• Cardiometabolic Biomarkers in Young Black Girls: Relations to Body Fatness and Aerobic Fitness, and Effects of a Randomized Physical Activity Trial

      Gutin, Bernard; Harris, Ryan A.; Howe, Cheryl A.; Johnson, Maribeth H.; Zhu, Haidong; Dong, Yanbin; Georgia Institute for Prevention of Human Diseases and Accidents; Department of Pediatrics; Department of Biostatistics and Epidemiology (2011-10-11)
      There is little evidence from randomized trials showing that physical activity alone influences biomarker profiles in youths. This study tested two hypotheses: (i) that elevated body fatness and poor fitness would be associated with unfavorable levels of cardiometabolic biomarkers in 8- 12-y-old black girls (n = 242) and (ii) that a 10-mo PA intervention would have favorable effects on the fatness-related cardiometabolic biomarkers. At baseline, all fatness indices (i.e., percent body fat, visceral adipose tissue, BMI, and waist circumference) were significantly (P < 0.05) associated with unfavorable levels of insulin, glucose, systolic BP, diastolic BP, triglycerides, C-reactive protein (CRP), and fibrinogen. Aerobic fitness was significantly (P < 0.05) associated with favorable levels of insulin, CRP, fibrinogen, and HDL2. The PA intervention had significant and favorable effects on fitness, fatness, and two biomarkersâ resting heart rate and LDL cholesterol. More research is needed to clarify what types of interventions can enhance the cardiometabolic health of youths.
    • Cell drinking: a closer look at how macropinocytosis drives cholesterol uptake in atherosclerotic vessels

      Lin, Huiping; Vascular Biology Center (Augusta University, 2020-05)
      Atherosclerotic vascular disease is the underlying cause of myocardial infarction, stable and unstable angina, stroke, peripheral artery disease and sudden cardiac death. Collectively, these cardiovascular diseases are responsible for the majority of deaths worldwide. Internalization of modified apolipoprotein B–containing lipoproteins by macrophages through scavenger receptor (SR)-mediated pathways is generally viewed as an essential step for the initiation and progression of atherosclerosis. Our studies were designed to investigate the contribution of receptor-independent LDL macropinocytosis to arterial lipid accumulation and atherosclerosis. We developed novel genetic and pharmacological approaches, utilized high resolution imaging techniques and employed unique in vivo lipid quantification assays to investigate the role of macrophage macropinocytosis in the pathogenesis of atherosclerosis. My results demonstrate that the macropinocytosis inhibitor EIPA and selective deletion of a key pathway regulating macropinocytosis in myeloid cells substantially decreased lesion size in both hypercholesterolemic wild type (WT) and SR knockout (CD36-/-/SR-A-/-) mice. Stimulation of macropinocytosis using genetic and physiologically relevant approaches promotes lipoprotein internalization by WT and CD36-/-/SR-A-/- macrophages, leading to foam cell formation. Serial section high-resolution imaging of murine and human atherosclerotic arteries identified for the first time subendothelial macrophages for the first time that demonstrate plasma membrane ruffling, cupping and macropinosome internalization. Immunoelectron microscopy, 3D reconstruction of macrophage foam cells and in vivo LDL tracking demonstrate macrophage internalization of LDL in human and murine atherosclerotic arteries via macropinocytosis. We next performed a large, unbiased-screen of an FDA-approved drug library to identify clinically relevant therapeutic agents that can be repurposed as pharmacological inhibitors of macropinocytosis. Our studies identified a low MW compound (imipramine) that inhibits macrophage macropinocytosis in vitro and in vivo. Imaging, toxicity and selectivity studies demonstrated that imipramine is a potent (IC50 = 130.9 nM), non-toxic (selectivity index CC50/IC50 > 300) and selective inhibitor of macropinocytosis. Repurposing of imipramine to inhibit macropinocytosis in hypercholesterolemic mice substantially decreased plaque development compared with control treatment. Taken together, our findings challenge the SR paradigm of atherosclerosis and identify inhibition of receptor-independent macrophage macropinocytosis as a new therapeutic strategy that may be beneficial in the treatment of atherosclerosis and its cardiovascular consequences.
    • Cellular and Molecular Players in Neuromuscular Junction (NMJ) Formation and Function

      Barik, Arnab; Institute of Molecular Medicine and Genetics (2014-04)
      There are three distinct segments in this dissertation. First, I attempted to address the role of Schwann cells in mammalian neuromuscular junction (NMJ) development and function. Schwann cells at the NMJs do not form myelin sheaths and are known as terminal Schwann cells. Terminal Schwann cells are thought to be analogous to astrocytes in the central nervous system. Schwann cells (as described in details in the next section) provide trophic support to motor axons and modulate synaptic activity by sensing neurotransmitter release at the nerve terminal. However, the role of Schwann cells in synapse formation and maintenance remains unknown. Second, during NMJ formation, anterograde signals from nerve to muscle, and retrograde signals from muscle to nerve are critical for the establishment of a functional synapse. Research over the last three decades has contributed to our understanding of the role of the anterograde signaling at NMJ. However, identification of muscle-derived retrograde signals involved in motoneuron terminal differentiation remains scarce. Recent work from our laboratory suggests that genes that are transcriptionally regulated by p-catenin in muscles might play a crucial role in pre-synaptic differentiation at the NMJ.2 Third, Agrin-LRP4-MuSK signaling is critical for NMJ formation. At the NMJ, LRP4-mediated activation of MuSK by neural Agrin is required for post-synaptic differentiation. Mice that lack any one of the three genes fail to form NMJs and die at birth. Due to perinatal lethality of these null mice, less is known about how Agrin-LRP4-MuSK might regulate NMJ maintenance. Moreover, mutations in Agrin, LRP4, and MuSK have been reported in patients diagnosed with congenital myasthenic syndrome (CMS), and autoantibodies against MuSK and LRP4 have been detected in patients with myasthenia gravis (MG). However, the role of Agrin-LRP4-MuSK in the etiology of these neuromuscular disorders is not clear.
    • Ceramide in Stem Cell Differentiation and Embryo Development: Novel Functions of a Topological Cell-Signaling Lipid and the Concept of Ceramide Compartments

      Bieberich, Erhard; Institute of Molecular Medicine and Genetics (2010-12-29)
      In the last two decades, the view on the function of ceramide as a sole metabolic precursor for other sphingolipids has completely changed. A plethora of studies has shown that ceramide is an important lipid cell-signaling factor regulating apoptosis in a variety of cell types. With the advent of new stem cell technologies and knockout mice for specific steps in ceramide biosynthesis, this view is about to change again. Recent studies suggest that ceramide is a critical cell-signaling factor for stem cell differentiation and cell polarity, two processes at the core of embryo development. This paper discusses studies on ceramide using in vitro differentiated stem cells, embryo cultures, and knockout mice with the goal of linking specific developmental stages to exciting and novel functions of this lipid. Particular attention is devoted to the concept of ceramide as a topological cell-signaling lipid: a lipid that forms distinct structures (membrane domains and vesicles termed â sphingosomeâ ), which confines ceramide-induced cell signaling pathways to localized and even polarized compartments.
    • Ceramide-mediated Regulation of Cell Polarity in Primitive Ectoderm Cells: A novel role for sphingolipids in morphogenesis

      Krishnamurthy, Kannan; Institute of Molecular Medicine and Genetics (2009-01)
      Ceramide is considered a key sphingolipid, regulating a variety of critical cellular processes. To facilitate the study of ceramide localization and its interaction with cellular proteins, we have developed a novel antibody against ceramide, raised in rabbit (rabbit IgG). The novel antibody specifically recognizes ceramide in lipid overlay assays and detects ceramide containing different fatty acid chain lengths (i.e. C2-, C16-, C18-, C20- and C24 ceramide). The new antibody was compared with the commercially available anti-ceramide mouse IgM antibody in immunocytochemistry experiments to study the localization of ceramide. Although both antibodies stain similar regions on the cell membrane, the rabbit IgG reveals the distribution of ceramide in intracellular compartments that are not well identified with the commercially available antibody. Pharmacological depletion or increase of ceramide levels results in a corresponding change in staining intensity, confirming the specificity of the antibody. These results indicate that the rabbit IgG is a suitable antibody to determine both the localization of ceramide, and its interaction with proteins by immunocytochemistry. To investigate the role of ceramide in early embryonic development, we used embryoid bodies (EBs) differentiated from mouse embryonic stem cells as a model. The primitive ectoderm cell layer of EBs represents the primitive ectoderm of the early embryo. In mammals, the primitive ectoderm is an epithelium of polarized cells that undergoes gastrulation and differentiates into all embryonic tissues. We find that in primitive ectoderm cells, ceramide was elevated and asymmetrically distributed to the apico-lateral cell membrane, where it was co-distributed with Cdc42 and F-actin. Pharmacological or siRNAmediated inhibition of ceramide biosynthesis impaired primitive ectoderm formation and concomitantly increased apoptosis in EBs. Primitive ectoderm formation was restored by incubation with ceramide or a ceramide analog, indicating that the observed defect was due to loss of ceramide. Ceramide depletion also prevented membrane translocation of atypical PKC (aPKC), interaction of aPKC with Cdc42, and phosphorylation of GSK-3|3. Recombinant aPKC, when bound to ceramide-containing lipid vesicles, formed a complex with the polarity protein Par6 and Cdc42. Taken together, our data suggest a novel mechanism by which a ceramide-induced, apico-lateral polarity complex with aPKC regulates primitive ectoderm cell polarity and morphogenesis.
    • Characterization o f the DNA Ligase IV and XRCC4 complex in the DNA double-strand break repair

      Lee, Kyung-Jong; Institute of Molecular Medicine and Genetics (2002-11)
      DNA double-strand breaks (DSBs) are among the most lethal forms of DNA damage. The nonhomologous end-joining (NHEJ) pathway is the principal mechanism for repairing DSBs in mammalian cells. It is also required for V(D)J recombination. There are at least four essential proteins in this pathway. These include Ku protein, DNA PKcs, and the DNA Ligase IV/XRCC4 (DNL IV/XRCC4) complex. This dissertation reports the determination of the quaternary structure of the DNL IV/XRCC4 complex, the mapping of a major human autoimmune epitope in XRCC4, the identification of DNAPKcs phosphorylation sites in XRCC4, and an investigation of the biochemical significance of XRCC4 phosphorylation. Biochemical characterization shows that DNA Ligase IV and XRCC4 form a stable mixed heterotetramer. This is the active form of the enzyme and is essential for in vitro DNA end joining in the presence of additional factors derived from cell extracts. Data shown here also demonstrate that the DNL IV/XRCC4 complex is a human autoantigen. The major autoimmune epitope maps to amino acids 251-266. This epitope coincides with several sites where XRCC4 is potentially modified in response to radiation or inflammation, including a DNA-PKcs phosphorylation site at serine 260. Results raise the possibility that radiation-induced post-translational modifications contribute to development of an autoimmune response in susceptible individuals. Previous work has shown that DNA-PKcs kinase activity is required for NHEJ, but the critical physiological target of this enzyme is not yet known. Current work shows that DNA-PKcs phosphorylates serine 318 of XRCC4, in addition to the serine 260 site described above. The presence of serine 260 increases phosphorylation at serine 318, suggesting that phosphorylation can occur sequentially. Mutation o f serine 260 reduced DNA end-joining activity and sensitivity to the PI3 kinase inhibitor (LY294002). These data provide preliminary evidence that phosphorylation of XRCC4 by DNA-PKcs contributes to regulation of DNA repair.
    • Characterization of Cervical and Head and Neck Squamous Cell Carcinomas by Proteomic Analysis

      Merkley, Mark A.; Institute of Molecular Medicine and Genetics (2009-06)
      (First Paragraph) The oral cavity, oropharynx, larynx, esophagus, and ano-genital orifices are lined with stratified squamous nonkeratinized epithelium, which forms the barrier between the underlying tissue and the external environment. The proliferative nature of this epithelium, together with its potential exposure to environmental insults such as tobacco carcinogens, alcohol, or oncogenic viruses, makes it susceptible to carcinogenesis. Indeed, carcinomas of stratified squamous nonkeratinized epithelium are among the most common and deadly cancers worldwide. In particular head and neck squamous cell carcinoma and cervical squamous cell carcinoma together account for about 1 million new cases annually, worldwide. These cancers arise from similar tissues and share common risk factors, although they differ in that effective population-based screening exists only for cervical cancer.
    • Chemokine (C-C Motif) Ligand 2 (CCL2) in Sera of Patients with Type 1 Diabetes and Diabetic Complications

      Guan, Ruili; Purohit, Sharad; Wang, Hongjie; Bode, Bruce; Reed, John Chip; Steed, R. Dennis; Anderson, Stephen W.; Steed, Leigh; Hopkins, Diane; Xia, Chun; et al. (2011-04-12)
      Background: Chemokine (C-C motif) ligand 2 (CCL2), commonly known as monocyte chemoattractant protein-1 (MCP-1), has been implicated in the pathogenesis of many diseases characterized by monocytic infiltration. However, limited data have been reported on MCP-1 in type 1 diabetes (T1D) and the findings are inconclusive and inconsistent.
    • Chetomin as a Potent Hsp90 Inhibitor

      Leibou, Stav; Lu, Sumin; Debbab, Abdssamad; Chadli, Ahmed; Georgia Cancer Center (2017-03)
      Molecular chaperones have been the focus of intense research for their important role in cancer cell homeostasis. Heat shock protein 90 (Hsp90) promotes metastasis, evasion of apoptosis, and proliferative angiogenesis in tumors through preserving the stability and functionality of its client proteins [1]. While the first generation of Hsp90 inhibitors has proven effective in hindering Hsp90 function, they have shown low clinical efficacy in part due to the induction of anti-apoptotic proteins Hsp27, Hsp40, and Hsp70 [2,3]. It is therefore our objective to develop novel efficacious Hsp90 inhibitors without these detrimental effects. During our screen for novel Hsp90 inhibitors, we found that the natural product, Chetomin, is a potent inhibitor of the Hsp90 machine chaperoning activity. Our in vitro data using human and murine mammary carcinoma cell lines suggest that Chetomin is effective in causing degradation of several known Hsp90 physiological client proteins that are crucial to cancer cell proliferation and survival. While the molecular mechanism by which Chetomin inhibits the Hsp90 function is still unclear, our data suggests that Chetomin is highly efficacious in killing cancer cells without induction of the anti-apoptotic proteins as does the first generation of Hsp90 inhibitors making Chetomin a promising new therapeutic agent.
    • Citizen Perceptions of Police in the Post-Ferguson Era: A Survey in Partnership with the Richmond County Sheriff's Office

      Hendricks, Austin; Kelley, Johnna; Gordon, Paxton; Foley, Alison, PhD; Pamplin College of Arts, Humanities, and Social Sciences (2015-08-11)
      The current study reports the results of a survey designed in conjunction with Richmond County Sheriff’s Office (RCSO) to assess citizen satisfaction with local police. Research suggests that overall satisfaction with and perceptions about police are shaped by individual’s personal experiences with police as well as their perception of neighborhood safety, sex, race, education level, and age. Based on prior research, we hypothesized that general attitudes about local police would be influenced by attitudes about American police as a whole. This is particularly important given the current national conversation revolving around police use-of-force in the wake of the highly-publicized events in places such as Ferguson, Missouri, and Baltimore, Maryland. While results are limited by the small and relatively homogenous sample of participants, linear regression models show that when race, neighborhood safety, and gender are controlled for, attitudes towards American police and attitudes towards the officer involved in their most recent encounter were significant predictors of attitudes toward RCSO. Attitudes towards the American police were the strongest predictor of satisfaction with RCSO. The majority of open-ended responses reflected negative attitudes towards police, which varied by race and sex, though many participants offered suggestions for improvement.
    • Cleft palate is caused by CNS dysfunction in Gad1 and Viaat knockout mice.

      Oh, Won-Jong; Westmoreland, Joby J; Summers, Ryan; Condie, Brian G.; Institute of Molecular Medicine and Genetics (2010-03-24)
      BACKGROUND: Previous studies have shown that disruption of GABA signaling in mice via mutations in the Gad1, Gabrb3 or Viaat genes leads to the development of non-neural developmental defects such as cleft palate. Studies of the Gabrb3 and Gad1 mutant mice have suggested that GABA function could be required either in the central nervous system or in the palate itself for normal palatogenesis. METHODOLOGY/PRINCIPAL FINDINGS: To further examine the role of GABA signaling in palatogenesis we used three independent experimental approaches to test whether Gad1 or Viaat function is required in the fetal CNS for normal palate development. We used oral explant cultures to demonstrate that the Gad1 and Viaat mutant palates were able to undergo palatogenesis in culture, suggesting that there is no defect in the palate tissue itself in these mice. In a second series of experiments we found that the GABA(A) receptor agonist muscimol could rescue the cleft palate phenotype in Gad1 and Viaat mutant embryos. This suggested that normal multimeric GABA(A) receptors in the CNS were necessary for normal palatogenesis. In addition, we showed that CNS-specific inactivation of Gad1 was sufficient to disrupt palate development. CONCLUSIONS/SIGNIFICANCE: Our results are consistent with a role for Gad1 and Viaat in the central nervous system for normal development of the palate. We suggest that the alterations in GABA signaling lead to non-neural defects such as cleft palate as a secondary effect due to alterations in or elimination of fetal movements.
    • Clinical Guide for Intraosseous Pathology

      Malik, M; Kalathingal, S; Cullum, A; Buchanan, A; Abdelsayed, A; Kurago, Z; Department of Oral Biology & Diagnostic Sciences, Center for Instructional Innovation (Augusta University, 2019)
      To provide a reference database for dental students to describe and analyze intra osseous pathology that aid to develop a list of differential diagnoses for various diseases affecting the maxillofacial region on patients treated in student clinics. The online database will serve as a resource for descriptive terminology and samples to demonstrate the origin of the lesion, radiographic appearance, borders, contents, effects on adjacent structures, etc. which are the fundamental elements that guide a clinician in developing an impression and formulate the differential diagnosis. Histopathologic evaluation that provides the final diagnosis of each disease process will also be included to demonstrate that radiographic presentation of various disease categories may be similar, however, clinical management is ultimately decided by the tissue sample from the biopsy specimen. The interactive database will have various features that enable the user to access a comprehensive glossary list, word- defined searches, a brief overview of the most common diseases affecting dento-alveolar regions and learn about the management strategies.
    • The Clinical Phenotype of An Extended Pedigree with Late-onset Alzheimer's Disease

      Fennell, Eleanor M.; Institute of Molecular Medicine and Genetics (2009-03)
      Alzheimer's disease (AD) is a devastating neurodegenerative disease with a multifaceted etiology. This retrospective exploratory study used the family history method to develop a phenotype based on the personal history of cognitive decline, changes in behavioral characteristics, and the medical, social and environmental history of the five AD affected family members in one large extended family. Group family interviews provided a description of the phenotype and identified medical and environmental risk modifiers. Average age-of-onset for AD in 5 of 12 siblings, (all homozygous APOE4/4) was 69.2 years (range 66-72 years). Fisher's exact test identified neuropsychiatric behaviors as common phenotypic manifestations; delusions (p = 0.0455), hallucinations (p = 0.0101), irritability (p = 0.0455), personality change (p = 0.0013), pacing (p = 0.0013), aggressiveness (p = 0.0455), and poor judgment (p = 0.0013). Environmental variables that emerged as significant were a less than an eighth grade education (p — 0.0152), presence of stroke/TIA (p = 0.0455), presence of osteoarthritis (p = 0.0455), and vitamin B12 deficiency (p = 0.0013). Risk of stroke/TIA may be related to the increased risk from APOE4 while vitamin B12 deficiency may be associated with advanced age. There was no significant protective benefit from the management of hypertension (p = 0.5758), use of statins to control cholesterol (p = 0.9545), use of Vitamin C (p = 1.000), and/or Vitamin E (p = 0.9899) among family members. Potential modifiable health practices among the AD unaffected siblings demonstrated that 85% continued to engage in a sedentary lifestyle (p = 0.6818), 57% were overweight (p = 0.6894), 57% (p = 0.6894) consumed an unhealthy diet, and 56% smoked (p = 0.1591). Single nucleotide polymorphism (SNP) microarray Clinical Phenotype of Alzheimer's Disease analysis indicated that several SNPs in the gene, transient receptor potential cation channel, subfamily C, member 4 associated protein, (TRPC4AP), were significant. All five of the affected siblings and three unaffected siblings exhibited one haplotype; the unaffected siblings with the same haplotype as affected siblings were younger in age and did not have any cognitive problems at the time of the study (Poduslo, Huang, Huang, Smith, 2008).
    • Cloning, Expression, and Purification of Nanoluc

      DuPlain, Holly; Parks, Jasmine; Blocker, Brittany; Spencer, Angie; College of Science and Mathematics (2015-08-07)
      Bioluminescence is a natural phenomenon that occurs in bacteria, insects, fungi and some marine species whereby light is emitted by a living organism. This emitted light is generated by a chemical reaction that occurs when a substrate is reacted upon by a class of enzymes called luciferases. Bioluminescence Resonance Energy Transfer (BRET) is a technique that relies on the use of a luciferase (energy donor) to transfer energy to a nearby fluorescent protein or dye (energy acceptor). If the donor and acceptor molecules are in close proximity and their emission and absorbance spectra overlap, the acceptor absorbs the energy from the donor (luciferase) which results in the emission of light at a longer wavelength (i.e. different color). This spectral shift can be measured and quantified. Because of the widespread applications and utility of luciferase enzymes, many assay systems have been developed that make use of various luciferases as energy donors. One such luciferase is Nanoluc (Nluc), a genetically engineered enzyme from the deep sea shrimp, Oplophorus gracilirostris. In order to explore the use of Nluc as an energy donor in BRET, we cloned the gene for Nluc into the plasmid vector, pET21c(+). The formation of the recombinant DNA was verified by agarose gel electrophoresis. After transformation of the recombinant plasmid into E. coli BL21 cells, the Nluc protein containing a C-terminal His6 tag was over-expressed and purified using affinity chromatography. The purification yielded a relatively pure protein with a molecular weight of 19 kDa as judged by SDS-PAGE. The activity of the protein was assessed by measuring its ability to generate light in the presence of the substrate coelenterazine.
    • Cloning, Over-expression and Purification of Nanoluc

      DuPlain, Holly; Parks, Jasmine; Blocker, Brittany; College of Science and Mathematics; Spencer, Angie (2015-11-13)
      Bioluminescence is a natural phenomenon that occurs in bacteria, insects, fungi and some marine species whereby a living organism emits light via a chemical reaction catalyzed by enzymes called luciferases. Luciferases are utilized in various applications including Bioluminescence Resonance Energy Transfer (BRET). BRET uses a luciferase (energy donor) to transfer energy to a fluorescent protein or dye (energy acceptor). If the donor and acceptor are close together and their emission and absorbance spectra overlap, the acceptor absorbs the energy from the donor and light is emitted at a longer wavelength. This spectral shift is measured. One such luciferase is Nanoluc (Nluc), a genetically engineered enzyme from the deep sea shrimp Oplophorus gracilirostris. To explore the use of Nluc as a BRET energy donor, the Nluc gene was cloned into the plasmid vector pET21c(+). Recombinant DNA formation was verified by agarose gel electrophoresis. After transformation of the recombinant plasmid into E. coli BL21 cells, C-terminal His6 tagged Nluc protein was over-expressed and purified using affinity chromatography. Purification yielded a relatively pure protein with a molecular weight of 19 kDa as judged by SDS-PAGE. Protein activity was assessed by measuring its ability to generate light in the presence of the substrate coelenterazine. Begin Time: 08:25 End Time: 31:16
    • Colorectal Cancer Screening Trends in Georgia: 1997-2012

      Smith, Selina A.; Ansa, Benjamin E.; Augusta University (2016-03)
    • A community-engaged approach to promoting adherence to diet-related cancer prevention guidelines

      Smith, Selina A.; Sheats, Joyce Q.; Whitehead, Mary S.; School of Medicine; Georgia Regents University; Morehouse College (2015-11-09)
    • Comparative Study of Aggression in Captive Western Lowland Gorillas with Their Wild Counterparts

      Dixon, Megan K.; Katherine Reese Pamplin College of Arts, Humanities, and Social Sciences; Trunzo, Jennifer (2015-11-13)
      Observational research on captive populations of Western Lowland Gorillas has been used to identify and understand the social patterns of captive, as well as wild, gorilla groups. My research focuses on identifying aggressive and competitive behaviors such as biting, slapping, threatening, pushing, etc. in the Western Lowland Gorilla population at Zoo Atlanta. This research is in its preliminary stages, focusing on the existing literature and studies of both wild and captive gorillas used to gain insight into the social dynamic of primates. The literature reviewed for my research focuses on the impact of aggressive behaviors on a gorilla family group, the situation the target behavior occurred in, and the types of responsive behaviors elicited from the initial aggression. Begin Time: 31:17 End Time: 59:58
    • Comparing attitudes toward addressing patient drug use among health professionals in training

      Johnson, J. Aaron; Kelly, Ursula; Georgia Regents University; Emory University (2015-06)
    • Comparison and Avoidance of Toxicity of Penetrating Cryoprotectants

      Szurek, Edyta A.; Eroglu, Ali; Institute of Molecular Medicine and Genetics; Department of Medicine; Department of Obstetrics and Gynecology; GHSU Cancer Center (2011-11-16)
      The objective of this study was to elucidate the toxicity of widely used penetrating cryoprotective agents (CPAs) to mammalian oocytes. To this end, mouse metaphase II (M II) oocytes were exposed to 1.5 M solutions of dimethylsulfoxide (DMSO), ethylene glycol (EG), or propanediol (PROH) prepared in phosphate buffered saline (PBS) containing 10% fetal bovine serum. To address the time- and temperature-dependence of the CPA toxicity, M II oocytes were exposed to the aforementioned CPAs at room temperature (RT, ,23uC) and 37uC for 15 or 30 minutes. Subsequently, the toxicity of each CPA was evaluated by examining post-exposure survival, fertilization, embryonic development, chromosomal abnormalities, and parthenogenetic activation of treated oocytes. Untreated oocytes served as controls. Exposure of MII oocytes to 1.5 M DMSO or 1.5 M EG at RT for 15 min did not adversely affect any of the evaluated criteria. In contrast, 1.5 M PROH induced a significant increase in oocyte degeneration (54.2%) and parthenogenetic activation (16%) under same conditions. When the CPA exposure was performed at 37uC, the toxic effect of PROH further increased, resulting in lower survival (15%) and no fertilization while the toxicity of DMSO and EG was still insignificant. Nevertheless, it was possible to completely avoid the toxicity of PROH by decreasing its concentration to 0.75 M and combining it with 0.75 M DMSO to bring the total CPA concentration to a cryoprotective level. Moreover, combining lower concentrations (i.e., 0.75 M) of PROH and DMSO significantly improved the cryosurvival of MII oocytes compared to the equivalent concentration of DMSO alone. Taken together, our results suggest that from the perspective of CPA toxicity, DMSO and EG are safer to use in slow cooling protocols while a lower concentration of PROH can be combined with another CPA to avoid its toxicity and to improve the cryosurvival as well.