Introduction and Objective: Neutral sphingomyelinase 2 (nSMase2) is a crucial enzyme involved in several important cellular processes, including exosome formation, ceramide production, cellular stress responses, and inflammatory signaling responses. However, impacts on beta cell health are unknown.Methods: INS-1 823/13 nSMase2 knockdown cells were generated by transfecting plasmids with rat nSMase2 shRNA containing a GFP reporter. A non-effective 29-mer scrambled shRNA cassette in a pGFP-C-shLenti Vector was used as a control. Protein expression was quantified using western blotting and/or flow cytometry. Insulin content and secretion was quantified using ELISA. RNA sequencing was performed and analyzed using Qiagen IPA.Results: Knockdown of nSMase2 led to a notable upregulation of three critical transcriptional regulators of insulin synthesis – PDX1, Neurogenin-3, and MAFA. Knockdown also increased GLUT2 protein levels, and significantly enhanced total cellular insulin content and basal insulin secretion. Pathway analysis identified insulin secretion signaling, insulin processing, insulin receptor signaling, and IGF/IGF1R regulation as key pathways impacted by nSMase2 knockdown.Conclusion: nSMase2 inhibits proteins regulating insulin synthesis and may also influence glucose uptake and insulin secretion. Our data suggest that nSMase2 plays a critical role in insulin production and secretion. Inhibitors of nSMase2 could serve as potential therapeutic agents for the treatment of diabetes.
J. Xu: None. E.K. Sims: Consultant; Sanofi. Speaker’s Bureau; Med Learning Group. Other Relationship; American Diabetes Association.
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