mir-124-3p/SPOCK2在急性呼吸窘迫综合征早产儿血浆中表达的临床研究

The clinical values of mir-124-3p and SPOCK2 protein detection in serums of preterm infants with RDS

目的 :探讨测定急性呼吸窘迫综合征早产儿血浆中mir-124-3p和SPOCK2蛋白的临床意义,为早产儿严重肺损伤寻找新的高危因子和治疗靶标。方法:根据早产儿呼吸窘迫综合征(respiratory distress syndrome,RDS)诊断标准,早产儿分成RDS组17例及对照组(无RDS等并发症的普通早产儿)21例。采用ELISA和Western blot法测定外周血浆中和体外培养的人肺成纤维细胞中的SPOCK2蛋白。采用Target Scan分析与SPOCK2基因3′末端非编码区结合的可能mi RNA,并用荧光实时定量RTPCR扩增候选mi RNA。采用Pearson统计方法对血浆中SPOCK2蛋白和mi RNA进行相关分析。构建mir-124-3p慢病毒载体,并转染人肺成纤维细胞,研究mir-124-3p过表达对SPOCK2蛋白表达的影响。结果:对照组早产儿血浆中SPOCK2蛋白浓度为(7.24±0.43)μg/L,而RDS组升高为(16.43±0.54)μg/L,两组间比较差异有统计学意义(t=12.81,P<0.01)。SPOCK2基因3′末端存在mir-124-3p、mir-25-3p、mir-122-5p等候选的mi RNA结合位点。荧光定量RT-PCR证实外周血中不能测到mir-25-3p、mir-122-5p。mir-124-3p表达在对照组中较高,在RDS组中显著降低,差异有统计学意义(P<0.05)。并且mir-124-3p的表达量与SPOCK2蛋白血浆水平呈负相关(r=-0.645 9,P=0.012 6)。mir-124-3p慢病毒载体转染可以显著抑制人肺成纤维细胞中SPOCK2蛋白的表达。结论:测定早产儿血浆中SPOCK2蛋白,可能对RDS等肺疾病的病情评估有一定临床意义。利用mir-124-3p调控SPOCK2蛋白表达,可能对早产儿RDS等严重肺损伤的治疗有潜在应用价值。

Objective：To study the clinical values of detection for SPOCK2 protein in serums of preterm infants and to determine a novel risk factor and therapeutic target against serious lung injuries of premature neonates. Methods：Upon diagnostic criteria of Respiratory Distress Syndrome（RDS）,premature infants were divided into Group RDS（17 cases）and control（21 cases without RDS）.ELISA and Western blot were performed to investigate the serum levels of SPOCK2 protein in every groups and expressions of SPOCK2 in human lung fibroblasts,respectively. The noncoding region in 3 terminal of SPOCK2 gene was analyzed to scan candidate binding sites of micro RNA with software Target Scan online and followed verification was performed by fluorescence real-time quantification PCR. Person method was used to analyze the correlation between SPOCK2 and micro RNA. The lentiviral vector of mir-124-3p was constructed and transfected into human lung fibroblasts. The regulatory effect of mir-124-3p on SPOCK2 expression was studied by Western blotting. Results：The serum levels of SPOCK2 protein in Group RDS were 16.43 ± 0.54 μg / L,which was higher than that of control（7.24 ± 0.43 μg / L,t=12.81,P 〈 0.01）. There were several micro RNAs located in the 3 terminal of SPOCK2 gene,including mir-124-3p,mir-25-3p,and mir-122-5p. However,mir-25-3p and mir-122-5p failed to be observed in serums of preterm infants. The expression of mir-124-3p was negatively correlative with that of SPOCK2 protein in preterm infants with lung injury（r=-0.645 9,P=0.012 6）. Mir-124-3p overexpression markedly suppressed the expression of SPOCK2 protein in human fibroblasts. Con-clusion：The detection of SPOCK2 protein in serum benefitted evaluating severity of preterm infants with RDS. Mir-124-3p down-regulated the expression of SPOCK2 protein,which might be helpful to build a novel therapeutic strategy against lung injury of premature neonates.