hsa-Let-7a-5p靶向结合TGF-βR1抑制局限性硬皮病成纤维细胞纤维化

doi:10.16352/j.issn.1001-6325.2022.12.1841

基础医学与临床 ›› 2022, Vol. 42 ›› Issue (12): 1841-1849.doi: 10.16352/j.issn.1001-6325.2022.12.1841

hsa-Let-7a-5p靶向结合TGF-βR1抑制局限性硬皮病成纤维细胞纤维化

王立铨, 黄久佐, 俞楠泽, 马旭达, 李天浩, 龙笑^*

中国医学科学院北京协和医学院北京协和医院整形外科,北京 100032

收稿日期:2022-09-26 修回日期:2022-10-21 出版日期:2022-12-05 发布日期:2022-11-23
通讯作者: ^* pumclongxiao@126.com
基金资助:
中国医学科学院医学与健康科技创新工程“十四五”项目中国罕见病的精准诊疗研究(2021-I2M-1-003);科技部战略性国际科技创新合作重点专项(2020YFE0201600);北京协和医院中央高水平医院临床科研专项青年培优计划(2022-PUMCH-A-210)

hsa-Let-7a-5p inhibits fibrosis of fibroblasts from localized scleroderma through targeted binding of TGF-βR1

WANG Li-quan, HUANG Jiu-zuo, YU Nan-ze, MA Xu-da, LI Tian-hao, LONG Xiao^*

Department of Plastic Surgery, Peking Union Medical College Hospital, CAMS & PUMC, Beijing 100032, China

Received:2022-09-26 Revised:2022-10-21 Online:2022-12-05 Published:2022-11-23
Contact: ^* pumclongxiao@126.com

摘要/Abstract

摘要： 目的探讨hsa-Let-7a-5p对局限性硬皮病成纤维细胞(LSFs)纤维化过程的影响及其与转化生长因子-β受体(TGF-βR)和TGF-β/Smad信号通路的关系。方法将携带hsa-Let-7a-5p和shTGF-βR1的慢病毒载体及空白慢病毒载体感染LSFs。CCK-8法和划痕实验检测各组LSFs的增殖和迁移情况。RT-qPCR、免疫荧光和Western blot检测转染的效率以及各组LSFs的Ⅰ、Ⅲ型胶原、α-SMA、TGF-βR1、TGF-β、p-Smad2/3的mRNA和蛋白表达水平。结果病毒感染LSFs后,shTGF-βR1组的TGF-βR1表达量明显低于对照组(P＜0.05);shTGF-βR1组的细胞增殖率、迁移能力均有所降低;shTGF-βR1组的Ⅰ、Ⅲ型胶原、α-SMA、TGF-β、p-Smad2/3的mRNA和蛋白表达水平均明显低于对照组(P＜0.05)。生物信息学分析表明hsa-Let-7a-5p可以与TGF-βR1 3′-UTR的靶区位置配对,hsa-Let-7a-5p慢病毒转染组的hsa-Let-7a-5p表达量明显高于对照组(P＜0.05),TGF-βR1表达量明显降低(P＜0.05)。hsa-Let-7a-5p组的细胞增殖率、迁移能力均有所降低;hsa-Let-7a-5p组的Ⅰ、Ⅲ型胶原、α-SMA、TGF-β、p-Smad2/3的mRNA和蛋白表达水平均明显低于对照组(P＜0.05)。结论 hsa-Let-7a-5p在体外通过靶向结合TGF-βR1调控TGF-β/Smad通路,抑制LSFs增殖与迁移,降低LSFs的纤维化标志物,从而抑制局限性硬皮病的纤维化。

关键词: hsa-Let-7a-5p, 局限性硬皮病, 纤维化

Abstract: Objective To investigate the effect of hsa-Let-7a-5p on the fibrosis process of localized scleroderma fibroblasts (LSFs) and its correlation with transforming growth factor-β receptor (TGF-βR) and TGF-β/Smad signaling pathway. Methods LSFs were infected with lentiviral vectors carrying hsa-Let-7a-5p and shTGF-βR1 and blank lentiviral vectors. The proliferation and migration of LSFs in each group were detected by CCK-8 method and scratch test. Real-time quantitative PCR (RT-qPCR), immunofluorescence and Western blot were used to detect the transfection efficiency, and the mRNA and protein levels of collagen type Ⅰ & Ⅲ, α-SMA, TGF-βR1, TGF-β, p-Smad2/3 in LSFs in each group. Results After virus infection of LSFs, the expression of TGF-βR1 in the shTGF-βR1 group was significantly lower than that in the control groups (P＜0.05); the cell proliferation rate and migration of shTGF-βR1 group were inhibited(P＜0.05); The mRNA and protein expression of collagen type Ⅰ & Ⅲ, α-SMA, TGF-β, p-Smad2/3 were significantly lower than those of the control groups(P＜0.05). Bioinformatics analysis showed that hsa-Let-7a-5p can be matched with the target region of TGF-βR1 3′-UTR, and the expression of hsa-Let-7a-5p in the lentivirus transfection group was significantly increased compared with the control groups(P＜0.05), the expression of TGF-βR1 was significantly decreased(P＜0.05). The cell proliferation rate and migration ability of the hsa-Let-7a-5p group were decreased; the mRNA and protein expression of collagen type Ⅰ & Ⅲ, α-SMA, TGF-β, p-Smad2/3 in the hsa-Let-7a-5p group were significantly lower than those of the control groups(P＜0.05). Conclusions hsa-Let-7a-5p regulates TGF-β/Smad pathway by targeting at TGF-βR1,inhibits proliferation and migration of LSFs, decreases fibrotic markers in LSFs thereby inhibiting fibrosis in localized scleroderma.

Key words: hsa-Let-7a-5p, localized scleroderma, fibrosis

中图分类号:

王立铨, 黄久佐, 俞楠泽, 马旭达, 李天浩, 龙笑. hsa-Let-7a-5p靶向结合TGF-βR1抑制局限性硬皮病成纤维细胞纤维化[J]. 基础医学与临床, 2022, 42(12): 1841-1849.

WANG Li-quan, HUANG Jiu-zuo, YU Nan-ze, MA Xu-da, LI Tian-hao, LONG Xiao. hsa-Let-7a-5p inhibits fibrosis of fibroblasts from localized scleroderma through targeted binding of TGF-βR1[J]. Basic & Clinical Medicine, 2022, 42(12): 1841-1849.

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参考文献

[1]Careta MF, Romiti R. Localized scleroderma: clinical spectrum and therapeutic update[J].An Bras Dermatol,2015, 90: 62-73.doi: 10.1590/abd1806-4841.20152890.
[2]Peng L, Chen Y, Shi S, et al. Stem cell-derived and circulating exosomal microRNAs as new potential tools for diabetic nephropathy management[J].Stem Cell Res Ther,2022, 13: 25.doi: 10.1186/s13287-021-02696-w.
[3]Jinnin M. Various applications of microRNAs in skin diseases[J].J Dermatol Sci,2014, 74: 3-8.doi: 10.1016/j.jdermsci.2014.01.004.
[4]Kim KK, Sheppard D, Chapman HA. TGF-β1 signaling and tissue fibrosis[J].Cold Spring Harb Perspect Biol,2018, 10.doi: 10.1101/cshperspect.a022293.
[5]Luo Z, Sun Y, Qi B, et al. Human bone marrow mesenchymal stem cell-derived extracellular vesicles inhibit shoulder stiffness via let-7a/Tgfbr1 axis[J].Bioact Mater,2022, 17: 344-359.doi: 10.1016/j.bioactmat.2022.01.016.
[6]Vuorio TK, Kähäri VM, Lehtonen A, et al. Fibroblast activation in scleroderma[J].Scand J Rheumatol,1984, 13: 229-37.doi: 10.3109/03009748409100391.
[7]Chu H, Wu T, Wu W, et al. Involvement of collagen-binding heat shock protein 47 in scleroderma-associated fibrosis[J].Protein Cell,2015, 6: 589-598.doi: 10.1007/s13238-015-0171-3.
[8]Meng XM, Nikolic-Paterson DJ, Lan HY. TGF-β: the master regulator of fibrosis[J].Nat Rev Nephrol,2016, 12: 325-38.doi: 10.1038/nrneph.2016.48.
[9]Li T, Yan Y, Wang B, et al. Exosomes derived from human umbilical cord mesenchymal stem cells alleviate liver fibrosis[J].Stem Cells Dev,2013, 22: 845-54.doi: 10.1089/scd.2012.0395.
[10]Hu J, Chen Y, Huang Y, et al. Human umbilical cord mesenchymal stem cell-derived exosomes suppress dermal fibroblasts-myofibroblats transition via inhibiting the TGF-β1/Smad 2/3 signaling pathway[J].Exp Mol Pathol,2020, 115: 104468.doi: 10.1016/j.yexmp.2020.104468.
[11]Lu J, Liu Q, Wang L, et al. Increased expression of latent TGF-β-binding protein 4 affects the fibrotic process in scleroderma by TGF-β/SMAD signaling[J].Lab Invest,2017, 97: 591-601.doi: 10.1038/labinvest.2017.20.
[12]Makino K, Jinnin M, Hirano A, et al. The downregula-tion of microRNA let-7a contributes to the excessive expression of type I collagen in systemic and localized scleroderma[J].J Immunol,2013, 190: 3905-3915.doi: 10.4049/jimmunol.1200822.

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hsa-Let-7a-5p靶向结合TGF-βR1抑制局限性硬皮病成纤维细胞纤维化

hsa-Let-7a-5p inhibits fibrosis of fibroblasts from localized scleroderma through targeted binding of TGF-βR1

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