Später Thomas, Frueh Florian S, Nickels Ruth M, Menger Michael D, Laschke Matthias W
1Institute for Clinical & Experimental Surgery, Saarland University, 66421 Homburg/Saar, Germany.
Division of Plastic Surgery and Hand Surgery, University Hospital Zürich, University of Zürich, 8091 Zürich, Switzerland.
J Biol Eng. 2018 Nov 13;12:24. doi: 10.1186/s13036-018-0118-3. eCollection 2018.
The seeding of scaffolds with the stromal vascular fraction (SVF) of adipose tissue is a common prevascularization strategy in tissue engineering. Alternatively, adipose tissue-derived microvascular fragments (ad-MVF) may serve as vascularization units. In contrast to SVF single cells, they represent a mixture of intact arteriolar, capillary and venular vessel segments. Therefore, we herein hypothesized that the ad-MVF-based prevascularization of scaffolds is superior to the conventional SVF single cells-based approach.
SVF single cells and ad-MVF were enzymatically isolated from epididymal fat pads of green fluorescent protein (GFP) donor mice to assess their viability and cellular composition using fluorescence microscopy and flow cytometry. Moreover, collagen-glycosaminoglycan matrices (Integra®) were seeded with identical amounts of the isolates and implanted into full-thickness skin defects within dorsal skinfold chambers of GFP recipient mice for the intravital fluorescent microscopic, histological and immunohistochemical analysis of implant vascularization and incorporation throughout an observation period of 2 weeks. Non-seeded matrices served as controls. While both isolates contained a comparable fraction of endothelial cells, perivascular cells, adipocytes and stem cells, ad-MVF exhibited a significantly higher viability. After in vivo implantation, the vascularization of ad-MVF-seeded scaffolds was improved when compared to SVF-seeded ones, as indicated by a significantly higher functional microvessel density. This was associated with an enhanced cellular infiltration, collagen content and density of CD31/GFP microvessels particularly in the center of the implants, demonstrating a better incorporation into the surrounding host tissue. In contrast, non-seeded matrices exhibited a poor vascularization, incorporation and epithelialization over time.
The present study demonstrates that ad-MVF are highly potent vascularization units that markedly accelerate and improve scaffold vascularization when compared to the SVF.
用脂肪组织的基质血管成分（SVF）接种支架是组织工程中一种常见的预血管化策略。另外，脂肪组织来源的微血管片段（ad-MVF）可作为血管化单元。与SVF单细胞不同，它们代表完整的小动脉、毛细血管和小静脉血管段的混合物。因此，我们在此假设基于ad-MVF的支架预血管化优于传统的基于SVF单细胞的方法。
从绿色荧光蛋白（GFP）供体小鼠的附睾脂肪垫中酶解分离出SVF单细胞和ad-MVF，使用荧光显微镜和流式细胞术评估它们的活力和细胞组成。此外，用等量的分离物接种胶原-糖胺聚糖基质（Integra®），并植入GFP受体小鼠背部皮褶腔的全层皮肤缺损处，在2周的观察期内对植入物的血管化和整合进行活体荧光显微镜、组织学和免疫组织化学分析。未接种的基质用作对照。虽然两种分离物都含有相当比例的内皮细胞、血管周围细胞、脂肪细胞和干细胞，但ad-MVF的活力明显更高。体内植入后，与接种SVF的支架相比，接种ad-MVF的支架的血管化得到改善，这表现为功能性微血管密度显著更高。这与细胞浸润增强、胶原含量增加以及CD31/GFP微血管密度增加有关，特别是在植入物中心，表明更好地整合到周围宿主组织中。相比之下，未接种的基质随着时间的推移血管化、整合和上皮化较差。
本研究表明，与SVF相比，ad-MVF是高效的血管化单元，可显著加速并改善支架的血管化。
