Phenotypic Analysis of Nanofat-Derived Stromal Vascular Fraction and Mesenchymal Stem Cells Reveals Improved Progenitor Subpopulations When Compared to Standard Lipoaspirate

Introduction: Optimization of fat grafting, especially with the introduction of cell-assisted lipotransfer, is a technique that continues to gain increasing attention in the field of regenerative medicine. One form that was first described by Tonnard et al. in 2013, “nanofat grafting,” implements mechanical emulsification and injection of standard lipoaspirate (LA) for the correction of rhytides and pigmentation in superficial regions such as the eyelids and lips (1). Tonnard’s group demonstrated good clinical outcomes, safety, and the ability to generate Adipose-derived stem cells (ADSCs) from this starting material; however, little work has been done to further characterize the stem and progenitor cell populations of the stromal vascular fraction (SVF) generated by this technique.

Methods: Standard vacuum-assisted liposuction was used to obtain 100 ml LA from 2 patients undergoing routine elective procedures. 50 ml of each LA was subjected to nanofat processing. The nanofat and remaining 50 ml LAs were subjected to collagenase digestion, red blood cell lysis and the resulting SVF pellets were then stained with propidium iodide and a multicolor fluorescence antibody panel (CD13-APC-Vio770, CD31-FITC, CD34-PerCP-Vio700, CD45-VioBlue, CD73-PE and CD146-APC). The cells were then immediately subjected to flow cytometry analysis for absolute cell count and population characterization (T0). Cells from the two treatment groups were then placed into standard culture conditions and then subjected to flow cytometry analysis at the end of passages 0 (P0) and 1 (P1).

Results: While the MSC population (CD45-/CD34+) was similar between nanofat and standard LA (1.96% ± 0.004 vs 2.27% ± 0.009, respectively), the freshly isolated SVF derived from nanofat was found to be much more abundant in CD34+ cells (16.8% ± 0.044 vs 6.56% ± 0.017, respectively). This CD34+ advantage was also observed at P0 and P1 of the cultured cells (49.1% ± 0.106 vs 40.0% ± 0.259 and 4.87% ± 0.02 vs 2.14% ± 0.009, respectively). Subpopulation analysis at T0 also revealed higher concentrations of ADSCs (CD45-/CD34+/CD13+/CD73+ - 14.1% ± 0.007 vs 5.02% ± 0.019, respectively) as well as endothelial progenitor cells (CD45-/CD31+/CD34+/CD146+ [EPCs] - 7.25% ± 0.011 vs 0.93% ± 0.002, respectively).

Conclusions: Nanofat has proven useful for the amelioration of superficial rhytides and pigmentation and this study is the first to demonstrate possible mechanisms of action for tissue regenerative effects conferred by the presence of increased progenitor activity in the form of ADSCs and EPCs. More research is needed to validate these findings and further define the underlying regenerative mechanisms of mechanically processed lipoaspirate, but this is a promising first step.

1.           Tonnard, P., Verpaele, A., Peeters, G., Hamdi, M., Cornelissen, M., Declercq, H. Nanofat grafting: basic research and clinical applications. Plastic and reconstructive surgery 2013;132:1017-1026.