论文部分内容阅读
目的分析活动性肺结核患者(TB)外周血CD14+单核细胞mi RNA表达谱,筛选一组用于结核诊断及鉴别诊断的分子标志物。方法选取健康对照(HD)、结核菌潜伏感染者(LTBI)、TB各6例,采集外周抗凝血并分离外周单个核细胞(PBMC),再用免疫磁珠技术分选CD14+单核细胞,提取总RNA,利用mi RNA芯片检测CD14+单核细胞中mi RNA表达谱。筛选差异表达的mi RNA分子,利用Taq Man探针q PCR技术在大样本人群中进行验证(HD、LTBI、TB各25例)。结果基因芯片结果显示mi R-487家族可以将三组人群进行有效区分,依据其在不同人群中表达趋势共分为4个基因簇。从每个基因簇中筛选具有代表性的4个mi RNA分子进行Taq Man q PCR验证,包括mi R-487b-3p、mi R-134-5p、mi R-487a-3p、mi R-539-3p,其表达趋势与芯片结果一致。其中TB人群mi R-487b-3p表达量显著低于HD、LTBI,差异有统计学意义(P<0.01,P<0.01);mi R-487a-3p在LTBI、TB人群中表达水平显著低于HD(P<0.01,P<0.05);TB人群mi R-539-3p表达水平显著高于HD、LTBI(P<0.05,P<0.05)。结论 mi R-487家族与结核菌感染、发病的关系密切,其中mi R-487b-3p联合mi R-539-3p可以作为活动性结核诊断标识,而mi R-487a-3p可以作为结核菌感染诊断标识。
Objective To analyze the miRNA expression profile of CD14 + monocytes in peripheral blood of patients with active pulmonary tuberculosis (TB) and to screen a group of molecular markers for the diagnosis and differential diagnosis of tuberculosis. Methods The peripheral blood mononuclear cells (PBMCs) were collected from 6 cases of healthy control (HD), LTBI and TB, and peripheral blood mononuclear cells (PBMCs) were collected. Immunomagnetic beads were used to sort CD14 + monocytes. Total RNA was extracted and miRNA expression profiling in CD14 + monocytes was detected by mi RNA microarray. The differentially expressed miRNA molecules were screened and validated in a large sample population using Taq Man probe q PCR (25 in HD, LTBI, and TB). Results Gene microarray results showed that the mi R-487 family can effectively distinguish three groups of people, based on their expression trends in different populations is divided into four gene clusters. Taq Man q PCR validation of 4 representative mi RNA molecules was screened from each gene cluster, including mi R-487b-3p, mi R-134-5p, mi R-487a-3p, mi R-539- 3p, the expression trend consistent with the chip results. The expression of mi R-487b-3p in TB patients was significantly lower than that in HD and LTBI (P <0.01, P <0.01). The expression of mi R-487a-3p was significantly lower in LTBI and TB HD (P <0.01, P <0.05). The expression of mi R-539-3p in TB patients was significantly higher than HD and LTBI (P <0.05, P <0.05). Conclusion mi R-487b-3p combined with mi R-539-3p can be used as a marker for diagnosis of active tuberculosis, while mi R-487a-3p can be used as a marker of tuberculosis infection Diagnostic identification.