Prediabetes uncovers differential gene expression at fasting and in response to oral glucose load in immune cells.

Summary Background and Objective Metabolic disorders including diabetes are associated with immune cell dysfunction. However, the effect of daily glucose metabolism or impairment thereof on the immune cell gene expression is not well known. Hence, in this cross-sectional study, we sought out to determine the differences in gene expression in the peripheral blood mono-nuclear cells (PBMCs) of normal glucose tolerant (NGT) and insulin resistant pre-diabetic (PD) Asian Indian men, at fasting and in response to 75 g oral glucose load. Methods Illumina HT12 bead chip-based microarray was performed on PBMCs at fasting and 2-hour post load conditions for NGT (N=6) and PD (N=9) subjects. Following normalization and due quality control of the raw data, differentially expressed genes (DEGs) under different conditions were compared within and across the two groups using GeneSpring GX V12.0 software. Paired and unpaired Student’s t-tests were applied along with fold change cut-offs for appropriate comparisons. Validation of the microarray data was carried out through real-time qPCR analysis. Significantly de-regulated biological pathways were analyzed by employing DEGs and DAVID resource. Deconvolution of the DEGs between NGT and PD subjects at fasting was performed using CIBERSORT and genes involved in regulatory T-cell (Treg) function were further analyzed for biological significance. Results Glucose load specifically de-regulated 112 probe sets in NGT and 356 probe sets in insulin resistant PD subjects. Biological significance analysis revealed transient up-regulation of innate and adaptive response related genes following oral glucose load in NGT individuals, which was not observed in PD subjects. Instead, in the PD group, glucose load led to pro-atherogenic and anti-angiogenic gene expression changes. Comparison of gene expression at fasting state in PD versus NGT revealed 21,707 differentially regulated probe sets. Biological significance analysis of the immune function related genes between the two groups (at fasting) revealed increased gene expression for members of the TLR signaling, MHC class II, T-cell receptor, chemotaxis and adhesion pathways. Expression of interferon-γ (IFN-γ) and TNFα was upregulated and that of type-1 interferons and TGF-β was down-regulated at fasting state for PD subjects. Additionally, multiple proteasome subunits and protein arginine methyl transferase genes (PRMTs) were up-regulated and Treg specific genes were significantly de-regulated at fasting in PD subjects. Conclusion Systemic insulin resistance uncovers immune cell plasticity, constitutive TLR activation, enhanced INF-γ signaling, and Treg dysfunction at fasting along with altered gene expression response to oral glucose load.