Analysis of Allele-Specific Expression Highlights Novel Participants of Empagliflozin-Driven Effects on T2DM-Associated Regulatory Pathways

Abstract

It is well-known that the morbidity and clinical burden of type 2 diabetes mellitus (T2DM) are predominantly associated with its chronic complications, in which fibrosis is a significant contributor. Recently, sodium–glucose cotransporter 2 (SGLT2) inhibitors have made a pivotal advancement in the therapeutic landscape not only improving glycemic control, but also demonstrating high effectiveness in the prevention and treatment of T2DM complications. In this work, we aimed to assess the transcription factors (TFs) mediating the effects of SGLT2 inhibitor empagliflozin (EMPA) treatment by a comprehensive analysis of the allele-specific expression (ASE) events utilizing the RNA-seq data. Initial logistic regression analysis of the in vitro transcriptomic data for EMPA-treated peripheral blood mononuclear cells (PBMCs) of three healthy donors revealed a significant inter-individual variation in ASE for 240 genes linked to EMPA treatment beyond the glucose-lowering effects. Then, 146 TFs were predicted to regulate the expression of the corresponding targets using motifbreakR and DESeq2. Among these, multiple TFs (including ATF3, ATF4, E2F1, EGR1, FOS, JUN, JUNB, IRF8, KLF6, KLF11, SNAI1, TWIST1, and ZEB1) were involved in the TGF-β/SMAD3 canonical profibrotic signaling cascade, pertinent to diabetes-related fibrosis, playing a significant role in the development of diabetic complications. Further analysis of the in vivo data for the PBMCs from ten T2DM patients initiating EMPA therapy identified 98 TFs related to the ASE variation in both in vitro and in vivo cohorts. To conclude, our integrative allele-specific approach enables the prediction of novel EMPA-responsive regulatory interactions and suggests the important mediators of the mechanisms underlying the effects of EMPA on human PBMCs.