NR5A2/LRH-1 regulates the PTGS2-PGE2-PTGER1 pathway contributing to pancreatic islet survival and function

LRH-1/NR5A2 is implicated in islet morphogenesis postnatally, and its activation using the agonist BL001 protects islets against apoptosis, reverting hyperglycemia in mouse models of Type 1 Diabetes Mellitus. Islet transcriptome profiling revealed that the expression of PTGS2/COX2 is increased by BL001. Herein, we sought to define the role of LRH-1 in postnatal islet morphogenesis and chart the BL001 mode of action conferring beta cell protection. LRH-1 ablation within developing beta cells impeded beta cell proliferation, correlating with mouse growth retardation, weight loss, and hypoglycemia leading to lethality. LRH-1 deletion in adult beta cells abolished the BL001 antidiabetic action, correlating with beta cell destruction and blunted Ptgs2 induction. Islet PTGS2 inactivation led to reduced PGE levels and loss of BL001 protection against cytokines as evidenced by increased cytochrome c release and cleaved-PARP. The PTGER1 antagonist—ONO-8130—negated BL001-mediated islet survival. Our results define the LRH-1/PTGS2/PGE/PTGER1 signaling axis as a key pathway mediating BL001 survival properties.The authors are supported by grants from the Consejería de Salud, Fundación Pública Andaluza Progreso y Salud, Junta de Andalucía (PI-0727-2010 to B.R.G., PI-0085-2013 to P.I.L., PI-0247-2016 to F.J.B.S.), the Consejería de Economía, Innovación y Ciencia (P10.CTS.6359 to B.R.G.), the Ministerio de Ciencia e Innovación co-funded by Fondos FEDER (PI10/00871, PI13/00593 and BFU2017-83588-P to B.R.G and PI17/01004 to F.J.B.S.), Vencer el Cancer (B.R.G), DiabetesCero (B.R.G.) and the Juvenile Diabetes Research Foundation Ltd (17-2013-372 and 2-SRA-2019-837-S-B to B.R.G.). E.M.V. is recipient of a Fellowship from the Ministerio de Ciencia e Innovación co-funded by Fondos FEDER (PRE2018-084907). F.J.B.S. is a recipient of a "Nicolás Monardes" research contracts from Consejería de Salud Junta de Andalucía, (C-0070-2012). A.M.M. is supported by CPII19/00023 and PI18/01590 from the Instituto de Salud Carlos III co-funded by Fondos FEDER. V.C. is supported by a AECC investigator award. CIBERDEM is an initiative of the Instituto de Salud Carlos III

Pancreatic ductal cells may have a negative effect on human islet transplantation.

AimTo evaluate the effect of pancreatic ductal cells on experimental human islet transplantation.Materials and methodsIsolated islets were additionally purified by handpicking. Ductal cells were purified by magnetic cell sorting and then clustered into ductal pancreatospheres (DPS). Islets, DPS, and islets + DPS (100 islets + 75 DPS, or 100 islets + 200 DPS) were cultured and glucose-stimulated insulin secretion, β-cell apoptosis, and gene expression was determined. Islets and islets + DPS preparations (800 islets + 600 DPS) were transplanted to streptozotocin-treated immunodeficient mice and glycemia, graft morphometry, and gene expression were determined.ResultsInsulin stimulation index was higher in islets than in islets co-cultured with DPS (5.59 ± 0.93 vs 4.02 ± 0.46; p0.05), and the ratio β-/endocrine non-β-cell mass was lower in islets + DPS grafts (islets: 2.05 ± 0.18, islets + DPS: 1.35 ± 0.15; pConclusionsIslet preparations enriched with ductal cells have a lower insulin stimulation index in vitro and achieved a worse metabolic outcome after transplantation. Inflammation may mediate the deleterious effects of ductal cells on islet cells

A functional genomic approach to identify reference genes for human pancreatic beta cell real-time quantitative RT-PCR analysis

Exposure of human pancreatic beta cells to pro-inflammatory cytokines or metabolic stressors is used to model events related to type 1 and type 2 diabetes, respectively. Quantitative real-time PCR is commonly used to quantify changes in gene expression. The selection of the most adequate reference gene(s) for gene expression normalization is an important pre-requisite to obtain accurate and reliable results. There are no universally applicable reference genes, and the human beta cell expression of commonly used reference genes can be altered by different stressors. Here we aimed to identify the most stably expressed genes in human beta cells to normalize quantitative real-time PCR gene expression. We used comprehensive RNA-sequencing data from the human pancreatic beta cell line EndoC-beta H1, human islets exposed to cytokines or the free fatty acid palmitate in order to identify the most stably expressed genes. Genes were filtered based on their level of significance (adjusted P-value >0.05), fold-change (|fold-change| <1.5) and a coefficient of variation <10%. Candidate reference genes were validated by quantitative real-time PCR in independent samples. We identified a total of 264 genes stably expressed in EndoC-beta H1 cells and human islets following cytokines - or palmitate-induced stress, displaying a low coefficient of variation. Validation by quantitative real-time PCR of the top five genes ARF1, CWC15, RAB7A, SIAH1 and VAPA corroborated their expression stability under most of the tested conditions. Further validation in independent samples indicated that the geometric mean of ACTB and VAPA expression can be used as a reliable normalizing factor in human beta cells

GLP-1R agonists demonstrate potential to treat Wolfram syndrome in human preclinical models.

AIMS/HYPOTHESIS: Wolfram syndrome is a rare autosomal recessive disorder caused by pathogenic variants in the WFS1 gene. It is characterised by insulin-dependent diabetes mellitus, optic nerve atrophy, diabetes insipidus, hearing loss and neurodegeneration. Considering the unmet treatment need for this orphan disease, this study aimed to evaluate the therapeutic potential of glucagon-like peptide 1 receptor (GLP-1R) agonists under wolframin (WFS1) deficiency with a particular focus on human beta cells and neurons. METHODS: The effect of the GLP-1R agonists dulaglutide and exenatide was examined in Wfs1 knockout mice and in an array of human preclinical models of Wolfram syndrome, including WFS1-deficient human beta cells, human induced pluripotent stem cell (iPSC)-derived beta-like cells and neurons from control individuals and individuals affected by Wolfram syndrome, and humanised mice. RESULTS: Our study shows that the long-lasting GLP-1R agonist dulaglutide reverses impaired glucose tolerance in WFS1-deficient mice, and that exenatide and dulaglutide improve beta cell function and prevent apoptosis in different human WFS1-deficient models including iPSC-derived beta cells from people with Wolfram syndrome. Exenatide improved mitochondrial function, reduced oxidative stress and prevented apoptosis in Wolfram syndrome iPSC-derived neural precursors and cerebellar neurons. CONCLUSIONS/INTERPRETATION: Our study provides novel evidence for the beneficial effect of GLP-1R agonists on WFS1-deficient human pancreatic beta cells and neurons, suggesting that these drugs may be considered as a treatment for individuals with Wolfram syndrome