Dataset Information

A mitochondrial ROS-dependent antiviral response promotes β cell resilience and is diminished in donors with type 1 diabetes

ABSTRACT: Type 1 diabetes (T1D) is a multifactorial disease driven by genetic and environmental factors, including, potentially, viral infection. However, the mechanisms linking infection-associated cytokines to human β cell loss are poorly understood. Here, we coupled in vivo and in vitro imaging with genetic analysis to investigate the impact of interferon α (IFN-α), a cytokine produced during the immune response to viral infection or detection of unedited endogenous double-stranded RNAs, on human β cell physiology. We identified a subset of human β cells that acutely produce reactive oxygen species (ROS) in response to IFN-α and were more prevalent in islets from donors with lower body mass index and HbA1c. RNA sequencing of flow-sorted ROS+ and ROS- populations identified a gene signature predisposing some cells to IFN-α-stimulated ROS production, including genes involved in inflammatory and immune response. IFN-α treatment of human islets in vitro similarly elicited a heterogeneous increase in superoxide production. Parallel analysis of a human β cell line demonstrated that this ROS originated in the mitochondria. Rapid stimulation of key antiviral response genes by IFN-α in human islets was dependent on mitochondrial ROS elevation. Comparison with single-cell RNA sequencing datasets showed that genes up-regulated in ROS-producing cells were enriched in β cells from nondiabetic and autoantibody-positive donors rather than donors with T1D. Overall, our data demonstrate that IFN-α-induced mitochondrial ROS production in healthy human β cells is critical for the acute antiviral response, and impairment of this heterogeneous adaptive response may predict β cell loss during T1D pathogenesis.

ORGANISM(S): Homo sapiens

PROVIDER: GSE318506 | GEO | 2026/02/27

REPOSITORIES: GEO

ACCESS DATA

Dataset's files

Source:

			Action	DRS
		Other

Items per page:

1 - 1 of 1

Similar Datasets

Project description:Type 1 diabetes (T1D) is characterized by autoimmune destruction of pancreatic β-cells, insulin insufficiency, and hyperglycemia. Receptor interacting protein kinase 1 (RIPK1) is a multifunctional regulator of cell fate with kinase and scaffolding functions. We previously identified RIPKs as regulators of beta-cell cytotoxicity in vitro, and Ripk1 expression is increased in islets from aged non-obese diabetic (NOD) mice and β-cells from T1D donors. We hypothesized that RIPK1 regulates cytokine- and autoimmune-mediated β-cell demise in a mouse model of T1D. Using NIT-1 beta-cells derived from NOD mice, we tested control (CTL) and Ripk1 gene-edited (Ripk1delta, CRISPR/Cas9 KO) beta-cells treated with TNFα+IFNγ or co-cultured with self-reactive splenocytes from diabetic NOD mice. We performed live cell imaging, RNAseq, kinome profiling, and flow cytometry, tested small molecule RIPK1 inhibitors, and evaluated resistance to beta-cell autoimmunity in vivo. We found TNF alpha +IFN gamma increase RIPK1 phosphorylation, caspase 3/7 activity, and cell death in NIT-1 CTL cells. In contrast, cytotoxicity was blocked with small molecule RIPK1 inhibition or in NIT-1 Ripk1delta cells. Co-labeling of caspase 3/7 activation and cell death in single cells revealed protection from caspase-dependent and -independent death in Ripk1Δ cells. RNAseq uncovered differential cell death-, immune-, and identity-related gene expression between genotypes, and kinome profiling identified changes in MAPK, Eph, JAK, and other kinase activity that were associated with protection from cell death. Furthermore, NIT-1 Ripk1delta cells were protected from destruction following co-culture with self-reactive splenocytes in vitro, and NIT-1 implant graft luminescence was 399-fold greater in Ripk1delta versus CTL cells 19 days post splenocyte administration in vivo. Collectively, our findings indicate RIPK1 regulates beta-cell fate in a model of autoimmune diabetes via actions on gene expression and kinase signaling. Additional studies are needed to characterize RIPK1 signaling in primary islets and human beta-cells, and to determine whether RIPK1 can be targeted to protect beta-cells in T1D. In this data set, NIT-1 cells (control and Ripk1 KO), vehicle treated or treated with IFN-gamma and TNF alpha were used for kinome profiling.

Project description:Aims/Hypothesis: Type 1 diabetes (T1D) is an autoimmune disease marked by the destruction of beta cells in the pancreatic islets, with an incomplete picture of disease pathogenesis/progression and a lack of definitive cure. A recent finding linked pancreatic ductal cells of T1D donors with elevated levels of human leukocyte antigen (HLA) Class II molecules; however, the causal relationship and functional significance of this finding remain unknown. Because HLA Class II molecules are typically expressed by professional antigen presenting cells (APCs), this raises the possibility of ductal cells functioning as non-professional APCs. In this study, we test the hypothesis that non-T1D ductal cells are responsive to T1D-associated proinflammatory cytokines, TNF-α, IL-1β, and IFN-γ, and can act as non-professional APCs. Methods: Pancreatic exocrine cells, after the removal of islets, were obtained from cadaveric donors without apparent diseases and with appropriate consent. Cells were cryopreserved, thawed into a defined culture medium tailored to support ductal cell survival in a 3D suspension culture system. Ductal cells were exposed to various doses of cytokines for 48 hours and analyzed for gene and protein expression, using qRT-PCR, bulk-RNAseq, flow cytometry and Western blot analyses. Functional ability of cytokine-treated ductal cells to present an exogenous autoantigen (glutamate decarboxylase 65kD isoform [GAD65]) to T cells was tested using a GAD65-specific autoreactive CD4+ T cell clone (named BRI4.13) isolated from a T1D donor. Results: We found that within 48 hours a combination of TNF-α, IL-1β, and IFN-γ stimulated the expression of HLA Class II, co-stimulatory, and antigen-processing molecules at mRNA and protein levels in non-T1D ductal cells. Bulk RNAseq analysis showed that cytokines most significantly upregulated biological pathways in “antigen processing and presentation” and “T1D”. When cytokine-treated ductal cells were pulsed with an exogenous GAD65 peptide and cocultured with the BRI4.13 T cells, these T cells became activated and proliferated. Unexpectedly, we also found ~1% of KRT19+ ductal cells express GAD protein endogenously. Conclusions/interpretation: These results demonstrate that non-diseased primary ductal cells are sensitive to TNF-α, IL-1β, and IFN-γ stimulation, and can upregulate APC molecules and function in antigen presentation to autoreactive CD4+ T cells. To the best of our knowledge, our results provide the first evidence demonstrating antigen presentation by non-T1D human ductal cells to T cells, which implicate ductal cells in contributing to T1D progression.

Dataset Information

A mitochondrial ROS-dependent antiviral response promotes β cell resilience and is diminished in donors with type 1 diabetes

Dataset's files

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets