基于miRNA-29c/白血病抑制因子轴探讨针刺调节脾虚大鼠肠道炎症机制研究  

Exploring Mechanism of Intestinal Inflammation Regulation by Acupuncture in Rats with Spleen Deficiency Based on miRNA-29c/LIF Axis

作　　者：张家豪 王建波[1,2] 薛亚楠[1,2] 曲怡[1,2] 董佳梓 王天朗 金璐 刘继东[1,2] ZHANG Jiahao;WANG Jianbo;XUE Yanan;QU Yi;DONG Jiazi;WANG Tianlang;JIN Lu;LIU Jidong(Liaoning University of Traditional Chinese Medicine,Shenyang 110847,Liaoning,China;Key Laboratory of The Theory and Application of Viscera in Chinese Medicine,Ministry of Education,Liaoning University of Traditional Chinese Medicine,Shenyang 110847,Liaoning,China)

机构地区：[1]辽宁中医药大学,辽宁沈阳110847 [2]辽宁中医药大学中医脏象理论及应用教育部重点实验室,辽宁沈阳110847

出　　处：《中华中医药学刊》2024年第5期98-102,I0024-I0027,共9页Chinese Archives of Traditional Chinese Medicine

基　　金：国家自然科学基金项目(81603704);辽宁省教育厅高等学校基本科研项目(LJKZ0886);辽宁省科学技术计划项目(2021-MS-248)。

摘　　要：目的观察针刺调节miRNA-29c/白血病抑制因子(leukemia inhibitory factor,LIF)轴改善脾虚大鼠结肠炎症机制,探讨“健脾益气”针刺疗法改善脾虚便溏症状现代作用机理。方法将60只SD雄性大鼠以随机数字表法分为正常组、模型组、非经非穴组、针刺组,每组15只。除正常组外,其余3组采用劳倦过度和不规律饮食复合方法建立脾虚证大鼠模型。造模成功后,针刺组电针“足三里”穴及“脾俞”穴干预治疗,非经非穴组给予电针尾尖作为对照干预,每日1次/20 min,共治疗14 d。治疗结束后,取大鼠结肠组织及全血。qPCR检测大鼠结肠组织miRNA-29/LIF轴相关基因的表达;Western blot检测大鼠白介素-6(interleukin6,IL-6)、肿瘤坏死因子α(tumornecrosisfactor-α,TNF-α)的表达;ELISA检测结肠组织中白介素-1β(interleukin1β,IL-1β)、白介素-18(interleukin18,IL-18)的含量;HE染色法观察大鼠结肠组织病理形态变化;TUNEL染色观察各组结肠组织凋亡情况;流式细胞术观察外周血单个核细胞(PBMC)凋亡状况。结果与正常组相比,模型组大鼠结肠组织LIF的表达水平明显下降(P<0.05),miRNA-29、IL-1β、IL-6、白介素-8(interleukin18,IL-8)、TNF-αmRNA表达以及IL-6、TNF-α蛋白表达水平有所升高(P<0.05);与模型组相比,针刺组大鼠结肠组织LIF表达水平明显升高(P<0.05),miRNA-29c、IL-1β、IL-6、IL-8、TNF-αmRNA表达以及IL-6、TNF-α蛋白表达水平有所降低(P<0.05),非经非穴组差异无统计学意义(P>0.05);与正常组相比,模型组大鼠结肠组织中促炎因子IL-1β、IL-18表达水平明显升高(P<0.05),与模型组相比,针刺组大鼠结肠组织中促炎因子IL-1β、IL-18的表达水平有所降低(P<0.05)。另外HE染色结果显示,针刺治疗可有效减轻结肠黏膜组织中炎症细胞浸润和杯状细胞的损失;TUNEL染色结果显示,针刺可以抑制miRNA-29c/LIF轴,改善脾虚大鼠结肠炎症,抑制结肠黏膜细胞凋亡;流式细胞术�Objective To observe the mechanism of miRNA-29c/leukemia inhibitory factor(LIF)axis regulation by acupunc⁃ture to improve colonic inflammation in rats with spleen deficiency and explore the modern mechanism of“strengthening spleen and benefiting Qi”acupuncture therapy to improve the symptoms of spleen deficiency and loose stool.Methods Sixty SD male rats were divided into normal group,model group,non-acupoint group and acupuncture group by random number table method,with 15 rats in each group.Except for the normal group,the remaining three groups were used to establish the rat model of spleen deficiency syndrome by the compound method of overwork and irregular diet.After successful modelling,the acupuncture group was treated with electro-acupuncture at the“Zusanli(ST36)”and“Pishu(BL20)”,while the non-acupoint group was treated with electro-acupuncture at the tail tip as a control,once a day for 20 min for a total of 14 d.After treatment,colon tissues and whole blood were collected from the rats.The expressions of miRNA-29/LIF axis-related genes in rat colon tissues were detec⁃ted by qPCR.The expressions of interleukin-6(IL-6)and tumor necrosis factor-α(TNF-α)in rat colon tissues were de⁃tected by Western blot.The contents of interleukin-1β(IL-1β)and nterleukin-18(IL-18)in colon tissues were detected by ELISA.The pathological morphological changes in rat colon tissues were observed by HE staining.The apoptosis of colon tis⁃sues in each group was observed by TUNEL staining.Flow cytometry was used to observe the apoptotic status of peripheral blood mononuclear cells(PBMC cells).Results Compared with those of the normal group,the expression of LIF in colon tissues of rats in the model group decreased significantly(P<0.05),while the expressions of miRNA-29,IL-1β,IL-6,IL-8 and TNF-αmRNA and IL-6 and TNF-αprotein increased(P<0.05).Compared with those of the model group,the expression of LIF in the colon tissue of rats in the acupuncture group was significantly increased(P<0.05),and the express

关 键 词：足三里 脾俞 脾虚 肠道炎症 miRNA-29c/LIF 

分 类 号：R245.31[医药卫生—针灸推拿学]