力生长因子E肽在严重低氧条件下调控前交叉韧带损伤后修复的机理研究

Due to the angiorrhexis and other factors, oxygen concentration is dramatically reduced after anterior cruciate ligament (ACL) rupture, whereas severe hypoxic environmental conditions often induce cell apoptosis and lead to the lack of progenitor cells participating in the repair process. Our previous studies demonstrated that high-expression of HIF-1α and remarkable cell apoptosis could be observed at the injured sites after ACL rupture. Moreover, we found that mechano-growth factor (MGF) E peptide could protect ACL fibroblasts from severe hypoxia-induced cell apoptosis through in vitro and in vivo analysis, but the relevant mechanisms are still unknown. Based on these preliminary studies, in this project, we will examine relevant apoptotic pathways and molecular mechanisms involved in MGF E peptide mediated regulation of the cell apoptosis of ACL fibroblasts under severe hypoxia. In addition, New Zealand white rabbits will be used to establish the ACL rupture models. Effects of MGF E peptide on angiogenesis, cell recruitment and mechanical functions of the injured ACL will be analyzed in vivo. This subject will provide a feasible therapeutic strategy for regeneration and repair of ACL, and imply significance of basic research and clinical application value.

前交叉韧带(anterior cruciate ligament, ACL)损伤后由于血管破裂等因素导致膝关节腔内氧浓度急剧下降，而严重低氧环境将引起细胞发生凋亡,导致损伤位点缺乏充足的前体细胞参与修复。我们前期的研究发现ACL组织撕裂后损伤位点处HIF-1α表达显著升高，并伴随大量的细胞凋亡。进一步在体外和体内实验中发现，力生长因子(mechano-growth factor, MGF)E肽能够保护ACL成纤维细胞免于严重低氧引起的细胞凋亡，但具体机理未知。本项目中将分别在基因水平、蛋白水平考察MGF E肽在严重低氧条件下调控ACL成纤维细胞凋亡所涉及到的凋亡途径与分子机制。同时建立兔ACL撕裂损伤模型，在体分析MGF E肽在ACL损伤修复过程中对血管生成、细胞招募及力学功能的影响。本课题为ACL再生修复提供了一条可行性策略，具有重要的研究意义与应用价值。

膝关节是人体最大的关节结构，控制了人体运动与力学稳定。力学失稳或过度负荷可能会引起关节腔内前交叉韧带(ACL)的损伤，更进一步引起半月板降解，甚至骨关节炎。病理过程中，由于炎症反应和血供不足会导致缺损部位产生缺氧微环境，进一步阻碍了ACL和骨关节炎的恢复与组织再生。力生长因子E肽（MGF E肽）能够在严重低氧条件下降低新生海马细胞、大鼠心肌细胞等的凋亡。MGF还被证实可以有效促进肌肉肥大、活化卫星细胞以及促进细胞迁移等，表现出调控组织修复过程中细胞增殖的效能。本项目中，我们将验证MGF在严重低氧条件下对ACL细胞和软骨细胞的活力、增殖、凋亡等的影响，并探索相关机制。同时分别建立韧带、软骨损伤动物模型，在体水平上进一步验证相关假设。结果显示，严重低氧条件下ACL成纤维细胞活力、增殖能力下降，细胞凋亡显著增加。MGF处理后能够明显地促进低氧条件下ACL成纤维细胞的增殖能力并增加细胞存活能力。该过程MGF通过调控caspase家族相关分子表达，并且依赖PI3K-Akt和MEK-ERK1/2信号通路调控了严重低氧条件下的ACL成纤维细胞的凋亡。在体水平上不仅证实MGF能够通过降低损伤部位HIF-1α的表达抑制了细胞凋亡，同时还通过促进血管生成促进了ACL损伤后再生。骨关节炎课题中，结果证实MGF能够显著促进严重低氧环境下软骨细胞的增殖能力，抑制了软骨细胞凋亡并平衡了细胞外基质的表达。同样在体实验表明，MGF能够有效促进软骨缺损后修复和软骨基质表达。以上结论表明，MGF能够有效促进膝关节内组织损伤后的组织修复，是一种潜在的可用于临床韧带损伤、骨关节炎治疗的可行方式。

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