Department of Biosciences and Biomedical Engineering, Indian Institute of Technology, Indore, India.
Materials and Device Laboratory, Department of Physics, Indian Institute of Technology, Indore, India.
Front Immunol. 2023 Oct 9;14:1192032. doi: 10.3389/fimmu.2023.1192032. eCollection 2023.
EBV infection has long been postulated to trigger multiple sclerosis (MS) and anti-EBV antibodies showed a consistent presence in MS patients. Previous reports from our group have shown that the EBV infects different brain cells. Entry of the virus in neuronal cells is assisted by several host factors including membrane cholesterol. By using an inhibitor, methyl-β-cyclodextrin (MβCD), we evaluated the role of membrane cholesterol in EBV infection and pathogenesis.
The membrane cholesterol depleted cells were infected with EBV and its latent genes expression were assessed. Further, EBV-mediated downstream signalling molecules namely STAT3, RIP, NF-kB and TNF-α levels was checked at protein level along with spatial (periphery and nucleus) and temporal changes in biomolecular fingerprints with Raman microspectroscopy (RS).
Upon treatment with MβCD, lmp1 and lmp2a suggested significant downregulation compared to EBV infection. Downstream molecules like STAT3 and RIP, exhibited a decrease in protein levels temporally upon exposure to MβCD while NF-kB levels were found to be increased. Further, the intensity of the Raman spectra exhibited an increase in triglycerides and fatty acids in the cytoplasm of EBV-infected LN-229 cells compared to MβCD+EBV. Likewise, the Raman peak width of cholesterol, lipid and fatty acids were found to be reduced in EBV-infected samples indicates elevation in the cholesterol specific moieties. In contrast, an opposite pattern was observed in the nucleus. Moreover, the ingenuity pathway analysis revealed protein molecules such as VLDLR, MBP and APP that are associated with altered profile of cholesterol, fatty acids and triglycerides with infection-related CNS disorders.
Taken together, our results underline the important role of membrane cholesterol over EBV entry/pathogenesis in astroglia cells which further trigger/exacerbate virus-associated neuropathologies. These results likely to aid into the prognosis of neurological disease like MS.
EBV 感染长期以来被认为是引发多发性硬化症（MS）的原因，抗 EBV 抗体在 MS 患者中持续存在。我们小组之前的报告显示，EBV 感染不同的脑细胞。病毒进入神经元细胞需要多种宿主因子的辅助，包括膜胆固醇。我们使用一种抑制剂甲基-β-环糊精（MβCD），评估了膜胆固醇在 EBV 感染和发病机制中的作用。
用 MβCD 处理耗竭膜胆固醇的细胞，评估 EBV 及其潜伏基因的表达。进一步，通过蛋白质水平检查 EBV 介导的下游信号分子，如 STAT3、RIP、NF-κB 和 TNF-α，同时使用拉曼微光谱（RS）检查生物分子指纹的空间（外周和核）和时间变化。
在用 MβCD 处理后，与 EBV 感染相比，lmp1 和 lmp2a 显示出显著下调。下游分子如 STAT3 和 RIP 在暴露于 MβCD 时在时间上表现出蛋白水平下降，而 NF-κB 水平则升高。此外，与 MβCD+EBV 相比，感染 EBV 的 LN-229 细胞的细胞质中拉曼光谱的强度显示出甘油三酯和脂肪酸的增加。同样，在感染 EBV 的样本中，胆固醇、脂质和脂肪酸的拉曼峰宽减小表明胆固醇特异性部分的增加。相比之下，在核中观察到相反的模式。此外，通路分析揭示了与胆固醇、脂肪酸和甘油三酯的改变有关的 VLDLR、MBP 和 APP 等蛋白分子与感染相关的中枢神经系统疾病。
综上所述，我们的结果强调了膜胆固醇在星形胶质细胞中对 EBV 进入/发病机制的重要作用，这进一步引发/加重了与病毒相关的神经病理学。这些结果可能有助于 MS 等神经疾病的预后。
