甲氨蝶呤对炎性细胞因子及丝裂原活化蛋白激酶信号通路的影响

中国临床药理学与治疗学 ›› 2014, Vol. 19 ›› Issue (12): 1347-1351.

甲氨蝶呤对炎性细胞因子及丝裂原活化蛋白激酶信号通路的影响

蔡质其¹, 张育², 张学增³, 沈维干²

¹扬州文峰社区卫生服务中心,扬州 225003,江苏;
²扬州大学临床医学院,扬州 225001,江苏;
³山东省青岛卫生学校,青岛 266071,山东

收稿日期:2014-04-02 修回日期:2014-08-21 发布日期:2020-07-20
通讯作者: 张育,通信作者,男,主任医师,教授,研究方向:炎性关节病的基础与临床研究。Tel:0514-7978805 E-mail:yzzy10182001@aliyun.com
作者简介:蔡质其,男,副主任医师,研究方向:脊柱关节病变的基础与临床研究。Tel:13013708518 E-mail:377586392@qq.com

Study on the effect of methotrexate on inflammatory cytokines and MAPK signal pathway

CAI Zhi-qi¹, ZHANG Yu², ZHANG Xue-zeng³, SHEN Wei-gan¹

¹Wenfeng Community Health Service Center of Yangzhou, Yangzhou 225003, Jiangsu, China;
²Clinic Medical College of Yangzhou University, Yangzhou 225001, Jiangsu, China;
³Qingdao Health School, Qingdao 266071, Shandong, China

Received:2014-04-02 Revised:2014-08-21 Published:2020-07-20

摘要/Abstract

摘要： 目的: 探讨甲氨蝶呤(methotrexate,MTX)对类风湿关节炎(RA)发病起重要作用的炎性细胞因子IL-1β和TNF-α的作用及其可能的机制。方法: 用II型胶原建立类风湿关节炎(CIA)动物模型,以MTX(0.2 mg/kg/W)进行处理,6周后处死大鼠,取血清进行IL-1β和TNF-α检测;分离大鼠膝关节取关节滑膜细胞(FLS)进行培养、鉴定,检测脂多糖(LPS)诱导的FLS培养上清中IL-1β和TNF-α的含量,并观察MTX对FLS分泌IL-1β和TNF-α的影响;在FLS的培养中加入不同剂量的MTX,用Westen Blot方法检测FLS中ERK蛋白及磷酸化蛋白的表达。结果: 关节炎指数评分、关节肿胀度测定及关节病理显示造模成功,分离关节滑膜细胞经流式细胞仪检测FLS的血管细胞黏附分子-1(VCAM-1)的表达为 85.5%。造模后,CIA大鼠血清IL-1β和TNF-α明显增加,与空白对照组比较有统计学差异(P<0.05),MTX能有效减少CIA大鼠血清中IL-1β和TNF-α的含量(P<0.05),但未能恢复至正常水平。MTX能抑制LPS诱导的CIA大鼠关节FLS分泌IL-1β和TNF-α。Western Blot检测显示,不同浓度的MTX对CIA大鼠FLS中ERK蛋白的表达与模型对照组相比无统计学差异(P>0.05)。CIA大鼠 FLS中p-ERK蛋白的表达明显高于空白对照组(P<0.01),MTX干预后,低剂量组对p-ERK蛋白表达无明显影响,而中、高剂量组可降低p-ERK蛋白的表达(P<0.05)。结论: MTX既有免疫抑制作用,同时还通过抑制炎性细胞因子IL-1β和TNF-α而具有抗炎作用,其机制可能部分与其抑制MAPK信号通路中的ERK蛋白磷酸化有关。

关键词: 类风湿关节炎, 甲氨蝶呤, IL-1β, TNF-α, ERK/p-ERK

Abstract: AIM: To observe the role and its possible mechanism of inflammatory cytokines of IL-1 β and TNF α which plays an important role in the etiology of MTX on RA. METHODS: The establishment of animal model of rheumatoid arthritis with collagen type II, with MTX (0.2 mg/kg/W) treatment, the rats were killed after six weeks, serum for detection of IL-1β and TNF-α; separation of knee joint synovial cells in rats (FLS) were cultured, identification,detection of LPS induced FLS in the culture supernatant of IL-1β and TNF -α, and observes the effect of MTX on the secretion of IL-1β and TNF-α of FLS; adding different doses of MTX on the FLS culture, expression by Western Blot method for detection of FLS ERK protein and phosphorylation of protein. RESULTS: Arthritis index score and joint swelling and joint pathology showed determination modeling success, isolated synovial cells by flow cytometry FLS expression of VCAM-1 was 85.5%. After modeling, CIA rat serum IL-1 β and TNF- α increased obviously, there was significant difference compared with the blank control group (P<0.05), MTX could effectively reduce the content of serum CIA in rats of IL-1 β and TNF -α (P<0.05), but failed to recover to normal.In rats with CIA FLS MTX could inhibit LPS induced secretion of IL-1 β and TNF-α. Displayed in Western Blot detection, different concentrations of MTX on ERK protein expression of FLS in CIA rats compared with the model group showed no significant difference (P>0.05). Expression of p-ERK CIA in FLS of rats was significantly higher than that in control group (P<0.01), after the treatment of MTX, low dose group had no effect on the expression of p-ERK protein, while in the medial and high dose group could reduce the p-ERK protein (P<0.05). CONCLUSION: MTX has immunosuppressive effects at the same time through the inhibition of inflammatory cytokines IL-1 β and TNF -α, it also has anti-inflammatory effect. The mechanism may be partly related to inhibition of MAPK signaling pathway in ERK phosphorylation relate.

Key words: rheumatoid arthritis, methotrexate, IL-1β, TNF-α, ERK/p-ERK

中图分类号:

R965.2

蔡质其, 张育, 张学增, 沈维干. 甲氨蝶呤对炎性细胞因子及丝裂原活化蛋白激酶信号通路的影响[J]. 中国临床药理学与治疗学, 2014, 19(12): 1347-1351.

CAI Zhi-qi, ZHANG Yu, ZHANG Xue-zeng, SHEN Wei-gan. Study on the effect of methotrexate on inflammatory cytokines and MAPK signal pathway[J]. Chinese Journal of Clinical Pharmacology and Therapeutics, 2014, 19(12): 1347-1351.

[1]	范杰, 金永明, 江孝龙, 蒋国华. lncRNA HOTAIR调控miR-206影响类风湿关节炎滑膜细胞增殖和凋亡的分子机制研究[J]. 中国临床药理学与治疗学, 2023, 28(7): 736-742.
[2]	王鑫昱, 陈亦鹏, 马永力, 何君妃. “杜仲-当归”调控NLPR3炎症小体对骨关节炎大鼠改善作用的研究[J]. 中国临床药理学与治疗学, 2023, 28(7): 751-757.
[3]	潘淑, 吴义金, 张洒洒, 王启海, 罗婷婷, 殷勤. 基于OAT3介导的MTX治疗佐剂诱导性关节炎大鼠的药动学研究[J]. 中国临床药理学与治疗学, 2022, 27(5): 516-525.
[4]	陈宇罡, 屠艳琼, 王聪庆, 陈娟, 张博宏. 应用网络药理学和分子对接技术探讨金虎通胆汤治疗胆石症的作用机制及实验验证[J]. 中国临床药理学与治疗学, 2022, 27(10): 1090-1098.
[5]	杜娟，徐玲燕，吴伟，沈青青，江振洲，张陆勇，黄鑫. 类风湿关节炎下转运体表达及活性的变化[J]. 中国临床药理学与治疗学, 2018, 23(2): 223-229.
[6]	陈兰芳，强孚勇，徐亮. 托珠单抗联合改善病情抗风湿药治疗中重度类风湿关节炎短期临床观察[J]. 中国临床药理学与治疗学, 2018, 23(12): 1386-1391.
[7]	伍嘉琪，杨俏雯，陈秀敏，黄闰月，黄清春，赵越，吴晓东. 艾拉莫德联合甲氨蝶呤治疗类风湿关节炎有效性和安全性的系统评价和Meta分析[J]. 中国临床药理学与治疗学, 2018, 23(10): 1132-1140.
[8]	杨红，林杉，杨宏，谢晓薇，王新. 艾拉莫德对类风湿关节炎患者血管内皮生长因子及色素上皮衍生因子表达的影响[J]. 中国临床药理学与治疗学, 2017, 22(1): 68-71.
[9]	陶绍能，王莹莹，戴云海，程光华，吕坤. 体外沉默血管内皮生长因子对D-(-)-MTX/A549耐药细胞株迁移侵袭能力的影响[J]. 中国临床药理学与治疗学, 2016, 21(7): 745-748.
[10]	李小心, 王田玲, 余菲, 邵体红, 陈兰芳, 毛桐俊, 陆进明, 徐亮. 类风湿关节炎缓解后维持治疗方案的研究[J]. 中国临床药理学与治疗学, 2015, 20(7): 797-802.
[11]	刘东阳, 苏金梅, 宋瀚麟, 陈嘉, 刘洋, 幺雪婷, 宋玲, 江骥, 胡蓓. 甲氨蝶呤及其活性代谢产物在类风湿关节炎患者体内的药动学模型研究[J]. 中国临床药理学与治疗学, 2015, 20(5): 571-577.
[12]	姚锦英. 积雪草总苷抗A549肺癌细胞诱导的肿瘤的实验研究[J]. 中国临床药理学与治疗学, 2015, 20(4): 384-387.
[13]	陈菲菲, 钱家树, 王良荣, 金立达, 熊响清. 参麦注射液对单肺通气患者肺损伤的保护作用[J]. 中国临床药理学与治疗学, 2015, 20(2): 199-202.
[14]	曾凡湘, 史道华, 宋一一, 孙蓬明, 郑翠仙, 邓婕, 王长连. 甲氨蝶呤治疗异位妊娠疗效预测因素分析[J]. 中国临床药理学与治疗学, 2015, 20(10): 1156-1160.
[15]	张思, 顾兵, 李华南, 王烁宇, 张水印. 甲氨蝶呤对大鼠脊髓挫伤急性期氧化损伤相关蛋白的影响[J]. 中国临床药理学与治疗学, 2015, 20(1): 1-6.