ROLE OF MICRORNAS IN THE MOLECULAR PATHOGENESIS OF HIV/SIV ENTEROPATHY

微生物在 HIV/SIV 肠病分子发病机制中的作用

• 批准号: 8173006
• 负责人: Mahesh Mohan
• 金额: $ 3.94万
• 依托单位: TULANE UNIVERSITY OF LOUISIANA
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-05-01 至 2011-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8173006
• 关键词: Animals; Autopsy; Cells; Chronic; Colitis; Colon; Complex; Computer Retrieval of Information on Scientific Projects Database; Diarrhea; Disease; Epithelium; Funding; Gastrointestinal tract structure; Gene Expression; Grant; Histopathology; Immune system; Immunofluorescence Immunologic; In Situ Hybridization; In Vitro; Infection; Inflammation; Institution; Intestines; Lamina Propria; Macaca; Macaca mulatta; Mass Spectrum Analysis; Messenger RNA; Methods; MicroRNAs; Modeling; Molecular; Nuclear; Nucleotides; Pathogenesis; Regulator Genes; Research; Research Personnel; Resources; Role; Source; Structure; Tissues; Translational Repression; United States National Institutes of Health; Untranslated RNA; improved; intestinal epithelium; locked nucleic acid; macrophage; novel; response; wasting

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Background & Aims: The Gastrointestinal tract (GIT) is a major target of HIV/SIV infection. Although our understanding of HIV/SIV enteropathy has greatly improved, the recent discovery of miRNAs has added yet another novel and complex regulator of gene expression with potential roles in the molecular pathogenesis of this disorder. microRNAs (miRNAs) are genomically transcribed, ~21-23 nucleotide noncoding RNAs that are highly conserved and suppress gene expression by targeting mRNAs for translational repression or degradation. We investigated the contribution of 11 miRNAs to GIT disease and inflammation using the SIV-infected rhesus macaque model. Methods: Colon tissue was collected at necropsy from 10 SIV-infected (G1) with chronic diarrhea and 5 uninfected control macaques (G2). The contributions of miR-21, miR-125b, miR-132, miR-142-3p, miR-142-5p, miR-146a, miR-155, miR-203, miR-212, miR-223 and miR-338, previously known to be associated with inflammation were investigated using QRT-PCR, In situ hybridization/Immunofluorescence, histopathology and mass spectrometry. Results: All G1 macaques had chronic diarrhea, wasting and colitis. Significant to moderate increase in the expression of miR-142-3p, miR-142-5p (5-6-fold), miR-212 (3-fold), miR-21 (2-fold) was observed in the colon of G1 macaques compared to G2 animals. Interestingly, miR-125b and miR-203 were downregulated (2-3 fold) in G1 animals. No change in miR-132, miR-146a, miR-155 , miR-223 and miR-338 expression was observed between the groups. In situ hybridization using Locked-nucleic acid modified miR-212 probes revealed cytoplasmic and nuclear localization in the colonic epithelium and lamina propria cells (macrophages). Using mass spectrometry, we validated several predicted mRNA targets of miR-142-3p and miR-212 following over expression of both miRNAs in in vitro cultured primary intestinal macrophages. Conclusion: Our findings suggest that deregulation in cell/tissue-specific expression of miRNAs occurring in response to HIV/SIV infection may disrupt the functional relationship between the intestinal epithelium and the mucosal immune system causing alterations in intestinal structure and function.

该子项目是利用该技术的众多研究子项目之一 资源由 NIH/NCRR 资助的中心拨款提供。子项目及 研究者 (PI) 可能已从 NIH 的另一个来源获得主要资金， 因此可以在其他 CRISP 条目中表示。列出的机构是 对于中心来说，它不一定是研究者的机构。 背景与目的：胃肠道 (GIT) 是 HIV/SIV 感染的主要目标。尽管我们对 HIV/SIV 肠病的了解已大大提高，但最近对 miRNA 的发现增加了另一种新型且复杂的基因表达调节因子，在这种疾病的分子发病机制中具有潜在作用。 microRNA (miRNA) 是基因组转录的约 21-23 个核苷酸的非编码 RNA，高度保守，通过靶向 mRNA 进行翻译抑制或降解来抑制基因表达。我们使用感染 SIV 的恒河猴模型研究了 11 种 miRNA 对胃肠道疾病和炎症的影响。方法：在尸检时收集 10 只患有慢性腹泻的 SIV 感染者 (G1) 和 5 只未感染的对照猕猴 (G2) 的结肠组织。 miR-21、miR-125b、miR-132、miR-142-3p、miR-142-5p、miR-146a、miR-155、miR-203、miR-212、miR-223 和 miR-338 的贡献，以前已知与炎症相关，使用 QRT-PCR、原位杂交/免疫荧光进行了研究，组织病理学和质谱分析。结果：所有 G1 期猕猴均患有慢性腹泻、消瘦和结肠炎。在结肠中观察到 miR-142-3p、miR-142-5p（5-6 倍）、miR-212（3 倍）、miR-21（2 倍）的表达显着至中度增加G1 猕猴与 G2 动物的比较。有趣的是，miR-125b 和 miR-203 在 G1 动物中下调（2-3 倍）。各组之间未观察到 miR-132、miR-146a、miR-155、miR-223 和 miR-338 表达的变化。使用锁核酸修饰的 miR-212 探针进行的原位杂交揭示了结肠上皮和固有层细胞（巨噬细胞）中的细胞质和核定位。在体外培养的原代肠道巨噬细胞中过度表达 miR-142-3p 和 miR-212 后，我们使用质谱法验证了 miR-142-3p 和 miR-212 的几个预测 mRNA 靶点。结论：我们的研究结果表明，HIV/SIV 感染引起的细胞/组织特异性 miRNA 表达失调可能会破坏肠上皮和粘膜免疫系统之间的功能关系，从而导致肠道结构和功能的改变。