Laboratory of Vascular Biology, Department of Biotechnology, Bhupat and Jyoti Mehta School of Biosciences, Indian Institute of Technology Madras, Chennai, India.
Bionivid Technology Pvt. Ltd., Bangalore, India.
Clin Nutr. 2021 Mar;40(3):1247-1259. doi: 10.1016/j.clnu.2020.08.007. Epub 2020 Aug 13.
Metabolic disorders including diabetes are associated with immune cell dysfunction. However, the effect of normal glucose metabolism or impairment thereof on immune cell gene expression is not well known. Hence, in this cross-sectional pilot study, we sought to determine the differences in gene expression in the peripheral blood mono-nuclear cells (PBMCs) of normal glucose tolerant (NGT) and prediabetic (PD) Asian Indian men, at fasting and in response to 75 g oral glucose load.
Illumina HT12 bead chip-based microarray was performed on PBMCs at fasting and 2-h 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 within and across the two groups were identified 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 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.
Glucose load specifically altered the expression of 112 genes in NGT and 356 genes in PD subjects. Biological significance analysis revealed transient up-regulation of innate and adaptive immune 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 an increase in the expression of pro-atherogenic and anti-angiogenic genes. Comparison of gene expression at fasting state in PD versus NGT revealed 21,707 differentially expressed genes. Biological significance analysis of the immune function related genes between these two groups (at fasting) revealed higher gene expression of members of the TLR signaling, MHC class II molecules, and T-cell receptor, chemotaxis and adhesion pathways in PD subjects. Expression of interferon-γ (IFN-γ) and TNFα was higher and that of type-1 interferons and TGF-β was lower at fasting state in PD subjects compared to NGT. Additionally, expression of multiple proteasome subunits and protein arginine methyl transferase genes (PRMTs) were higher and that of Treg specific genes was significantly distinct at fasting in PD subjects compared to NGT.
Prediabetes uncovers constitutive TLR activation, enhanced IFN-γ signaling, and Treg dysfunction at fasting along with altered gene expression response to oral glucose load.
代谢紊乱包括糖尿病与免疫细胞功能障碍有关。然而，正常葡萄糖代谢或受损对免疫细胞基因表达的影响尚不清楚。因此，在这项横断面初步研究中，我们旨在确定空腹和口服 75g 葡萄糖负荷后，正常葡萄糖耐量（NGT）和糖尿病前期（PD）亚洲印度男性外周血单核细胞（PBMC）中的基因表达差异。
对空腹和负荷后 2 小时的 NGT（N=6）和 PD（N=9）受试者的 PBMC 进行 Illumina HT12 珠芯片微阵列分析。在对原始数据进行归一化和适当的质量控制后，使用 GeneSpring GX V12.0 软件识别不同条件下两组内和组间的差异表达基因（DEG）。应用配对和非配对学生 t 检验以及适当比较的倍数变化截止值。通过实时 qPCR 分析验证微阵列数据。通过使用 DEG 和 DAVID 资源分析显著调节的生物学途径。使用 CIBERSORT 对空腹时 NGT 和 PD 受试者之间的 DEG 进行去卷积，并进一步分析参与调节性 T 细胞（Treg）功能的基因的生物学意义。
葡萄糖负荷特异性改变了 NGT 中的 112 个基因和 PD 中的 356 个基因的表达。生物学意义分析表明，NGT 个体口服葡萄糖负荷后，先天和适应性免疫反应相关基因短暂上调，而 PD 个体则未观察到这种情况。相反，在 PD 组中，葡萄糖负荷导致致动脉粥样硬化和抗血管生成基因的表达增加。PD 与 NGT 空腹时的基因表达比较显示 21707 个差异表达基因。对这两组（空腹时）的免疫功能相关基因的生物学意义分析显示，PD 组中 TLR 信号、MHC Ⅱ类分子和 T 细胞受体、趋化和粘附途径的成员的基因表达更高。与 NGT 相比，PD 患者空腹时干扰素-γ（IFN-γ）和 TNFα 的表达更高，而 1 型干扰素和 TGF-β 的表达更低。此外，PD 患者空腹时多个蛋白酶体亚基和蛋白质精氨酸甲基转移酶基因（PRMTs）的表达更高，而 Treg 特异性基因的表达明显不同。
糖尿病前期揭示了空腹时 TLR 持续激活、增强的 IFN-γ 信号传导和 Treg 功能障碍，以及口服葡萄糖负荷后基因表达的改变。
