低分子量透明质酸通过CD44调控S100A4核转移促进心肌纤维化
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1.南京医科大学第一附属医院老年医学科;2.江苏省中医院

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国家自然科学基金项目(面上项目,重点项目,重大项目)


Low molecular weight hyaluronic acid regulates the nuclear translocation of S100A4 to promote myocardial fibrosis via CD44
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Department of Geriatrics, the First Affiliated Hospital with Nanjing Medical University

Fund Project:

The National Natural Science Foundation of China (General Program, Key Program, Major Research Plan)

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    摘要:

    目的:研究低分子量透明质酸(Low molecular weight hyaluronic acid, LMW-HA)对小鼠心脏成纤维细胞表型转化的促进作用,以及探讨CD44和S100A4在此过程中的作用。方法:从ICR乳小鼠的心脏中分离提取心脏成纤维细胞(Cardiac fibroblasts, CFs)并培养, 使用LMW-HA进行刺激,采用CCK-8和EdU染色测细胞增殖程度,免疫印迹(Western Blotting,WB)、实时定量PCR、免疫荧光确定纤维化程度,对CD44和S100A4蛋白质核分离并通过WB和免疫荧光明确核转移。CD44抑制剂BRIC-235处理后再次检测上述指标。结果:CCK-8增殖与EdU染色实验确定了LMW-HA的最佳刺激浓度为0.8mg/ml。WB、PCR和IF表明LMW-HA的刺激使心脏纤维化标志物(α-SMA和Collagen 3)的水平显著增加,CD44保持不变,S100A4随时间逐渐增加,同时CD44和S100A4蛋白转移到细胞核内。CD44抑制剂BRIC-235能抑制这些改变。结论:LMW-HA的刺激能够促进小鼠心脏成纤维细胞的表型转化从而促进心肌纤维化,该过程是通过与CD44结合并促进S100A4转移入细胞核内激活下游通路实现的,因此可能为心肌纤维化的治疗筛选新靶点。

    Abstract:

    Objective: This study aimed to investigate the promotion effect of low molecular weight hyaluronic acid (LMW-HA) on phenotypic transformation of mice cardiac fibroblasts and the role of CD44 and S100A4 in this process. Methods: Cardiac fibroblasts (CFs) were isolated and cultured from neonatal ICR mice and then stimulated by LMW-HA. The proliferation of CFs was measured by CCK-8 and EdU, and we used Western Blotting, quantitative RT-PCR and Immunofluorescence to determine myocardial fibrosis. We extracted nuclear and cytoplasmic protein of CD44 and S100A4 to specified the nuclear translocation by WB and Immunofluorescence. To measure aforesaid indicators once again after treatment of the CD44 inhibitor BRIC-235. Results: CCK-8 and EdU determined that the optimal concentration of LMW-HA stimulation was 0.8mg/ml. WB, qRT-PCR and IF showed that LMW-HA stimulation significantly increased the level of myocardial fibrosis markers (α-SMA and Collagen 3), but CD44 remained unchanged, and S100A4 gradually increased over time. The protein of CD44 and S100A4 were transferred into the nucleus at the same time. BRIC-235 can inhibit these changes. Conclusions: LMW-HA can promote the differentiation of mice cardiac fibroblasts, which is regulated by the nuclear translocation of CD44 and S100A4 and the activation of downstream signaling pathway. This may be a new therapeutic target of myocardial fibrosis in the future.

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  • 收稿日期:2020-12-13
  • 最后修改日期:2021-04-04
  • 录用日期:2021-06-21
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