM) was also used to determine if there were auto-antibodies, which are inflammatory molecules in Alzheimer brain tissue. These experiments usually revealed certain areas that were specific for staining, proving that there were some AD related molecules in that section. The lipid peroxidation measurements on plasma samples didn't show a difference in the amount of oxidative stress between the two age groups. EM and light microscopy showed immunostained neurons and cells, which could be indicators of AD related molecules. The molecules found in the brain sections could be responsible for the development or progression of AD.
9:15 EFFECTS OF BUTORPHANOL ON NEURONAL ACTIVITIES OF THE RAT LOCUS COERULEUS
Matthew Burford* and Hong Zhu, University of Mississippi, University, MS 38677 and University of Mississippi Medical Center Jackson, MS 39216
The brain noradrenergic locus coeruleus (LC) has been shown to play an important role in the development of physical dependence on opioids. Butorphanol is a mixed agonist/antagonist opioid analgesic. The dependence liability of butorphanol has been considered to be low when used within the therapeutic dose range. However, a marked physical dependence liability has been observed in humans and animals when larger doses were administered for a prolonged period. In the present study, using a multiple-electrode recording technique, we examined the effects of burtorphanol on LC neuronal activities in adult Sprague-Dawley rats. A multi-wire electrode was implanted into the nucleus LC. This technique allows us to monitor several LC neurons simultaneously. A cannula was implanted into the lateral cerebroventricle for drug injections. LC neuronal activities were recorded before and after butorphanol injection under halothane anesthesia. We found that a relative high dose of burtorphanol (78 nmol) significantly inhibited the firing rates of LC neurons. In a subpopulation of the LC neurons, butorphanol also induced synchronous oscillatory burst activities. The inhibitory action and synchronous burst activities can be reversed by opioid receptor antagonist naloxone, indicating the involvement of opioid receptors. These results suggest that changes in LC neuronal activity may be involved in the development of butorphanol dependence. (supported by NIDA 016440 and Center for Psychiatric Neuroscience in the UMC)
9:30 ROLE OF CYTOCHROME P-450 3A4 (CYP 3A4) GENES IN THE DEVELOPMENT OF BREAST CANCER
Crystal Berry (1*), Angela Lewis (1), Steven Bigler (1), Paula Smith (1), Henry Barber (1), Teandria Burns (2), Joseph Cameron (2), and lga McDaniel (1), (1) Univ. of MS Medical Center, Jackson, MS 39216 and (2) Jackson State University, Jackson, MS 39217
Background: Cytochrome P-450 3A4 has been implicated in the etiology of breast cancer. A transition of A to G at position -290 of the CYP 3A4 gene has an effect on the level of gene transcription and has been associated with several disease conditions. Goal of Study: We sought to determine the frequency distribution of this variant and its effect on gene expression in breast cancer. Methods: Patients undergoing lumpectomy, needle localization, simple mastectomy and modified radical mastectomy were recruited. Genotypes were detected by a PCR-base approach using SNP analysis of human CYP 3A4 gene. Expression levels of mRNA transcripts were determined by RT-PCR. Results: The homozygous GG allele had a 2-fold increase in patients with malignant tumors. The AA allele was present in 50% and 33% of patients in stages 0 and I, respectively, while 67-100% in stages II and III carried AG or GG alleles. The expression levels of CYP 3A4 transcript appeared to be higher in PBMCs of patients with benign tumors as compared to malignant tumors (p<0.03). Conclusions: Patients who were homozygous GG genotype may have an increased risk of developing breast cancer. CYP 3A4 genotype analysis may predict likelihood of developing breast carcinoma, and might allow earlier detection and more effective treatment of breast cancer. (Crystal Berry, 2nd year medical student, is recipient of the 200 Dean's Summer Research Fellowship Program.)
9:45 DEVELOPING NEW CLASSES OF ANTIDEPRESSANT MEDICATION
John Stoker (1*), Mark Hamann (2), Rae Matsumoto (2), Jamal Shaikh (2), Jia Jia Wang (2), and Matt Brammer (3), (1) Alcorn State University, Lorman, MS 39096, (2) University of Mississippi, University, MS 38677 and (3) University of Oklahoma, Norman, OK 73019
Antidepressant drugs that are used to treat depression take up to several weeks before becoming effective. Not only do they take too long before they become active, but they have some major or minor side effects. This research is being conducted to identify new and better antidepressant medications for treating depression with less side effects. The new compounds were chosen because they presently show potential new treatments for cancer and infectious diseases. Two of the new compounds show some resemblance in their chemical structure to those of current antidepressant medications or to serotonin, the brain chemical that they target. Even though the others have different chemical structures, we feel that if antidepressant-like actions are displayed, then these compounds may target different receptors. The new compounds were tested for antidepressant-like activity in mice using the forced swim test, the most established animal model for predicting antidepressant-like activity. Reductions in immobility time served as the indicator of antidepressant-like activity. The results from the new compounds were compared to the compounds that were used as the positives controls. The positive controls used in the experiment were fluvoxamine and desipramine. Two of the new compounds had significant antidepressant-like actions, while the others did not. These two compounds represent potential new antidepressant drugs.
10:00 EFFECTS OF HUMAN UMBILICAL CORD BLOOD REGULATORY T CELLS ON EFFECTOR CD8+ T CELL DIFFERENTIATION
Pamela L. Ruffin (1,2), Young-June Kim (2), and Hal E. Broxmeyer (2), (1) Jackson State University, Jackson, MS 39217 and (2) Indiana University, School of Medicine and the Walther Cancer Institute, Indianapolis, IN 46202
CD[8.sup.+] T cells are crucial for host defense against virus and tumor cells. Upon antigen recognition, CD[8.sup.+] T cells acquire effector functions. CD[4.sup.+]CD[25.sup.+] regulatory T cell (Treg) has been known to suppress CD[8.sup.+] T cell-mediated anti-tumor immune responses. The mechanisms which Treg suppresses CD[8.sup.+] T cell-mediated effector responses remain unclear. Contrast to Treg, signal through NKG2D receptor promotes CD[8.sup.+] T cell effector responses to tumor cells as an activating co-stimulatory receptor. We investigated whether Treg and NKG2D counteract the effector CD[8.sup.+] T cell activity using naive CD[8.sup.+] T cells isolated from human umbilical cord blood (CB). Induction of granzyme B is a hallmark of CD[8.sup.+] T cell effector differentiation. By coculturing Treg with CD[8.sup.+] T cells, we determined an effect of Treg on induction of granzyme B expression in the CD[8.sup.+] T cells through flow analysis. Unexpectedly, we found that Treg did not suppress but promoted induction of granzyme B. NKG2D has been known to be downregulated by tumor cell-derived ligand binding in cancer patients. Thus, we hypothesized that Treg might inhibit induction of NKG2D in CD[8.sup.+] T cells. We mimicked the ligand-induced downregulation of NKG2D by culturing CD[8.sup.+] T cells on anti-NKG2D-coated plates. Downregulated NKG2D expression was gradually restored. We examined whether Treg affected restoration of NKG2D on CD[8.sup.+] T cells by culturing anti-NKG2D on CD[8.sup.+] T cells with/without Treg. Similar to Treg on granzyme B, Treg enhanced the NKG2D expression. In summary, Treg isolated from CB enhanced induction of granzyme B and NKG2D expression in CD[8.sup.+] T cells. Supported in part by NIH R25 GM067592-02 and P01 AI56097-02S1.
Symposium II: Drug Delivery, Chairs: Kenneth Butler & Felix Adah
11:00 CONVENTIONAL AND SUSTAINED DELIVERY L-DOPA AND THYMOQUINONE ON SH-SY5Y HUMAN NEUROBLASTOMA CELLS
Tina Mitchell Martin*, Hamed A. Benghuzzi, and Michelle Tucci, University of Mississippi Medical Center, Jackson, MS 39216
L-dihydroxyphenylalanine-(L-dopa) toxicity has been associated with the production of high levels of quinones. The reactive oxygen or nitrogen species generated in the enzymatical oxidation or auto-oxidation of L-dopa induce apoptotic or non-apoptotic cell death or neuronal damage. Thymoquinone is the major active component in N. sativa and has significant cytoprotective, anti-inflammatory and antioxidant actions. It is thought that thymoquinone may act on the oxygen free radicals and quinones from L-dopa cytotoxicity and/or auto-oxidation thereby decreasing L-dopa neurotoxicity. OBJECTIVE: The objective of this study was to determine the effects of thymoquinone on the viability and metabolic activity of SHSY5Y human neuroblastoma cells alone or challenged with levodopa (L-dopa) using conventional and sustained drug delivery. RESULTS: Cell numbers were relatively maintained with conventional delivery thymoquinone at 24 and 48 hours and in sustained thymoquinone delivery routes at 24, 48 and 72 hours. A reduction in cell number in thymoquinone/L-dopa cells was seen at 72 hours in the conventional delivery group. Cell numbers, however, were relatively maintained in conventional at 24 and 48 hours and in sustained drug delivery routes at 24, 48 and 72 hours suggesting a possible protective effect of thymoquinone. CONCLUSION: When thymoquinone and L-dopa were administered together using sustained drug delivery, cell counts were similar to thymoquinone given alone at 24 and 48 hours Thymoquinone has proven to have cytoprotective and antioxidative properties and therefore, may prove beneficial in neuroprotective strategies of Parkinson's disease.
11:15 THE EFFECTS OF CORTISOL AND ENDOTOXIN EXPOSURE ON RC/4B PITUITARY ADENOMA CELLS
Lisa A. Haynie*, Michelle Tucci, and Hamed A. Benghuzzi, University of Mississippi Medical Center, Jackson, MS 39216
Pituitary adenomas are benign tumors that arise exclusively within the anterior pituitary. Many of these tumors can respond to cortisol levels that are physiologic or supraphysiological. The objective of this study was to develop a model system to study the effects of hypocortisolism, hypercortisolism alone or at different points of endotoxin administration and compare those with physiological cortisol concentration. RC/4B cells were challenged with LPS either 30 minutes prior to or post cortisol exposure. The cells were evaluated at 24, 48 and 72 hours following the challenge. Regardless of the cortisol dose + the pre or post exposure to LPS, RC/4B cells were viable throughout all experimental periods. Also, the conventional delivery of cortisol 30 minutes after LPS exposure on the RC/4B cell line appeared to reduce the production of hydroxyl ion radical, which suppressed the membrane lipid peroxidation. Additionally, future considerations are warranted to investigate whether LPS directly triggers reactive nitrogen intermediates.
11:30 EFFECT OF EXOGENOUS STEROID HORMONES ON CYTOKINE EXPRESSION IN FIBROUS TISSUE SURROUNDING TCP BIOCERAMIC IMPLANTS
Kenneth R. Butler and Hamed A. Benghuzzi, University of Mississippi Medical Center, Jackson, MS 39216
Cytokines are the mediators of the inflammatory response. Previous studies have indicated that certain cytokines are expressed during acute and chronic phases of inflammation. The purpose of this study was to study the effect of testosterone, dihydrotestosterone, and androstenedione on the expression of IL-1[beta], IL-2, IL-6, and TNF[alpha] at 90 days post-implantation. Twenty animals in four experimental groups (n=5/group) were implanted with one tri-calcium phosphate (TCP) device each. Group I animals were implanted with sham TCP ceramics (control) containing no hormone. Group II animals received a ceramic loaded with testosterone. Group III animals were implanted with a dihydrotestosterone-loaded TCP ceramic. Group IV animals received the TCP bioceramic loaded with androstenedione. At 90 days post-implantation, the animals were euthanized. The fibrous tissue surrounding the implants were evaluated microscopically after staining with hemotoxylin and eosin and antibodies to IL-1[beta], IL-2, IL-6, and TNF[alpha]. The results of this study indicate that cytokine expression in Groups I, II, & III were similar. Specifically, testosterone and dihydrotestosterone did not adversely effect expression of these cytokines. In Group IV, androstenedione inhibited the expression of IL-1[beta], IL-2, and IL-6. TNF[alpha] was the only cytokine that was positive in this group. These findings suggest that the effect of exogenous hormones on thickness and cellular composition of the fibrous tissue may be dependent on cytokine expression.
11:45 EFFECTS OF SUSTAINED RELEASE OF STATIN BY MEANS OF TRICALCIUM PHOSPHATE LYSINE DELIVERY SYSTEM IN A DEFECT AND SEGMENTAL FEMORAL INJURIES ON CERTAIN BIOCHEMICAL MARKERS IN VIVO
Felix Adah (1), Hamed A. Benghuzzi (1), Michelle Tucci (1), George Russell (1), Audrey Tsao (1), and Barry England (2), (1) University of Mississippi Medical Center, Jackson, MS 39216 and (2) University of Michigan Medical School, Ann Arbor, MI 48109
Statins, which are 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors are widely used for the treatment of hyperlipidemia, and recent studies and animal data suggest that statins promote osteogenesis and increase bone strength. However, little is known about the effects of statins delivered by sustained delivery system to a target site of a defect and segmental bone fractures on certain biochemical markers including reproductive hormones. The purpose of this study was to develop a targeted statin delivery system using Tricalcium Phosphate Lysine (TCPL) for defect and segmental femoral injuries and evaluate the effects on alkaline phosphatase, total protein, malinodialdehyde, glutathione, total cholesterol, testosterone, luteinizing hormone, statins, and follicle-stimulating hormone. Because of the influence oral intake of statins might have on certain body organs, we also examine the histomorphology of the vital and reproductive organs of the animals receiving statins for a period of 30 days and 12 weeks post surgery. Simvastatin used in this study significantly increased fracture healing and without significant influence on the body weights and the weights and morphology of the vital and reproductive organs. There was a significant reduction in the cholesterol levels on the 3rd week in both phases of the study and at the conclusion of the study the difference in the cholesterol levels was no more significant in both phases. Other biochemical markers including plasma LH, FSH and testosterone levels were not affected by active treatment with simvastatin. In conclusion, short and long-term simvastatin treatment delivered at a fracture target site did not influence vital and reproductive organs, the systemic levels of the biochemical markers studied, but was able to effectively stimulate bone formation in simple and complicated segmental fractures.
Exhibit Hall A1
1:00 First Aid Panel Discussion
2:15 Divisional Business Meeting & Awards
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|Publication:||Journal of the Mississippi Academy of Sciences|
|Date:||Jan 1, 2006|
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