Browsing by Subject "Endocrine System"
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Item Review of Blunt Pancreatic Trauma and Its Outcome(2007-12-01) Sanghvi, ChiragSanghvi, Chirag. Review of Blunt Pancreatic Trauma and Its Outcome. Master of Public Health (Biostatistics), December 2007, 37 pp., 4 tables, bibliography, 42 titles. Blunt pancreatic trauma (BPT) is an uncommon injury involving 1%-2% of blunt abdominal traumas but it can be associated with a high complication rate. Various studies have shown complication rate to range from 30%-64% following blunt pancreatic injury. With Institutional Review Board (IRB) approval a retrospective chart review study was performed for last 12 years. Chart review failed to support the hospital assigned diagnosis of BPT in 3 patients, leaving 37 cases for analysis. BPT had an overall mortality rate of 22%. In the patients surviving the injury, early surgical intervention (≤12 hours) had a complication rate of 31% versus 57% for delayed surgical intervention ([greater than] 12 hours). BPT has a high mortality rate but is usually secondary to other organs involved in the injury. Delayed surgical intervention has a higher complication rate compared to early intervention.Item The Role of Calcineurin and NFAT in the Regulation of Insulin Gene Transcription(2001-12-01) Lawrence, Michael C.; Richard Easom; Julian Borejdo; Ladislav DoryLawrence, Michael C., The Role of Calcineurin and NFAT in the Regulation of Insulin Gene Transcription. Doctor of Philosophy (Biomedical Sciences), December 2001, 185 pp., 41 illustrators, references, 222 titles. In an effort to understand glucose and hormone regulated insulin gene transcription elicited by increased intracellular calcium, a novel pathway has been identified. This pathway involves the calcium/calmodulin-dependent phosphatase 2B (calcinuerin) and nuclear factor activated T-cells (NFAT), which in the studies herein, have been determined to up-regulate insulin gene transcription in response to glucose and glucagon-like peptide-1 (GLP-1) in pancreatic β-cells. Three NFAT elements within the first 410 base pairs of the rat I insulin gene promoter were first identified, two of which are conserved (by presence and location) among mammals including dogs, mice, and humans. The presence of NFAT in rat insulinoma β-cells (INS-1) and rat pancreatic islets was detected by immumobotting, immunofluorescence, and RT-PCR. Electrophoretic mobility shift assays displayed NFAT-specific DNA-binding activity that could be competed with unlabeled NFAT probe when incubated with INS-1 cells or rat islet nuclear extracts and shifted with extracts pre-incubated in the presence of either anti-calcineurin or anti-NFAT antibodies. Transfection experiments with either the -410 rat I (rIsnI-Luc) or the NFAT-Luc promoter-reported showed increased promoter activity when stimulated by glucose or cell depolarization (increases intracellular calcium) and displayed a synergistically enhanced response when co-stimulated with glucose and GLP-1. The GLP-1 induced responses were mimicked by forskolin and concentration-dependently inhibited by each of two selective but distinct protein kinase A (PKA) inhibitors, H-89 and myristoylated PKI (14-22) amide. The selective calcineurin-inhibitor FK506, as well as the chelatin of intracellular Ca2+ by BAPTA, also abolished the effects of high glucose and GLP-1. Moreover, co-transfection experiments with a constitutively active form of calcineurin and the promoter-reporters (rISnI-Luc and NFAT-Luc) showed increased reporter activity over controls. Furthermore, two-point base pair mutations in any of the three identified NFAT sites within the rat insulin I promoter resulted in a significant (p [less than] 0.05) reduction in the combined effect of glucose and GLP-1. These studies establish the presence of NFAT in insulin-secreting cells, its ability to bind elements within the insulin gene promoter, and show that glucose and GLP-1 synergistically enhance NFAT-mediated insulin gene transcription by PKA- and calcineurin-dependent pathways in pancreatic β-cells.Item The Role of Estrogen and Estrogen Analogues in Friedreich’s Ataxia Cytoprotection(2014-05-01) Richardson, Timothy E.; Simpkins, James W.Friedreich’s ataxia (FRDA) is the most common form of inherited ataxia in the world, affecting roughly 1:50,000 people in the United States. It is inherited in an autosomal recessive manner due to a GAA trinucleotide repeat expansion in the first intron of the FXN gene on chromosome 9q13-21, causing gene silencing and a functional absence of the mitochondrial-localizing protein frataxin. The frataxin protein is responsible for the assembly of iron-sulfur centers in mitochondrial proteins, including the electron transport chain complex I-III, heme synthesis, as well as removing iron from around the mitochondria, preventing the formation of reactive oxygen species (ROS). The loss of FXN function causes an accumulation of mitochondrial iron and ROS, as well as impaired function of Fe-S centers in mitochondrial proteins, leading to mitochondrial damage and a decrease in activity of mitochondrial complexes I-III. The damaged mitochondria are unable to match ATP production to the cell’s energy requirements, resulting in cell death. High energy use cell types, such as neurons and cardiac myocytes, depend almost entirely on oxidative phosphorylation, leaving them especially vulnerable to the mitochondrial damage caused, and it is for this reason that these tissues are the most severely affected by the pathogenesis of FRDA. Cellular models of Friedreich’s Ataxia have employed L-buthionine (S,R)-sulfoximine (BSO), a chemotherapeutic agent which blocks the rate limiting step of de novo glutathione (GSH) synthesis, catalyzed by gamma-glutamylcysteine synthetase. Studies have shown that donor fibroblasts from Friedreich’s Ataxia patients are extremely susceptible to this BSO induced oxidative stress, while fibroblasts from healthy patients are not, due the presence of functional frataxin to absorb the increased load of cellular ROS when GSH is inhibited. Currently, there are few effective treatment modalities for FRDA. Historically, treatment has been focused on palliative care: patient counseling, genetic counseling for prospective parents, speech therapy, physical therapy, wheelchair and other ambulatory device use, propranolol for tremors, dantrolene sodium for muscle spasms and symptomatic treatment for heart disease and diabetes. Recently, antioxidant and mitochondria specific iron chelation therapy have both been proposed as possible therapies to treat the root cause of FRDA. Iron chelation therapy works by a similar principal, removing the iron from around the mitochondria, preventing the formation of free radicals and preventing the associated mitochondrial damage. The neuroprotective effects of 17β-estradiol (E2) have been clearly documented for more than a decade in a variety of disease states involving mitochondrial disruption, but the exact mechanism of action is currently poorly understood. Although the neuroprotective effects of estrogens have never been tested in an FRDA model and FRDA shows no gender-bias in incidence, some epidemiologic studies of FRDA have shown a better prognosis in female patients. Since there is a simple genetic test to determine the presence of FRDA in the children of silent FRDA carriers, it is possible to determine the presence of Friedreich’s ataxia in newborns, years before the cardio- and neurodegeneration and clinical symptoms begin, a time window during which nonfeminizing estrogens and other antioxidants could potentially be clinically useful. Estrogens are putative candidate drugs to provide a neuroprotective effect in Friedreich’s ataxia. The ability of phenolic estrogens to protect against the oxidative damage of ROS, coupled with the possibility that they maintain the integrity of the oxidative phosphorylation process makes them ideal for the treatment of the underlying cellular dysfunction, not just the symptoms of FRDA. This study will determine if E2 and estrogen-like compounds can protect human FRDA fibroblasts from oxidative insults in vitro. In addition, we will attempt to determine the exact mechanism by which E2 acts and investigate the possibility of any synergistic effects with other compounds proposed as putative treatments for FRDA.Item The Role of Fibroblast Growth Factor-2 in the Expansion of Pancreatic Islets IN VITRO(2004-05-01) Ettinger, Eve Sari; S. Dan Dimitrijevich; Julian Borejdo; Richard EasomEttinger, Eve Sari, The Role of Fibroblast Growth Factor-2 in the Expansion of Pancreatic Islets In Vitro. Doctor of Philosophy (Biomedical Sciences, Biochemistry and Molecular Biology), May 2004, 181 pp., 7 tables, 46 figures, 109 references. The focus of these studies was to identify a pool of progenitor cells within the adult islet, which can be expanded in vitro and aggregated into glucose responsive neoislets as a potential source for transplantation in diabetic patients. We hypothesize that normal adult islets contain an unspecialized proliferative progenitor cell population that can be expanded, aggregated by FGF-2 via PKC mechanism in 2-D culture or in a Rotating Wall Vessel Cell Culture System and matured into glucose-sensitive neoislets through the differential expression or activation of one or more PKC isoform(s). It was shown by immunofluorescence that normal adult islets from several species (rat, porcine, human and canine) contain proliferative endocrine precursor (endocrine epithelial) cells, which coexpress insulin, glucagon and somatostatin. These cells also express PDX-1, a marker of mature B cells and endocrine precursors as well as glut 2 and glucokinase, which are necessary components of glucose metabolism. Endocrine epithelial cells were shown to aggregate into neoislets either spontaneously in 2-D culture or via a rotating wall vessel cell culture system in the presence of fibroblast growth factor-2. FGF2 has been implicated in the aggregation of endocrine epithelial cells into neoislets via the FGFR-cadherin-catenin complex or through PKC signal transduction. Due to the variations in islet isolation, Ins-1 cells were used as a model of B cell aggregation. PKC profiles of endocrine epithelial cells, neoislets, native islets, Ins-1 cells and Ins-1 neoislets were determined. PKC-α, BII, ε and ζ were the only isoforms expressed in these cells and are potential targets for modulation in the differentiation of endocrine epithelial cells. Insulin secretion in response to a glucose challenge was examined in normal islets, neoislets, endocrine epithelial cells, and Ins-1 cells. Although endocrine epithelial cells express insulin, show dense secretory granules and contain two critical proteins in the glucose-sensing cascade, they are immature with respect to glucose-responsive insulin secretion. Neoislets show insignificant insulin secretion in response to elevated glucose when compared to normal islets. Aggregation of endocrine epithelial cells may be the first step in differentiation into mature β cells, however glucose-responsive insulin secretion must be achieved in vitro for use as an alternate source of islets for transplantation in diabetic patients.