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Health Sciences.

Chair: D. Olga McDaniel, University of Mississippi Medical Center

Vice-chair: Michelle Tucci, University of Mississippi Medical Center

THURSDAY MORNING

Emerald

9:30 Welcome: D. Olga McDaniel, Chair of Health Sciences Division

Mini-symposium--Selected Topics I

Moderator: L. Margaret Drake, University of Mississippi Medical Center

9:45 THE RISE OF HEALTHCARE INSTITUTIONS FOR SPECIFIC DISEASE IN MISSISSIPPI

L. Margaret Drake, University of Mississippi Medical Center, Jackson, MS 39216

In the 19th and early 20th century, healthcare institutions developed for citizens of Mississippi. The phenomenon of such institutions started in the Middle East. After the Crusades, Europeans adopted ideas about congregating people with the same disease together. Alms houses and pest houses were the first such European institutions followed by asylums for the mentally ill. Later sufferers of infectious diseases were similarly concentrated as the understanding of contagion arose. In the New World, mental health hospitals were the first large institutions to be built with the first in Virginia predating the Revolutionary War. In Mississippi, the Asylum for the Insane predated the Civil War. The Sanatorium predated World War I. As the outskirts of the city of Jackson reached the borders of the Mississippi Asylum for the Insane in the 1930s, the government provided a more remote place for the mentally ill at Whitfield. In the 1940s the Rapid Treatment Center buses traveled to provide penicillin for syphilis sufferers. The Hill Burton Act PL 79725, Hospital Survey and Construction Act of 1946 provided funds for hospitals which changed the way healthcare was delivered in the last half of the 20th century. Mississippi exemplifies society's response to healthcare. Citizens in the 19th and early 20th century were more willing to renounce their individual freedoms to a healthcare institution than after WWII. With more education through the GI Bill and more post secondary education after WWII, people claimed increasing decision making power in healthcare. Availability of more effective treatments also contributed to the demise of large state run institutions.

10:00 PHYSICAL THERAPY PIONEERS IN MISSISSIPPI--THE FOUNDING OF A PROFESSIONAL LEGACY

Cynthia Scott*, Ruth M. Burgess, W.T. Johnson, J.H. Pierce, S.L. Scruggs, A.L. Stegall, and L.D. Welch, University of Mississippi Medical Center, Jackson, MS 39216

Background and Significance: The history of physical therapy in Mississippi is not well documented. Many of the pioneers of the profession are elders, increasing the urgency of gaining their perspectives of the profession. Purpose: The purpose of this presentation is to record the geographical distribution and personal accomplishments of 7 pioneers of physical therapy in Mississippi. Methods: An oral history methodology was used to interview 5 of the earliest Mississippi practitioners. The interview instrument was validated by two content experts familiar with the methodology as well as professional history. The sampling frame consisted of thirty three physical therapists who were the first licensees in the state. Five therapists were recruited from this list, each participating in an oral interview, which was transcribed verbatim. Transcripts were analyzed for common themes and elements. Two therapists who were deceased were also investigated through interviewing colleagues and searching archival materials. Data was triangulated with the archives of the Mississippi Physical Therapy Association (MPTA). Results: Major themes that emerged from interviews included arrival of personnel in the state, development of the MPTA. Discussion and Conclusions: This presentation focuses on one of the major themes, the early personnel, their entrance into the state, and the accomplishments of these pioneer physical therapists. The importance of the city of Vicksburg as a polio center, infantile paralysis foundation scholarships, and the individual characteristics and accomplishments of these individuals will be highlighted.

10:15 WOMEN'S TRUST OF THE HEALTHCARE SYSTEM

Amal Khoury, Nedra Lisovicz, Mandy Avis*, and Deonna Allen, University of Southern Mississippi, Hattiesburg, MS 39406

Objective: People's trust of the healthcare system affects their use of services and, therefore, their health outcomes. There is concern that some Mississippians underutilize healthcare services because of their mistrust of the healthcare system. This study examined the level of Mississippians' trust of healthcare providers and perceptions of racism within the healthcare system. Method: A statewide telephone survey of a representative sample of women was conducted in the summer of 2003. We focused on women, because they make the healthcare decisions for their families. Respondents reported their use of specific healthcare services, beliefs regarding healthcare, and demographic characteristics. Results: 1054 women 40 years of age and older participated. More than half of the women had a high school degree or less; 69% were White, non-Hispanic, and 27% were African American. The analysis showed that 62% of women believed that rich people received better medical care than poor people, and 77% believed that health insurance affected the kind of care that a person received. Also, 44% agreed that hospitals sometimes do not tell patients the truth. Women's perceptions regarding racism in healthcare delivery were also assessed. Data analysis by subgroup of women is underway. Conclusion: A majority of women appeared to mistrust the healthcare system. To increase the appropriate use of healthcare services, providers, and policymakers should directly address women's beliefs.

10:30 PHYSICAL THERAPY PRACTITIONERS AND AN AGING POPULATION

Neva F. Greenwald, University of Mississippi Medical Center, Jackson, MS 39216

Within the last quarter of a century, the older segment of the US population has increased significantly. The changes in the composition of the population and public policy have resulted in alterations in educational and clinical practice for health care practitioners including physical therapy practitioners. This historical presentation identifies milestone events related to physical therapy services for older adults. The changes in practice and the various processes that physical therapy practitioners have taken to address this client/patient population over the past 25 years are discussed.

10:45 Break

Podium Presentation--Session I

Moderators: Ibrahim Farah and Walter Brehm, Jackson State University and Kessler Air Force Base

11:00 TREATMENT OF POSTMENOPAUSAL OSTEOPOROSIS IN SOUTH MISSISSIPPI

Tommy Winston* and Benito Velasquez, University of Southern Mississippi, Hattiesburg, MS 39406

Osteoporosis is a serious and growing public health problem due to its association with skeletal fracture. Physical activity in the post-menopausal years has been recommended as a primary treatment strategy against osteoporosis and fractures. Both aerobic exercises and strength training provide the mechanical stimuli important for the maintenance and improvement of bone health. The purpose of this study was to determine the preferred method of rehabilitation used to treat postmenopausal osteoporotic women in south Mississippi. The study involved physical therapists, occupational therapists, kinesiotherapists, and athletic trainers from south Mississippi and compared the use of traditional aerobic exercise versus the more contemporary resistive-training method. A survey was developed by the researchers to determine which method each group of therapists preferred using in treatment of postmenopausal osteoporotic women. Criterion for participation was each therapist must treat at least five osteoporotic patients a year. Surveys were distributed via U.S. Mail. Data collected from completed surveys were analyzed using Student Sample Paired T-Test and Multivariate Analysis of Variance (MANOVA). Data analysis from both Student Paired Sample T-Test and Multivariate Analysis of Variance showed a statistically significant difference between the preferences of resistive-training exercise over aerobic exercise. In conclusion, therapists in south Mississippi preferred the use of resistive-training exercise as opposed to aerobic exercise in the treatment of postmenopausal osteoporosis.

11:10 ASSESSMENT AND SPATIAL DISTRIBUTION OF PESTICIDE USE, SOCIOECONOMIC STATUS, HEALTH CARE LEVEL AND DISPARITIES IN BREAST CANCER MORTALITY IN MISSISSIPPI

Ibrahim Farah* and Mohamed H. Abdalla, Jackson State University, Jackson, MS 39217

A combination of risk factors makes Mississippi suitable for studying the pathways of breast cancer etiology. The purpose of this study was to analyze pesticide exposure and the risk of breast cancer mortality. Data for this study consisted of secondary analyses of the Mississippi age-adjusted breast cancer mortality aggregated by two periods (1970-1994 & 1996-1999). Statistically significant linear associations were found between level of pesticide exposure (acres planted/crop) and breast cancer mortality rate in Mississippi women per state economic area (SEA.) For the 1970-1994 period, significant associations (p<0.05) using Pearson correlation analysis (r) were found for the rice crop with white females at Greenville SEA (r = 0.88819) and black females at the same SEA (r = 0.67844); the soy crop at Hattiesburg SEA for black females (r = 0.72034) and at the same SEA for black females with wheat (r = 0.57018); for the catfish crop, a significant correlation was found for white females in Yazoo (r = 0.59200) and black females in Columbus SEA (r = 0.70945). For the 1996-1999 period, there were significant findings (p<0.05) for the black females with cotton in Meridian SEA (r = 0.41878) and black females with rice in Greenville SEA (r = 0.63370). A positive significant association was found for white females with soy crop in Columbus SEA (r = 0.79209) and with black females in Yazoo SEA (r = 0.50667). In comparing the two periods of study, black females and white females showed different frequencies of positive and significant correlations.

11:20 A LINGUISTIC ANALYSIS OF PATIENT/PHYSICIAN COMMUNICATION AND ITS EFFECTS ON PATIENT HEALTH AND TREATMENT OUTCOMES

Derrick Spires* and Elizabeth Heitman, Tougaloo College, Tougaloo, MS 39174, and Vanderbilt University, Nashville, TN

This observational study is intended to identify and analyze specific linguistic aspects of patient/physician communication and to explore what implications they present for the patient/physician relationship. The linguist aspects include the use of paralanguage, the use of the indirect passive voice by physicians, the use of "dehumanizing" language when talking about patients, the effects of cultural perception on language use, and methods of improving patient/physician communication. The methods of this study include interviewing physicians and patients, a review of medical literature in order to assess how physician talk about and to patients, observing hospital culture and power structures and observing patient/physician, physician/physician, and patient/patient interactions, listening for linguistic cues and indicators of person perceptions and attitudes. Preliminary results have shown that while physicians believe that empathy is a key component in patient/physician communication, they frequently use dehumanizing terms such as "case" and indirect passive phrases such as: "the patient was diagnosed with ..." in their everyday discourse. There also appears to be a hierarchical power structure that is shown in language. A positive correlation between extended conversations between patients and physicians before and after surgery and case outcomes may also exist. Additionally, as Mississippi's immigrant population continues to grow and there continues to be a visible social separation along racial, class, gender lines, cultural competence training is becoming as important to health care professionals' ability to serve the population as scientific competence.

11:30 THE ROLE OF THE STATISTICIAN IN THE PROTECTION OF HUMAN RESEARCH SUBJECTS

Walter Brehm, U.S. Air Force, Keesler Air Force Base, Biloxi, MS

Clinical studies have been around as long as scientific medicine and the physician-investigators have always been bound by their Oath. Present concepts for the protection of research subjects are traditionally held to have begun with the Nuremberg Code following the exposure of wartime atrocities committed in the name of medical research. The Declaration of Helsinki, the Belmont Report, and the Common Rule further refined these ideas. Although the Belmont reports distills the essence of human subjects protection down to two words and a sentence fragment, "Beneficence, Justice, and Respect for Persons," a whole industry, with its own literature, legends, and heroes, has grown up around the subject. The involvement of the statistician early in a study design will help meet all three requirements for the protection of the subjects. Respect for Persons is concerned mostly with informed consent and the statistician's role may be limited to presenting the existing data in an understandable way. The Justice requirement that benefits and burdens of research be distributed fairly is a sampling problem with statistical solutions. Beneficence demands that possible benefits of the study are reasonable compared to the risks. The statistician's role is to ensure that the study is properly designed to answer the question in a way that avoids undue risks to the subjects. Design considerations include collection of appropriate data, proper statistical tests, and adequate sample size.

THURSDAY AFTERNOON

Emerald

1:00 Divisional Poster Presentation

I Disease Studies and Clinical Evaluations

THE EFFECTS OF STATIC STRETCHING AND VARIED LOCATION OF ULTRASOUND APPLICATION ON GASTROCNEMIUS FLEXIBILITY

William Woodall*, Paula Stubbs, Jason Greer, Amanda Ehrhardt, Janet Greer, Robert Stallings, Brandi White, and Mark D. Weber, University of Mississippi Medical Center, Jackson, MS 39216

This study's purpose was to determine how the location of application of ultrasound impacts the gaining of muscle flexibility when static stretching is performed. Both legs of 20 subjects were used in the study. No subject had more than 20[degrees] of active dorsiflexion at their ankle. Each leg in the study was randomly assigned to one of three groups: (1) static stretch only, (2) ultrasound to muscle belly of gastroc, followed by static stretch, (3) ultrasound to Achilles tendon, followed by static stretch. The same examiner performed the measurement using the same specially designed measuring apparatus for accuracy. Using a repeated measures ANOVA it was found that dorsiflexion ROM improved significantly for both right (p < 0.001) and left ankles (p < 0.001) in all three groups over time. There was no difference between treatment groups (p = 0.8 for left and p = 0.6 for right) in relation to the amount of ROM gained. Much of the literature that reports changes in ROM measures following a program of heat and stretch has performed the heat and stretch simultaneously. This is often not the strategy utilized in the clinic, where often the stretching is performed following heat application. While one study is not enough to call into question a generally accepted physical therapy intervention, it does suggest that continued research involving specific interventions should be continued in this area.

IT BAND Z-PLASTY FOR RECALTRICANT BURSITIS AND TENDONITIS

Phani Tumu, Audrey K. Tsao*, and Charity Peacock, University of Mississippi Medical Center, Jackson, MS 39216

Trochanteric bursitis and IT tendinitis classically presents with tenderness over the greater trochanter and increased discomfort on active abduction and passive adduction. Patients typically have problems with walking long distances, climbing stairs, or simple weightbearing in extreme cases. Treatment is centered on physical therapy, use of ice, heat, ultrasound, NSAIDs, and steroid injections into the affected area. Effective non-operative treatment with this approach is better than 90%. A minority of patients are refractory and unable to tolerate the pain and limitations of this condition requiring surgical intervention. In this study, twelve patients (three males, nine females) refractory to non-operative care were identified. Each patient presented with increasing hip pain with a duration of months to years. Patients were treated with iliotibial band z-plasty for their refractory symptoms. The preoperative Harris Hip Score averaged 54.4 (range 31 to 73). Two out of twelve patients were lost to follow-up, leaving ten patients available for clinic or telephone follow-up. The average post-operative follow-up was 46.9 months (range 10 to 81). The average post-operative Harris Hip Score was 73(range 54 to 96). The majority of patients were satisfied with the results of the procedure, and only one patient reported needing a cane post-operatively. One patient had continued pain post-operatively due to a subcutaneous hematoma at the incision site. Another patient had developed sciatica on the non-operative side approximately three years after the procedure. This patient had reported significant improvement in her operated hip up until the sciatica had developed. These two patients, however, were able to resume everyday activities. One patient had passed away as a result of a gastrointestinal pathology unrelated to surgery.

THE EFFECT OF VISION ON FUNCTIONAL REACH SCORES IN NORMAL YOUNG, NORMAL AND BALANCE IMPAIRED OLDER SUBJECTS

Min Huang (1)*, Ruth M. Burgess (1), Mark D. Weber (1), Kimberly N. Shirley (2), A. Ken Causey (3), Stephanie R. Gorrell (3), Kim L. Bartusek (3), and Neva F. Greenwald (1), (1) University of Mississippi Medical Center, Jackson, MS 39216; (2) Rehab Plus of Mississippi, Jackson, MS 39211; and (3) Mississippi Baptist Medical Center, Jackson, Clinton, MS 39056

The purpose of this study was to determine the effect of altering vision on functional reach (FR) scores in normal young, normal and balance impaired older subjects. Subjects participating in the study were in

two age group categories: The younger group was 20-35 years of age, and the older group was 60-80 years of age. For the study, subjects were divided into four groups: normal young (NY) (n = 61), normal non-retired old (NNRO) (n = 30), community retired old (CRO) (n = 30) and balanced impaired old (BIO) (n = 15). After being instructed, each subject performed FR under 2 different visual conditions: eyes open (EO) and vision blurred (VB). All data were analyzed using repeated ANOVA, and are expressed as mean [+ or -] S.D. Alpha level of 0.05 was used to test for significance. The mean FR scores in NY group with EO and VB were 13.7[+ or -]2.7 and 13.0[+ or -]2.8 inches, respectively. The FR in NNRO group with EO and VB were 11.1[+ or -]2.3 and 10.3[+ or -]2.7 inches, respectively. The mean FR scores in CRO group were 8.9[+ or -]2.3 and 8.5[+ or -]2.9 inches, respectively. The mean FR scores in BIO group with EO and VB were 10.1[+ or -]2.8 and 9.7[+ or -]2.4 inches, respectively. Blurred vision significantly decreased FR scores in all groups (p < 0.01). The FR scores between the groups were also significantly different (p < 0.01). FR scores declined when vision was impaired and younger subjects performed better than older subjects did under either visual condition. There were no significant differences in FR scores between the subjects in CRO and BIO groups. These results suggest that FR test alone is inadequate to predict the severity of balance impairment in older subjects with visual impairment.

INTERFERENTIAL CURRENT AT ACUPUNCTURE POINTS FOR AN OLDER ADULT WITH KNEE PAIN: A CASE REPORT

Min Huang*, Felix I. Adah, and Neva F. Greenwald, University of Mississippi Medical Center, Jackson, MS 39216

It has been demonstrated that acupuncture is an effective tool in the management of pain under a variety of conditions. The purpose of this case report is to describe the effectiveness of interferential current (IFC) at acupuncture points around the knee for chronic pain caused by osteoarthritis. The patient was a 92-year old woman who was blind. After receiving physical therapy in the hospital for 3 months following right total hip replacement (THR), the patient was able to ambulate 20 feet with a standard walker and moderate assistance. The patient was referred to a skilled nursing facility for further rehabilitation. Upon admission to the nursing home, the patient complained of her right knee collapsing during ambulation due to severe knee pain. To facilitate a decrease in pain, the patient received IFC stimulation with surface electrodes placed on acupuncture points around her right knee (eyes of knee, ST-34, and SP-10) plus ice 5 x/week for 8 weeks in addition to the standard physical therapy treatment interventions. At the end of 8 weeks, the patient subjectively rated pain 5/10 as compared to 10/10 initially. Functional Independence Measure (FIM) score in sit << stand, and wheelchair << bed transfer increased from 3 to 7 and ambulation distance increased from 20 feet with moderate + 1 assist to 240 feet with standby + 1 assist. This case report shows that IFC stimulation at the acupuncture points around the knee may be an effective tool in reducing chronic knee pain and enhancing functional mobility for patients who have osteoarthritis.

HOMOCYSTEINE LEVELS AS AN INDICATOR FOR CVD: AGE AND GENDER DEPENDENCY AMONG MISSISSIPPI POPULATION

Olivia Rahaim Henry*, Stacy Hull Vance, and Hamed Benghuzzi, University of Mississippi Medical Center, Jackson, MS 39216

Recently, homocysteine (HC) levels have shown to be an excellent biomarker for the development of cardiovascular disease (CVD). The specific objectives of this study were (i) to evaluate the age dependency of HC as a CVD marker, (ii) to study the gender prevalence in regard to HC levels, and (iii) to investigate the incidence of abnormal HC levels among random patients reported to the University of Mississippi Medical Center. A total of 95 volunteers (68 females, 27 males) were included in this investigation. The average ages of patients were 41 [+ or -] 16 for females and 40 [+ or -] 15 for males. Upon construction of database, the patients were subdivided into four categories ([less than or equal to]20, 21-40, 41-60, and [greater than or equal to]61) for each gender. The abnormal levels were recorded as low (<5 [micro]M/L) or high (>18 [micro]M/L). Data analysis was performed using standard computer software (Sigma Stat, Slide Write). The results obtained from this study suggest: (1) Regardless of gender, the HC levels were found to be directly proportional to age (7.53 [+ or -] 2.18, 8.23 [+ or -] 3.20, 10.00 [+ or -] 1.87, 14.44 [+ or -] 4.77 [micro]M/L for females and 8.53 [+ or -] 3.26, 8.98 [+ or -] 3.08, 10.52 [+ or -] 2.56, 11.80 [+ or -] 4.67 [micro]M/L for males, (2) HC levels among the high risk population demonstrated a slight increase in the females (27.80 [+ or -] 12.79 [micro]M/L) compared to the males (22.95 [+ or -] 1.97 [micro]M/L). In conclusion, data obtained from this investigation provided the literature with more insights (age and gender dependency) regarding the use of HC as a diagnostic tool for CVD. *Ph.D. Student, CHS Graduate Program, UMC

PERCEPTIONS OF THE QUALITY OF CARE PROVIDED BY ADVANCED PRACTICE NURSES IN AN ELEMENTARY SCHOOL-BASED HEALTH CENTER

Lisa A. Haynie*, Anne A. Norwood, Kathryn Kolar, and Theresa M. Doddato, University of Mississippi School of Nursing, Jackson, MS 39216

Development and management of School Based Health Center (SBHC's) by advanced practice nurses provide a quality, cost-effective, and caring method of delivering primary health care services for young children within a school setting. The purpose of this study was to evaluate the perceptions of effectiveness of the SBHC and satisfaction of care that students received by teachers and parents. The concepts of interest for this study were parents' and teachers' perceptions of the effectiveness of a nurse-managed SBHC and the delivery of care by advanced practice nurses, and their satisfaction with the services that the student(s) received. Data from the surveys were analyzed using descriptive statistics to obtain information on the impact of SBHC on parental work and student absenteeism, health care access for students, decreases in visits to other health care providers, and the use of emergency rooms. Findings: The majority of parental respondents gave permission for participation in the SBHC. More than one-half indicated that health care services that their child received at the SBHC were effective in decreasing the hours/days parents missed at work. A similar percentage responded that the number of emergency room visits decreased. Parents indicated satisfaction with the SBHC, as did the teachers in an overwhelming majority. Conclusions: Findings indicated that both parents and teachers were very satisfied wit the care that had been provided for students and believed that the SBHC was effective in the services provided.

ENDODONTIC DIAGNOSTIC AND TREAMENT DATA FROM AN UNDERGRADUATE DENTAL SCHOOL CLINIC

J.C. Rutz*, B.O. Gilbert, R.S. Gatewood, and C.F. Streckfus, University of Mississippi Medical Center, Jackson, MS 39216

The purpose of this study was to review data collected on endodontic patients at the University of Mississippi School of Dentistry. Methods: The following information was collected and summarized: (1) age, (2) sex, (3) tooth treated, (4) number of canals in tooth treated, (5) history of pain, (6) preoperative pulpal and periapical diagnoses. All data were analyzed using a statistical SPSS software package. Results: (1) the most frequent age group was 40-49 (27.7%), (2) 61.1% were females; 38.9% males, (3) the most frequently treated tooth was the mandibular first molar, (4) analysis of number of canals revealed: 9.5% of mandibular incisors with 2 canals; 59% of maxillary second premolars with 2 canals; 47.8% of maxillary first molars with 4 canals; 34% of mandibular first molars with 4 canals, (5) sixty-five percent of patients reported prior incidence of pain, (6) the most frequent preoperative pulpal diagnoses were irreversible pulpitis (37.2%) and non-vital pulp (36.7%). The most frequent preoperative periapical diagnoses were normal apex (41%) and chronic apical periodontitis (23%). Conclusion: Analysis of the results of this project give valuable information as to the scope of endodontics practiced in this undergraduate endodontic clinic and help characterize clinical experiences available to the students.

BREAST CANCER CONTROL PRACTICES OF PRIMARY CARE PROVIDERS IN MISSISSIPPI

Amal Khoury, Nedra Lisovicz, Mandy Avis*, and Deonna Allen, University of Southern Mississippi, Hattiesburg, MS 39406

Objective: Breast cancer is a leading cause of death in American women. However, many women in the Deep South do not participate in breast cancer screening and clinical trials. Providers have a key role in educating and referring women. This study identified knowledge, attitudes, and practices of providers in the Deep South toward breast cancer screening and clinical trials. Method: Qualitative research was conducted in 2002 involving semi-structured interviews with 34 key primary care providers, including physicians and nurse practitioners, in the rural Mississippi Delta and urban areas of the state. The interviews were audiotaped, and the tapes were transcribed. Data analysis identified cross-cutting issues and themes. Results: Although providers were overall knowledgeable about breast health and concerned about the welfare of their patients, their implementation of breast cancer screening guidelines and knowledge of clinical trials, mammography costs, and programs that cover screening and treatment was limited. Providers faced barriers to referring women to screening and clinical trials, including time constraints, patient factors, and other issues. Conclusion: Providers participation in screening programs and clinical trials could be improved by: (1) educating providers about risk factors, screening guidelines, and programs such as the Breast and Cervical Cancer Early Detection Program, (2) instituting systems to monitor missed screening appointments, (3) addressing time barriers, and (4) addressing the fragmentation in women's health care.

MOTOR FUNCTIONS BUT NOT ACQUISITION AND RETENTION OF ACTIVE AVOIDANCE RESPONSE ARE IMPAIRED IN METHYL PARATHION-TOLERANT RATS

Tingting Sun*, Ian A. Paul, and Ing Kang Ho, University of Mississippi Medical Center, Jackson, MS 39216

Previous work showed that repeated exposure to methyl parathion (MP) caused prolonged inhibition of acetylcholinesterase (AChE) activity, and down-regulation of muscarinic receptor subtypes in the brain regions, including frontal cortex, striatum, hippocampus, and thalamus. In the present study, we found that these changes were not accompanied by changes in drinking and eating behaviors, but by delayed body weight gain and progressive deterioration of motor function. Further, the neurochemical changes did not cause impairment in the performance of acquisition and retention in active avoidance responding. Our studies indicate that prolonged inhibition of AChE activity and down-regulation of central nervous muscarinic receptors produces selective behavioral effects. Motor function appears more sensitive to the effects of chronic exposure to methyl parathion than learning and memory behavior. This is consistent with our previous finding that the most extensive down-regulation of muscarinic receptors was found in the striatum (-30%), the area believed to relate to motor function whereas hippocampus is relatively resistant to this adaptation mechanism (-10%).

COMPARING INCIDENCES OF BREAST CANCER AND SURVIVAL RATES IN AFRICAN AMERICAN AND CAUCASIAN WOMEN

Brittany Pinkney* and Raymond Wynn, Mississippi Gulf Coast Community College, Jackson County Campus, National Aeronautics and Space Administration, and the Singing River Oncology Center, Pascagoula, MS 39563

The purpose of this literature review is to further document the differences in survival rates of African American and Caucasian women with breast cancer. The incidence of breast cancer is lower among African American women, but African American women have a higher rate of breast cancer deaths than Caucasian women. Breast cancer is the leading cause of cancer death among African American women. In any one year, 95 out of every 100,000 African American women are diagnosed with breast cancer. In comparison, 112 out of every 100,000 Caucasian women are diagnosed with breast cancer. This presentation will reveal several documented reasons for the lower survival rate in African American women such as diagnosis, poverty, under-treatment, and the nature of cancer.

II Technology, Genes, and Bacterial Genetics

EXTRACTION OF TOTAL RNA FROM LIVER AND MAMMARY TISSUES OF MICE TREATED WITH ALCOHOL AND/OR P-NITROPHENOL

Ameka Catchings (1,2)*, Joseph A. Cameron (1), and Jacqueline Stevens (1), (1) Jackson State University, Jackson, MS 39217, and (2) Hinds Community College, Jackson, MS 39154

The purpose of this research is to determine whether or not alcohol is a risk factors in the development of breast cancer. Our hypothesis is that alcohol-induced modulations of mRNA expression leads to initiation of cancer phenotype and that the inducers of that modulation are breast cancer risk factors. Modest evidence implicates alcohol as a breast cancer risk factor and alcohol specifically induces cytochrome P450-2E1 (CYP2E1). We expected that alcohol would induce CYP2E1 in MCF-7 cells, but preliminary data in our laboratory indicates that ethanol treatment also induces CYP3A4. Total RNA isolation and probing for specific CYP2E1 and CYP3A4 mRNA expression after treatment of Balb-c mice with alcohol and other CYP inducers will provide a model system of metabolic activation by alcohol and/or co-treatment and serve as an biomarker of breast cancer risk, placing women who drink alcohol at higher risk of breast cancer development than unexposed women. We demonstrate varying levels of expression of total cellular RNA isolated from mouse liver and mammary samples following treatment with our test inducers. Total RNA was denatured with 7% formaldehyde and then was separated on 1% agarose gels. The gels were stained with ethidium bromide and photographed under UV light. (Supported in part by NIGMS R25 GM50117).

EFFECTS OF SUSTAINED AND CONVENTIONAL DELIVERY OF ESTROGEN ON THE PROLIFERATION OF SiHa CERVICAL CARCINOMA CELLS IN CULTURE

Joyce Brewer*, Hamed Benghuzzi, and Michelle Tucci, University of Mississippi Medical Center, Jackson, MS 39216

The objectives of this investigation were (i) to evaluate the effect of estrogen on SiHa cervical cells delivered by TCPL delivery system versus conventional administration as a model for cervical cancer intervention, and (ii) to determine the estrogen dose effect on proliferation of these cells in culture. Estrogen was delivered to these cells for three time periods, 24, 48, and 72 hours both by conventional administration [direct administration] and by drug delivery system using TCPL ceramic capsules. Three Dosages of estrogen were selected for this study [2, 10, and 100 pg/mL]. Ceramic capsules were prepared in our laboratory using standard procedures [release profile 2, 10, and 100 pg/ml]. Capsules were then gas sterilized prior to being placed in the wells with cells. Data collected from this study indicated that, regardless of the route of administration, estrogen exposure did not induce major cellular injury at all concentrations. Initially, there were various responses to estrogen among all groups compared to control. At 72-hour phase all experimental groups induced greater proliferation rate in the conventional delivery than in the groups exposed to sustained estrogen delivery. Low and medium doses conventionally added to the cells resulted in a decrease in cell number at 48-hour phase compared to the initial response (24-hour phase). With the use of medium and high sustained delivery of estrogen rates, the data revealed that proliferation rate is directly proportional to duration of delivery compared to conventional means. The data suggest that the route of administration of estrogen is an instrumental tool in the prognosis of cervical cancer. *Ph.D. Student, CHS Graduate Program, UMC

ANTIBIOTIC RESISTANCE AMONG GRAM NEGATIVE BACTERIA IN A HOSPITAL ICU

G. Ballard (1)*, H. White (2), Susan Bender (1), Cindy Cook (1), and Donna Sullivan (2), (1) Base Pair Program/HHMI and Jackson Public Schools, and (2) University of Mississippi Medical Center, Jackson, MS 39216

Susceptibility data derived from surveillance studies can gauge emerging resistance problems. Several recent studies have shown increasing incidence of antibiotic resistance concomitant with increased use of these drugs. In this survey, 100 consecutive gram-negative aerobic isolates recovered from ICU patients were identified to species level. Susceptibility tests were performed with a standardized microtiter minimal inhibitory concentration (MIC) panel (Dade International MicroScan, CA). Testing was conducted according to the National Committee for Clinical Laboratory Standards and MICs validated with ATCC test strains. MICs were determined for 16 antibiotics. The most commonly isolated pathogens were Enterobacter cloacae (20%), Pseudomonas aeruginosa (16%), Klebsiella pneumoniae (14%), Serratia marcescens (11%), and Acinetobacter baumannii (10%). National surveys report similar percentages for E. cloacae, P. aeruginosa, and K. pneumoniae. Overall, the activity of ciprofloxacin to the most common isolates in this study was 91.4%, above the national average of 76% in 2000. P. aeruginosa susceptibility to ceftazidime was 56% vs 80%, cefotaxime 100% vs. 13%, and ciprofloxacin was 81% vs. 76% in is study compared to national data from 2000. Some of these differences may reflect differences in prescribing patterns. This study represents the first series in a five year ongoing study.

FRACTURE HEALING IN RESPONSE TO CONTINUOUS SUSTAINED DELIVERY OF DEMINERALIZED BONE MATRIX PROTEINS AND TOBRAMYCIN

Chris Galjour*, Sergey Dzugan, Matt Graves, Hamed Benghuzzi, George Russell, Michelle Tucci, and Audrey K. Tsao, University of Mississippi Medical Center, Jackson, MS 39216

Demineralized bone matrix (DBX) is an allogenous, bioabsorbable material that has long been used for its osteo-inductive and osteoconductive properties. A significant complication experienced by physicians who perform bone defect filling surgery is the risk of subsequent bacterial infections and the inefficiency of oral antibiotics to provide adequate prophylaxis against microorganisms, especially Staphylcoccus aureus, Pseudomonas aeruginosa, and Staphylcoccus epidermidis In order to deliver both demineralized bone matrix and an efficient antibiotic at high local concentrations without deleterious systemic effects, a ceramic sustained delivery system was implanted and monitored over the course of 30 days for bone regeneration, infection, and systemic effects. Twenty-five Sprague Dawley albino male rats were used in the experiment. They were randomly divided into five equal groups. Animals in group 1 were used as control, in group 2 had a created 5mm defect, in group 3 defect with the antibiotic alone, in group 4 had a created defect plus DBX, and group 5 had a created defect plus antibiotic and DBX. At 30 days post-implantation, the experimental animals showed no significant difference in weight when compared to the control and sham animals. X-rays taken at this time showed the experimental femurs to be totally healed and virtually indistinguishable from control. Initial dissections revealed that the implants were accepted by the hosts as shown by the fibrous, vascularized sheath that surrounded the femurs and capsules. The implants were found to be in close contact with the cancellous bone and none of the sheaths showed signs of infection. Macroscopically, no defect could be detected in the experimental animals, while little regeneration was observed in the femurs of the sham animals. *Medical student (second year), UMC

THE EFFECTS OF DEMINERALIZED BONE MATRIX ON BONE CELL FORMATION

Tabitha Hardy (1)*, Hamed Benghuzzi (2), George Russell (2), Michelle Tucci (2), and Joseph A. Cameron (1), (1) Jackson State University, Jackson, MS 39217, and (2)University of Mississippi Medical Center, Jackson, MS 39216

Demineralized bone matrix protein (DBM) has been used to reconstruct bone. The studies have shown that DBM induces new bone formation when it is implanted subcutaneously or intramuscularly. The effects of DBM at the cellular level have not been clearly defined. MG-63 cells were plated onto 24 well tissue culture plates at a density of 1X [10.sup.5]. Cells were exposed to 30% or 100% DBM proteins for periods of 24, 48 and 72 hours and compared with untreated controls. After each incubation period, cell morphology, cell damage, cell number, and protein concentrations were determined. Preliminary results indicate a significant increase in cell number at 24 hours in both DBM treated groups. The increase was still evident after 48 hours in the 100% DBM treated cells. Cellular protein levels at 24 and 48 hours for the DBM treated cells were also increased when compared to control. The treatments did not cause an insult to the integrity of the cell membrane as evidence by lack of difference in the maliondialdehyde levels. Morphological differences were observed in treated cells after 24 and 48 hours. The cells treated with both concentrations of DBM had structural differences as well as an increase in nuclear swelling and evidence of prominent nucleoli. The early increases in cell number and cellular protein content indicate the DBM is effective in stimulating cell growth. Closer evaluation of the morphology especially the changes occurring at the nuclear level need to be addressed. (Supported in part by NIGMS R256M067592) *Graduate Student, Jackson State University

THE EFFECTS OF SUSTAINED DELIVERY OF THYMOQUINONE ON BONE HEALING OF MALE RATS

Philemon K. Kirui (1)*, Hamed Benghuzzi (1), Joseph A. Cameron (1), and Michelle Tucci (2), (1) Jackson State University, Jackson, MS 39217, and (2) University of Mississippi Medical Center, Jackson, MS 39216

The use of natural products as an alternative to conventional treatment of various diseases is on the rise. Nigella sativa, a natural herb has been used for many acute as well as chronic conditions. The objectives of this study were (1) to successfully deliver the active component of black seed, thymoquinone (TMQ), at sustained level using a tricalcium phosphate delivery system (TCPL); (2) to evaluate the effects TMQ on bone healing (femur) in adult male rats. Fifteen animals were randomly divided into three equal groups. Group I animals served as controls, those in group II served as sham, while group III served as TMQ experimental group (0.2 g TCPL). Blood, x-rays and body weights were collected and recorded weekly. Metabolic biomarkers were also evaluated weekly. At the end of 30 days, animals were sacrificed and vital as well reproductive organs were collected and analyzed morphometrically. The results revealed the following: (i) gross anatomical observation indicated an increase in the healing pattern of animals in the TMQ group compared to those in group II (sham); (ii) no significant differences where detected in the levels of cholesterol, proteins, maliondialdehyde and alkaline phosphatase in all groups; and (iii) no significant differences were observed in the weights of vital as well as reproductive organs in all the groups. In conclusion, it appears that TMQ can enhance bone healing with little or no side effects on major vital and reproductive organs.

COMPARISON OF RADIOGRAPHIC TECHNIQUES TO OPTIMIZE IMAGE ANALYSIS FOR EVALUATING TRAUMATIZED BONE USING A RAT AS A MODEL

Rishi Roy*, Michelle Tucci, Ramesh Patel, George Russell, Audrey K. Tsao, and Hamed Benghuzzi, University of Mississippi Medical Center, Jackson, MS 39216

The search for an ideal, safe, efficient and cost-effective protocol to assess the daily progression in osteogenesis is in the rise. The specific objective of this investigation was to employ various combinations of radiographic techniques that may optimize image analysis in orthopedics setting. Surgically induced femoral fractures immobilized with metallic plates and screws of adult male laboratory rats were used as a model (n = 50). The techniques were utilized to study excised specimens with the induced fractures. Specifically, conventional radiographic methods as well as those that are used for specimen radiography in clinical methods were utilized. For routine radiography, a portable x-ray machine was used and a Faxitron was used for specimen radiography. Exposure factors (kv, time), type of film, and magnification was varied in experiments using Faxitron. Two clinical film mammography machines were employed for all selected excised femurs. To obtain object magnification the distance between the film and the object was varied to bring the object closer to the x-ray tube. An aluminum wedge [standard density of 1.98 gm/c[m.sup.3]] was included in the radiographs along with the specimens to obtain density measurements. The criteria of evaluation for the images included: (a) visualization of the immobilizing hardware; (b) evaluation of the fracture site; (c) evaluation of the rest of the bone specimen, and (d) evaluation of the soft tissues. Images obtained with the portable radiographic equipment showed the least amount of spatial and contrast resolution while the results obtained with Faxitron and the mammographic machines showed much higher contrast and spatial resolution. Optimal soft-tissue resolution was difficult to obtain with any of the techniques.

MODULATION OF CYTOCHROME P450 PROTEIN AND GENOTYPIC EXPRESSION IN MOUSE LIVER AND MAMMARY TISSUES IN RESPONSE TO TREATMENT WITH ALCOHOL AND/OR P-NITROPHENOL

Tyranna Lacey (1,2)*, Joseph A. Cameron (1), and Carolyn Howard (1), (1) Jackson State University, Jackson, MS 39217, and (2) Hinds Community College, Raymond, MS 39154

We propose that multiple exposures increase the probability of breast cancer development by modulating cytochrome P450 (P450) protein synthesis and redirecting the metabolism of P450 substrates to more toxic pathways in breast cells. This lead us to question whether or not co-treatment with alcohol and p-NP would lead to complex metabolic interactions of biotransformation products in breast cells and subsequent consequences, different than those due to treatment with each compound alone and different than those induced in liver. Modulation in CYP ratio is a critical determinant of breast cancer risk, and therefore, this system represents a critical focal point in our inquiry. We compared P450 protein expression profiles obtained following alcohol/p-NP co-treatment to those observed following treatment with each compound alone in mouse mammary and liver samples. This screen for synergistic, agonistic or antagonistic effects caused by combined treatment revealed that treatment with ethanol alone leads CYP3A4 protein and mRNA expression compared to untreated samples and samples treated with p-NP alone, and combined exposure. This study represents an examination of ethanol as a potential breast cancer risk factors and the effects of combined exposures, which is relevant toward understanding tissue-specific biotransformation events and perhaps drug-drug interactions. (Supported in part by NIGMS R25 GM50117).

THE EFFECTS OF SUSTAINED DELIVERY OF GLUCOCORTICOIDS ON THE KIDNEY OF FEMALE RATS

Russell F. Lyon*, Stevie D. Adams, Hamed Benghuzzi, and Michelle Tucci, University of Mississippi Medical Center, Jackson, MS 39216

Chronic increased levels of glucocorticoids may impair renal function. The use of antioxidants has been shown to reduce renal tubular injury in cases of increased glucocorticoid administration in vitro. The objectives of this study were: (1) to establish an animal model of increased glucocorticoid levels by sustained delivery and (2) to determine if sustained delivery of selenomethionine in combination with glucocorticiods can protect kidney tubular structures. Sixteen female rats were divided into 4 equal groups (control and 3 experimental groups implanted with tricalcium phosphate lysine drug delivery systems (TCPL) charged with either 50 mg selenomethionine (SE), 50 mg cortisol (C), or 50 mg of both C and SE). At the end of 30 days, the rats were sacrificed and both kidneys were removed for histological analysis. Quantitative analysis was performed on serum calcium levels, body weights and kidneys weights in all groups. Kidneys slides were evaluated for changes in kidney morphology namely change in area and width. Sustained release of C resulted in a significant reduction of glomerular area. SE administration alone did not alter kidney structure and was not able to protect against the changes caused by C when co-administered. C, Se and C + SE caused a significant (p < 0.05) reduction in serum calcium levels compared with control. The reduction may be in part to changes in calcium-filtered load, changes in glomerular filtration rates or interference of calcium absorption from the gut. In conclusion, high levels of cortisol will modify kidney structure and possibly alter blood pressure. *Allied Health Undergraduate, UMC

EFFECTS OF ANTIOXIDANTS ON ULTRAVIOLET LIGHT INDUCED MORPHOLOGICAL CELLULAR DAMAGE IN MRC-5 CELLS

Milledge Smalls (1,2,3)*, Jarrett Smith (1,2,3)*, Larkin Mitchell (1), Joseph A. Cameron (2), Michelle Tucci (1), and Hamed Benghuzzi (1), (1) University of Mississippi Medical Center, Jackson, MS 39216; (2) Jackson State University, Jackson, MS 39217; and (3) Hinds Community College, Raymond, MS 39154

Recent studies have indicated the use of antioxidants offers protection against various types of cancers. The goal of this study was to use an established tissue culture model (MRC-5 cells), to assess cellular damage after exposure to ultraviolet light (UVL), and to determine if vitamin E addition after exposure can alleviate cellular damage. The specific aims were to evaluate the morphological responses of MRC-5 fibroblast cells exposed to UVL followed by vitamin E supplementation. MRC-5 cells were divided into three groups. Group I cells served as control, group II cells were exposed to UVL for 60 minutes, and group III cells were exposed to UVL for 60 minutes followed by addition of vitamin E (100 [micro]L of 1000 IU). All groups were incubated for periods of 24, 48, and 72 hours. After each incubation period cellular damage was assessed using cell number determination and by determination of cellular and supernatant MDA levels. Vitamin E post exposure provided a protective role by demonstrating lower levels of MDA (p < 0.05) as well as normal structure appearance. (Supported in part by NIGMS R25 GM50117).

DEGRADATION OF VITAMIN E IN THE PRESENCE OF ULTRA-VIOLET LIGHT

Larkin Mitchell (1)*, Hamed Benghuzzi (2), Michelle Tucci (2), and James Hughes (2), (1) Millsaps College, Jackson, MS 39210, and (2)University of Mississippi Medical Center, Jackson, MS 39216

Vitamin E is a lipid-soluble antioxidant in the biological membrane that contributes to membrane stability by protecting critical cellular structures against damage from oxygen free radicals and reactive products of lipid peroxidation. Vitamin E is an important anti-oxidant. It acts as a free radical scavenger to prevent the byproducts of chemical-cell interaction that cause cell damage. Studies have reported vitamin E to protect against some of the toxicities of ionizing radiation. One source of ionizing radiation is UV light, and numerous products containing vitamin E are sold to block UV light. The goal of this investigation was to determine the effect of prolonged UV light exposure on vitamin E. Our hypothesis was that UV light will degrade vitamin E and diminish it's antioxidant capacity. To test our hypothesis, vitamin E was prepared at a concentration of 0.075 M and exposed to UV light for a duration of 0, 5, 15, 30, and 60 minutes. After each exposure period, vitamin E was determined spectrophotometrically and compared with the non-exposed sample. Decreases in peak absorbance were noted as early as 5 minutes post exposure and continued to decrease with each exposure time. Measurements of thiobarbituric reactive species were also higher in the UV exposed samples indicating a decrease in antioxidant capacity. In conclusion, UV light causes degradation of vitamin E rendering it an ineffective antioxidant. Undergraduate Student, Millsaps College

PATHOPHYSIOLOGICAL RESPONSE OF MRC-5 FIBROBLAST CELLS TO DEMINERALIZED BONE MATRIX PROTEINS

Anne Sory*, Hamed Benghuzzi, Michelle Tucci, and Zelma Cason, University of Mississippi Medical Center, Jackson, MS 39216

Demineralized bone matrix proteins (DMB) have been used recently as an osteogenic agent that promotes fracture healing. We hypothesize that increased osseous formation by DMB leads to less adverse effects such as non-union and underlying infection. However, the effect of varying concentrations of (DMB) on the cells of the surrounding soft tissue has not been investigated. The specific goal of this study was to evaluate the biochemical and morphological changes in MRC-5 fibroblast cells treated with either 30% DBM proteins or 100% DBM proteins [24, 48, 72 hours incubation]. Cell growth in both 100% and 30% treatments showed a significant increase after 24 hours (p < 0.05). However at 48 and 72 hours there was not a substantial difference in cell numbers when compared with control. Cellular damage was not evident as indicated by the MDA levels for the treated groups at 24 and 72 hours. There was a slight but insignificant increase in MDA levels compared to the control at 48 hours. Cellular proteins were reduced in both DBM treated groups for the duration of the experiment. Morphologically, significant changes were evident as early as 24 hours. The 100% DBM treated cells also exhibited a prominent nucleoli and moderate coarse chromatin at 48 hours. Cellular rounding and cytolysis were also observed in both treated groups. Overall, DBM proteins can alter fibroblast cellular metabolism and cellular morphology. The results show a need for further characterization of these compounds on soft tissues.

ROLE OF THE N-TERMINAL DOMAIN IN SIGNAL TRANSDUCER AND ACTIVATOR OF TRANSCRIPTION PROTEIN 4 ACTIVATION

Tammi M. Taylor (1,2)*, Hua-Chen Chang (1), Joseph A. Cameron (3), and Mark H. Kaplan (1), (1)Indiana University School of Medicine, Indianapolis, IN 46202; (2) Walther Cancer Institute, Indianapolis, IN 46208; and (3) Jackson State University, Jackson, MS 39217

Signal Transducer and Activator of Transcription 4 (STAT4) mediates responses to Interleukin (IL)-12. Following IL-12 binding to its receptor, the Janus kinases, Jak2 and Tyk2, are activated to phosphorylate tyrosine residues on the IL-12 receptor. The STAT4 SH2 domain binds to IL-12R phosphotyrosines allowing STAT4 to become phosphorylated by the Janus kinases. STAT4 can then homodimerize, move to the nucleus, bind DNA and activate transcription. All STAT proteins have a conserved N-terminal domain shown to be important for phosphorylation and gene transcription. Our previous studies suggested that the N-terminal 51 amino acids of STAT4 are required to mediate the activation of STAT4. To determine precisely which region of the N-terminal domain is required for phosphorylation, we have generated a series of N-terminal truncated STAT4 mutants. Mutants that lack the amino terminal 10 amino acids are phosphorylated similarly to full-length STAT4. However, Jak2 less efficiently phosphorylated mutants lacking the 44, 51, or 75 amino-terminal amino acids than full-length STAT4. In contrast, full-length and mutant STAT4 proteins were capable of interacting with a phosphorylated receptor peptide. Thus, an N-terminal segment of STAT4 between amino acids 10 and 44 is required for efficient phosphorylation by Janus kinases but is dispensable for STAT4 recruitment to the IL-12R complex. (Supported in part by NIGMS R25 GM067592 and R01 AI45515).

EXPRESSION OF VARIANT PSPAs IN STREPTOCOCCUS PNEUMONIAE WU2

Anderson Lampton* and Larry S. McDaniel, Murrah High School, Jackson, MS 39216, and University of Mississippi Medical Center, Jackson, MS 39216

PspA is a surface protein present on all pneumococci, is required for full virulence, and is capable of eliciting protective immune responses. In mice, some pneumococcal strains are more virulent and difficult to protect against than other strains even though the mice are immunized with PspA fragments from the homologous strain. To examine the role the the genetic background on protection in which a particular PspA resides, a mutant of Streptococcus pneumoniae WU2 was constructed by allelic replacement of pspA/WU2 with pspA/BG9737. An erythromycin resistance cassette was inserted downstream of pspA in S. pneumoniae BG9739. This was achieved by insertion duplication mutagenesis in which BG9739 was transformed with a plasmid containing homology to the region of pspA encoding the C-terminus. Genomic DNA from the resultant strain was used to transform WU2. Any resultant transformants should have replaced pspA/WU2 with pspA/BG9739 by homologous recombination at two sites. The expression of PspA/BG9739 in the WU2 mutant was confirmed by Western blot and flow cytometry. This work was supported (in part) by the Howard Hughes Medical Institute.

THROMBOCYTOSIS ASSOCIATED WITH ANTI-FUNGAL THERAPY AND CANDIDEMIA

Angela D. Saathoff (1,2)*, Stephanie Elkins (1), Stanley W. Chapman (1), and John D. Cleary (1), (1) University of Mississippi Medical Center; Jackson, MS 39216, and (2) Mississippi College, Clinton, MS 39058

Purpose: Platelet counts >400,000/m[m.sup.3] worrisome due to the associated increase in incidence of gastrointestinal tract bleeding and stroke. Secondary, "Reactive", thrombocytosis has been attributed to bacterial infection and multiple pharmaceuticals. Our purpose is to report a case series of thrombocytosis observed in patients treated with anti-fungal therapy for disseminated candidiasis. Methods: Three groups (candidemia with anti-fungal therapy, candidemia without anti-fungal therapy and anti-fungal therapy without candidemia) containing patients treated at university medical center and over the age of 18, were evaluated for presence of anti-fungal therapy and candidemia in a retrospective manner. Platelet administration, pharmacologic or pathologic contributors to thrombocytosis, and other pertinent details related to thrombocytosis were also evaluated. Results: Reactive thrombocytosis was observed in 10% of candidemia patients treated with an azole. Within the sub-group affected by reactive thrombocytosis associated anti-fungal therapy and candidemia, life-threatening thrombolytic complications were common. Mean baseline platelet counts were 354,500/m[m.sup.3] with a mean peak (695,000/m[m.sup.3]) occurring 13 days after initiation of therapy. The maximum peak (1,056,000/m[m.sup.3]) was observed in a patient 14 days post-therapy initiation. Mean time to onset was 4 days and duration of thrombocytosis was 4 days post therapy. Conclusions: Thrombocytosis occurs during the treatment of candidemia. The causative agent (drug vs disease) will need to be elucidated by a larger epidemiological trial addressing the extrapolatibility of these results, addressing risk associated with this reaction and whether treatment is necessary.

ETHNICITY INFLUENCES THE DISTRIBUTION OF CYTOCHROME P450 3A4 GENE POLYMORPHISM

David T. Arrington, Jr. (1)*, Brenda D. Mangilog (2), Sebron Harrison (2), and D. Olga McDaniel (2), (1) Murrah High School, Jackson, MS 39216, and (2) University of Mississippi Medical Center, Jackson, MS 39216

Cytochrome P450 (CYP 450) is a heme-containing enzyme, which plays an important role in the metabolism of a variety of drugs including cyclosporin. CYP3A4 exhibits an inter-individual variation in expression levels. It is hypothesized that such variations in the level of expression are caused by polymorphisms within the regulatory element of the gene. Thus, we are interested in investigating such polymorphisms in different ethnic population. We have isolated DNA from blood samples obtained from 142 African Americans (AFAM) and 99 Caucasians (CAU). DNA was tested in a PCR assay using sequence specific nucleotide primers and Taq polymerase. The primers are representing either nucleic acid A at position -290 (natural type) or nucleic acid G at position -290. The assay was performed in a 9600 Thermal Cycler using a procedure common in the laboratory. The amplification product was tested in a 2% agarose gel electrophoresis. Genotypes were detected based on the presence or absence of the amplification product. Any ambiguity in genotype detection was resolved by gene cloning and sequencing. CYP3A4 G genotype was present with a higher frequency in AFAM individuals as compared with CAU (AFAM 80% VS 3%, p < 0.0001). Whereas, the homozygous AA was present in the majority of CAU (97%) but only 17% in AFAM. Furthermore, females in both groups carry fewer homozygous AA as compared with males (AFAM Male:63%, Female:37%, CAU Male:66%, Female:34%). Such polymorphism in CYP3A4 gene might affect on the levels of CYP3A4 protein production and subsequent variations in the metabolic rates and drug clearance.

III Animal Models and Chemistry

SYNTHESIS AND CHARACTERIZATION OF COOPER (II), NICKLE (II), ZINC (II) COMPLEXES OF SCHIFF BASE DERIVED FROM 1,2-DIAMINOBENZENE AND 2,5-DIHYDROXY BENZALDEHYDE

Camekio Garrett (1,2)*, Melissa K. Thomas (1,2), Thelma M. Brown (1,2), Joseph A. Cameron (1), and Ramaiyer Venkatraman (1), (1) Jackson State University, Jackson, MS 39217, and (2) Hinds Community College, Raymond, MS 39154

Transition metal complexes derived from Schiff bases are found to have biological, clinical and analytical applications. The activity of the metal complexes is enhanced compared to their parent organic molecule due to coordination with metal ions. In view of this perspective, we have synthesized complexes of copper (II), nickel (II), and zinc (II) using Schiff base derived by the 1:2 molar condensation of 1,2-Diamino benzene and 2,4-dihydroxy benzaldehyde in methanol. The Schiff base formed complexes with the above metal ions in a 1:1 molar ratio through oxygen, nitrogen (ONNO) atoms. The complexes were characterized on the bases of their molar conductance, IR, UV-Vis, and (1) HNMR spectral studies. IR, UV-Vis spectral data suggested that all the complexes are square planar in nature. The metal complexes exist as monomers and behave as non-electrolytes in DMSO solution. The data obtained by this study will be helpful in designing new ligands with potential applications as an analytical reagent for metal ions. Further, the metal complexes can be used as potential candidates in the study of photo-induced electron transfer interaction with DNA (Supported in part by NIGMS R25 GM50117).

THE SYNERGISTIC EFFECT OF ANTIOXIDANTS IN COMBINATION ON LNCAP PROSTATE CANCER CELL LINE

La'Toya Ross Richards*, Hamed Benghuzzi, and Michelle Tucci, University of Mississippi Medical Center, Jackson, MS 39216

The most common male cancer in the United States is prostate cancer. It has been postulated that prostate cancer will be the leading cause of cancer deaths in men for the next decade unless new interventions are discovered. The goal of this study was to develop treatment and prevention methods for the invasive behavior of prostatic mutations by exploring the synergistic effects of several antioxidants in a combined manner. The specific aims were to examine the viability and proliferation of human prostate cells in culture upon combined treatment of low and high doses of vitamin E, selenium, and lycopene. Various groups were treated in combination to observe synergistic effects. Data demonstrated that the combination group containing lycopene, vitamin E, and selenium tended to portray the greatest decrease in PSA levels. One suggestion could be that vitamin E has no effect on the making of PSA, but has a greater effect on the cell number, which resulted in decreased PSA values.

DELIVERY OF STATIN AND VANCOMYCIN BY MEANS OF A TRICALCIUM PHOSPHATE LYSINE (TCPL) CERAMIC DELIVERY SYSTEM IN A RAT FEMORAL DEFECT MODEL

Felix I. Adah (1)*, Hamed Benghuzzi (1), Michelle Tucci (1), George Russell (1), Audrey K. 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

The potential role of statin in bone metabolism as well the effects of the drug on the reproductive organs has not been fully elucidated. Therefore, the specific aim of this study was to develop a statin delivery system for treatment of a femoral defect, and evaluate the vital and reproductive organs 30 days post-op. The study consisted of 14 rats divided into three groups. Group I animals (n = 5) served as control. Animals in groups II (n = 5) and III group (n = 4) received a 5 mm femoral defect and implanted with TCPL delivery containing antibiotic or statin + antibiotic, respectively. Blood, x-rays and body weights were evaluated weekly. All the animals were euthanized at 30 days. The reproductive and vital organs were collected, weighed and histopathologically evaluated. The data showed no structural damage. The result showed that the TCPL ceramics were capable of delivering statin at a sustained level for four weeks. Body, vital and reproductive organ weights were not significantly different. Total testosterone, LH, and FSH levels of the animals implanted with statin loaded TCPL were not significantly different (p > 0.05) from the sham and control groups. The observation in this study suggests that TCPL delivery system can be used to release statin at a sustained level for long duration without negatively affecting the reproductive function. *Clinical Health Sciences Graduate Student, UMC

INHIBITION OF VAGINAL KERATINIZATION IN ADULT FEMALE RATS EXPOSED TO PHYSIOLOGICAL LEVELS OF ESTROGEN

Zelma Cason*, Hamed Benghuzzi, and Michelle Tucci, University of Mississippi Medical Center, Jackson, MS 39216

The short length of the estrous cycle in rats (4 days) allows for rapid observations of changes that occur during the reproductive cycle. The aim of the present work was to provide the literature with helpful considerations regarding the distribution of cornified cell determinations during proestrus, estrus, metestrus and diestrus in rats receiving estrogen. Eight female rats (four control and four experimental) were (R1-R8) used in this study. Cyclic activity at 2, 4, 8, 12, 24, 36, 48, 72 hours were determined. Briefly, 0.5 ml Hank's solution was placed within the vaginal canal for few seconds followed by aspiration. This mixture was then smeared onto microscopic slides and stained using a routine PAP and Diff Quick (DQ) staining methods. Data obtained revealed that the PAP stain proved to be a better staining technique than the DQ stain in both nuclear and cytoplasmic details. Histologically, keratinization of the vaginal epithelium appeared to be evident at the estrus phase (day 4) of a 4-day cycle (3 rats out 4). This keratinization process is dependent on the endogenous estradiol secreted between the evening of diestrus 2 (day 2) and that of proestrus (day 3). In the second stage of this experiment, the rats labeled R1-R4 were used as controls, whereas lab rats R5-R8 had estrogen administered (2 mg/ml) to them for three days. The results showed a significant increase in the proliferation of degenerative cells in the E treated rats compared to control animals. Inhibition of vaginal keratinization was obvious and this protocol can be used as a rapid and convenient in vivo investigational model for screening the effects of agents that have antikeratinizing activity. *Clinical Health Sciences Graduate Student, UMC

THE EFFECTS OF CONTINUOUS ADMINISTRATION OF CORTICOSTERONE AND SELENOMETHIONINE ON THE HEART MUSCLE OF MALE AND FEMALE RATS

Shontell Credit (1)*, Hamed Benghuzzi (1), Michelle Tucci (1), Ibrahim Farah (2), and Joseph A. Cameron (2), (1) University of Mississippi Medical Center, Jackson, MS 39216, and (2) Jackson State University, Jackson, MS 39217

A direct comparison of heart tissue between males and females under continuous cortisol stimulation has not been fully elucidated. The aim of this study was to determine the effects of continuous corticosterone and antioxidant administration on the male and female rat myocardial tissues. A total of 24 rats were divided into six equal groups control (male-c and female c), TCPL implanted groups containing corticosteroid (M cort and F cort), corticosteroid + selenomethionine (M combo and F combo). At the end of 4 weeks the animals were sacrificed and the heart tissue was collected, weighed, divided into apex, left ventricle and right ventricle then processed for routine histological evaluations. Bundle lengths, width and myocyte numbers were determined. The data showed MC and FC had the highest number of myocytes present in the apex, and both ventricles. M-cort animals had higher numbers of myocytes present in the ventricles compared with M-combo group and M-combo had more myocytes in the apex. Interestingly, the female rats had the exact opposite scenario. Measurements of bundle lengths were also different between female and male animals. In the apex of the female rats the lengths were in the following order comb>cort>control, whereas in the male the order was control>comb>cort. The right and left ventricle bundle fiber measurements were also different between the males and females M-comb>M-cort> MC and in the female they were reversed. This information collected in this experiment showed the male and female hearts respond differently to continuous administration of both stress hormone as well as anti-oxidants.

SUSTAINED RELEASE OF OP-1 AND ANTIBIOTICS IN TREATMENT OF FEMORAL DEFECTS IN MALE RATS

Sergey Dzugan*, Chris Galjour, Joe Conflitti, Hamed Benghuzzi, Michelle Tucci, George Russell, and Audrey K. Tsao, University of Mississippi Medical Center, Jackson, MS 39216

Osteogenic proteins (OP-1) promote osteoinduction. Formation of new bone growth in patients receiving OP-1 is not consistent, and is possibly due to the short half-life of the drug. In order to test the capacity of OP-1 to consistently produce bone in a fracture model a drug delivery system was developed to prolong the action of OP-1. Fifteen Sprague Dawley male rats were randomly divided into three equal groups Animals in group 1 served as control. Animals in groups 2-3 had a 5 mm defect created in the left femur using a number six dental burr and a drug delivery capsule (TCPL) containing either antibiotic alone (sham) or antibiotic +OP-1. Body weights, blood, and X-rays were taken weekly. Femurs and organs were harvested 30 days post-op, and processed for histomorphometry. Data was analyzed using ANOVA and significant difference between the groups was determined using Student Newman Kuels (p<0.05). The results showed complete bone healing in the OP-1 group with an evident callus formation. The osteoid tissue exhibited a proliferation of osteoblasts, which differentiated from the vascularized mesenchymal tissue. The complete bone healing using OP-1 was sharply contrasted sham treatment, where an obvious injury was still seen at 30 days. Histologically sham animals exhibited the early stage of repair with evidence of blood clotting and mesenchyme with early formation of osteoblasts. Overall, Op-1 delivered in a sustained manner for 30 days caused increased bone in a defect model.

EVALUATION OF ANTIOXIDANT COMPONENTS OF FRACTIONATED BLACK SEED

Nourelhoda Farah (1)*, Larkin Mitchell (2), Hamed Benghuzzi (1), and Michelle Tucci (1), (1) University of Mississippi Medical Center, Jackson, MS 39216, and (2) Millsaps College, Jackson, MS 39210

Nigella sativa has been used for thousands of years in the Middle East for allergies, asthma and for treating immune disorders. The seeds are known to contain essential amino acids, phytosterols, lipase, fatty acids, tannins, and thymoquinones. The essential oil of Nigella sativa seeds has antioxidant properties. The active constituents of Nigella sativa oils are thymoquinone, dithymoquinone, thymohydroquinone, thymol, carvacrol, and 4-terpineol. The purpose of the following study was to fractionate black seed and evaluate the fractions for anti-oxidant properties and compare with vitamin E a potent antioxidant. Black seeds were pulverized and twenty milligrams were extracted by either hot water or ethanol over a period of four days. The water-extracted fraction appeared yellow-white to light brown, while the alcohol fraction appeared clear to light brown. The water fraction most likely contained tannins, which are esters of sugar, and the alcohol fraction most likely contained thymoquinone. The fractions were evaluated for their ability to scavenge lipid radicals in an in vitro lipid peroxidation assay. The results show that each of the isolated fractions were able to scavenge approximately 50% of the generated radicals where as pure vitamin E was able to scavenge in a dose dependent fashion between 75 to 90% of the radicals generated. Pure thymoquinone was able to quench radical formation in a dose dependent fashion between 50-72%. Overall, the water and lipid soluble fractions of black seed contain potent antioxidants. *Allied Health Undergraduate Student, UMC

PROTEOMIC ANALYSIS OF TYROSINE PHOSPHORYLATED PROTEINS IN THE FRONTAL CORTEX OF BUTORPHANOL-DEPENDENT RATS

Seong-Youl Kim*, Nuannoi Chudapongse, and Ing Kang Ho, University of Mississippi Medical Center, Jackson, MS 39216

Butorphanol (17-cyclobutylmethyl-3,14-dihydroxymorphinan) tartrate (Stadol) is a mixed agonist-antagonist opioid analgesic agent that produces about five to seven times as potent as morphine in analgesic effects. The chronic use of butorphanol produces physical dependence in humans as well as in animals. Post-translational modifications such as phosphorylation play a very important role in developing dependence. The aim of this study is to determine tyrosine phosphorylated proteins in the frontal cortex of butorphanol-dependent rats using proteomic approach. Dependence was produced by continuous intracerebroventricular (i.c.v.) infusion of butorphanol (26 nmol/[micro]m/hr) for 72 hours via osmotic minipumps in rats. Proteins were separated by two-dimensional gel electrophoresis (2-DE) and 90 tyrosine phosphorylated protein spots were detected by immunoblotting with anti-phosphotyrosine specific antibodies. About 35 phosphotyrosyl spots showed predominant changes in comparison with that of control rat brains. Some of these spots were identified by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). Proteins involved in cell cytoskeleton, cell intermediary metabolism, and cell signaling mechanism such as tubulin, pyruvate kinase, aldolase C, and glutamine synthetase were identified. This proteomic approach may provide useful information to understand complex mechanism of butorphanol dependence. This work was supported by DA05828.

DOPAMINE RECEPTOR BINDING AND APOMORPHINE-INDUCED LOCOMOTOR ACTIVITY IN [mu]-OPIOID RECEPTOR KNOCKOUT MICE

Lu-Tai Tien (1)*, Younjoo Park (1,2), Lir-Wan Fan (1,3), Tangeng Ma (1), Horace H. Loh (4), and Ing Kang Ho (1), (1) University of Mississippi Medical Center, Jackson, MS 39216; (2) National Institute of Toxicological Research, KFDA, Seoul 122-704, Korea; (3) Mayo Clinic, Jacksonville, FL 32224; and (4) University of Minnesota, Minneapolis, MN 55455

Previous studies from our laboratory have indicated possible interactions between opioidergic and dopaminergic neurons in the central nervous system. In this study, apomorphine-induced locomotor activity and the D1 and D2 subtype dopamine receptor binding were examined in mice lacking the [mu]-opioid receptor genes. The ambulatory time, vertical time and total motor distance of locomotor activity were measured after administration of apomorphine (2 mg/kg, i.p.) for a period of 90 min. The autoradiographic studies of D1 and D2 dopamine receptors were conducted using [[.sup.3]H] SCH23390 and [[.sup.3]H] raclopride as ligand, respectively. In wild type mice that received apomorphine, 2 mg/kg, i.p., the locomotor activity such as ambulatory time, vertical time and total motor distance were not significantly altered as compared with that of the saline control group. However, the locomotor activity measured was significantly increased in the same dose of apomorphine treated &mu;-opioid receptor knockout mice between 5 to 40 min after administration. The results obtained also show that the binding of D2 dopamine receptor in [mu]-opioid receptor knockout mice was significantly higher than that of the wild type in the caudate putamen. However, the binding of the D1 dopamine receptor in [mu]-opioid receptor knockout mice was not significantly different from that of the wild type. It appears that the apomorphine treated [mu]-opioid receptor knockout mice showed enhancement in locomotor activity. The enhanced locomotor activity may be related to the compensatory upregulation of D2 dopamine receptors in mice lacking [mu]-opioid receptor genes. This study was supported by the Human Science Grant Foundation of Japan.

THE EFFECTS OF VINCA ALKALOIDS ON TRYPANOSOMA LEWISI IN RATS

Jennifer Craft*, Samantha Morris, and Amanda Ruth, Belhaven College, Jackson, MS

Trypanosomiasis is one of the top 10 parasitic killers n the world and has surpassed AIDS as the leading cause of death in some African nations. Much research has been done on the causative agents of this disease, Trypanosoma brucei gambiense and T. brucei rhodesiense. Wosu and Ibe (1989) suggested that botanical compounds are effective in controlling these parasites. Based on their study, an experiment was set up to investigate the effects of Vinca alkaloids related to the compounds used in the original research. Sprague-Dawley rats were infected with a non-pathogenic trypanosome species, Trypanosoma lewisi. Varying doses of the Vinca alkaloids vinblastine and vincristine were administered to subgroups of the rats and the numbers of the trypanosomes were monitored to observe the treatment's effects. Compared to the control group, the rats that recieved a lower dosage of vinblastine had lower levels of parasitemia. High dosages of vinblastine and vincristine did not seem advantageous, as they appeared to increase parasitemia when compared to the control group and in most cases were lethal. This could be due to supression of the immune response. Further studies may confirm that low doses of Vinca alkaloids may be effective in treating trypanosomiasis.

POINT MUTATION ANALYSIS OF PTEN EXONS 1, 3, AND 9 IN TWELVE WILMS' TUMORS, TEN STOMACH CANCERS, ONE MELANOMA, AND ONE LIPOSARCOMA IN MUSCLE

Crystal A. Jones (1)*, Robin Shanta Williams (2), James Weston (3), Mary L. Haasch (3), Lizhen Gui (4), and Margot Kaelbling (4)*, (1) Tougaloo College, Tougaloo, MS 39174; (2) Alcorn State University, Alcorn State, MS 39096; (3) University of Mississippi, University, MS 38677; and (4) University of Mississippi Medical Center, Jackson, MS 39216

To detect hereditary mutations caused by the tumor suppressor gene PTEN, we recently analyzed 119 tumor and uninvolved tissue panels of 14 tumor types for point mutations. That analysis was limited to exons 5, 7, and 8 that contain active sites. Here we present the analysis of exons 1, 3, and 9 in twelve Wilms' tumors and ten stomach cancers, and exons 1 and 3 in one melanoma and one liposarcoma. We aimed to sequence tumor tissue and analyze uninvolved tissue only if a point mutation occurred in the matching tumor. Stored DNAs were quality-checked with an Agilent Technologies 2100 Bioanalyzer. Exons were PCR-amplified and purified using gel electrophoresis and QIAEX II gel extraction. Amplicons were sequenced and analyzed with the CEQ 8000 Genetic Analysis System. All exons amplified successfully except exon 3 of two stomach tumors that each failed in 4 of 5 amplifications; the faint band in one amplification probably derived from stromal tissue. Two Wilms' tumors contained the same insertion in exon 1 that, most likely represent artifacts. One Wilms' tumor and six stomach cancers yielded insufficient exon 9 sequence data. All 24 exon 1 sequences, 22 of the 24 exon 3 sequences, and the 16 analyzable exon 9 sequences are deemed normal. Two stomach tumors apparently had lost both copies of exon 3 and at least 247 surrounding bases. In summary, we did not find any point mutations; however, we detected deletions of both copies of exon 3 in two stomach tumors.

TISSUE SPECIFIC INCREASE IN ANTIOXIDANT ENZYMES AND LIPID PEROXIDATION PRODUCTS IN BRAIN OF RATS EXPOSED TO LEAD

Christopher D. Bennett (1)*, Yallapragada Prabhakara Rao (2), Bettaiya Rajanna (1), Levenia Baker (1), Jon J. Brice (1), Samuel L. White (1), and Kiran Kumar (2), (1) Alcorn State University Alcorn State, MS 39096 and (2) Visakhapatnam, India 54367

The brain has a high rate of oxidative metabolism, high content of unsaturated lipids and low levels of protective enzymes to eliminate free radicals. Therefore, the brain is prone to oxidative damage. Reactive oxygen species formation has been proposed to be the final common pathway for most of the neurotoxicants. Lead is a potent neurotoxicant. The purpose of this study is to determine the effect of lead on antioxidant enzymes and lipid peroxidation products in different regions of the rat brain. The rats (Wistar strain) were treated with lead acetate (500 ppm) through drinking water for a period of 8 weeks. Controls were maintained on sodium acetate. They were sacrificed by cervical dislocation at intervals of 1, 4, and 8 weeks and the brains were isolated immediately. The brains were washed in ice-cold normal saline solution and the cerebellum, the hippocampus, the frontal cortex and the brain stem were separated on ice. Antioxidant enzymes and lipid peroxidation products were determined at treatment intervals 1, 4, and 8 weeks. The results indicated a gradual increase in the activity of antioxidant enzymes, more in the cerebellum and hippocampus, and the response was time-dependent. The data suggests a region specific oxidative stress in the brain. The increase in lipid peroxidation products might be due to damage caused to membrane lipids by lead toxicity. Lead exposure might have resulted in the formation of excessive reactive oxygen species causing enhanced levels of antioxidant enzymes. (Supported by NIH/FIC/MIRT #T37 TW00132 and NIH/NIGMS/SCORE #GM 55356)

IV Cell Cultures and Immunology

THE EFFECT OF CORTISOL AND SELENOMETHIONINE ON KIDNEY EPITHELIAL TUBULAR CELLS

Stevie D. Adams*, Russell F. Lyon, Hamed Benghuzzi, and Michelle Tucci, University of Mississippi Medical Center, Jackson MS 39216

Glucocorticoids play an essential role in maintaining homeostasis and regulation of normal growth and functions of a variety of organs. In the kidney, it modulates renal glucocorticoid receptors and effects Na-K ATPase activity, but can have dampening effects on the body when in excess. Antioxidants are useful in combating the effects of glucocorticoid on the total body excess and its function is well documented. However, the effects of glucocorticoids on epithelial cell structure and function are not well defined. In our investigation we examined the viability, proliferation, and morphology of rhesus monkey kidney cells in culture upon treatment with doses of selenium alone or in combination with doses of cortisol in an attempt to prevent or treat epithelial damage of the kidneys. The experimental design for the cells consisted of exposure to low and high doses of selenium alone, low and high doses of cortisol alone, and a combination of selenium with high dose of cortisol at 24, 48, and 72-hours. Cells were evaluated for viability, cellular protein content, and cellular damage. Cellular viability was not altered in treated cells, however significant increases were observed at 24 hours in the Cortiso[l.sub.lo] and Combo treated cells. Cellular protein levels were not different for the duration of the experiment. Data obtained showed all treated groups resulted in decreased levels of lipid peroxide generation at 48 and 72 hours. The data suggests that the kidney tubular cells given a bolus dose of cortisol may metabolize the cortisol to an inactive form and are not exposed to toxic effects of continuous release of the drug that would ultimately result in cellular alterations and cellular damage. *Allied Health Undergraduate Student, UMC

PRIMARY RABBIT KIDNEY EPITHELIAL CELLS TREATED WITH SUSTAINED LEVELS OF ALDOSTERONE, AND SPIRONOLACTONE IN A CELL CULTURE

Brandy Blaylock*, Hamed Benghuzzi, Michelle Tucci, and Zelma Cason, University of Mississippi Medical Center, Jackson, MS 39216

Elevated plasma aldosterone level may be a contributor to cardiac and renal disease. Administration of sustained levels of aldosterone may alter the viability and integrity of rabbit kidney epithelial cells at 24, 48, and 72 hours. Spironolactone (S) is an antagonist to aldosterone (A) and may offer cellular protection. The main objectives of this study were: (1) to show that aldosterone has an effect on kidney cell viability and that the concentration of aldosterone is important; (2) to show that administration of spironolactone prior to the administration of aldosterone will protect the cells from cellular damage (MDA). Experimental design consisted of thirty tubes of rabbit kidney epithelial cells (Rk) divided into six equal groups (Control (C), S, S+[A.sub.lo], S+[A.sub.hi], [A.sub.lo], and [A.sub.hi],. After 24 hours there was a significant decrease in cell proliferation in the following groups S, [A.sub.lo] [A.sub.hi], S+[A.sub.lo]. At 48 hours a similar trend in cell proliferation was observed. However, S+[A.sub.lo] and S+[A.sub.hi] resulted in cell numbers that were higher than the treatments of aldosterone or spironolactone alone. After 72 hours there was a steady decrease in the proliferation in all groups compared to the control. Cellular damage was not evident in the treated groups after 24, 48, or 72 hours. Morphological differences were observed in all treated groups after 48 hours. Presence of nucleoli was most notable possibly indicating increased ribosomal RNA and an increase in protein synthesis. The increase ribosomal RNA also suggests the possibility that the treatments have a direct genomic effect on renal cells. *Allied Health Undergraduate Student, UMC

INHIBITORY EFFECTS OF BLACK SEED OIL (NIGELLA SATIVA) ON SURVIVAL OF THE HUMAN LIVER CARCINOMA (HEPG2) CELLS

Terrell Bradford (1,2)*, Waria Holmes (1,2), Jemeka Miller (1,2), Clement Yedjou (1), Joseph A. Cameron (1), and Ibrahim Farah (1), (1) Jackson State University, Jackson, MS 39217, and (2) Hinds Community College, Raymond, MS 39154

Black seed (N. sativa L.), an oriental spice of the family Ranunculaceae has long been used tradionally as a natural medicine for treatment of many acute as well as chronic conditions including cardiovascular disease and immunological disorders. It has also been used in the treatment of diabetes, hypertension, and dermatological conditions. There have been very few studies on the effects of N. sativa and/or its oil extracts as a chemoprevention of chronic diseases as well as in cancer prevention and/or therapy. Oxidative stress is a condition that underlies many acute as well as chronic conditions. The combination and role of oxidative stress and antioxidants in vivo is still a matter of conjecture. Our objective for the present study was to expose hepatocarcinoma (HepG2) cells in vitro (as a chronic disease example) to commercial seed oil extracts. Measurement of inhibitory effects as reflected by cell survival under various concentrations was conducted using standard cell culture techniques, exposure protocols in 96-well plates and the MTT cell survival assay. Following cellular growth to 90% confluencey, 24 h exposure to black seed oil extracts was performed. Cell survival index (LC50) was calculated from percent survival using regression analysis. Results showed there was a dose-dependent inhibitory effects at concentrations ranging from 1.95-1000 ppm. Concentrations of 500-1000 ppm were found to be deleterious to these cells. In conclusion, black seed oil was found to have very strong inhibitory effects on the survival of HepG2 Liver carcinoma cells, unveiling promising opportunities in the field of cancer chemoprevention and/or treatment and warranting further future studies to elucidate the underlying mechanisms. (Supported in part by the NIGMS R25 GM50117).

THE EFFECTS OF STEROID HORMONES ON THE VIABILITY AND METABOLISM OF A549 CELLS

Raegan Castle*, Hamed Benghuzzi, Zelma Cason, and Michelle Tucci, University of Mississippi Medical Center, Jackson, MS 39216

Surfactant synthesis by A549 cells has previously been shown in our lab to be stimulated by glucocorticoids. In addition, it has also been shown that sex steroids exert opposing effects, with estrogens accelerating and androgens inhibiting surfactant production. However, the viability, metabolism and morphology of these cells have not been fully characterized in the presence of steroid hormones. The aim of this investigation was to determine the effects of testosterone (T), estrogen (E), androstendione (AED), and a combination of E+T and E+AED on the viability and morphology of A549 cells after 24, 48, and 72 hours. A549 cells were divided into six groups (n=6) Control, T, E, AED, E+T and AED+E. The groups were evaluated after each incubation period for cellular viability, protein, and changes in morphology. The results of this study showed increased cell proliferation in all treated groups after 24 hours. Significant growth was seen in E, AED, E+T, and E+A (p < 0.001). After 72 hours in culture, the growth was not sustained, and a significant decrease in cell number was detected in all treated groups (p < 0.05). The decrease could possibly be explained by the need for greater nutritional support. At 24 hours slight decreases in protein levels were seen in all groups when compared with control. However, by 48 hours there was significant increase in cellular protein levels (p < 0.05) followed by a rapid decline in cell protein content at 72 hours, further suggesting the culture conditions were compromised and no longer able to support the rapid growth. Overall, the results suggest that gonadal and adrenal steroid hormones alone or in combination stimulate the growth of type II pneumocytes. *Allied Health Undergraduate Student, UMC

THE EFFECTS OF THE ESTROGEN, PROGESTERONE, AND CORTISOL, ON THE VIABILITY AND PROLIFERATION OF THE CERVICAL TUMOR CELL LINE, SW 756, IN CULTURE

Melissa Daniel* and Hamed Benghuzzi, University of Mississippi Medical Center, Jackson, MS 39216

Cervical cancer remains a major health threat to women worldwide and role of steroid hormones on cervical cancer cells is not clearly defined. This study investigated the effects of steroid hormones on the viability and proliferation of SW 756 cervical cell line. In this study, SW 756 cells were treated with physiological and supraphysiological of cortisol, estrogen, and progesterone. The cells were harvested at 24, 48, and 72 hours and cell numbers were determined. In addition, cellular damage, cellular protein content, and cellular morphological characteristics were determined at each time period and compared to a control. The data obtained from this investigation demonstrated the following: Cell counts revealed supraphysiological doses of cortisol, estrogen and progesterone caused marked decreases in cell numbers at 24 hours (p < 0.05), compared to the control. At 48 and 72 hours, decreases were still apparent for supraphysiological doses of cortisol and estrogen, while progesterone treated cells adapted. Assays for cell damage revealed marked cellular damage, at 24 hours following supraphysiological levels of progesterone (p < 0.05). Supraphysiological levels of cortisol and estrogen caused cellular damage at 24 hours and marked cellular damage at 72 hours (p< 0.05, Dunnett's test). Cellular protein levels were unremarkable; however they indicated cell viability at all time periods. Morphological changes were insignificant. This investigation provides significant information regarding the interrelationship between the steroid hormones cortisol, estrogen, and progesterone and the viability and proliferation of HPV containing SW 756 cells in culture. *Clinical Health Sciences Graduate Student, UMC

THE EFFECTS OF INSULIN-LIKE GROWTH FACTOR-1 ON A MG63 (OSTEOSARCOMA) CELL LINE

Laura Franklin*, Michelle Tucci, Hamed Benghuzzi, George Russell, Ashraf Ragab, and Audrey K. Tsao, University of Mississippi Medical Center, Jackson, MS 39216

Growth hormone secreted by the pituitary gland stimulates the liver to produce Insulin-like Growth Factor-1 (IGF-1), which is believed to play a valuable role in building lean muscle mass, maintaining bone density, and protecting nerve cells. The purpose of this experiment was to examine MG-63 cells after treatment with low (1 [micro]g), medium (5 [micro]g) and high (50 [micro]g) doses of IGF-1. MG-63 cells, were plated onto a 24 well tissue culture plate at a density of 1X [10.sup.5] cells per well. The experiment was designed to evaluate cell counts, MDA, protein levels, and the cell morphology after 24, 48, and 72 hour post incubation with IGF-1. IGF-1 stimulated cellular division as evidenced both morphologically as well as by an increase in cell numbers. There was inverse relationship between dose and cell number with the lower dosage of IGF-1 causing the most significant increase. Increases in MDA levels were seen at twenty-four hours in all treated groups. The increase was dose dependent with the highest dosage of IGF-1 yielding the highest MDA levels. No significant changes in cellular protein levels were found for the duration of the experiment. Morphological evaluation showed increased cellular division and prominent nucleoli for the duration of the experiment. This information suggests that IGF-1 has an anabolic effect on MG-63 and the effect is dose dependent with the lower dose being more effective, suggesting receptor mediated effects. * Undergraduate Student, Mississippi College

THE EFFECT OF ULTRAVIOLET RADIATION EXPOSURE TO FIBROBLAST CELLS

Pamala Jones*, Felicia Magee Tardy, Hamed Benghuzzi, and Michelle Tucci, University of Mississippi Medical Center, Jackson, MS 39216

Fibroblasts play an important role in healing and tissue damage repair. When there is tissue damage, fibrocytes are transformed into fibroblasts, which contains large amounts of organelles, which are necessary for the synthesis and excretion of proteins needed to repair the damaged tissue. The objective of this study was to determine the effect that ultraviolet radiation had on fibroblast cells and how well the fibroblasts responded to antioxidants in an attempt to prevent skin damage from the harmful UV rays of the sun. The cells were divided into control groups and treatment groups and evaluated after 24, 48, and 72 hours. The first treatment group were exposed to 45 minutes of ultraviolet radiation and evaluated for morphological damage using Image Pro Digital Analysis. The second group of cells were treated with antioxidants prior to and after exposure to UV radiation and evaluated. The results of this study showed significant findings. UV radiation produced dramatic changes and alterations in the cells such as pleomorphism, swelling, and mitosis as well as other changes. Antioxidant treatment caused significantly less cellular damage pre and post treatment. However, further investigation is needed to determine an appropriate dose of antioxidants that will offer complete cellular protection from ultraviolet radiation.

THE EFFECT OF AGONISTS AND ANTAGONISTS ON HEP-2 CELLS

LaToya Phillips*, Hamed Benghuzzi, Zelma Cason, and Michelle Tucci, University of Mississippi Medical Center, Jackson, MS 39216

Androgens may play an important role in promoting the growth of laryngeal carcinomas. The aims of this investigation were to investigate the effects of (testosterone (T) and androstendione (AED)) in the presence of the anti-androgen, spironolactone (S), on Hep-2 cellular proliferation and damage after 24, 48, and 72 hours. Hep-2 cells were divided into six groups (n = 5) control, S, T, AED, S+T, and S+AED, respectively. The cells were harvested after each incubation period into two different fractions: suspended cells and adhered cells. Cell counts and cellular damage determinations were performed on each fraction. Analysis of variance was used to determine significance at p < 0.05. Data for cell counts revealed an interesting phenomenon between the two fractions. Adhered cells showed decreased cell numbers in the presence of S and T for 24-48 hours followed by a significant increase at 72 hours. Cells in the adhered fraction incubated in the presence of AED or AED +S were significantly lower for the duration of the experiment. However AED or AED + S treatment caused significant increase in cell number in suspended fraction for the duration of the experiment. All treatments after 72 hours showed a slight reduction in MDA levels indicating treatments did not cause cell damage. Overall, the data suggests the possibility of two populations of cells that respond differently to the AED. T had no significant effect on either cell fraction for the first 48 hours followed by a significant increase in cell number at 72 hours suggesting T may need to be converted enzymatically to the more potent androgen, dihydrotestosterone. *Allied Health Undergraduate Student, UMC

SEMIQUANTITATIVE MEASUREMENT OF CYTOKINE MRNA IN PERIPHERAL BLOOD MONONUCLEAR CELLS FROM HEART TRANSPLANT RECIPIENTS.

Geeter Gilmore (1,2,3)*, Andi Barker (1), Laura Godfrey (1), Joseph A. Cameron (2), and D. Olga McDaniel (1), (1) University of Mississippi Medical Center, Jackson, MS 39216; (2) Jackson State University, Jackson, MS 39217; and (3) Hinds Community College, Raymond, MS 39154

Cytokines play a major role in the inflammation process and in specific immune responses triggered by alloantigens. Previous studies demonstrated that cardiac allograft rejection is associated with an individual's inflammatory cytokine gene polymorphisms. Based on these findings, it was hypothesized that possession of specific cytokine alleles, for TGF-[beta]1, IFN-[gamma] or IL-10 might be influential in predisposing the recipient to allograft rejection and/or tolerance. Because transplant patients have undergone immunosuppressive therapy, demonstration of the relationship between genotype/phenotype is not practical. Therefore, the relationship between cytokine gene polymorphisms and the level of cytokine production was tested in an in vitro assay, using the recipient's peripheral blood mononuclear cells (PBMCs) treated with phytohemagglutinin (PHA). We used the polymerase chain reaction to semiquantitatively measure changes in the amounts of messenger RNA from the TGF-[beta]1, IFN-[gamma] and IL-10 genes in the PBMCs before and 48 hours after PHA treatment. A total of 10 recipients with known cytokine genotype profile were studied. No elevation in expression level of TGF-[beta]1 was observed in resting or stimulated samples. The IL-10 was elevated after stimulation in the PBMCs from recipients with high producing genotypes. The IFN-[gamma] was 50% elevated in the PBMCs from recipients with high producing genotypes and in some patients even in low producing genotypes. In conclusion, because of the therapeutic effects of the immunosuppressant, the analysis of the mRNA, demonstrated in this study might not represent precise quantification of cytokines associated with cytokine genotypes. (Supported in part by NIGMS R25 GM50117).

ELECTROPHORETIC MOBILITY OF LDL USING MRC-5 FIBROBLASTS EXPOSED TO SEX HORMONES

Felicia Magee Tardy*, Pamala Jones, Hamed Benghuzzi, and Michelle Tucci, University of Mississippi Medical Center, Jackson, MS 39216

Several studies have shown that the oxidative modification of LDL is the key to the development of atherosclerotic cardiovascular disease. Although there is extensive evidence implicating the oxidatively modified form of LDL, the exact mechanisms by which LDL oxidation occurs is still unresolved. The objective of this investigation was to study the effects of LDL modification by MRC-5 fibroblasts exposed to sex hormones. MRC-5 fibroblasts were treated with low and high doses of LDL and exposed to estrogen and testosterone. Representative samples from each group were used for the determination of LDL modification using lipoprotein electrophoresis. Results from the lipoprotein electrophoresis revealed that the migration patterns of the treatment groups varied significantly from the native form of LDL. Results from this investigation indicate that LDL was modified during the incubation periods with estrogen, testosterone, and the MRC-5 cells. In addition, it was evident the MRC-5 cells have the ability to metabolize LDL. Further investigation is needed to determine the role of MRC-5 cells in LDL modification. * Clinical Health Sciences Graduate Student, UMC

THE EFFECTS OF (-) EPIGALLOCATECHIN-3-GALLATEON RHESUS MONKEY KIDNEY EPITHELIAL CELLS

Stacy Hull Vance*, Hamed Benghuzzi, and Michelle Tucci, University of Mississippi Medical Center, Jackson, MS 39216

Currently, it is estimated that approximately $5.1 billion dollars are spent annually on herbal supplements in the U.S. Herbal supplements have beneficial properties; however, these properties need to be clinically proven and any possible interactions need to be documented. The chemopreventative effects of green tea are contributed to its major polyphenolic constituent, (-) epigallocatechin-3-gallate (EGCG). EGCG has been shown to induce growth arrest and/or apoptosis in various cell lines. EGCG is thought to selectively inhibit growth of cancer cells without adversely effecting normal cells. The purpose of this study was to incubate Rhesus Monkey Kidney Epithelial cells (RMKEC) with various doses of EGCG (20, 2, 0.2, 0.002 [micro]M) and evaluate cellular viability and morphology after 24, 48, 72, and 96 hours. The cells treated with 20 [micro]M EGCG appeared in small clusters with more cytosol and large nucleoli in addition to hydrophic swelling. In cells treated with 2 and 0.2 [micro]M EGCG the cells appeared in a honeycomb pattern with hyperchromatic nuclei. Several cellular regions within the honeycomb showed evidence of anucleation. Overall the cells appeared round and swollen. Cells treated with 20, 2, and 0.2 [micro]M EGCG resulted in lysed cells and/or anucleated with frothy cytoplasm. There were few normal cells present with 0.002 [micro]M of EGCG however, the cells appeared swollen, round, and anucleated. The data obtained suggests that normal epithelial cells, like cancer cells, are adversely affected by EGCG at concentrations ranging from 20-0.002 [micro]M.

CA242 COMPARED WITH TEN OTHER TUMOR ANTIGENS FOR THE SERODIAGNOSIS OF PANCREATIC CANCER

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

The fourth commonest cause of cancer deaths, pancreatic cancer (CA) is a serious health problem in the United States. Due to its non-specific early symptoms, pancreatic cancer is often not diagnosed until late stage disease when the prognosis is poor. A noninvasive early detection method would be clinically useful. This study's goal was the comparison of CA242 with ten other tumor antigens for diagnostic efficacy in pancreatic CA. Sera from 554 patients (16 pancreatic CA, 128 other GI CA, 216 other CA, and 195 non-CA) were assayed for the presence of tumor antigens and the results correlated with diagnoses established pathologically. Immunoassay test kits from Diagnostic Automation (CA242), Hybritech (CEA, CA195), Centocor/Fujirebio 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 manufacturer's decision values the following diagnostic sensitivities were obtained for pancreatic cancer: CA242 66.7%, CA195 100.0%, CA19-9 66.7%, CA50 66.7%, CA125 40.0%, CA 27.29 40.0%, CEA 37.5%, CA72-4 31.3%, Cyfra21-1 26.7%, CA15-3 26.7%, and AFP 18.2%. Diagnostic specificities were >75%. From these data we conclude that CA242 is useful for the diagnosis of pancreatic CA.

Mini-Symposium--Selected Topics II

Moderator: D. Olga McDaniel, University of Mississippi Medical Center

2:30 Genomics to Health

D. Olga McDaniel, University of Mississippi Medical Center

2:50 Genomics to Biology

Micheal Hebert, University of Mississippi Medical Center

3:10 Genomics to Society

Elizabeth Heitman, University of Mississippi Medical Center and Vanderbilt University

3:45 Podium Presentations--Session II

Moderator: Larry S. McDaniel, University of Mississippi Medical Center

4:00 PURIFICATION OF RECOMBINANT PSPC

Marques Slaughter (1,2,3)*, Lashondra Johnson (1), Joseph A. Cameron (2), and Larry S. McDaniel (1), (1) University of Mississippi Medical Center, Jackson, MS 39216; (2) Jackson State University, Jackson, MS 39217; and (3) Hinds Community College, Raymond, MS 39154

Streptococcus pneumoniae, pneumococcus, is an important pathogen that causes disease worldwide. The pneumococcus has evolved different strategies to evade host responses including the capsular polysaccharide and surface proteins. PpC is a surface protein that binds human Complement Factor H (FH). FH is a serum protein that helps protect host tissue from Complement by regulating the deposition of Complement on host tissues. It appears that the pneumococcus binds FH on PspC to help this bacteria evade the Complement system. The purpose of our experiment was to purify PspC that has been expressed in Escherichia coli. E. coli was grown to late log phase in broth and induced to express recombinant PspC. Analysis of cultures demonstrated significant expression of PspC. We then purified the PspC using an affinity column. Fractions from the column were examined by SDS-PAGE to identify those factions that contained PspC and to assess the purity of the protein. A Western blot confirmed the presence of PspC which reacted with FH. An ELISA was carried out to determine the reactivity of the purified protein. The specificity of the interaction of FH with the purified PspC was examined by an inhibition assay. We demonstrated that the purified protein specifically interacted with FH. Currently this purified protein is being used in studies to produce monoclonal antibodies that are specific for PspC. (Supported in part by NIGMS Grant R25 GM50117).

4:10 CHARACTERIZATION OF COLIFORM BACTERIA FROM INFLUENT WASTEWATER OF THE UNIVERSITY OF MISSISSIPPI WASTEWATER TREATMENT PLANT

Trisha Weekley* and Al Mikell, University of Mississippi, University, MS 38677

Coliform bacteria are those bacteria used as an indicator organism in determining whether or not water has been contaminated. Escherichia coli is one example of coliform bacteria. It is one of the many bacteria that are a normal flora of the gastrointestinal tract of all warm-blooded terrestrial species. Its presence can be detected in the waste of these species. In this paper, wastewater samples were collected from the University of Mississippi Wastewater Treatment Plant in order to test for coliform bacteria found in humans. Specifically, bacteria having the characteristics of E. coli were selected for and tested. The bacteria were run through a series of tests to determine its identification. These tests included EC with MUG, gram staining, and API-20E test kit. After the identification of the samples using the API-20E test kits, there was found to be Klebsiella, Enterobacter, and unidentified bacterium. Why there was no presence of identified E. coli is still a part of ongoing research.

4:20 PHOTOMUTAGENICITY OF A HAIR DYE INGREDIENT 3'3-DICHLOROBENZIDINE (DCB)

William Hardy*, Charity Mosley*, Lei Wang, Jian Yan, and Hongtao Yu, Jackson State University, Jackson, MS 39217

Salmonella typhimurium bacteria strain TA 102 was used to examine the chemical toxicity, phototoxicity, and photomutagenicity of 3,3'-dichlorobenzidine (DCB), a chemical used in the production of pigments of hair dyes, paper, paint, rubber, leather, plastic, and other related industries. Up to a concentration of 625 [micro]M (160 mg/plate), DCB did not have chemical toxicity on TA 102. DCB is phototoxic for TA 102 when the bacteria is exposed to DCB and light at the same time. In a DCB concentration range of 125 [micro]M (32 mg/plat) to 625 [micro]M, DCB is phototoxic for TA 102 in a DCB dose dependent manner. Below 125 [micro]M, DCB is not phototoxic. Pre-incubation of DCB in the above concentration range with TA 102 followed by light irradiation (UVA, 6.5 J/c[m.sup.2] and visible light, 13 J/c[m.sup.2]) indicated a count of more than twice the revertant colonies of TA 102 as the negative control. This means that DCB is also photomutagenic. This research is supported by a grant from the National Institutes of Health (NIH-SCORE S06GM08047). We would like to thank the National Science Foundation for student support through LSMAMP and HBCU-UP STARGE programs.

4:30 PNEUMOLYSIN INDUCED INFLAMMATORY RESPONSES FOLLOWING ANTIBIOTIC TREATMENT OF STREPTOCOCCUS PNEUMONIAE

Justin Thornton* and Larry S. McDaniel, University of Mississippi Medical Center, Jackson, MS 39216

Pneumolysin (PLY) is an important virulence protein of Streptococcus pneumoniae. To examine the global effect of pneumolysin on cells of the immune system we used cDNA microarray analysis of human THP-1 cells exposed to S. pneumoniae. THP-1 cells were suspended in antibiotic free RPMI and co-incubated with either medium alone, capsular type 2 strain D39, or PLN, a Ply lacking isogenic mutant of D39. This allowed us to identify genes that were differentially regulated in response to pneumolysin. Since the cell wall of gram-positive organisms has been shown to lead to a potent inflammatory response, we hypothesized that pneumolysin played an additive role in this response. Human THP-1 cells were exposed to either D39 or a PLN in the presence of different antibiotics. THP-1 viability and bacterial cell counts were determined at 3 and 10 hours and supernatants were collected. Penicillin treatment resulted in a three log decrease in colony forming units of D39 and PLN at three hours. THP-1 cells exposed to D39 in the presence of penicillin had significantly higher levels of IFN-[gamma] than cells exposed to PLN. The viability of THP-1 cells remained above 70% for all exposures. We also examined the expression of cell adhesion molecules in response to pneumococci and pneumolysin using real time PCR. Our data suggest that specific antibiotic treatment of pneumococci results in pneumolysin release that can affect the subsequent inflammatory response.

FRIDAY MORNING

Emerald

9:00 Podium Presentations--Session III

Moderators: Robin Rockhold and Joseph Cameron, University of Mississippi Medical Center and Jackson State University

9:15 EVALUATION OF THE CORIN/ANP SYSTEM IN TRANSGENIC HEART FAILURE MODELS

Kristina Vaughn* and Guy Reed, Jackson State University, Jackson, MS 39217, and Harvard University, Boston, MA 02115

The high incidence of congestive heart failure is partially attributed to uncontrolled hypertension. The corin/ANP system is investigated in heart failure with respect to blood pressure and mortality. Atrial natriuretic peptides (ANP) possess the ability to lower blood pressure with vasodilatation and salt excretion once activated by corin. Previous data shows that this system is altered in heart failure models, with ANP expression increasing and corin expression decreasing. This difference in expressions may affect the overall illness. The mutated cAMP Response Element-Binding Protein, mCREB, was used in transgenic mice to simulate the physiological effects of heart failure. Overexpression of corin was used in transgenic and wild type mice to determine the affect of corin on blood pressure. During blood pressure monitoring, mice were anesthetized and then catheterized. Mortality rates were generated from transgenic mice from mCREB and ANP knockout/mCREB mice, to determine the role of ANP in heart failure mortality. In the blood pressure study, there was no significant differences in blood pressure between genotypes (p>0.05), while a difference was noted between heart rates of corin transgenic mice and wild type controls (p<0.05) for the females. Mortality data showed a significant decrease in lifespan for both mCREB and ANP knockout/mCREB mice with a p<0.01 for males and p<0.001 for females for ANP knockout/mCREB mice. Results for the blood pressure study suggest that corin may not have a direct relationship to lowered blood pressure, while the mortality study demonstrates that ANP provides a protective benefit for congestive heart failure.

9:25 CYTOKINE GENOTYPE POLYMORPHISM AND EXPRESSION CORRELATES WITH THE OUTCOME OF CARDIAC TRANSPLANTATION

Andrea Barker*, Laura Godfrey, Brenda D. Mangilog, and D. Olga McDaniel, University of Mississippi Medical Center, Jackson, MS 39216

Coronary vasculopathy (CV) is a major factor in long-term survival of heart transplantation. The pathogenesis of CV in part is contributed by the activated immune response of the recipient to the donor tissue. Evidence supports the role of cytokines in the inflammatory and immune responses that mediate allograft survival outcome. To test this hypothesis, peripheral blood mononuclear cells (PBMCs) from heart transplant patients with varying degrees of CV and/or grade 3A rejections were tested for their capacity to express different inflammatory cytokines at the mRNA level using RT-PCR. Previously we have demonstrated that the IFN-[gamma] intermediate, IL-10 low and IL-18 high producer genotypes were associated with the occurrence of rejection episodes following cardiac transplantation. In this study, cDNA from the PBMCs were tested for the levels of mRNA transcript of the IFN-[gamma], IL-10, and IL-18 by PCR. The IL-18 mRNA was elevated in the patients with mild to strong CV as compared with those without the CV. The level of gene expression for IL-18 correlated with the genotype assignment. The IFN-[gamma] expression level 70% was in agreement with the genotypes, but there was a stronger correlation between IFN-[gamma] high producing genotypes as compared with the intermediate or low producers. IL-10 expression was elevated after treatment with PHA, but this phenomenon was not observed in IL-18 genotypes. Analysis of such polymorphism in a greater number of patients might allow the identification of patients before transplantation who have a greater risk of developing graft rejection.

9:35 SYNTHETIC ALKALOIDS FROM SOLENOPSIS INVICTA (IMPORTED FIRE ANT) VENOM EXERT CARDIOVASCULAR ACTIONS

George Howell*, Jordan Butler*, and Rob Rockhold, University of Mississippi Medical Center, Jackson, MS 39216

Two structurally verified, synthetic S. invicta venom alkaloids, solenopsin A (trans-2-methyl-6-undecylpiperidine) and its cis-isomer, isosolenopsin A, were synthesized. Both have been shown previously to inhibit nitric oxide synthase (NOS) isoforms in vitro, with isosolenopsin A showing I[C.sub.50]S of 18 [+ or -] 3.9 (neuronal NOS), 156 [+ or -] 10 (endothelial NOS), and >1000 [micro]M (inducible NOS). Sprague-Dawley rats were anesthetized with isoflurane, paralyzed with gallamine, artificially ventilated and instrumented to record arterial blood pressure (BP; mm Hg), heart rate (HR; bpm) and left ventricular dP/dt (percent change from control; LVC; an index of cardiac contractile force). Solenopsin A elicited maximal percent changes ([+ or -] S.D.; n = 5) of -40 [+ or -] 12 (BP), -27 [+ or -] 8 (HR) and -41 [+ or -] 17 (LVC). Isosolenopsin A (15 mg/kg, i.v.) produced responses similar to solenopsin A (30 mg/kg). In a single spontaneously breathing rat, solenopsin A (30 mg/kg, i.v.) caused respiratory arrest. Superfusion of a working, isolated perfused heart with solenopsin A elicited a marked, reversible decrement in contractile function (dP/dt) at 10 [micro]M and cardiac arrest at 100 [micro]M. No overt cardiovascular responses were identified in conscious, freely moving rats chronically instrumented to record BP and HR and given solenopsin A (30 mg/kg, i.v.). The results demonstrate that these alkaloids possess robust depressant activity on the cardiac and respiratory systems, actions that are not consistent with the demonstrated in vitro inhibition of NOS. (Supported by the Howard Hughes Medical Institute.)

9:45 IMMUNOLOGICAL RESPONSES TO CYTOPLASMIC ANTIGENS: ROLE OF MHC CLASS II ANTIGENS

Jeremy Lott (1,2)*, Delu Zhou (1), Joseph A. Cameron (2), and Janice S. Blum (1), (1) Indiana University School of Medicine, Indianapolis, IN 46202, and (2) Jackson State University, Jackson, MS 39217

The Major Histocompatability Complex (MHC) encoded class I and II molecules are expressed on the surface of cells and function to display peptides derived from pathogens as well as self proteins for recognition by host T lymphocytes. Typically, class I displays peptides from cytoplasmic proteins, while class II presents peptides derived from extracellular pathogens and proteins that have been delivered into lysosomes. This research is focused on a novel alternate pathway for class II molecules, specifically their ability to present peptides generated in the cytoplasm. The project was designed to address whether peptides in the cytoplasm can be transported directly into acidic vacuoles containing class II molecules. As an assay system, MHC class I[I.sup.+] and MHC class I[I.sup.-] B cells were used to monitor peptide transport from the cytoplasm to vacuoles. Cells were electroporated to deliver labeled peptides into the cytoplasm. Western blots were performed to determine whether the labeled peptides had entered vacuoles to bind class II molecules. Studies demonstrated that only with delivery of peptides into the cytoplasm, could labeled peptides bind to class II. It is hoped that the results of this study will lead to novel insights concerning immune tolerance, and one day the development of new treatments for autoimmune diseases. (Supported in part by NIGMS R25 GM 067592).

9:55 SIGNAL TRANSDUCTION MECHANISM FOR VON WILLEBRAND FACTOR

John Kermode* and Qi Zheng, University of Mississippi Medical Center, Jackson, MS 39216

High shear stress in an arterial stenosis causes von Willebrand factor (VWF) to interact with its platelet receptor, glycoprotein Ib-IX-V (GpIb-IX-V). Such interaction leads to platelet activation by a poorly defined mechanism that may involve calcium mobilization and protein tyrosine phosphorylation. The present study examined the responses of human platelets to VWF. Intracellular calcium concentration ([C[a.sup.2+]]i) was assayed with Fura-PE3, platelet activation through serotonin secretion, and phospholipase A2 activity through measurement of thromboxane A2 generation. Treatment of platelets with the cyclo-oxygenase inhibitor aspirin abolished both the [C[a.sup.2+]]i transient and platelet activation in response to VWF. The influence of various phospholipase A2 inhibitors on these responses was also examined. One inhibitor, oleyloxyethyl phosphorylcholine, completely abrogated the platelet responses to VWF, but did not affect responses to [alpha]-thrombin or collagen. In contrast, methyl arachidonyl fluorophosphonate suppressed the platelet responses to [alpha]-thrombin and collagen, but not those to VWF. Differential inhibition was confirmed with other phospholipase A2 inhibitors. These findings imply that VWF induces platelet activation by stimulating a phospholipase A2 distinct from the cytosolic calcium-dependent type IV isozyme. Treatment of platelets with tyrphostin A23, a broad-spectrum inhibitor of protein tyrosine kinases, abrogated VWF-induced thromboxane A2 generation and platelet activation. The latter observation suggests that a tyrosine kinase might be responsible for activation of the novel phospholipase A2 when platelets are stimulated with VWF. [Supported by the American Heart Association (Southeast Affiliate).]

10:05 Break

Podium Presentations--Session IV

Moderators: Michelle Tucci and Jacqueline Stevenes, University of Mississippi Medical Center and Jackson State University

10:20 ANALYSIS OF CYP 3A4 GENOTYPE VARIATION IN RENAL TRANSPLANT RECIPIENTS AND THE ASSOCIATION WITH CYCLOSPORIN CLEARANCE

Sebron Harrison*, Brenda D. Mangilog, W. Henry Barber, Laura Godfrey, Andrea Barker, Xinchun Zhou, Jeffrey Dolittle, and D. Olga McDaniel, University of Mississippi Medical Center, Jackson, MS 39216

Most organ transplant recipients in addition to immunosuppressive drugs such as cyclosporin, tacrolimus or prednisone also take medications for diabetes, hypertension and other conditions. Frequently, when more than 2 drugs are concomitantly administered there is good chance of alteration in drug distribution and excretion following long-term drug treatment. Such alteration might affect metabolism of many drugs. We have demonstrated interindividual variation in CYP 3A4 expression in different individuals. CYP 3A4 is found in most adults with 10 to 40 fold variations in the level of expression. Such variations are due to the polymorphism within the promoter region of the genes, causing variation in the level of expression. The aim was to determine the allelic frequency of the CYP 3A4 variants in African-American (AFAM) patients and to examine a possible association with cyclosporin elimination in the transplant settings. Blood samples from 77 AFAM patients and 67 matched controls were studied by single nucleotide polymorphism (SNP) and PCR. CYP3A4 G genotype was present in 82.5% of renal transplant patient population. There was no difference between G frequency in the patients as compared with control samples. Frequency distribution of G genotype was significantly higher in our study population (82.5%) as compared with AFAM (53%) elsewhere. There was a trend towards higher cyclosporin clearance index and AA variant, indicating that the effect of CYP3A4 might be more evident in a GG genotype. Such information might allow strategies for immunosuppressive drugs such as cyclosporin and tacrolimus that are commonly used to prevent allograft rejections.

10:30 PRIME-BOOST IMMUNIZATION WITH DNA AND PROTEIN ELICITS PROTECTION AGAINST PNEUMOCOCCAL INFECTION

Quincy C. Moore III*, Joseph R. Bosarge, Xiangyun He, and Larry S. McDaniel, University of Mississippi Medical Center, Jackson, MS 39216

We have previously reported that immunization with the gene encoding the [alpha]-helical domain of PspA/EF5668 or immunization with the purified recombinant protein can elicit protection against pneumococcal challenge in mice. In previous studies mice were immunized with 50[micro]g pspA and challenged with pneumococcal strain WU2 (100% protection) and EF5668 (43% protection). Cross-protection was observed but we wanted to modulate the immune response by using prime-boost immunizations. In this study, we examined the effect of priming with pspA/EF5668 and boosting with rPspA/EF5668. Mice primed and boosted with 50 [micro]g pspA had anti-PspA serum specific antibodies of 5.96 [micro]g/ml [+ or -] 1.45 and a survival rate of 89%. Mice primed with 50 [micro]g pspA and boosted with 10 [micro]g of PspA without adjuvant had anti-PspA serum specific antibodies of 1.75 [micro]g/ml [+ or -] 0.55 and a survival rate of 90%. In mice primed with 10 [micro]g of PspA without adjuvant and boosted with 50 [micro]g pspA, the concentration of anti-PspA specific antibodies was 3.63 [micro]g/ml [+ or -] 1.02 and there was a 70% survival rate. We have demonstrated that priming with DNA (pspA) leads to an enhanced response to PspA when mice are boosted with protein (PspA).

10:40 THE EFFECT OF FRUCTOSE-1,6-DIPHOSPHATE ON IL-1RA,IL-1[beta], IL-6 AND TNF[alpha] FROM LPS ACTIVATED MACROPHAGES

Jay Wentworth*, Hari H.P. Cohly, Joe Lopez, John J Jenkins, and Angel K. Markov, University of Mississippi Medical Center, Jackson, MS 39216

Fructose 1,6-diphosphate (FDP) is a naturally occurring intracellular metabolite, which has been demonstrated to inhibit endotoxin shock in vivo. We wanted to determine the in vitro role of FDP on the mRNA expression of IL-1Ra, IL-6, IL-1[beta] and TNF-[alpha], lipid peroxidation and iNOS production in monocytic mouse cell line RAW 264.7 using LPS. RAW 264.7 cells were incubated for 20 hrs at 37[degrees]C, 5% C[O.sub.2] with LPS at 10 ng/ml. Then, cells were tested with decreasing concentrations of FDP (5000 [micro]g/ml to 5 [micro]g/ml) and Cyclosporin (500 ng/ml, to 0.5 ng/ml), for iNOS production, lipid peroxidation, and cytokine expression. Cells were plated in T-flasks to confluence and were then treated with 10 ng/ml of LPS. After 18-20 hr, the cells were treated with trypsin-EDTA, centrifuged at 12,000 g for 5 min and 100 [micro]l of the supernatant was removed and mixed with equal volumes of Greiss reagent to determine iNOS production. In addition, 500 [micro]l of the supernatant was mixed with equal volumes of thiobarbaturic acid to determine lipid peroxidation. RNA was extracted from cell pellets using trizol, then was analyzed by a spectrophotometer, and the RT-PCR was performed for the evaluation of IL-1Ra, TNF-[alpha], IL-6 and IL-1[beta]. Preliminary results show that LPS was non-stimulatory for lipid peroxidation and iNOS production. TNF-[alpha] activity was undetectable. IL-6 and IL-1Ra showed a dose response correlation at FDP concentrations of 500 [micro]g/ml, 250 [micro]g/ml, and 100 [micro]g/ml. These preliminary results present the possibility that FDP influences IL-6 and IL-1Ra mRNA expression.

10:50 EFFECTS OF PFT [alpha] (PIFITHRIN-[alpha], P53 INHIBITOR) ON CELL SURVIVAL AND TRANSACTIVATION OF KNOWN P53 RESPONSIVE GENES USING RAT LIVER CELLS AND HEPG2 CELL LINE

Ibrahim Farah* and Rowshan A. Begum, Jackson State University, Jackson, MS 39217

Pifithrin-[alpha] (PFT-[alpha]) is a reversible inhibitor of down stream function of p53. Responding to genotoxic agents, normal cells are instructed by p53 to either perform DNA repair or to commit suicide. Chemo and/or radiotherapy damage both normal and cancerous cells indiscriminately. The objective therefore, was (1) to evaluate PFT-[alpha] for differential protection in response to arsenic trioxide and cadmium chloride exposure of normal and neoplastic cells, and (2) to evaluate the transcriptional activation of p53 and p53-responsive genes in rat liver cells and HepG2 carcinoma cell line. Cells were cultured to 90% confluency and subsequently exposed to cytotoxic agents in presence or absence of PFT-[alpha] (10 ug/ml) for a 24 hour period. Cell survival was detected by fluorospectroscopy (FDA). Percent survival indices (LC50) were calculated using regression analysis. Mean LC50 and (SD) for HepG2 cells following exposure to arsenic were 13.7 ([+ or -]1.0) [micro]g/ml with PFT-[alpha] and ([+ or -]2.5) [micro]g/ml with PFT-[alpha] and 573.15 ([+ or -]1.0) [micro]g/ml without PFT-[alpha]; (p<0.5). With rat liver cells exposed to cadmium chloride the LC50 was found to be 57.72 ([+ or -]0.8) and 58.1 ([+ or -]5.5) [micro]g/ml; (p>0.5), in presence of PFT-[alpha] and in its absence respectively. Significant differences from controls upon exposure to arsenic trioxide in presence and absence of PFT-[alpha] were only seen in rat liver cells. PFT-[alpha] inhibited the transactivation of p53 in rat liver cells and resulted in repression of bc12, PCNA, mdm2, cyclin G and p21 genes in response to arsenic. HepG2 cells exposed to arsenic trioxide and PFT-[alpha]a showed extensive expression p53 and PCNA.

11:00 THE CONTRIBUTION OF IFN-[gamma] AND IL-18 GENOTYPE POLYMORPHISM TO THE ALLOGRAFT FUNCTION IN AFRICAN-AMERICAN RENAL TRANSPLANT RECIPIENTS

Xinchun Zhou*, W. Henry Barber, Donald E. Butkus, Larry S. McDaniel, Brenda D. Mangilog, and D. Olga McDaniel, University of Mississippi Medical Center, Jackson, MS 39216

It was hypothesized that a high producing IFN-[gamma] allele might be influential in predisposing the recipient to allograft rejection and the IL-18 cytokine genotype might play an important role in the induction of IFN-[gamma]. Thus, the purpose of this study is to characterize the IL-18 genotype, in relation to IFN-[gamma] production and the outcome of allograft function in kidney recipients. Cytokine gene polymorphism were evaluated in 76 African-American patients who undergone renal transplantation. The frequency distribution of cytokines were analyzed in respect to the clinical characterizations, including delayed graft function (DGF), rejection episodes (REs) and stable graft function (SGF). We have shown that the IFN-[gamma] T/A intermediate producer genotype was associated with allograft rejection (50% in REs, 20.5% in SGF, p < 0.01, RR = 0.4), whereas the IFN-[gamma] low producer genotype was significantly protective of the allograft (38.5% in REs and 74.4% in SGF, p < 0.005, RR = 2.85). The IL-18 CA/GC intermediate producer genotype was found in a higher frequency in recipients with REs as compared with SGF and controls (38.5%, 15.4%, and 23.4%, respectively). A combined effect of IFN-[gamma] T/A and IL-18 CA/GC genotypes was observed in seven recipients with REs, and was absent in recipients with SGF (p < 0.001). These finding support a role for IL-18 induction of IFN-[gamma] and might provide better understanding of the posttransplatation cytokine release and the management of allograft survival.

11:10 DEVELOPMENT OF A NOVEL IMAGE GUIDED TECHNIQUE FOR THE ASSESSMENT OF OSTEOGENESIS USING A RAT MODEL WITH INDUCED BONE TRAUMA IN PRE-CLINICAL MODE

Rishi Roy*, Michelle Tucci, Ramesh Patel, Ashraf Ragab, George Russell, and Hamed Benghuzzi, University of Mississippi Medical Center, Jackson, MS 39216

Evaluation of osteogenesis following surgically induced and immobilized fractures in laboratory animals over a period of time in a sequential fashion often involves enrollment of a large number of animals. A certain number of animals have to be periodically sacrificed to adequately assess the progress of the experiment. The goal of this project was to explore the feasibility of using imaging technique as a tool to serially evaluating surgically induced femoral fractures in a rat model. Adult male rats were (n = 50) were divided into experimental (5 mm femoral segmental defect [FSD], and FSD plus Delivery Capsules), and control groups. All surgical procedures were conducted following IACAUC approved protocol. Live animals were subjected to imaging setting following anesthesia (Xylazine, 10 mg/kg; Ketamine, 75 mg/kg). Digitized data were generated by rotating head angiographic unit as well as a multi-slice Computerized Tomographic scanner with helical method and processed in dedicated computer systems with algorithms to reconstruct 3-dimensional images in various angles with the capabilities of subtracting overlying soft tissues or bones. The radiographic findings [3-dimensional mode] were analyzed for definition of the immobilizing hardware, appearance of the fracture sites, proximity and appearance of the implanted bone-growth enhancing delivery capsules contiguous to the fracture sites. Results of this project suggest that (I) the use of imaging methods may offer an advantage by reducing the number of animals that have to be involved in the experiment, and (II) the use of imaging methods could provide an accurate assessment of the status of the healing process (termination of treatment vs additional treatment) without sacrificing the animals (predicting prognosis). *Undergradute Student Award Category Competition

FRIDAY AFTERNOON

Gulf Hall

1:00-2:15 Workshop on CPR (Family and Friends)

Martha Howard, Karen Bell, and Chris Powell, University of Mississippi Medical Center, Jackson, MS 39216; American Heart Association, Mississippi Chapter, Hattiesburg, MS; and AMR, Gulfport, MS

FRIDAY AFTERNOON

Emerald

Special Symposium

GIS and Remote Sensing in Health Sciences

Organized by: The University of Mississippi Medical Center

Geospatial information technology, such as GIS and Remote Sensing, provide powerful analytical, visual, and operational tools for understanding of our world. The complex analysis of the intersection of environmental, economic, political, medical, and social conditions that affect our well-being is one of the many benefits of such tools. As health organizations strive to meet the demands of a resource-constrained world, the use of GIS in service logistics has taken on even greater value. Such technology has been a key factor in developing intelligent solutions that help health professionals eradicate and control infectious disease, allocate scarce government and private resources, and increase the efficiency of the resources that are available

Mississippi is a leader in applying GIS and RS in many areas, including health. This year, as an academic partner of NASA, UMMC hosted two collaborative meetings between NASA and CDC with aims of exploring efficient utilization NASA's technology in studying diseases, particularly vector-born diseases, as well as in tracking public health environmental factors. This proposed mini-symposium will provide an opportunity for a wide group of audience from this state to hear national experts, which hopefully will enhance a broader collaboration in Mississippi.

1:00 Welcome and Introduction

D. Olga McDaniel and Hamed Benghuzzi, University of Mississippi Medical Center

1:05 Speakers' Introduction

Faruque Fazlay, Director of GIS, University of Mississippi Medical Center

1:15 William Davenhall

Manager, Health and Human Services Solutions Group ESRI, Inc., Redlands, California

2:00 Dr. Frances J. Mather

Assistant Dean for School of Public Health and Tropical Medicine Tulane University

2:25 Break

2:35 Dr. Robert A. Venezia

Program Manager for Public Health Applications, NASA Headquarters

3:00 Dr. Tim Orsi

Project Coordinator-Harmful Algal Blooms Observing System (HABSOS); NOAA National Coastal Data Development Center (NCDDC)

3:25 TBA--Stennis/ATSDR

3:50 Dr. David Dzielak

Associate Vice Chancellor for Research, University of Mississippi Medical Center

4:05 Health Sciences Division Business Meeting, Election of New Officers, and Awards

4:30 Meeting Adjourns
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Date:Jan 1, 2004
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