生物力学通过HDAC5介导的微管骨架去乙酰化调控病理性瘢痕形成的机制研究

The local disturbed mechanical environment is an important factor in hypertrophic scar formation. Mechanical stimulation promotes the activation of scar fibroblasts. Microtubule deacetylation of the cell skeleton is closely related to fibroblasts activation. It has been reported that HDAC5 is capable of the deacetylation of α-tubulin through its nuclear export thereby promoting cell repairing. Studies have also shown that biomechanics can promote the phosphorylation of HDAC5, resulting export of HDAC5, thus exerting biological activity. In our early stage of the study, we conducted the high-throughput sequencing of many pathological scar tissue. Results revealed that the expression of HDAC5 in pathological scar tissue was significantly higher than that in normal skin. Meanwhile, we found after mechanical stimulation of scar fibroblasts in vitro, HDAC5 phosphorylation increased with deacetylation of α-tubulin. Based on these grounds, we hypothesize that biomechanics may activate HDAC5 to down-regulate the acetylation level of microtubules thus promoting fibroblast activation. So this project intends to first explore the effects of biomechanics on the biological function of fibroblasts and the deacetylation of α-tubulin. Next, through HDAC5 knockdown/overexpression cells and HDAC5 knockout mice to testify the role of HDAC5 in fibroblast activation. Finally, HDAC5 inhibitors are used to verify the effective efficacy of targeting HDAC5 to treat pathological scars, providing new ideas for the treatment of pathological scars.

局部皮肤张力是病理性瘢痕形成的重要因素。张力刺激可促进瘢痕成纤维细胞活化。细胞微管骨架是细胞感受张力的重要元件，其乙酰化修饰与张力诱导的成纤维细胞活化有密切关系。研究表明，在损伤修复过程中，HDAC5能够出核调控微管骨架乙酰化促进细胞修复。同时有报道，张力刺激能够激活HDAC5，促使其出核发挥功能。项目组前期通过高通量测序发现，病理性瘢痕组织中HDAC5表达显著高于正常皮肤。同时，瘢痕成纤维细胞经张力刺激后，HDAC5磷酸化增多并伴随微管骨架去乙酰化。据此提出假说: 张力通过激活HDAC5，下调微管骨架乙酰化水平，促进成纤维细胞活化。本项目拟首先探索张力对HDAC5及微管骨架去乙酰化的影响；进一步通过HDAC5基因敲减/过表达研究，明确HDAC5介导了张力对成纤维细胞活化的作用；最后通过HDAC5抑制剂进行转化研究，验证靶向HDAC5治疗病理性瘢痕的切实疗效，为病理性瘢痕的治疗提供新思路。

张力刺激在皮肤病理性瘢痕形成过程中起到重要作用，然而具体调控机制尚不明确。本项目率先发现一种IIa类家族HDAC成员HDAC5在人及小鼠的病理性瘢痕组织高表达，且HDAC5具有力学敏感性，张力能够上调HDAC5磷酸化水平并使其出核。HDAC5基因敲除小鼠结果显示，HDAC5 KO小鼠瘢痕增生情况较WT小鼠明显减轻。体外研究显示，shRNA敲低HDAC5后，Smad7蛋白表达增加。而Smad7 敲低能够缓解HDAC5缺失介导的Smad2/3磷酸化下调和瘢痕增生抑制。荧光素酶报告基因实验和ChIP-qPCR分析进一步表明，HDAC5与MEF2A相互作用，从而抑制了MEF2A与Smad7启动子区域的两个位点(-1340-1330和-398-388)结合，从而导致Smad7启动子活性抑制。更重要的是，我们发现LMK235 (HDAC4/5抑制剂)局部应用可以显著抑制增生性瘢痕形成。综上所述，本研究的结果为病理性瘢痕的临床治疗提供了新靶点与新方法。

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