Mechanical stimulation promotes fibrochondrocyte proliferation by activating the TRPV4 signaling pathway during tendon–bone insertion healing: CCN2 plays an important regulatory role

IF 6.3 1区 医学 Q1 DERMATOLOGY Burns & Trauma Pub Date : 2024-10-20 DOI:10.1093/burnst/tkae028
Xuting Bian, Xiao Liu, Mei Zhou, Hong Tang, Rui Wang, Lin Ma, Gang He, Shibo Xu, Yunjiao Wang, Jindong Tan, Kanglai Tang, Lin Guo
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Abstract

Background We previously confirmed that mechanical stimulation is an important factor in the repair of tendon–bone insertion (TBI) injuries and that mechanoreceptors such as transient receptor potential ion-channel subfamily V member 4 (TRPV4; also known as transient receptor potential vanilloid 4) are key to transforming mechanical stimulation into intracellular biochemical signals. This study aims to elucidate the mechanism of mechanical stimulation regulating TRPV4. Methods Immunohistochemical staining and western blotting were used to evaluate cartilage repair at the TBI after injury. The RNA expression and protein expression of mechanoreceptors and key pathway molecules regulating cartilage proliferation were analyzed. TBI samples were collected for transcriptome sequencing to detect gene expression. Calcium-ion imaging and flow cytometry were used to evaluate the function of TPRV4 and cellular communication network factor 2 (CCN2) after the administration of siRNA, recombinant adenovirus and agonists. Results We found that treadmill training improved the quality of TBI healing and enhanced fibrochondrocyte proliferation. The transcriptome sequencing results suggested that the elevated expression of the mechanistically stimulated regulator CCN2 and the exogenous administration of recombinant human CCN2 significantly promoted TRPV4 protein expression and fibrochondrocyte proliferation. In vitro, under mechanical stimulation conditions, small interfering RNA (siRNA)-CCN2 not only inhibited the proliferation of primary fibrochondrocytes but also suppressed TRPV4 protein expression and activity. Subsequently, primary fibrochondrocytes were treated with the TRPV4 agonist GSK1016790A and the recombinant adenovirus TRPV4 (Ad-TRPV4), and GSK1016790A partially reversed the inhibitory effect of siRNA-CCN2. The phosphoinositide 3-kinase/ protein kinase B (PI3K/AKT) signaling pathway participated in the above process. Conclusions Mechanical stimulation promoted fibrochondrocyte proliferation and TBI healing by activating TRPV4 channels and the PI3K/AKT signaling pathway, and CCN2 may be a key regulatory protein in maintaining TRPV4 activation.
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在肌腱骨插入愈合过程中,机械刺激通过激活 TRPV4 信号通路促进纤维软骨细胞增殖:CCN2 发挥着重要的调节作用
背景 我们之前证实,机械刺激是肌腱-骨插入(TBI)损伤修复的一个重要因素,而瞬时受体电位离子通道 V 亚家族成员 4(TRPV4;又称瞬时受体电位香草素 4)等机械感受器是将机械刺激转化为细胞内生化信号的关键。本研究旨在阐明机械刺激调控 TRPV4 的机制。方法 采用免疫组化染色法和 Western 印迹法评估损伤后 TBI 处的软骨修复情况。分析机械感受器和调控软骨增殖的关键通路分子的 RNA 表达和蛋白表达。收集 TBI 样本进行转录组测序,以检测基因表达。使用钙离子成像和流式细胞术评估了服用 siRNA、重组腺病毒和激动剂后 TPRV4 和细胞通讯网络因子 2(CCN2)的功能。结果 我们发现跑步机训练提高了创伤性脑损伤的愈合质量,并增强了纤维软骨细胞的增殖。转录组测序结果表明,机理刺激调节因子 CCN2 表达升高,外源性给予重组人 CCN2 能显著促进 TRPV4 蛋白表达和纤维软骨细胞增殖。在体外,在机械刺激条件下,小干扰 RNA(siRNA)-CCN2 不仅能抑制原代纤维软骨细胞的增殖,还能抑制 TRPV4 蛋白的表达和活性。随后,用 TRPV4 激动剂 GSK1016790A 和重组腺病毒 TRPV4(Ad-TRPV4)处理原代纤维软骨细胞,GSK1016790A 可部分逆转 siRNA-CCN2 的抑制作用。磷酸肌酸 3- 激酶/蛋白激酶 B(PI3K/AKT)信号通路参与了上述过程。结论 机械刺激通过激活TRPV4通道和PI3K/AKT信号通路促进纤维软骨细胞增殖和创伤性脑损伤愈合,而CCN2可能是维持TRPV4激活的关键调控蛋白。
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来源期刊
Burns & Trauma
Burns & Trauma 医学-皮肤病学
CiteScore
8.40
自引率
9.40%
发文量
186
审稿时长
6 weeks
期刊介绍: The first open access journal in the field of burns and trauma injury in the Asia-Pacific region, Burns & Trauma publishes the latest developments in basic, clinical and translational research in the field. With a special focus on prevention, clinical treatment and basic research, the journal welcomes submissions in various aspects of biomaterials, tissue engineering, stem cells, critical care, immunobiology, skin transplantation, and the prevention and regeneration of burns and trauma injuries. With an expert Editorial Board and a team of dedicated scientific editors, the journal enjoys a large readership and is supported by Southwest Hospital, which covers authors'' article processing charges.
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