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Section V: Biomedical Sciences.

8:15 PRELIMINARY DATA ON THE LONG-TERM SURVIVAL AND FUNCTION OF EMBRYONIC CHICK HEARTS MAINTAINED IN VITRO **, Lindsey Parks *, Terry Archer-Liefde-Chance and Army Lester, Kennesaw State University, Kennesaw, GA 30144. Cardiovascular diseases are a major health problem for the US and many other parts of the world. Scientists are constantly looking for effective and efficient ways to study this class of diseases. This study is based on the hypothesis that maintaining the appropriate culture environment will enhance the normal survival and development of embryonic chick hearts maintained in vitro, thus making the system more suitable for cardiovascular studies. Fertilized chicken eggs were incubated for 4-11 days and the embryonic hearts quickly dissected out. The hearts were placed in varying culture media supplemented with 10% fetal calf serum, and antibiotics. The hearts were placed in an [O.sub.2] or [CO.sub.2] enriched incubator and maintained at 37.5[degrees]C. The hearts were monitored daily for survival rate, growth, development, and pattern of contraction. Results indicate no significant difference in heart survival rates between the two varied gas concentrations. Hearts taken from 5-day-old embryos appeared to survive better than older and younger hearts. Strength of heartbeat appeared highly varied among organs of the same age. The strength of contractions appeared to diminish with length of culture. Contractions appeared to completely cease within the ventricles, while the beating became highly reduced to the atria region. Additionally, there was very little noticeable growth in the any of the hearts, however in most cases, cells from the hearts heavily populated the culture dishes. These results indicate that it is possible to culture embryonic hearts for up to two weeks while maintaining a beat. Modifications are needed to promote heart growth and uniformity in heart rate and strength.

8:30 IS OBESITY A PROBLEM IN COLLEGE STUDENTS? **, Katie Rousseau and Deepa Arora, Middle Georgia College, Cochran, GA 31014. Obesity is a growing problem in the US. According to a report by the Centers for Disease Control, in 2008, the prevalence of obesity was greater than or equal to 25% in thirty-two states and below 20% in only one state. Since obesity is a predisposing factor for diabetes and cardiovascular problems, it is important to combat this growing menace. The objective of this study is to examine the prevalence of obesity with its contributing factors in a rural community college in the southeast. At least fifty students will be randomly selected from the college campus to participate in the study. After obtaining informed consent, they will be asked to record their socio-demographic information, educational level, physical activity status, and their detailed dietary intake for five consecutive days. Each respondents average dietary intake of energy, proteins, carbohydrates, and fat will be calculated. Results obtained will be compared to the Dietary Reference Intakes proposed by the Food and Nutrition Board of the Institute of Medicine, National Academy of Sciences. This preliminary study will help to determine if college students in a rural area meet the objectives of "Healthy People 2010."

8:45 DECREASED PRESSURE AND INCREASED DEHYDRATION AS INDICATORS OF THE PROBLEMS ASSOCIATED WITH SHELL-LESS CHICK CULTURES **. Terry Archer-Liefde-Chance *, Lindsey Parks, Army Lester, Kennesaw State University. Kennesaw, GA 30144. Shell-less chick cultures offer much promise in a myriad of biological studies. However, shell-less embryos with no prior in ovo incubation typically die during the first 10 days of culture. Embryos that survive longer experience reduced growth and development and all die prior to hatching. This study investigates the hypothesis that removing the eggshell leads to low atmospheric pressure around the embryo, thus decreasing available oxygen and promoting dehydration. After 0, 24, 48, or 72 hours of in ovo incubation, embryos were cultured on a sheet of gas permeable plastic wrap, placed in a clear plastic cup-like culture vessel at various oxygen levels, saturated humidity levels and 37.5[degrees]C. Embryos were monitored daily for survivability, growth and development. Preliminary results showed that mortality rates increased as the hours of in ovo incubation decreased, while embryos cultured in an oxygen-enriched environment experienced no increase in mortality. Embryos cultured at high pressures survived similarly to embryos cultured under high oxygen. The yolk of shell-less embryos often appeared very viscous with a greenish bile color. These findings suggest that the open culture system led to a reduced partial pressure of oxygen and thus inhibited survival of embryos before the development of a vascular system. The decreased pressure also led to dehydration thus causing the yolk to become too viscous for absorption, which may have caused the observed lower rate of growth and development. The liver may have attempted to compensate by increasing bile production and hence accounting for the greenish color of the yolk and liver.

9:00 Break

9:15 TARGETING TRYPANOSOMA BRUCEI CALCIUM ATPASES AND CHANNELS: A POTENTIAL STRATEGY FOR DISRUPTING CALCIUM HOMEOSTASIS, Kiantra Ramey * (1), Zuzana Kucerova (2), Winston Thompson (1) and Jonathan K. Stiles (1), (1) Morehouse School of Medicine, Atlanta, GA 30310 and (2) Centers for Disease Control and Prevention, Atlanta, GA 30333. Trypanosoma brucei causes Human African Trypanosomiasis (HAT). 200-300 million people are affected by this disease with an estimated 50,000 deaths annually in sub-Saharan Africa. Existing drugs for HAT are toxic and often times lethal and parasite resistance is common due to having an impervious membrane and antigenic variation. As a result vaccine development against HAT has been unsuccessful, thus there is a need for an effective vaccine that targets the accessible flagellar pocket membrane proteins. We previously synthesized T. brucei [Ca.sup.2+], ATPase and channel peptides and determined their expression and localization in the flagellar pocket of parasites where they are involved in [Ca.sup.2+] homeostasis. Commercial Ca2+ pump inhibitors and channel blockers were tested against parasites which decreased parasite survival. We hypothesized that inhibiting T. brucei [Ca.sup.2+] ATPases and channels together will disrupt intracellular [Ca.sup.2+] ion ([Ca.sup.2+].sub.i]) levels and arrest growth in parasites. To test this hypothesis we performed drug and antibody inhibition assays using [Ca.sup.2+] pump and channel inhibitors and antibodies respectively on blood stage parasites. Intracellular [Ca2+]i was determined by assaying for levels of fura-2, a [[Ca.sup.2+].sub.i] indicator. Results indicate that combining [Ca.sup.2+] pump inhibitors and channel blockers decreased intracellular [[Ca.sup.2+].sub.i] and inhibited parasite growth greater than when treated individually. Combining pump and channel antibodies also reduced parasite growth more effectively than alone. Therefore, targeting both [Ca.sup2+] pump and channel proteins simultaneously reduces the capability of parasites to utilize alternative strategies for [[Ca.sup.2+]] homeostasis. Thus, this is a plausible approach for developing new drugs and/or vaccines targeting [[Ca.sup.2+.+]] homeostasis in African trypanosomes. This study was supported by the Minority Biomedical Research Support Program, NIH-NIGM-MBRS/RISE (GM58268), at Morehouse School of Medicine and the National Institutes of Health, NIH-RCMI (RR033062).

9:30 CHLAMYDIA TRACHOMATIS CRYPTIC PLASMID ANTIGENS IN RECOMBINANT pGKVAX CHLAMYDIA VACCINE DEVELOPMENT **, A. Campbell *(1), E. Ekong (2), G. Ifere (1), K. Joseph (3), T. Belay (1), E. Barr (1) F. Eko, (2) C. Black (3), J. Igietseme (2), (3) and G. Ananaba (1), (1) Clark Atlanta University, (2) Morehouse School of Medicine and (3) Centers for Disease Control & Prevention, Atlanta, GA. Chlamydia trachomatis genital infection is a prevalent sexually transmitted disease that is often asymptomatic. Developing a vaccine is an ideal way to protect against this disease and its pathology. A strategy is the development of a vaccine that utilizes Lactobacillus as a live delivery vehicle of chlamydia antigens to the immune system. Many chlamydial species contain a 7.5-kb cryptic plasmid (pCT). The role of pCT in the pathogenesis of Chlamydia trachomatis genital infection is unknown. We hypothesize that cryptic plasmid antigens may be used in the development of an effacious vaccine against chlamydial genital infection. We have evaluated the effect of chlamydia cryptic plasmid in fertility, and found that plasmid deficient Chlamydia did not cause pathology. Additionally, we have isolated the eight open reading frame genes. The pCT antigens, p-glycoproteins or pgps were isolated from pGEX/pgp plasmid construct using a series of different restriction enzyme digestions. The pgp 2 and pgp3 have been ligated with pGKVAX and cloned via JM109 transformation. Positive JM109 E. coli transformants were cultured on LB chloramphenicol selective media plates. Qiagen Miniprep was performed to isolate the plasmids from each culture. Lactobacillus fornicalis was transformed with pGKVAXpgp by electroporation and cultured on Lactobacillus MRS chloramphenicol selective media plates. Live recombinant Lactobacillus has the potential to produce an efficacious vaccine against Chlamydia genital infection pathology. Supported by NIH grants GM08247 and A141231.

10:00 Section business meeting

10:30 ENTEROCOCCUS ISOLATES FROM COMMERCIAL MEATS, Michael W. Reeves, Perimeter College, Covington, GA 30014. In previous studies, Enterococcus have proven an excellent indicator of fecal contamination of commercial foods, particularly processed meats. To examine if this problem still exists, commercial and local meats were purchased from four stores in the Athens, GA, area over a period of six months. Samples were tested for contamination in tryptic soy broth. Enterococcus were identified by growth on bile-esculin-azide agar, by gram stain and catalase reaction, and growth in 6.5% salt. Species were identified by sugar fermentation patterns. All meats examined contained a low level of bacterial contamination with the exception of a single sliced ham product. Of thirty-four meats tested, fifteen contained Enterococcus, and these meats were all locally ground beef and pork. None of the national commercial brands contained Enterococcus. The most common species isolated was E. faecalis. Antibiotic testing showed that none of the isolates were resistant to vancomycin. Isolates from one store were all E. faecalis with similar antibiotic patterns, suggesting a single point source of contamination. Isolates from other stores were more varied, suggesting multiple sources of contamination. These results show that Enterococcus contamination is specifically a local problem, and that protection programs must be organized at that level.

10:45 PLASMODIUM BERGHEI ANKA INFECTION UP REGULATES FOXP3 AND IL-10, AND DOWN REGULATES TGF-[beta]l IN IP-10 DEFICIENT C57BL/6 MICE, Bismark Sarfo (1), Nana Wilson * (1), Danielle Whittaker * (2), Vincent Bond (1) and Jonathan Stiles (1), (1) Dept. Microbiology, Biochemistry, and Immunology, Morehouse School of Medicine, Atlanta, GA 30310 and (2) Vanderbilt University, Nashville, TN 37235. The mechanism mediating cerebral malaria (CM) is not clear, although sequestration of infected red blood cells in the brain and high production of pro-inflammatory factors such as IP 10, have been implicated. C57BL/6 mice deficient for IP-10 are less susceptible to experimental CM. Depletion of IP-10 enhanced the production of regulatory T cells (T regs), IL-10 and TGF-[beta]1 which regulate excessive production of pro-inflammatory factors. We hypothesized that deletion of IP-10-/- protects against ECM due to modified expressions of T regs, IL-10 and TGF-[beta]1. To test this hypothesis, IP-10-/- and WT mice were infected with Plasmodium berghei ANKA, and brain, peripheral blood mononuclear cells (PBMCs) and plasma were analyzed for Foxp3, IL-10 and TGF-[beta]1. T-cells were isolated from non T cells from PBMCs using antibody coated magnetic beads, and T regs (CD4+CD25+) and non-T regs (CD4+CD25-) were subsequently isolated, and restimulated with P. berghei antigens with co-stimulants PMA and ionomycin in vitro. The supernatants from this restimulation assay were analyzed for IL-10 and TGF-[beta]1. Infected WT but not IP-10-/-mice exhibited CM symptoms. P. berghei induced high Foxp3 mRNA (p < 0.05) in brain and PBMC's of infected IP-10-/- at days 2 and 4 compared with WT. Plasma IL-10 in infected IP-10-/- mice was up regulated (p < 0.05) at days 2 and 4 than in infected WT. In contrast, at day 2 and 4, TGF-[beta]1 in infected WT was significantly up regulated (p < 0.05) compared with infected IP-10-/- mice. Ex-vivo CD4+CD25+ and CD4+CD25- T cells re-stimulated with P. berghei antigens produced higher amounts of IL-10 and TGF-pi than those re-stimulated with supernatants from uninfected cells. P. berghei antigen re-stimulated CD4+CD25+ T cells from IP-10-/- produced higher IL-10 and TGF-pl than WT CD4+CD25+ T cells. Deleting IP-10 and early activation of T-regs in conjunction with IL-10 are important in preventing ECM.

POSTERS

EPIGENOMIC REGULATION OF VEGFR2 BY LEPTIN-OTCH SIGNALING CROSS-TALK IN MAMMARY CANCER CELLS, Shanchun Guo *, Yanbo Xu, Miles Fuller and Ruben R. Gonzalez-Perez, Microbiology, Biochemistry & Immunology, Morehouse School of Medicine, Atlanta, GA 30310. The vascular endothelial growth factor (VEGF) receptor family in mammals contains three members, VEGFR1 (Flt-1), VEGFR2 (KDR/Flk-1) and VEGFR3 (Flt-4), which are transmembrane tyrosine kinase receptors and directly regulate the formation of blood and lymphatic vessels. VEGFR2 is generally recognized as the major form that mediates VEGF-induced response and the earliest marker for endothelial cell development. More importantly, VEGFR2 directly regulates tumor angiogenesis. We have previously reported that leptin signaling plays an important role in the growth of both ER+ and ER- breast cancer that is associated with the leptin regulation of pro-angiogenic and pro-proliferative molecules, i.e., VEGF/VEGFR2. Disruption of leptin signaling with pegylated leptin peptide receptor antagonist (PEG-LPrA2) markedly reduced the growth of tumors and the expression of VEGFR2 in mouse models of syngeneic and human breast cancer xenografts. On the other hand, it is generally believed that Notch signaling is essential for tumor angiogenesis. We hypothesized that Notch and leptin signaling crosstalk could impact the expression of pro-angiogenic molecules, especially VEGFR2. In the present studies, the mouse VEGFR2 5'-end transcription region was cloned and used to establish VEGFR2-Luc assay in mouse mammary tumor (MT) cells. We found that VEGFR2 expression is heavily depend on VEGFR2 gene methylation and histone acetylation status in MT cells that maybe linked to leptin's regulatory effects. Interestingly, leptin signaling affects several important members of Notch family (Notch 2 and Notch 3) in MT cells. This suggests leptin and Notch signaling crosstalk could impact on MT angiogenesis and growth. Our results provide novel evidence on how VEGFR2 could be regulated in MT cells. Present data reinforce the idea that disruption of leptin signaling could reduce tumor angiogenesis/growth by inhibiting leptin-mediated upregulation of VEGFR2 and/or by negatively impacting on signals from some key members of Notch family. This might help to design new pharmacological strategies aimed at controlling breast cancer growth and angiogenesis. [This work was supported in part by NIH/NCI 1SC1CA138658-02; NIH/ARRA/3SC1CA138658-02S1; NIH/UAB Breast SPORE Career Development Award and the Georgia Cancer Coalition Distinguished Cancer Scholar Award (to RRGP)].

MUTUAL EXCITATION AMONG OLFACTORY BULB MITRAL CELLS REVEALED BY RECURRENCE TIME HISTORY MAPPING (RTHM) **, Alexandra Radu *, Maame Boateng *, Henaa Razzak * and Barry K. Rhoades, Wesleyan College, Macon, GA 31210. In the mammalian olfactory bulb coordinated neural activity associated with odor discrimination is dominated by narrow-band oscillations in the gamma EEG range (30-80 Hz), gated by respiratory inspiration. Mutual excitation among the mitral cell projection neurons is a required feature of successful bulbar models, but has not been conclusively demonstrated by either direct histological or electrophysiological evidence. In the present study tungsten-steel microelectrodes were lowered to the mitral cell layer of the olfactory bulb in urethane- or pentobarbital-anesthetized rats, and positioned using electrophysiological response criteria. Ten to twenty minute samples of resting neuronal activity were recorded to analog tape. A template matching system was used to isolate and extract multiple, simultaneous single-unit spike trains. Temporal linkages between spike trains were evaluated using recurrence-time history mapping (RTHM), a algorithm for enhancing conventional conditional cross-correlograms. In the RTHM analysis plots of some pairs of mitral cells apparent mutual excitation was evidenced by the presence of high-density, short-latency linkage bands and a within-band pattern suggesting activity-dependent gain. The validity of inferences based on RTHM was verified using artificially-generated spike trains with stochastically-defined firing patterns and inter-neuronal linkages.

A SURVEY OF THE ANTIBIOTIC RESISTANCE OF BACTERIAL SPECIES IN THE EAR OF CANIS FAMILIARIS, Kristin S. Timmons *, Christopher S. Bates and Richard D. Griner, Department of Biology, Augusta State University, Augusta, GA 30904. Recurring ear infections in canines are a common problem encountered in veterinary medicine and are usually associated with high bacterial growth in the ear canal. For this reason, we chose to examine the bacterial flora in the external ear canal of healthy canines and canines with infected ears. A survey was conducted using 19 randomly selected infected and non-infected canine ears. From these, over 30 bacterial colonies were isolated and were subjected to gram staining and disk diffusion antibiotic susceptibility testing. Most of the bacterial species isolated from the ears were gram positive and demonstrated increased resistance to several antibiotics. Specifically, 44% of the bacteria were resistant to sulfisoxazole, 31% were resistant to vancomycin, and 16% were resistant to ciprofloxacin. A lower incidence of resistance to streptomycin, doxycycline, and erythromycin was observed. The large percentage of vancomycin resistant bacterial species was highly unexpected since this antibiotic is not commonly used to treat ear infections in canines.

Cunningham Center, room 315

Seyed H. Hosseini, Presiding
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Author:Hosseini, Seyed H.
Publication:Georgia Journal of Science
Geographic Code:1U5GA
Date:Mar 22, 2010
Words:2825
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