Individual islet respirometry reveals functional diversity within the islet population of mice and human donors

Taddeo, Evan P.; Stiles, Linsey; Sereda, Samuel B.; Ritou, Eleni; Wolf, Dane M.; Abdullah, Muhamad Rais; Swanson, Zachary; Wilhelm, Joshua J.; Bellin, Melena D.; McDonald, Patrick L. L.; Caradonna, Kacey L.; Neilson, Andrew P.; Liesa, Marc; Shirihai, Orian S.

doi:10.1016/j.molmet.2018.07.003

Cited by 37 publications

(54 citation statements)

References 39 publications

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“…Islet respirometry was measured using an XF96 extracellular flux analyzer (Seahorse Bioscience) according to manufacturer's instructions. Briefly, 20 similarly sized islets were plated per well of a Seahorse spheroid plate pre-treated with CellTak (Corning), similar to previously published approaches (52). Prior to respirometry profiling, islets were incubated for 1 hour in an atmospheric…”

Section: Mitochondrial Respirometrymentioning

confidence: 99%

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

Sidarala

Zhu

Pearson

et al. 2021

Preprint

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Mitochondria are vital for β-cell function, yet the importance of mitochondrial fusion for glucose homeostasis is uncertain. Here, we report that the dynamin-like GTPases Mitofusin 1 and 2 (Mfn1 and Mfn2) are critical for glucose-stimulated insulin secretion (GSIS). Whereas Mfn1 and Mfn2 individually were dispensable for glucose control, combined Mfn1/2 deletion in β-cells induced mitochondrial fragmentation, reduced mtDNA content, and impaired respiratory function, ultimately resulting in severe glucose intolerance. Importantly, gene dosage studies revealed that Mfn1/2 control of glucose homeostasis is dependent on maintenance of mtDNA content, rather than mitochondrial structure. Mfn1/2 maintain mtDNA content by regulating the expression of the crucial mitochondrial transcription factor Tfam, as Tfam overexpression ameliorated the reduction in mtDNA content and GSIS in Mfn1/2-deficient β-cells. Further, mitofusin agonists rescued mtDNA content and GSIS in islets from db/db mice, a model of type 2 diabetes. Thus, Mfn1 and 2 act collectively to promote both optimal mitochondrial dynamics and mtDNA copy number in β-cells, and may be promising therapeutic targets to improve insulin release in diabetes.

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Section: Mitochondrial Respirometrymentioning

confidence: 99%

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

Sidarala

Zhu

Pearson

et al. 2021

Preprint

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“…Islets were isolated from mice via collagenase P injection in the bile duct, as previously described (13,14). The islets were cultured overnight at 37°C and 5% CO 2 in islet medium [11 mM glucose Roswell Park Memorial Institute (RPMI) 1640 medium, 10% fetal bovine serum (FBS), 100 U/ml penicillin, and 100 mg/ ml streptomycin] prior to respirometry experiments.…”

Section: Isolation and Culture Of Mouse Isletsmentioning

confidence: 99%

“…A Seahorse extracellular flux analyzer XF96 (Agilent) was used to assess oxygen consumption rate in intact mouse islets as previously described in (13,14) with minor modifications. Islet respirometry was performed in poly-D-lysine-coated XF96 cell culture plates.…”

Section: Measurement Of Oxygen Consumption Rate In Isletsmentioning

confidence: 99%

Nanoparticle‐mediated lysosomal reacidification restores mitochondrial turnover and function in β cells under lipotoxicity

et al. 2018

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Chronic exposure of pancreatic β cells to high concentrations of free fatty acids leads to lipotoxicity (LT)‐mediated suppression of glucose‐stimulated insulin secretion. This effect is in part caused by a decline in mitochondrial function as well as by a reduction in lysosomal acidification. Because both mitochondria and lysosomes can alter one another's function, it remains unclear which initiating dysfunction sets off the detrimental cascade of LT, ultimately leading to β‐cell failure. Here, we investigated the effects of restoring lysosomal acidity on mitochondrial function under LT. Our results show that LT induces a dose‐dependent lysosomal alkalization accompanied by an increase in mitochondrial mass. This increase is due to a reduction in mitochondrial turnover as analyzed by MitoTimer, a fluorescent protein for which the emission is regulated by mitochondrial clearance rate. Mitochondrial oxygen consumption rate, citrate synthase activity, and ATP content are all reduced by LT. Restoration of lysosomal acidity using lysosome‐targeted nanoparticles is accompanied by stimulation of mitochondrial turnover as revealed by mitophagy measurements and the recovery of mitochondrial mass. Remarkably, re‐acidification restores citrate synthase activity and ATP content in an insulin secreting β‐cell line (INS‐1). Furthermore, nanoparticle‐mediated lysosomal reacidification rescues mitochondrial maximal respiratory capacity in both INS‐1 cells and primary mouse islets. Therefore, our results indicate that mitochondrial dysfunction is downstream of lysosomal alkalization under lipotoxic conditions and that recovery of lysosomal acidity is sufficient to restore the bioenergetic defects.—Assali, E. A., Shlomo, D., Zeng, J., Taddeo, E. P., Trudeau, K. M., Erion, K. A., Colby, A. H., Grinstaff, M. W., Liesa, M., Las, G., Shirihai, O. S. Nanoparticle‐mediated lysosomal reacidification restores mitochondrial turnover and function in β cells under lipotoxicity. FASEB J. 33, 4154–4165 (2019). http://www.fasebj.org

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“…Islet respirometry was measured using an XF96 extracellular flux analyzer (Seahorse Bioscience) according to manufacturer's instructions. Briefly, 20 similarly sized islets were plated per well of a Seahorse spheroid plate pre-treated with CellTak (Corning), similar to previously published approaches (63). Prior to respirometry profiling, islets were incubated for 1 hour in an atmospheric CO 2 incubator at 37°C and supplemented with mouse islet culture media (64,65)…”

Section: Mitochondrial Respirometrymentioning

confidence: 99%

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

Sidarala

Zhu

Pearson

et al. 2021

Preprint

View full text Add to dashboard Cite

The dynamin-like GTPases Mitofusin 1 and 2 (Mfn1 and Mfn2) are essential for mitochondrial function, which has been principally attributed to their regulation of fission/fusion dynamics. Here, we report that Mfn1 and 2 are critical for glucose-stimulated insulin secretion (GSIS) primarily through control of mtDNA content. Whereas Mfn1 and Mfn2 individually were dispensable for glucose homeostasis, combined Mfn1/2 deletion in β-cells reduced mtDNA content, induced mitochondrial fragmentation, and impaired respiratory function, ultimately resulting in severe glucose intolerance. Importantly, gene dosage studies unexpectedly revealed that Mfn1/2 control of glucose homeostasis was dependent on maintenance of mtDNA content, rather than mitochondrial structure. Indeed, pharmacologic mitofusin agonists rescued islet mtDNA depletion due to mitofusin deficiency independent of changes on mitochondrial structure. Mfn1/2 maintain mtDNA content by regulating the expression of the crucial mitochondrial transcription factor Tfam, as Tfam overexpression ameliorated the reduction in mtDNA content and GSIS in Mfn1/2-deficient β-cells. Thus, the primary physiologic role of Mfn1 and 2 in β-cells is coupled to preservation of mtDNA content rather than mitochondrial architecture, and Mfn1 and 2 may be promising targets to overcome mitochondrial dysfunction and restore glucose control in diabetes.

show abstract

Individual islet respirometry reveals functional diversity within the islet population of mice and human donors

Cited by 37 publications

References 39 publications

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

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

Nanoparticle‐mediated lysosomal reacidification restores mitochondrial turnover and function in β cells under lipotoxicity

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

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