Browsing by Author "Colvin, Stephanie C."

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    An Animal Model of Combined Pituitary Hormone Deficiency Disease
    (2010-08) Colvin, Stephanie C.; Konieczny, Stephen F.; Rhodes, Simon J.; Walvoord, Emily C.; Belecky-Adams, Teri; Herring, B. Paul; Roper, Randall
    LHX3 is a LIM-homeodomain transcription factor that has essential roles in pituitary and nervous system development in mammals. Children who are homozygous for recessive mutations in the LHX3 gene present with combined pituitary hormone deficiency disease (CPHD) characterized by deficits of multiple anterior pituitary hormones. Most LHX3 patients also present with additional defects associated with the nervous system including a characteristic limited head rotation and sometimes deafness. However, of the 10 types of LHX3 mutation described to date, one mutation type (W224ter) does not result in the limited head rotation, defining a new form of the disease. W224ter patients have CPHD but do not have nervous system symptoms. Whereas other mutations in LHX3 cause loss of the entire protein or its activity, the W224ter mutation causes specific loss of the carboxyl terminal of the LHX3 protein—a region that we have shown to contain critical regulatory domains for pituitary gene activation. To better understand the molecular and cellular etiology of CPHD associated with LHX3 gene mutations, I have generated knock-in mice that model the human LHX3 W224ter disease. The resulting mice display marked dwarfism, thyroid disease, female infertility, and reduced male fertility. Immunohistochemistry, real-time quantitative polymerase chain reaction (PCR), and enzyme-linked immunosorbant assays (ELISA) were used to measure hormones and regulatory factor protein and RNA levels, an approach which is not feasible with human patients. We have generated a novel mouse model of human pediatric CPHD. Our findings are consistent with the hypothesis that the actions of the LHX3 factor are molecularly separable in the nervous system and pituitary gland.
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    Deoxyhypusine synthase promotes differentiation and proliferation of T helper type 1 (Th1) cells in autoimmune diabetes
    (ASBMB, 2013-12-20) Colvin, Stephanie C.; Maier, Bernhard; Morris, David L.; Tersey, Sarah A.; Mirmira, Raghavendra G.; Department of Pediatrics, IU School of Medicine
    In type 1 diabetes, cytokines arising from immune cells cause islet β cell dysfunction even before overt hyperglycemia. Deoxyhypusine synthase catalyzes the crucial hypusine modification of the factor eIF5A, which promotes the translation of a subset of mRNAs involved in cytokine responses. Here, we tested the hypothesis that deoxyhypusine synthase and, secondarily, hypusinated eIF5A contribute to the pathogenesis of type 1 diabetes using the non-obese diabetic (NOD) mouse model. Pre-diabetic NOD mice that received injections of the deoxyhypusine inhibitor N1-guanyl-1,7-diaminoheptane (GC7) demonstrated significantly improved glucose tolerance, more robust insulin secretion, and reduced insulitis compared with control animals. Analysis of tissues from treated mice revealed selective reductions in diabetogenic T helper type 1 (Th1) cells in the pancreatic lymph nodes, a primary site of antigen presentation. Isolated mouse CD90.2(+) splenocytes stimulated in vitro with anti-CD3/anti-CD28 and IL-2 to mimic autoimmune T cell activation exhibited proliferation and differentiation of CD4(+) T cell subsets (Th1, Th17, and Treg), but those treated with the deoxyhypusine synthase inhibitor GC7 showed a dose-dependent block in T cell proliferation with selective reduction in Th1 cells, similar to that observed in NOD mice. Inhibition of deoxyhypusine synthase blocked post-transcriptional expression of CD25, the high affinity IL-2 receptor α chain. Our results suggest a previously unrecognized role for deoxyhypusine synthase in promoting T cell proliferation and differentiation via regulation of CD25. Inhibition of deoxyhypusine synthase may provide a strategy for reducing diabetogenic Th1 cells and preserving β cell function in type 1 diabetes.
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    Hypusinated eIF5A is expressed in the pancreas and spleen of individuals with type 1 and type 2 diabetes
    (Public Library of Science, 2020) Mastracci, Teresa L.; Colvin, Stephanie C.; Padgett, Leah R.; Mirmira, Raghavendra G.; Biochemistry and Molecular Biology, School of Medicine
    The gene encoding eukaryotic initiation factor 5A (EIF5A) is found in diabetes-susceptibility loci in mouse and human. eIF5A is the only protein known to contain hypusine (hydroxyputrescine lysine), a polyamine-derived amino acid formed post-translationally in a reaction catalyzed by deoxyhypusine synthase (DHPS). Previous studies showed pharmacologic blockade of DHPS in type 1 diabetic NOD mice and type 2 diabetic db/db mice improved glucose tolerance and preserved beta cell mass, which suggests that hypusinated eIF5A (eIF5AHyp) may play a role in diabetes pathogenesis by direct action on the beta cells and/or altering the adaptive or innate immune responses. To translate these findings to human, we examined tissue from individuals with and without type 1 and type 2 diabetes to determine the expression of eIF5AHyp. We detected eIF5AHyp in beta cells, exocrine cells and immune cells; however, there was also unexpected enrichment of eIF5AHyp in pancreatic polypeptide-expressing PP cells. Interestingly, the presence of eIF5AHyp co-expressing PP cells was not enhanced with disease. These data identify new aspects of eIF5A biology and highlight the need to examine human tissue to understand disease.
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    Peroxisome Proliferator-activated Receptor-γ Activation Augments the β-Cell Unfolded Protein Response and Rescues Early Glycemic Deterioration and β Cell Death in Non-obese Diabetic Mice
    (American Society for Biochemistry and Molecular Biology, 2016-10-21) Maganti, Aarthi V.; Tersey, Sarah A.; Syed, Farooq; Nelson, Jennifer B.; Colvin, Stephanie C.; Maier, Bernhard; Mirmira, Raghavendra G.; Biochemistry and Molecular Biology, School of Medicine
    Type 1 diabetes is an autoimmune disorder that is characterized by a failure of the unfolded protein response in islet β cells with subsequent endoplasmic reticulum stress and cellular death. Thiazolidinediones are insulin sensitizers that activate the nuclear receptor PPAR-γ and have been shown to partially ameliorate autoimmune type 1 diabetes in humans and non-obese diabetic (NOD) mice. We hypothesized that thiazolidinediones reduce β cell stress and death independently of insulin sensitivity. To test this hypothesis, female NOD mice were administered pioglitazone during the pre-diabetic phase and assessed for insulin sensitivity and β cell function relative to controls. Pioglitazone-treated mice showed identical weight gain, body fat distribution, and insulin sensitivity compared with controls. However, treated mice showed significantly improved glucose tolerance with enhanced serum insulin levels, reduced β cell death, and increased β cell mass. The effect of pioglitazone was independent of actions on T cells, as pancreatic lymph node T cell populations were unaltered and T cell proliferation was unaffected by pioglitazone. Isolated islets of treated mice showed a more robust unfolded protein response, with increases in Bip and ATF4 and reductions in spliced Xbp1 mRNA. The effect of pioglitazone appears to be a direct action on β cells, as islets from mice treated with pioglitazone showed reductions in PPAR-γ (Ser-273) phosphorylation. Our results demonstrate that PPAR-γ activation directly improves β cell function and survival in NOD mice by enhancing the unfolded protein response and suggest that blockade of PPAR-γ (Ser-273) phosphorylation may prevent type 1 diabetes.
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    Protective effects of polyamine depletion in mouse models of type 1 diabetes: implications for therapy
    (Springer, 2014-03) Tersey, Sarah A.; Colvin, Stephanie C.; Maier, Bernhard; Mirmira, Raghavendra G.; Department of Pediatrics, IU School of Medicine
    The underlying pathophysiology of type 1 diabetes involves autoimmune-mediated islet inflammation, leading to dysfunction and death of insulin-secreting islet β cells. Recent studies have shown that polyamines, which are essential for mRNA translation, cellular replication, and the formation of the hypusine modification of eIF5A may play an important role in the progression of cellular inflammation. To test a role for polyamines in type 1 diabetes pathogenesis, we administered the ornithine decarboxylase inhibitor difluoromethylornithine to two mouse models--the low-dose streptozotocin model and the NOD model--to deplete intracellular polyamines, and administered streptozotocin to a third model, which was haploinsufficient for the gene encoding the hypusination enzyme deoxyhypusine synthase. Subsequent development of diabetes and/or glucose intolerance was monitored. In the low-dose streptozotocin mouse model, continuous difluoromethylornithine administration dose-dependently reduced the incidence of hyperglycemia and led to the preservation of β cell area, whereas in the NOD mouse model of autoimmune diabetes difluoromethylornithine reduced diabetes incidence by 50%, preserved β cell area and insulin secretion, led to reductions in both islet inflammation and potentially diabetogenic Th17 cells in pancreatic lymph nodes. Difluoromethylornithine treatment reduced hypusinated eIF5A levels in both immune cells and islets. Animals haploinsufficient for the gene encoding deoxyhypusine synthase were partially protected from hyperglycemia induced by streptozotocin. Collectively, these studies suggest that interventions that interfere with polyamine biosynthesis and/or eIF5A hypusination may represent viable approaches in the treatment of diabetes.