Mitophagy protects beta cells from inflammatory damage in diabetes

Abstract

Inflammatory damage contributes to β-cell failure in type 1 and 2 diabetes (T1D and T2D).Mitochondria are damaged by inflammatory signaling in β-cells, resulting in impaired bioenergetics and initiation of pro-apoptotic machinery. Hence, the identification of protective responses to inflammation could lead to new therapeutic targets. Here we report that mitophagy serves as a protective response to inflammatory stress in both human and rodent β-cells. Utilizing in vivo mitophagy reporters, we observed that diabetogenic pro-inflammatory cytokines induced mitophagy in response to nitrosative/oxidative mitochondrial damage. Mitophagy-deficient β-cells were sensitized to inflammatory stress, leading to the accumulation of fragmented dysfunctional mitochondria, increased β-cell death, and hyperglycemia. Overexpression of CLEC16A, a T1D gene and mitophagy regulator whose expression in islets is protective against T1D, ameliorated cytokine-induced human β-cell apoptosis. Thus, mitophagy promotes β-cell survival and prevents diabetes by countering inflammatory injury. Targeting this pathway has the potential to prevent βcell failure in diabetes and may be beneficial in other inflammatory conditions.

Mitofusin 1 and 2 regulation of mitochondrial DNA content is a critical determinant of glucose homeostasis

“…Secondly, we examined mitochondrial networking utilizing high-resolution deconvolution imaging of the mitochondrial marker succinate dehydrogenase A (SDHA). We initially used Imaris® imaging software to generate three-dimensional (3D) reconstructions of β-cell mitochondria to grossly evaluate mitochondrial networking 21, 22 . Consistent with our findings using TEM, the number of 3D mitochondrial networks appeared similar between Ctrl and single β-Mfn1 KO or β-Mfn2 KO β-cells (Figure S2c).…”

“…Mitochondrial morphology and subcellular localization analyses were performed on immuno-stained paraffin-embedded mouse pancreas tissue sections and dissociated islet cells as previously described 21 . Briefly, pancreatic tissue sections were fixed in 4% PFA overnight, embedded in paraffin and sectioned.…”

“…Following isolation and culture, live mouse islets were exposed to 100 nM Mtphagy dye (Dojindo Molecular Technologies) for 3 hours to assess time-dependent accumulation of mitochondria to acidic organelles by the relative fluorescence intensity of the dye per cell as described 91, 92 . Islets were then dissociated into single cells, stained with DAPI (Thermo Fisher Scientific) and Fluozin-3 (Thermo Fisher Scientific), and resuspended in phenol red–free islet culture medium as previously described 21 . Samples were analyzed on an LSR Fortessa flow cytometer (BD Biosciences).…”

“…Samples were analyzed on an LSR Fortessa flow cytometer (BD Biosciences). Single cells were gated using forward scatter and side scatter (FSC and SSC, respectively) plots, DAPI staining was used to exclude dead cells, and Fluozin-3 was used to identify β-cells as previously described 21, 93 . Mtphagy measurements in β-cells were made using 488 nm excitation laser with a 710 nm emission filter and analyzed using FlowJo (Tree Star Inc.).…”

“…All assays were performed as previously described 21, 85, 86 . Mouse pancreatic islets were lysed with radioimmunoprecipitation assay buffer containing protease and phosphatase inhibitors (Calbiochem), and insoluble material was removed by centrifugation.…”