TYK2, a Candidate Gene for Type 1 Diabetes, Modulates Apoptosis and the Innate Immune Response in Human Pancreatic β-Cells

Abstract

Pancreatic b-cells are destroyed by an autoimmune attack in type 1 diabetes. Linkage and genome-wide association studies point to >50 loci that are associated with the disease in the human genome. Pathway analysis of candidate genes expressed in human islets identified a central role for interferon (IFN)-regulated pathways and tyrosine kinase 2 (TYK2). Polymorphisms in the TYK2 gene predicted to decrease function are associated with a decreased risk of developing type 1 diabetes. We presently evaluated whether TYK2 plays a role in human pancreatic b-cell apoptosis and production of proinflammatory mediators. TYK2-silenced human b-cells exposed to polyinosinicpolycitidilic acid (PIC) (a mimick of double-stranded RNA produced during viral infection) showed less type I IFN pathway activation and lower production of IFNa and CXCL10. These cells also had decreased expression of major histocompatibility complex (MHC) class I proteins, a hallmark of early b-cell inflammation in type 1 diabetes. Importantly, TYK2 inhibition prevented PICinduced b-cell apoptosis via the mitochondrial pathway of cell death. The present findings suggest that TYK2 regulates apoptotic and proinflammatory pathways in pancreatic b-cells via modulation of IFNa signaling, subsequent increase in MHC class I protein, and modulation of chemokines such as CXCL10 that are important for recruitment of T cells to the islets.

Interferon-α mediates human beta cell HLA class I overexpression, endoplasmic reticulum stress and apoptosis, three hallmarks of early human type 1 diabetes

“…Western blot analysis, immunofluorescence and flow cytometry Western blotting was performed as described [23]. Briefly, cells were washed with cold PBS and lysed in Laemmli or RIPA buffer.…”

“…Quantitative real-time PCR was performed using SYBR Green and the data were compared with a standard curve [31]. Expression values were corrected by the housekeeping gene β-actin, as its expression is not modified under the conditions used in this study [23] (data not shown). A list with the primers used in this study is provided in ESM Table 3.…”

“…mRNA extraction and real-time PCR Poly(A) + mRNA extraction was performed using Dynabeads mRNA DIRECT kit (Invitrogen) in accordance with the manufacturer's instructions; reverse transcription was carried out as described [23]. Quantitative real-time PCR was performed using SYBR Green and the data were compared with a standard curve [31].…”

“…Human islets or EndoC-βH1 cells were exposed to cytokines or other agents as described in ESM Methods [23,26].…”

“…Neutralisation of the IFNα/β receptor (IFNAR1) with monoclonal antibodies prevents diabetes in NOD mice [13,19] and self-reactive antibodies targeting type I IFNs-particularly IFNα-are associated with protection against type 1 diabetes in patients with mutations in the thymus transcription factor autoimmune regulator (AIRE) [20]. It is surprising that very few studies have investigated the direct effects of this cytokine in pancreatic beta cells [21][22][23].…”