新型机械敏感离子通道Piezo1和TRPV4蛋白siRNA双沉默模式对骨关节炎大鼠模型的修复作用研究

doi:10.13418/j.issn.1001-165x.2021.06.014

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中国临床解剖学杂志 ›› 2021, Vol. 39 ›› Issue (6) : 692-699. DOI: 10.13418/j.issn.1001-165x.2021.06.014

实验研究

新型机械敏感离子通道Piezo1和TRPV4蛋白siRNA双沉默模式对骨关节炎大鼠模型的修复作用研究

周勇伟¹，章才华¹，李晓飞²，蔡迅梓¹

作者信息 +

The repair effect of novel mechanical sensing ion channels Piezo1 and TRPV4 protein siRNA double silencing on osteoarthritis rat model

Zhou Yongwei¹, Zhang Caihua¹, Li Xiaofei², Cai Xunzi¹

Author information +

文章历史 +

摘要

目的　探究新型机械敏感离子通道Piezo1和TRPV4蛋白siRNA双沉默模式对骨关节炎大鼠模型的修复作用。方法　构建SD大鼠的OA动物模型，根据处理方案的不同，将SD大鼠分成4组，即空白对照组，siRNA-Piezo1组，siRNA-TRPV4组和双基因沉默组。通过HE染色和番红-固绿染色等方法，利用改良Minkin’s评分和OARSI评分进行评估。利用免疫组织化学染色和RT-PCR检测膝关节软骨组织中Aggrecan蛋白和Collagen II的表达。结果　HE染色和番红-固绿染色等结果显示，siRNA-Piezo1组和siRNA-TRPV4组膝关节软骨组织的评分要明显优于模型组，siRNA-Piezo1组和siRNA-TRPV4组（P<0.05）。免疫组织化学染色和RT-PCR结果显示，双基因沉默组中Aggrecan和Collagen II蛋白的表达量明显高于siRNA-Piezo1组和siRNA-TRPV4组（P<0.05）。结论　Piezo1和TRPV4蛋白siRNA双沉默模式可以促进Aggrecan和Collagen II的表达，进而对骨关节炎大鼠模型起到一定的修复作用。

Abstract

Objective To explore the repair effect of siRNA double silencing of new mechanosensitive ion channels Piezo1 and TRPV4 on osteoarthritis rat model. Methods The osteoarthritis animal (OA) model of SD rats was established. According to the different treatment on rats, SD rats were divided into the following four groups: a blank control group, a siRNA-Piezo1 group, a siRNA-TRPV4 group and a double gene silencing group. HE staining and safranin solid green staining were used to evaluate the quality of the samples by modified Minkin's score and OARSI score. The expression of Aggrecan protein and Collagen II in articular cartilage were detected by immunohistochemical staining and RT-PCR. Results The result of the HE staining and safranin green staining showed that the scores of knee cartilage tissue in the siRNA-Piezo1 group and siRNA-TRPV4 group were significantly higher than that in the model group, the siRNA-piezo1 group and the siRNA-TRPV4 group (P<0.05). The results of immunohistochemical staining and RT-PCR showed that the expression of Aggrecan and Collagen II in the double gene silencing group was significantly higher than that in siRNA-piezo1 group and siRNA-TRPV4 group (P<0.05). Conclusions Piezo1 and TRPV4 protein siRNA double silencing mode can promote the expression of Aggrecan and Collagen II, and then play a certain role in the repair of osteoarthritis rat model.

导出引用

周勇伟, 章才华, 李晓飞, 蔡迅梓. 新型机械敏感离子通道Piezo1和TRPV4蛋白siRNA双沉默模式对骨关节炎大鼠模型的修复作用研究[J]. 中国临床解剖学杂志. 2021, 39(6): 692-699 https://doi.org/10.13418/j.issn.1001-165x.2021.06.014

Zhou Yongwei, Zhang Caihua, Li Xiaofei, Cai Xunzi. The repair effect of novel mechanical sensing ion channels Piezo1 and TRPV4 protein siRNA double silencing on osteoarthritis rat model[J]. Chinese Journal of Clinical Anatomy. 2021, 39(6): 692-699 https://doi.org/10.13418/j.issn.1001-165x.2021.06.014

中图分类号： R684.3

参考文献

[1] Bishnoi M, Jain A, Hurkat P, et al. Chondroitin sulphate: a focus on osteoarthritis[J]. Glycoconj J, 2016, 33(5):693-705. DOI:10.1007/s10719-016-9665-3.
[2] Mandl LA. Osteoarthritis year in review 2018: clinical[J]. Osteoarthritis Cartilage, 2019, 27(3):359-364. DOI:10.1016/j.joca.2018.11.001.
[3] Dobson GP, Letson HL, Grant A, et al. Defining the osteoarthritis patient: back to the future[J]. Osteoarthritis Cartilage, 2018, 26(8):1003-1007. DOI:10.1016/j.joca.2018.04.018.
[4] Rosenberg JH, Rai V, Dilisio MF, et al. Damage-associated molecular patterns in the pathogenesis of osteoarthritis: potentially novel therapeutic targets[J]. Mol Cell Biochem, 2017, 434(1-2):171-179. DOI:10.1007/s11010-017-3047-4.
[5] Schmidt TW. Approach to osteoarthritis management for the primary care provider[J]. Prim Care, 2018, 45(2):361-378. DOI:10.1016/j.pop.2018.02.009.
[6] Lawrence KM, Jones RC, Jackson TR, et al. Chondroprotection by urocortin involves blockade of the mechanosensitive ion channel Piezo1[J]. Sci Rep, 2017, 7(1):5147-5157. DOI: 10.1038/s41598-017-04367-4.
[7] Lee W, Leddy H A, Chen Y，et al. Synergy between Piezo1 and Piezo2 channels confers high-strain mechanosensitivity to articular cartilage[J]. Proc Natl Acad Sci Am, 2014, 111(47)：5114-5122. DOI: 10.1073/pnas.1414298111.
[8] Li XF, Leng P , Zhang Z, et al. The Piezo1 protein ion channel functions in human nucleus pulposus cell apoptosis by regulating mitochondrial dysfunction and the endoplasmic reticulum stress signal pathway[J]. Exp Cell Res, 2017, 358(2)：377-389.DOI： 10.1016/j.yexcr.2017.07.010.
[9] Xing R, Wang P, Zhao L, et al. Mechanism of TRPA1 and TRPV4 Participating in mechanical hyperalgesia of rat experimental knee osteoarthritis[J]. Arch Rheumatol, 2017, 32(2):96-104. DOI:10.5606/ArchRheumatol.2017.6061.
[10]李晓飞, 张钊, 李晓东,等. 新型机械激活离子通道蛋白Piezo1通过MAPK/ERK1/2信号通路介导软骨细胞凋亡的机制[J]. 中华医学杂志, 2016,12(96):2477-2484. DOI: 10.3760/cma.j.issn.0376-2491.2016. 31.007.
[11] Jiang L, Zhao YD, Chen WX. The function of the novel mechanical activated ion channel Piezo1 in the human osteosarcoma cells[J]. Med Sci Monit, 2017, 23:5070-5082. DOI: 10.12659/msm.906959.
[12] 李晓飞, 张钊, 王天宝, 等. Piezo1蛋白经MAPK/ERK5信号通路介导软骨细胞凋亡的机制研究[J]. 中华骨科杂志, 2016, 36(12):795-803.
[13] Atobe M, Nagami T, Muramatsu S, et al. Discovery of novel transient receptor potential vanilloid 4 (TRPV4) agonists as regulators of chondrogenic differentiation: identification of quinazolin-4(3 H)-ones and in vivo studies on a surgically induced rat model of osteoarthritis[J]. J Med Chem, 2019,62(3):1468-1483. DOI:10.1021/acs.jmedchem. 8b01615.
[14] Walter BA, Purmessur D, Moon A, et al. Reduced tissue osmolarity increases TRPV4 expression and pro-inflammatory cytokines in intervertebral disc cells[J]. Eur Cell Mater, 2016, 32:123-136. DOI:10.22203/ecm.v032a08.
[15] Kumai T, Yui N, Yatabe K, et al. A novel, self-assembled artificial cartilage-hydroxyapatite conjugate for combined articular cartilage and subchondral bone repair: histopathological analysis of cartilage tissue engineering in rat knee joints[J]. Int J Nanomedicine, 2019, 14:1283-1298. DOI:10.2147/IJN.S193963.