Effect of Stromal Vascular Fractions on Angiogenesis of Injected Diced Cartilage

Abstract

Objectives: This study explored the effect of adipose-derived stromal vascular fractions (SVFs) on angiogenesis in injected autologous diced cartilage. Methods: Stromal vascular fractions were extracted by enzymatic digestion. Cartilage grafts were harvested from 1 side of the auricular cartilage of New Zealand rabbit and then diced to a size of 1.0 mm 3 . The grafts were divided into 2 groups. The control group was diced cartilage mixed with culture medium, and the experimental group was diced cartilage mixed with SVFs. The 2 groups of composite grafts were subcutaneously implanted on both sides of the back of each rabbit. After 4, 12 and 24 weeks, the tissue structure, number of blood vessels, and angiogenic factors in the grafts were observed. Results: The SVFs conformed to the current standard of the biological evaluation. Under an inverted microscope, the number of layers of chondrocytes in the experimental group was higher than that in the control group at 4 weeks. A small number of inflammatory cells and blood vessels were observed around the cartilage grafts. At 12 and 24 weeks, the volume of tissue was increased gradually by general observation. And a large number of chondrocytes were observed microscopically, whereas the number of inflammatory cells decreased. And meanwhile additional new blood vessels were observed. Immunohistochemical analysis of CD31 showed that the number of capillaries in the control group was significantly lower than that in the experimental group at 4, 12 and 24 weeks. Further, the expression of Hypoxia inducible factor-1 (HIF-1) and vascular endothelial growth factor (VEGF) mRNA and protein were measured by RT-PCR and Western bloting, respectively. The results showed that the mRNA expression of VEGF and HIF-1a in the experimental group was increased. The mRNA level remained higher than that of the control group at 24 weeks (P < 0.05). And the relative expression levels of VEGF and HIF-1a protein in the experimental group were higher than those in the control group at 4, 12 and 24 weeks (P < 0.05). Conclusion: Autologous diced cartilage mixed with adiposederived SVFs can promote angiogenesis when transplanted by injection. Further research showed that SVFs could increase the expression levels of VEGF and HIF-1a in the grafts, which may be part of the mechanism that SVFs promoted the angiogenesis of diced cartilage.

SVF cell sheets: A new multicellular material-based strategy for promoting angiogenesis and regeneration in diced cartilage grafts

“…Currently, adipose-derived stem cells and EPCs are important components of the SVF [21] . Our previous studies also con rmed that rabbit adiposederived SVF contains these two types of cells and that the injection of SVF cell suspensions promoted angiogenesis and the survival of diced cartilage grafts [12] . This study demonstrated for the rst time that using SVF cell sheets to wrap diced cartilage grafts could more effectively promote angiogenesis and cartilage regeneration in cartilage grafts; notably, this method improved the survival rate of the grafts.…”

“…In this study, for SVF cell sheets, the expression of the pro-angiogenic factors Ang-1 and VEGF was stronger than that using autologous perichondrium, and more peripheral adipose tissue and neovasculature formation around the grafts were observed at 3 and 6 months after surgery, suggesting that the SVF cell sheets better promoted the vascularization of cartilage grafts. The reason may be that the SVF cell sheets contain a large number of EPCs, which are conducive to neovasculature formation [12] .…”

“…1b). Flow cytometry revealed that the SVF included adipose stem cells (ADSCs) and EPCs [12]. At day 2 of culture in cell sheet-induction medium, observation under an inverted phase-contrast microscope showed that the cells in the temperature-sensitive culture dish had reached 100% con uence.…”

“…Rabbit adipose-derived SVF cells were extracted and identi ed following the method reported in our previous study [12] . The experiments were approved by The Experimental Animal Department of Nanhua University (SYXK (Xiang 2020-0002)), and reported in accordance with ARRIVE guidelines.…”

“…Studies on cartilage regeneration have shown that SVFs can promote cartilage regeneration and repair [10,11] . In our previous study, rabbit adipose-derived SVFs cultured in vitro were mixed with autologous diced cartilage and then injected for transplantation in vivo, and the results showed that SVFs can promote angiogenesis in and the survival of diced cartilage grafts [12] . Takeuchi et al reported that, compared to cell injection, cell sheet transplantation consistently increased the cell survival time, and there were fewer apoptotic cells in the cell sheet than in the cell suspension [13] .…”