Abstract

This study elucidates the immunological implications of methylglyoxal (MGO)-modified low-density lipoprotein (LDL) in type 2 diabetes mellitus (T2DM) patients. Under in vitro modifications, MGO altered the tertiary structure of LDL. 2,4,6-Trinitrobenzene sulfonic acid (TNBS) and phenanthrenequinone assays confirmed lysine and arginine residues as main targets of MGO in LDL. High-performance liquid chromatography (HPLC) and Liquid chromatography-mass spectrometry (LC-MS) studies confirmed the generation of N^ϵ-(carboxymethyl) lysine in the modified protein. Comet assay showing increased tail length of DNA in lymphocytes inferred the cytotoxicity of MGO-LDL. The easy penetration of MGO-LDL into the nucleus is possibly a consequence of its reduced size, postmodification, as observed from hydrodynamic radii studies in Dynamic light scattering (DLS) experiments. MGO-LDL was found to be more immunogenic, as compared to native LDL, in immunological studies conducted on experimental rabbits. Our results reflect the presence of neo-antigenic determinants on modified LDL. Competitive inhibition enzyme-linked immunosorbent assay suggested the presence of neo-epitopes with marked immunogenicity eliciting specific immune response. Binding studies on purified immunoglobulin G confirmed the enhanced and specific immunogenicity of MGO-LDL. Studies on interaction of MGO-LDL with the circulating auto-antibodies from T2DM patients showed high affinity of serum antibodies toward MGO-LDL. This study suggests a potent role of glycoxidatively modified LDL in the generation of auto-immune response in T2DM patients.

中文翻译：

---

低密度脂蛋白的糖氧化产生细胞毒性加合物并引发 2 型糖尿病的体液反应

 抽象的

本研究阐明了甲基乙二醛 (MGO) 修饰的低密度脂蛋白 (LDL) 对 2 型糖尿病 (T2DM) 患者的免疫学影响。在体外修饰下，MGO 改变了 LDL 的三级结构。 2,4,6-三硝基苯磺酸 (TNBS) 和菲醌测定证实赖氨酸和精氨酸残基是 LDL 中 MGO 的主要靶标。高效液相色谱 (HPLC) 和液相色谱-质谱 (LC-MS) 研究证实修饰蛋白中生成了 N ^ϵ -(羧甲基)赖氨酸。彗星试验显示淋巴细胞中 DNA 尾部长度增加，推断出 MGO-LDL 具有细胞毒性。正如动态光散射 (DLS) 实验中的流体动力学半径研究所观察到的，MGO-LDL 容易渗透到细胞核中可能是其尺寸减小和修饰后的结果。在对实验兔进行的免疫学研究中发现，与天然 LDL 相比，MGO-LDL 更具免疫原性。我们的结果反映了修饰 LDL 上新抗原决定簇的存在。竞争性抑制酶联免疫吸附测定表明存在具有明显免疫原性的新表位，可引发特异性免疫反应。对纯化免疫球蛋白 G 的结合研究证实了 MGO-LDL 增强的特异性免疫原性。 MGO-LDL 与 T2DM 患者循环自身抗体相互作用的研究表明，血清抗体对 MGO-LDL 具有高亲和力。这项研究表明，糖氧化修饰的 LDL 在 T2DM 患者自身免疫反应的产生中发挥着重要作用。