干细胞归巢促进自体胶原蛋白脂肪体系稳定成脂及微血管化动态追踪

The apoptosis of mature adipocytes and insufficient vascularizatin after fat grafting, result in calcification, chronic inflammation, and low retention, increase the treatment times and patient’s suffering. The research in recent years indidated that the adipogenesis of autologous Adipose stem cells (ADSCs) were promoted with Stromal Vascular Fraction (SVF) derived form fat tissue. But The early depletion of macrophages remarkably hindered SVF reconstruction process and adipogenesis by down-regulating the expression of collagen proteins, and the extraction of SVF waste a great quantity of fat tissue, which not fulfill the Large volume fat transplantation. The prophase research of our research team indicated that EPCs with stable proliferation and microvascularizaion ability could be harvested from peripheral blood, which can promote the retention of fat grafting and construct the microcirculation system, and a large amount fibroblasts were stable co-cultured and harvested. In this research, the collagen gel, which were harvested from autologous fibroblasts, were combined with autologous adipose tissue to format autologous collagen-adipose system (ACAS) , which with ability to promote autologous ADSCs and (Epithelial progenitor cells) EPCs homing. In vitro and vivo, the ADSCs and EPCs homing, microvascularization and whole process of adipogenesis of ACAS were dynamic traced, to provide a new method for large amount adipose tissue transplantation applying to clinical surgery. This research is to provide the systematized technology for constructing an adipose grafting system with high effectively microvascularization and stable ability of adipogenesis by promoting ADSCs and EPCs homing, which can be fit for clinic plastic surgery application.

脂肪移植术后难以避免成熟脂肪细胞凋亡及血管化不良，导致钙化、感染、成活率低等，增加治疗次数和患者痛苦。近年研究热点脂肪基质血管组分（SVF）可促进脂肪干细胞（ADSCs）成脂，但此过程中巨噬细胞早期耗竭下调胶原蛋白，显著阻碍SVF成脂，且SVF提取损耗大量脂肪，不利于大容量脂肪移植。课题组前期研究表明，外周血可获取稳定增殖及微血管化的EPCs，可促进移植脂肪的成活，且成功培养扩增大批量成纤维细胞。本研究采用自体成纤维细胞提取胶原蛋白，与脂肪组织构建研发自体胶原蛋白脂肪体系（ACAS），诱导干细胞ADSCs及外周血内皮祖细胞（EPCs）归巢。体外、体内持续动态追踪ACAS体系中干细胞ADSCs和EPCs归巢和近、中、远期稳定成脂,，以及微血管化情况，将成为大容量脂肪移植临床应用新思路。本课题为研发能诱导自体干细胞归巢进行高效率微血管化及稳定成脂的微环境脂肪体系应用于临床提供系统化技术支持。

提高大容量脂肪移植的容留率是整形外科面临的挑战之一。大体积移植脂肪容易导致成熟脂肪细胞凋亡，移植物因缺血导致移植失败，利用自身ADSCs进行成脂、早期微血管化以及提供良好的细胞微环境是提高脂肪移植容留率的关键，课题组前期研究发现EPCs和ADSCs能够有效生成新生微血管并促进移植脂肪组织存活。本研究采用成纤维细胞提取自体胶原蛋白，构建胶原蛋白脂肪体系，促进机体ADSCs和EPCs动态归巢，稳步提升脂肪移植物微血管化和稳定成脂的效率。课题研究针对解决大容量脂肪组织移植血管化困难导致的坏死液化、吸收、成活困难、需要反复治疗等问题，对自体胶原蛋白提取及鉴定，I型胶原蛋白对于ADSCs以及EPCS的体外细胞增殖、趋化以及功能表达检测，胶原蛋白脂肪体系中干细胞体内归巢效应的监控以及胶原蛋白脂肪体系移植后微血管及脂肪存活情况进行研究。研究表明，在体外自体胶原蛋白能够从大量成纤维细胞中获取，I型胶原蛋白能够促进ADSCs增殖以及趋化能力和EPCs增殖能力，并通过上调PI3K/Akt信号通路及粘附分子促进EPCs成管，而ADSCs在I型胶原蛋白的作用下能够保持多向分化的潜能；在体内，尾静脉注射ADSCs及EPCs归巢至移植脂肪组织效率有限，I型胶原蛋白能够促进ADSCs及EPCs归巢至移植脂肪组织，胶原蛋白脂肪联合体系刺激移植脂肪组织血管重建，提高成脂效率，减轻炎症，降低空泡形成，同时提供了较好的细胞微环境，有效提高大体积脂肪移植的容留率。表明本研究构建的胶原蛋白脂肪联合体系在促进干细胞归巢、改善血供以及提高移植脂肪成活等方面，具有单一脂肪移植无法比拟的优点。课题研究使整体脂肪移植过程更加高效，降低不良反应及减少治疗次数，使研究成果更适宜临床推广及运用。