Ultrasound-assisted liposuction provides a source for functional adipose-derived stromal cells

Abstract

Background aims: Regenerative medicine employs human mesenchymal stromal cells (MSCs) for their multi-lineage plasticity and their pro-regenerative cytokine secretome. Adipose-derived mesenchymal stromal cells (ASCs) are concentrated in fat tissue, and the ease of harvest via liposuction makes them a particularly interesting cell source. However, there are various liposuction methods, and few have been assessed regarding their impact on ASC functionality. Here we study the impact of the two most popular ultrasound-assisted liposuction (UAL) devices currently in clinical use, VASER (Solta Medical) and Lysonix 3000 (Mentor) on ASCs.

Methods: After lipoaspirate harvest and processing, we sorted for ASCs using fluorescent-assisted cell sorting based on an established surface marker profile (CD34⁺CD31^-CD45^-). ASC yield, viability, osteogenic and adipogenic differentiation capacity and in vivo regenerative performance were assessed.

Results: Both UAL samples demonstrated equivalent ASC yield and viability. VASER UAL ASCs showed higher osteogenic and adipogenic marker expression, but a comparable differentiation capacity was observed. Soft tissue healing and neovascularization were significantly enhanced via both UAL-derived ASCs in vivo, and there was no significant difference between the cell therapy groups.

Conclusions: Taken together, our data suggest that UAL allows safe and efficient harvesting of the mesenchymal stromal cellular fraction of adipose tissue and that cells harvested via this approach are suitable for cell therapy and tissue engineering applications.

Keywords: ASCs; Lysonix; VASER; adipose-derived stromal cells; adult mesenchymal stromal cells; cell therapy; regenerative medicine; ultrasound-assisted liposuction.

Figure 1

Vaser and Lysonix lipoaspirates yield similar amounts of ASCs with comparable viability. (A): FACS analysis for identification of ASCs (defined as CD45−/CD31−/CD34+) of Vaser and Lysonix derived lipoaspirates; (B): CD45−/CD31−/CD34+ASC-quantification in Vaser and Lysonix derived SVF showed no significant difference in ASC yield across samples (C): MTT assay showed no significant difference in cell viability. n=3; data are given as mean ±SEM. Abbreviations: SVF: Stromal Vascular Fraction; ASC: Adipose derived Stromal Cells.

Figure 2

Assessment of Osteogenic Differentiation Capacity of Vaser and Lysonix-derived samples. (A) and (B): Alizarin red staining and quantification showing a trend towards higher mineralization for Vaser samples; (C), (D), and (E): Real-time-PCR quantifying the expression of early (RUNX-2), intermediate (OPN) and late (OCN) osteogenic marker. RUNX-2 showed significantly higher expression in Vaser samples on day 7 and 14 and OCN was significantly higher expressed in Vaser samples on day 14, while OPN was significantly higher on day 0 in samples of Lysonix ASCs. Abbreviations: PCR: polymerase chain reaction, RUNX2: runt related transcription factor 2, OPN: osteopontin, OCN: osteocalcin.

Figure 3

Adipogenic Differentiation Capacity of Vaser and Lysonix derived samples. (A), (B) and (C): Real-time-PCR for quantification of key-adipogenic markers (PPAR-γ, FABP-4 and LPL) showed a higher expression of all markers in Vaser harvested samples at day 7. (D) and (E): Oil Red O staining showed no differences in lipid accumulation. Abbreviations: PPAR-γ: peroxisome proliferator-activated receptor-γ, FABP4: fatty acid binding protein 4, and LPL: lipoprotein lipase.

Figure 4

ASCs harvested with either VASER or Lysonix liposuction showed similar ability to enhance murine wound healing. There was a significant enhancement of wound closure compared with control in both cell therapy groups. (A) wound appearance, (B) kinetics of wound size and (C) closure time of CTRL, Vaser and Lysonix groups. n= 8; p< 0.01. All data are given as mean ±SEM. Abbreviations: CTRL: control.

Figure 5

Hydrogels seeded with Vaser and Lysonix derived ASCs resulted in improvement of vascular density of wounds. (A) CD 31 (red) and DAPI (blue) staining were performed and showed increased neovascularization in both ASC seeded hydrogel groups. (B) Quantification showed no significant difference between ASC loaded hydrogels. Scale bar= 100 μm. n=8. Asterisk indicates p≤0,05. All data are mean ± SEM, Abbreviations: CTRL: control; HPF: high power field.