Effects of Frozen Stromal Vascular Fraction on the Survival of Cryopreserved Fat Tissue

Abstract

Background Nowadays, the use of cryopreserved fat tissue for soft tissue augmentation is common, except for its unpredictable fat graft absorption, and the toxicity of the cryoprotective agent remains a limitation. In this study, the effects of freezing stored fat tissue without a cryoprotector, and the addition of the stromal vascular fraction (SVF) on the survival of cryopreserved transplants was studied. Methods Lipoaspirates from six donors were processed and cryopreserved at − 20 °C, − 80 °C and − 196 °C, respectively. The authors evaluated the lipoaspirates in vitro, on the basis of fat tissue and SVF viability after cryopreservation. In vivo fat grafting was performed in nude mice. Six trenches were injected on the backs of mice. Cryopreserved tissues (− 20 °C, − 80 °C and − 196 °C) were injected on the right side, and the other side received the SVF combination. At 4 and 8 weeks after transplantation, the authors examined the weight, volume and morphology of the tissue and analyzed histochemical staining and immunohistochemistry (i.e., DIL, CD31 and VWF) to evaluate the survival of the fat grafts. Results After cryopreservation without the cryoprotective agent, adipose tissue maintained its morphology better in − 80 °C than − 20 °C and − 196 °C. SVF cells can retain their adhesive and proliferative properties after cryopreservation. Although cryopreservation caused damage to fat tissue, all explants showed intact adipocytes and vascular ingrowth. Most of all, the − 80 °C group had less graft resorption and fibrosis than the other temperature groups. There was increased survival of fat grafts in the SVF group compared with the control group. Conclusion In this study, the authors demonstrated that the storage temperature of − 80 °C was promising for 3 months of adipose tissue cryopreservation without a cryoprotective agent, and SVF could increase the survival rate of cryopreserved fat tissue. No Level Assigned 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 .

Cryopreservation of lipoaspirates: in vitro measurement of the viability of adipose‐derived stem cell and lipid peroxidation

“…However, by using commercial freezers, cryopreservation at −20°C or −80°C may be accessible for surgeons to cryopreserve fat tissue effectively, so we focused on comparing the effects of cryopreservation in those temperatures 27 . Consistent with other studies, cryopreservation in −80°C was most effective for 3 months of lipoaspirate cryopreservation compared with −20°C or −196°C in terms of cell viability and proliferation ability 5,22,26,27 …”

“…However, in general, nevertheless about 66.2% to 77% of cell necrosis is inevitable in all subzero samples; controlled freezing and rapid thawing method is gaining consensus as a most effective method 3,25,29,31 . Especially without any CPAs, fat tissues that traverse through −196°C undergo devastating changes and produce significantly more free oil droplets compared with −20°C and −80°C samples as in our study 5,22 . Storing fat tissue at liquid nitrogen requires a complex facility, so liquid nitrogen is limited in clinics.…”

“…Furthermore, proliferation of few survived stem cells or transformation of exogenous SVF into adipocytes in frozen fat grafts may contribute. Our cell yield was very low after freezing/thawing and the graft was filled with dead cells 22 . Clinically, Butterwick et al showed equivalent to improved result of frozen fat graft compared with fresh fat graft by side‐by‐side two‐hand comparison pilot study 23 .…”

“…Our cell yield was very low after freezing/ thawing and the graft was filled with dead cells. 22 Clinically, Butterwick et al showed equivalent to improved result of frozen fat graft compared with fresh fat graft by side-by-side two-hand comparison pilot study. 23 These results may delude the meaning of dead cells in fat graft.…”

“…Unviable adipocyte could act as extracellular matrix filler and induce subclinical inflammation reaction that resulted in positive tissue rejuvenation effects. But histologic examination of cryopreserved fat with high necrotic cell proportion shows high rate of graft absorption as well as oil cyst formation, chronic inflammation, and progressive calcification 22 . As compared with other currently available cryogenic banking of various tissues such as skin, heart valve, and dura mater, which the cryopreserved tissue serves only as tissue matrix scaffold, fat graft requires elaborate strategy to retain its volume without complications.…”