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Mercado A.A.,Pancreatic Islet Development and Regeneration Unit | Cobo-Vuilleumier N.,Pancreatic Islet Development and Regeneration Unit | Martin E.S.,Office of Technology Transfer | Gauthier B.R.,Pancreatic Islet Development and Regeneration Unit
Recent Patents on Regenerative Medicine | Year: 2013

In recent years, the concept of preserving and/or replenishing the functional β-cell mass vital to sustain insulin output and normalized blood glucose levels has gained much interest as a therapeutic approach in regenerative medicine for the treatment of Diabetes Mellitus. Herein, we surveyed the diabetes area patent literature published in recent years to identify novel uprising therapeutic targets specifically implicated in regeneration and survival. One hundred and sixty nine international patent applications filed under the Patent Cooperation Treaty (PCT) (hereinafter, patents or applications) were highlighted from which 8 particular targets stood out with more than 4 patents published within the last few years. Not surprisingly, GLP-1 analogues and DPP-4 inhibitors along with GPR119 agonists and SGLT2 inhibitors were among the top ranked candidates. However, new emerging targets into the field of regenerative medicine for the treatment of diabetes include: 1) BACE-2; a protease that was recently shown to cleave the plasma membrane glycoprotein TMEM27 (also called collectrin) resulting in the inhibition of pancreatic β cell proliferation and insulin secretion, 2) GIP; a 42 amino acid incretin hormone that potentiates glucose induce insulin secretion and protect β-cells against cytokine-mediated apoptosis, 3) neurturin; a neurotrophic factor capable of improving blood glucose levels in high fat diet treated animals, and 4) LRH-1, an orphan nuclear receptor that improves islet viability. These novel targets along with GPR119 are further discussed in this review. © 2013 Bentham Science Publishers.

Baquie M.,University Hospitals Geneva Medical Center | Baquie M.,Pancreatic Islet Development and Regeneration Unit | Cobo-Vuilleumier N.,Pancreatic Islet Development and Regeneration Unit | Lorenzo P.I.,Pancreatic Islet Development and Regeneration Unit | And 11 more authors.
Human Molecular Genetics | Year: 2011

Liver receptor homolog (LRH-1) is an orphan nuclear receptor (NR5A2) that regulates cholesterol homeostasis and cell plasticity in endodermal-derived tissues. Estrogen increases LRH-1 expression conveying cell protection and proliferation. Independently, estrogen also protects isolated human islets against cytokine- induced apoptosis. Herein, we demonstrate that LRH-1 is expressed in islets, including β-cells, and that transcript levels are modulated by 17β-estradiol through the estrogen receptor (ER)α but not ERβ signaling pathway. Repression of LRH-1 by siRNA abrogated the protective effect conveyed by estrogen on rat islets against cytokines. Adenoviral-mediated overexpression of LRH-1 in human islets did not alter proliferation but conferred protection against cytokines and streptozotocin-induced apoptosis. Expression levels of the cell cycle genes cyclin D1 and cyclin E1 as well as the antiapoptotic gene bcl-xl were unaltered in LRH-1 expressing islets. In contrast, the steroidogenic enzymes CYP11A1 and CYP11B1 involved in glucocorticoid biosynthesis were both stimulated in transduced islets. In parallel, graded overexpression of LRH-1 dose- dependently impaired glucose-induced insulin secretion. Our results demonstrate the crucial role of the estrogen target gene nr5a2 in protecting human islets against-stressed-induced apoptosis. We postulate that this effect is mediated through increased glucocorticoid production that blunts the pro-inflammatory response of islets. © The Author 2011. Published by Oxford University Press. All rights reserved.

Mellado-Gil J.M.,Pancreatic Islet Development and Regeneration Unit | Jimenez-Moreno C.M.,Pancreatic Islet Development and Regeneration Unit | Martin-Montalvo A.,Pancreatic Islet Development and Regeneration Unit | Alvarez-Mercado A.I.,Pancreatic Islet Development and Regeneration Unit | And 18 more authors.
Diabetologia | Year: 2016

Aims/hypothesis: A strategy to enhance pancreatic islet functional beta cell mass (BCM) while restraining inflammation, through the manipulation of molecular and cellular targets, would provide a means to counteract the deteriorating glycaemic control associated with diabetes mellitus. The aims of the current study were to investigate the therapeutic potential of such a target, the islet-enriched and diabetes-linked transcription factor paired box 4 (PAX4), to restrain experimental autoimmune diabetes (EAD) in the RIP-B7.1 mouse model background and to characterise putative cellular mechanisms associated with preserved BCM. Methods: Two groups of RIP-B7.1 mice were genetically engineered to: (1) conditionally express either PAX4 (BPTL) or its diabetes-linked mutant variant R129W (mutBPTL) using doxycycline (DOX); and (2) constitutively express luciferase in beta cells through the use of RIP. Mice were treated or not with DOX, and EAD was induced by immunisation with a murine preproinsulin II cDNA expression plasmid. The development of hyperglycaemia was monitored for up to 4 weeks following immunisation and alterations in the BCM were assessed weekly by non-invasive in vivo bioluminescence intensity (BLI). In parallel, BCM, islet cell proliferation and apoptosis were evaluated by immunocytochemistry. Alterations in PAX4- and PAX4R129W-mediated islet gene expression were investigated by microarray profiling. PAX4 preservation of endoplasmic reticulum (ER) homeostasis was assessed using thapsigargin, electron microscopy and intracellular calcium measurements. Results: PAX4 overexpression blunted EAD, whereas the diabetes-linked mutant variant PAX4R129W did not convey protection. PAX4-expressing islets exhibited reduced insulitis and decreased beta cell apoptosis, correlating with diminished DNA damage and increased islet cell proliferation. Microarray profiling revealed that PAX4 but not PAX4R129W targeted expression of genes implicated in cell cycle and ER homeostasis. Consistent with the latter, islets overexpressing PAX4 were protected against thapsigargin-mediated ER-stress-related apoptosis. Luminal swelling associated with ER stress induced by thapsigargin was rescued in PAX4-overexpressing beta cells, correlating with preserved cytosolic calcium oscillations in response to glucose. In contrast, RNA interference mediated repression of PAX4-sensitised MIN6 cells to thapsigargin cell death. Conclusions/interpretation: The coordinated regulation of distinct cellular pathways particularly related to ER homeostasis by PAX4 not achieved by the mutant variant PAX4R129W alleviates beta cell degeneration and protects against diabetes mellitus. The raw data for the RNA microarray described herein are accessible in the Gene Expression Omnibus database under accession number GSE62846. © 2016, The Author(s).

He K.H.H.,University of Geneva | Lorenzo P.I.,Pancreatic Islet Development and Regeneration Unit | Brun T.,University of Geneva | Jimenez Moreno C.M.,Pancreatic Islet Development and Regeneration Unit | And 11 more authors.
Diabetes | Year: 2011

OBJECTIVE - To establish the role of the transcription factor Pax4 in pancreatic islet expansion and survival in response to physiological stress and its impact on glucose metabolism, we generated transgenic mice conditionally and selectively overexpressing Pax4 or a diabetes-linked mutant variant (Pax4R129W) in β-cells. RESEARCH DESIGN AND METHODS - Glucose homeostasis and β-cell death and proliferation were assessed in Pax4- or Pax4R129W-overexpressing transgenic animals challenged with or without streptozotocin. Isolated transgenic islets were also exposed to cytokines, and apoptosis was evaluated by DNA fragmentation or cytochrome C release. The expression profiles of proliferation and apoptotic genes and β-cell markers were studied by immunohistochemistry and quantitative RT-PCR. RESULTS - Pax4 but not Pax4R129W protected animals against streptozotocin-induced hyperglycemia and isolated islets from cytokine-mediated β-cell apoptosis. Cytochrome C release was abrogated in Pax4 islets treated with cytokines. Interleukin-1β transcript levels were suppressed in Pax4 islets, whereas they were increased along with NOS2 in Pax4R129W islets. Bcl-2, Cdk4, and c-myc expression levels were increased in Pax4 islets while MafA, insulin, and GLUT2 transcript levels were suppressed in both animal models. Long-term Pax4 expression promoted proliferation of a Pdx1-positive cell subpopulation while impeding insulin secretion. Suppression of Pax4 rescued this defect with a concomitant increase in pancreatic insulin content. CONCLUSIONS - Pax4 protects adult islets from stress-induced apoptosis by suppressing selective nuclear factor-κB target genes while increasing Bcl-2 levels. Furthermore, it promotes dedifferentiation and proliferation of β-cells through MafA repression, with a concomitant increase in Cdk4 and c-myc expression. © 2011 by the American Diabetes Association.

Lorenzo P.I.,Pancreatic Islet Development and Regeneration Unit | Jimenez Moreno C.M.,Pancreatic Islet Development and Regeneration Unit | Delgado I.,Pancreatic Islet Development and Regeneration Unit | Cobo-Vuilleumier N.,Pancreatic Islet Development and Regeneration Unit | And 6 more authors.
Histochemistry and Cell Biology | Year: 2011

The paired box transcription factor Pax8 is critical for development of the eye, thyroid gland as well as the urinary and reproductive organs. In adult, Pax8 overexpression is associated with kidney, ovarian and thyroid tumors and has emerged as a specific marker for these cancers. Recently, Pax8 expression was also reported in human pancreatic islets and in neuroendocrine tumors, identifying Pax8 as a novel member of the Pax family expressed in the pancreas. Herein, we sought to provide a comprehensive analysis of Pax8 expression during pancreogenesis and in adult islets. Immunohistochemical analysis using the most employed Pax8 polyclonal antibody revealed strong nuclear staining in the developing mouse pancreas and in mature human and mouse islets. Astonishingly, Pax8 mRNA in mouse islets was undetectable while human islets exhibited low levels. These discrepancies raised the possibility of antibody crossreactivity. This premise was confirmed by demonstrating that the polyclonal Pax8 antibody also recognized the isletenriched Pax6 protein both by Western blotting and immunohistochemistry. Thus, in islets polyclonal Pax8 staining corresponds mainly to Pax6. In order to circumvent this caveat, a novel Pax8 monoclonal antibody was used to re-evaluate whether Pax8 was indeed expressed in islets. Surprisingly, Pax8 was not detected in neither the developing pancreas or in mature islets. Reappraisal of pancreatic neuroendocrine tumors using this Pax8 monoclonal antibody exhibited no immunostaining as compared to the Pax8 polyclonal antibody. In conclusion, Pax8 is not expressed in the pancreas and cast doubts on the value of Pax8 as a pancreatic neuroendocrine tumor marker. © The Author(s) 2011.

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