保留双层肌膜的功能性肌肉移植中S1P/S1PR1轴调节巨噬细胞迁移及分化对移植肌肉粘连与功能的影响

Functioning free muscle transplantation (FFMT) is an important method to restore the function of the limbs after brachial plexus injury and muscle defect. However, adhesion and fibrosis between the transplanted muscle and the recipient area seriously affect the contraction function. Applicants improved the harvest technique of transplanted muscle and emphasized the concept of preserving the intact "visceral-parietal layer" double-layer epimysium. After that, adhesion was alleviated and function recovery was improved after protecting the transplanted muscle by the double epimysium. In previous animal researches, we found that preserving the intact "visceral-parietal layer" double-layer epimysium could reduce the macrophage infiltration around the donor muscle. The literature suggested that the S1P/S1PR1 axis is an important pathway regulating the migration and differentiation of macrophage, and activation of the S1P/S1PR1 axis can improve muscle repair. Our pre-experimental results indicated that the down regulation of S1P pathway in the “double-layer epimysium” is associated with reduction of macrophage infiltration and collagen deposition. Base on this, we hypothesize the regulation of macrophage infiltration and differentiation by S1P/S1PR1 axis was associated with muscle adhesion and fibrosis after FFMT. We will further investigate the relation between S1P/S1PR1 and macrophage infiltration and differentiation and the effect of macrophage differentiation on proliferation and differentiation of fibroblasts, and explore the molecular mechanism in vitro and vivo. This project will provide experimental basis and a therapeutic target for alleviating the adhesion and fibrosis after functional muscle transplantation and improving the function recovery.

功能性肌肉移植是重建肌肉缺损和臂丛损伤后肢体功能的重要方法，但移植肌肉与受区的粘连、纤维化严重影响其重获神经再支配后的收缩功能。申请人发现保留完整“脏-壁层”双层肌膜，以壁层肌膜与受区基床接触而保护脏层肌膜下的移植肌肉，粘连减轻，临床疗效提高；动物实验发现保留双层肌膜可减少移植肌肉巨噬细胞浸润。文献表明S1P/S1PR1轴是调节巨噬细胞迁移和分化的重要通路，通过调节S1PR1可促进肌肉损伤修复。我们预实验发现在肌肉移植动物实验中双层肌膜内侧移植肌肉中巨噬细胞S1PR1表达下调，伴随炎性细胞浸润减少，胶原沉积减少。拟提出S1P/S1PR1轴调节巨噬细胞迁移及分化影响移植肌肉粘连及纤维化的假设，深入研究双层肌膜中巨噬细胞S1PR1受体与其迁移及分化的关系，以及巨噬细胞分化对局部成纤维细胞增殖分化的影响，探讨其分子机制，这将为减轻功能性肌肉移植术后粘连及纤维化，提高疗效提供理论依据与治疗靶点。

近年来，保留肌外膜完整性与肌肉移植术后粘连纤维化的关系逐渐引起临床医生的重视。本项目首先顺利建立了三种股薄肌原位移植模型（保留双侧肌膜、单层肌膜及去肌膜），探讨肌膜对股薄肌周围炎症细胞浸润和术后功能的影响、S1P/S1PR1通路在保留双层肌膜肌肉移植中对局部巨噬细胞迁移分化的影响及抑制肌肉粘连和纤维化的影响、以及S1P/S1PR1通路对成纤维细胞增殖分化的影响。阐明了功能性肌肉移植过程中，保留双层肌膜能减轻炎症细胞对股薄肌的浸润，对股薄肌起到保护作用；明确了保留双层肌膜联合S1PR1激动剂FTY720能进一步减轻股薄肌与邻近肌肉炎症反应，降低胶原沉积；探明了在肌肉移植中S1P/S1PR1通路不仅通过调节巨噬细胞的极化，促进M1型巨噬细胞向M2型巨噬细胞转化来促进抗炎作用，还增强M2型巨噬细胞的抗炎作用；此外，该通路还抑制巨噬细胞对成纤维细胞的增殖和分化，抑制胶原形成，促进组织的修复，减少粘连形成。