微小RNA在慢性咀嚼肌疼痛中作用的初步探索

Preliminary exploration of the role of microRNA in chronic masticatory myalgia

目的:研究微小RNA(miRNA)在慢性咀嚼肌疼痛(CMM)中的调控作用。方法:收集北京大学口腔医院颞下颌关节病及口颌面痛门诊CMM患者23名,以性别及年龄匹配的健康志愿者作为对照,静脉采血2 mL;以可能通过丝裂原活化蛋白激酶激酶3(MEK3)基因调控CMM的miR-15a-5p和miR-19b-3p作为目标miRNA,用qRT-PCR进一步检测可能相关的miRNA。进行人单核细胞白血病细胞(THP-1)转染和miRNA过表达实验,并通过qRT-PCR及Western blot验证miRNA与MEK3基因的关系。结果:qRT-PCR检测发现miR-19b-3p在CMM患者组显著下调(P<0.05),miR-15a-5p在两组间无显著性差异(P>0.05)。与miR-NC组比较,miR-19b-3p过表达组THP-1细胞的MEK3 mRNA和蛋白的表达无显著性差异(P>0.05)。结论:miR-19b-3p可能参与了CMM,但可能不是通过与MEK3基因直接结合发挥作用。

Objective:To investigate the regulatory effect of miRNA on chronic masticatory myalgia(CMM).Methods:Twenty-three CMM patients were recruited from the Center for TMD and Orofacial Pain of Peking University School and Hospital of Stomatology,23 gender-and age-matched healthy people recruited as control group.A total of 2 mL peripheral blood was collected from each participant of two groups.MiR-15a-5p and miR-19b-3p were preliminarily screened out as target miRNAs which are associated with CMM by regulating MEK3 according to Targetscan database and DIANA database in the previous experiments.QRT-PCR was used to select the possible miRNA further.Subsequently,the cell transfection and miRNA-overexpression experiment were conducted to verify the relationship of miRNA and MEK3 gene with qRT-PCR and Western blot.Results:MiR-19b-3p expressions were significantly decreased in CMM group compared to healthy control group using qRT-PCR assays(P<0.05),and miR-15a-5p expressions were not significantly different between these two groups(P>0.05).MiR-19b-3p was selected to verify the relationship between miRNA and MEK3 gene in the following experiments.The results showed that the mRNA and protein of MEK3 gene had no significant difference between miR-19b-3p-overexpression group and miR-NC group in human leukemia monocytic cell line(THP-1)at 24,48 and 72 h(P>0.05).Conclusions:MiR-19b-3p may be involved in CMM and its mechanism is likely to be complicated.It may not work by binding to the MEK3 gene directly.