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课题组前期研究发现宿州地方特色药材王枣子中总黄酮(total flavonoids of Isodon amethystoides,TFIA)对佐剂性关节炎(adjuvant arthritis,AA)具有一定的治疗作用,并且这种治疗作用可能是通过对miR-152表达的上调实现的,该文进一步研究TFIA对AA大鼠病理机制影响的分子机制。采用完全弗氏佐剂制备AA大鼠,TFIA灌胃治疗,采用Real-time qPCR检测TFIA灌胃治疗对AA大鼠关节滑膜成纤维样滑膜细胞(fibroblast like synoviocytes,FLS)中miR-152、甲基化酶DNMT1及甲基化结合蛋白MeCP2构成的负调控环路的影响,对miR-152下游经典Wnt信号通路的影响,对AA大鼠病理基因fibronectin表达的影响。TFIA灌胃治疗显著抑制DNMT1表达,逆转了AA大鼠病理中存在的由miR-152,DNMT1和MeCP2构成的负调控环路,向治疗组FLS转染miR-152 inhibitors后,DNMT1表达显著恢复。TFIA灌胃治疗显著上调SFRP4表达,抑制经典Wnt信号通路关键基因β-catenin,C-myc和ccnd1表达,显著抑制AA病理基因fibronectin表达,向治疗组FLS转染miR-152 inhibitors后,逆转了TFIA灌胃治疗对SFRP4,β-catenin,C-myc,ccnd1和fibronectin表达的影响。TFIA可能通过miR-152表达的上调抑制DNMT1表达,上调SFRP4表达,抑制经典Wnt信号关节基因β-catenin,C-myc,ccnd1表达,抑制RA基因fibronectin表达。
Our previous study found that the total flavonoids of Isodon amethystoides (TFIA), a local medicinal herb in Suzhou, had a certain therapeutic effect on adjuvant arthritis (AA), and this therapeutic effect may be achieved by miR-152 expression is up-regulated, this paper further studies the molecular mechanism of TFIA on the pathological mechanism of AA rats. AA rats were treated with complete Freund’s adjuvant and TFIA was given intragastrically. Real-time qPCR was used to detect the effect of TFIA on the expression of miR-152 in synovial fibroblast synoviocytes (FLS) of AA rats , Methylating enzyme DNMT1 and methylating binding protein MeCP2, on the canonical Wnt signaling pathway downstream of miR-152, and on the expression of fibronectin in AA rats. TFIA gavage treatment significantly inhibited the expression of DNMT1 and reversed the negative regulatory loop consisting of miR-152, DNMT1 and MeCP2 in the pathological changes of AA rats. The expression of DNMT1 was significantly restored after transfection with miR-152 inhibitors in FLS treated group. TFIA gavage treatment significantly upregulated the expression of SFRP4, inhibited the expression of β-catenin, C-myc and ccnd1, the key genes of canonical Wnt signaling pathway, significantly inhibited the expression of fibronectin in AA. Effect of intragastric administration on the expression of SFRP4, β-catenin, C-myc, ccnd1 and fibronectin. TFIA may inhibit the expression of DNMT1, upregulate the expression of SFRP4, downregulate the expressions of β-catenin, C-myc and ccnd1, and inhibit the expression of fibronectin of RA gene by upregulation of miR-152.