摘要
目的探讨长链非编码RNA （lncRNA）在潜伏性结核菌感染中差异性表达。
方法选择2015年1至6月在珠海市慢性病防治中心诊断的潜伏性结核菌感染患者血浆33例（病例组），纳入标准为年龄范围18～40岁，确诊的涂阳肺结核病例密切接触者；胸部影像学检查正常；皮肤结核菌素试验强阳性；6个月后随访未患活动性肺结核。对照组选自同一机构体检科健康人群血浆33名，纳入标准为年龄范围18～40岁，排除肺结核或其他肺部疾病，皮肤结核菌素试验阴性。病例组和对照组各3例用于lncRNA芯片实验，30例用于RT-PCR验证实验。Trizol法抽提总RNA后，利用lncRNA芯片技术进行高通量检测，建立潜伏性结核菌感染者的lncRNA基因表达谱。筛选6个差异表达明显的lncRNA，利用实时定量PCR技术，在30例扩大化样本中进行验证，获得高灵敏度、高特异性的生物标志物。利用基因功能分析、基因通路分析初步分析差异基因的表达功能。数据的正态性检验采用Kolmogorov-Smirov（K-S），正态分布资料两个样本均数比较采用t检验，多个样本均数比较采用单因素方差分析，两样本率的比较用χ2检验。
结果 潜伏性结核感染的特异性差异表达lncRNA共1 485个，占所检测lncRNA总数的4.9%，其中上调819个，下调666个。ENST00000566054（t＝10.92，P〈0.001）、TCONS_00016510（t＝2.98，P＝0.004）在潜伏性结核菌感染者血浆中表达水平显著上调，NR_034120表达水平显著下调（t＝－16.63，P〈0.001）。基因功能分析中，上调表达的mRNA中，有382个基因富集于生物学进程、60个基因富集于细胞组件、43个基因富集于分子功能；下调表达的mRNA中，有276个基因富集于生物学进程、55个基因富集于细胞组件、55个基因富集于分子功能；基因通路分析上调的mRNA主要富集于7条生物学途径，下调的mRNA主要富集于7条生物学途径�
ObjectiveTo investigate the differential expression of long non-coding RNA （lncRNA） in latent tuberculous infection （LTBI）.
MethodsA total of 33 diagnosed latent tuberculous infection patients and 33 controls were collected from Zhuhai center for chronic disease and control during January to June in 2015. The age of participates were from 18 to 40 years old. The cases were closely contacted with confirmed smear positive pulmonary tuberculosis and had normal chest radiograph. Meanwhile, the cases behaved strongly positive tuberculin skin test and without suffered active tuberculosis after six month follow-up. The controls were randomly recruited from health examination persons who were negative with tuberculin skin test and without any lung diseases. Among them, 3 patients and 3 controls were used in lncRNA microarray experiments and others were used in RT-PCR validating experiments. RNA in plasma was extracted by Trizol and lncRNA microarray was used to establish the expression spectrum of lncRNA in latent tuberculous infection. Six differentially expressed lncRNA were collected for validating in the thirty latent tuberculous infection cases by using real time （RT）-PCR. Moreover, gene functional analysis and pathway analysis were used to explore the function of differentially expressed lncRNA.SPSS22.0 were used to conduct correlation analysis, the value of P〈0.05 was considered to be statistically significant.
ResultsCompare with healthy controls, 1485 lncRNAs were differentially expressed in LTBI （819 up-regulated and 666 down-regulated）, which accounted for 4.9% of total lncRNAs tested. The expressions of ENST00000566054 （t=10.92, P〈0.001） and TCONS_00016510（t=2.98, P=0.004）in plasma of LTBI was significantly up-regulated, while that of NR_034120 was significantly decreased（t=-16.63, P〈0.001）. The gene functional analysis found that 382 genes were enriched in biological process, 60 in cell composition and 43 in molecular function among the up-regulated mRNAs. In addition,
出处
《中华检验医学杂志》
CAS
CSCD
北大核心
2016年第11期857-863,共7页
Chinese Journal of Laboratory Medicine
基金
基金项目：国家自然科学基金（K16026）
深圳市科技研发资金（SGLH20121008144756945,JCYJ20120618172144495）
