Elevated CD4 + T-cell glucose metabolism in HIV+ women with diabetes mellitus

Abstract

Objective: Immune dysfunction and chronic inflammation are characteristic of HIV infection and diabetes mellitus, with CD4 + T-cell metabolism implicated in the pathogenesis of each disease. However, there is limited information on CD4 + T-cell metabolism in HIV+ persons with diabetes mellitus. We examined CD4 + T-cell glucose metabolism in HIV+ women with and without diabetes mellitus.

Design: A case-control study was used to compare CD4 + T-cell glucose metabolism in women with HIV with or without diabetes mellitus.

Methods: Nondiabetic (HIV+DM-, N = 20) or type 2 diabetic HIV+ women with (HIV+DM+, N = 16) or without (HIV+DMTx+, N = 18) antidiabetic treatment were identified from the WIHS and matched for age, race/ethnicity, smoking status and CD4 + cell count. CD4 + T-cell immunometabolism was examined by flow cytometry, microfluidic qRT-PCR of metabolic genes, and Seahorse extracellular flux analysis of stimulated CD4 + T cells.

Results: HIV+DM+ displayed a significantly elevated proportion of CD4 + T cells expressing the immunometabolic marker GLUT1 compared with HIV+DMTx+ and HIV+DM- ( P = 0.04 and P = 0.01, respectively). Relative expression of genes encoding key enzymes for glucose metabolism pathways were elevated in CD4 + T cells of HIV+DM+ compared with HIV+DMTx+ and HIV+DM-. T-cell receptor (TCR)-activated CD4 + T cells from HIV+DM+ showed elevated glycolysis and oxidative phosphorylation compared with HIV+DM-.

Conclusion: CD4 + T cells from HIV+DM+ have elevated glucose metabolism. Treatment of diabetes mellitus among women with HIV may partially correct CD4 + T-cell metabolic dysfunction.

Figure 1.

The proportions of total CD4+ T cells and CD4+ T cell subpopulations expressing GLUT1. GLUT1 expressing T cells were identified by light scatter, AQUA Live/Dead and CD3 and CD4 expression. CD4+ T cell subpopulations were then identified by CD45RA and CCR7 expression. (A) Number of CD4+ T cells as a proportion of cells. (B) Proportions of naïve (CD45RA+CCR7+), effector memory (CD45RA-CCR7-), central memory (CD45RA-CCR7+) and terminally differentiated (TemRA) (CD45RA+CCR7-) live CD4+ T cells. (C) Proportion of GLUT1-expressing CD4+ T cells and (D) proportion of GLUT1-expressing CD4+ T cell subpopulations. Open squares represent women with viral loads >200 copies/mL and not on ART at the time of sampling. Closed squares represent women on ART with viral load >200 copies/mL at the time of sampling. Blue circles represent women not on ART with viral load <200 copies/mL at the time of sampling. Groups were compared using the Mann Whitney U test. A p-value <0.05 was considered significant. HIV+DM+ women, n = 13; HIV+DMTx+ women, n = 15; HIV+DM− women, n = 17.

Figure 2.

CD4+ T cell expression of glucose metabolism genes. Relative gene expression of 82 genes encoding enzymes and isozymes in glucose metabolism pathways assessed by Fluidigm HD Biomark RT-PCR of RNA extracted from CD4+ T cells purified by magnetic bead negative selection. (A) Heatmaps depicting the relative expression of metabolic pathway gene sets as listed in Table S2. (B) CD4+ T cell glucose metabolism pathways highlighting enzymes encoded by genes with increased relative expression in HIV+ women with DM compared to controls. (C) Gene encoding enzymes with elevated relative expression in HIV+ women with DM compared to controls. Fold-change in gene expression calculated using 2^−ΔΔCt with RPL30 as reference gene and HIV+DM− as reference samples. Genes with 95% C.I. >1 or 95% C.I. <1 were assessed for differences between groups. Fold change was compared between groups using one-way ANOVA with Tukey post-hoc test for multiple comparison. A p < 0.05 was considered significant. HIV+DM+ women, n = 5; HIV+DMTx+ women, n = 10; HIV+DM− women, n = 6. Created with BioRender.com

Figure 3.

Glycolytic metabolism and oxidative phosphorylation of stimulated CD4+ T cells. CD4+ T cells were purified by magnetic bead negative selection and stimulated using anti-CD3/CD28 Dynabeads. (A) Extracellular acidification rate (ECAR) and (B) oxygen consumption rate (OCR) were evaluated. All measurements were normalized to cell count. HIV+DM+ women, n = 4; HIV+DMTx+ women, n = 4; HIV+DM− women, n = 4. Each sample had 3 technical replicates.