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中华临床医师杂志(电子版) ›› 2021, Vol. 15 ›› Issue (03) : 194 -199. doi: 10.3877/cma.j.issn.1674-0785.2021.03.008

临床研究

外周血外泌体及RhoA/ROCK信号通路与系统性红斑狼疮血小板减少患者出血症状的相关性
冯鸿雁1, 孙雪霞1, 赵磊1, 王丽芹1, 张媛媛1, 董静1,()   
  1. 1. 266003 山东青岛,青岛大学附属医院风湿免疫科
  • 收稿日期:2020-10-10 出版日期:2021-03-15
  • 通信作者: 董静
  • 基金资助:
    青岛大学附属医院临床医学+X科研项目资助(QDFY+X2021041)

Relationship of peripheral blood exosomes and RhoA/ROCK signaling pathway with bleeding symptoms in systemic lupus erythematosus patients with immune thrombocytopenia

Hongyan Feng1, Xuexia Sun1, Lei Zhao1, Liqin Wang1, Yuanyuan Zhang1, Jing Dong1,()   

  1. 1. Department of Rheumatology and Clinical Immunology, the Affiliated Hospital of Qingdao University, Qingdao 266003, China
  • Received:2020-10-10 Published:2021-03-15
  • Corresponding author: Jing Dong
引用本文:

冯鸿雁, 孙雪霞, 赵磊, 王丽芹, 张媛媛, 董静. 外周血外泌体及RhoA/ROCK信号通路与系统性红斑狼疮血小板减少患者出血症状的相关性[J]. 中华临床医师杂志(电子版), 2021, 15(03): 194-199.

Hongyan Feng, Xuexia Sun, Lei Zhao, Liqin Wang, Yuanyuan Zhang, Jing Dong. Relationship of peripheral blood exosomes and RhoA/ROCK signaling pathway with bleeding symptoms in systemic lupus erythematosus patients with immune thrombocytopenia[J]. Chinese Journal of Clinicians(Electronic Edition), 2021, 15(03): 194-199.

目的

研究系统性红斑狼疮(SLE)免疫性血小板减少患者有出血症状组及无出血症状组外周血外泌体的表达,以及外周血单个核细胞(MNC)RhoA/ROCK信号通路的变化,探讨影响出血症状的相关机制。

方法

收集2018年1月至2020年1月在青岛大学附属医院风湿免疫科诊断为SLE并免疫性血小板减少22例患者的资料,按照有无出血症状分为2组,收集患者外周血,分离血浆外泌体,通过透射电镜观测外泌体,蛋白免疫印迹法检测外泌体标志物CD63、CD81表达。Pull-down法检测外周血MNC的RhoA/ROCK信号通路的活化,采用RT-PCR检测MNC的RhoA/ROCK信号通路mRNA的表达,并分析mRNA的表达与临床指标的相关性。

结果

与无出血症状组比较,SLE免疫性血小板减少患者有出血症状组血浆外泌体减少;与无出血症状组比较,有出血症状组RhoA/ROCK信号通路活化及mRNA表达升高,2组差异有统计学意义(P均<0.05)。SLE免疫性血小板减少患者RhoA/ROCK信号通路mRNA的表达与血小板计数呈负相关(RhoA:r=-0.333,P=0.032;ROCK:r=-0.475,P=0.046),与SLEDAI评分呈正相关(RhoA:r=0.435,P=0.042;ROCK:r=0.462,P=0.047)。

结论

血浆外泌体及外周血MNC的RhoA/ROCK信号通路可能参与了SLE免疫性血小板减少患者的出血症状调控。

Objective

To investigate the expression of plasma exosomes and the activation of the RhoA/ROCK signaling pathway in peripheral blood of systemic lupus erythematosus (SLE) patients with immune thrombocytopenia with or without bleeding symptoms, and to explore the related mechanism of bleeding symptoms.

Methods

The data of 22 SLE patients with immune thrombocytopenia diagnosed at the Department of Rheumatology and Clinical Immunology, the Affiliated Hospital of Qingdao University from January 2018 to January 2020 were collected. The patients were divided into two groups according to whether bleeding symptoms were present. Peripheral blood was collected and plasma exosomes were separated. The exosomes were observed by transmission electron microscopy, and the exosomes markers CD63 and CD81 were detected by Western blot. Pull-down method was used to detect the activation of RhoA/ROCK signaling pathway in mononuclear cells (MNC), and RT-PCR was used to detect the mRNA expression of RhoA/ROCK. The correlation between RhoA/ROCK mRNA expression and clinical indicators was analyzed.

Results

The number of plasma exosomes was decreased in patients with bleeding symptoms compared to that of the patients without bleeding symptoms. The activation of RhoA/ROCK signal pathway as well as the mRNA expression of RhoA/ROCK was significantly elevated in patients with bleeding symptoms compared to those without (P<0.05). The expression of RhoA/ROCK mRNA was negatively correlated with platelet count (RhoA: r=-0.333, P=0.032; ROCK: r=-0.475, P=0.046) and positively correlated with SLEDAI score (RhoA: r=0.435, P=0.042; ROCK: r=0.462, P=0.047).

Conclusion

Plasma exosomes and the RhoA/ROCK signaling pathway in peripheral blood MNC may participate in the regulation of bleeding symptoms in SLE patients with immune thrombocytopenia.

图1 外泌体的形态及鉴定。图a为透射电镜下无出血症状组外泌体形态;图b为透射电镜下有出血症状组外泌体形态;图c为WB法检测外泌体标志物CD63、CD81表达情况
图2 Pull-down法检测RhoA及ROCK总蛋白及磷酸化蛋白在系统性红斑狼疮免疫性血小板减少患者外周血单个核细胞的表达情况
图3 实时荧光定量PCR法检测RhoA及ROCK mRNA在系统性红斑狼疮免疫性血小板减少患者外周血单个核细胞的表达情况
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