An Optimized Method for Adipose Stromal Vascular Fraction Isolation and its Application in Fat Grafting.

Abstract

The stromal vascular fraction (SVF) derived from adipose tissue contains heterogeneous cell populations and has enormous potential for clinical therapy. There are two main methods for SVF isolation: enzymatic isolation and mechanical isolation, both of which have shortcomings. In this study, optimized conditions for the isolation of high-quality SVF were established, and applications in fat grafting were evaluated. Adipose tissue was chopped into small pieces and then ground into an erosive shape using a syringe. The pieces were digested with 0.15% type II collagenase for 35 min at 37°C. After centrifugation, the pellets were resuspended in DMEM and passed through a 100-μm strainer. The filtered cells were analyzed by flow cytometry. The fat graft was enriched with isolated SVF and subcutaneously transplanted into nude mice. Three weeks after transplantation, grafts were isolated, and H&E staining, immunocytochemistry, and western blotting were conducted. The harvested SVF cells reached > 2 × 106/ml of adipose tissue within 90 min of operation. The number of CD34+ ADSCs in our SVF pellets was > 6 × 105/ml of adipose tissue, which has the potential for differentiating into osteoblasts, adipocytes, and chondrocytes. Freshly collected adipose tissue is better for SVF isolation, and isolated SVF should also be kept at 4°C and used as soon as possible. SVF may promote revascularization after fat grafting. The adipose tissue of an SVF co-transplanted group had an integral structure, clear capillaries, and higher VEGF expression. SVF co-transplantation inhibited adipose cell apoptosis. Our study provides an efficient procedure for SVF isolation, its application in fat grafting, and possible underlying mechanisms. This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266 .