Isolation of adipose tissue derived regenerative cells from human subcutaneous tissue with or without the use of an enzymatic reagent

Background Freshly isolated, unmodified autologous adipose derived regenerative cells (ADRCs) (also named Stromal Vascular Fraction; SVF) have emerged as a promising tool for regenerative cell therapy. Due to regulatory concerns recently the use of cells recovered by non-enzymatic processing of adipose tissue has been suggested. The Transpose RT system (InGeneron, Inc., Houston, TX, USA) is a commercially available system for isolating ADRCs from adipose tissue. This system makes use of the proprietary, enzymatic Matrase Reagent for isolating cells. It is the aim of this study to address the question how and to which extent the use of Matrase Reagent would influence cell yield, cell recovery, cell viability, biological characteristics, physiological functions or structural properties of the ADRCs in the final cell suspension when compared to isolation of ADRCs from adipose tissue under otherwise identical conditions, but without the use of the Matrase Reagent. Methods Subcutaneous adipose tissue from 12 subjects undergoing elective lipoplasty was processed using the Transpose RT system. Identical samples of 25 g tissue were processed either with or without the use of Matrase Reagent. Then, characteristics of the ADRCs in the respective final cell suspensions were evaluated with SVF yield statistics, microscopic inspection, CFU assays, embryoid body formation, differentiation assays and quantitative PCR. Results Cell yield in numbers of viable cells/gram of processed lipoaspirate was approximately twelve times higher in the final cell suspension when ADRCs were isolated from lipoaspirate with the use of Matrase Reagent (p<0.001), compared to the final cell suspension that was obtained when ADRCs were isolated from lipoaspirate just mechanically without the use of enzyme. Furthermore, cells isolated from lipoaspirate with the use of Matrase Reagent formed on average 16 times more CFUs per g lipoaspirate compared to cells processed without enzyme. On the other hand, cells isolated with the use of Matrase Reagent exhibited no statistically significant differences in the expression of regenerative cell-associated genes such as Oct4, Hes1 and Klf4 compared to cells isolated without enzyme. Also, cells isolated from lipoaspirate both with and without the use of Matrase Reagent were independently able to differentiate into cells of all three germ layers (i.e., into the adipogenic, osteogenic, hepatogenic and neurogenic lineages). Conclusion The process of isolating ADRCs from lipoaspirate with the use of Matrase Reagent significantly increased the yield in cell numbers and cell viability compared to just mechanical processing. Biological characteristics, physiological functions or structural properties relevant for the intended use (i.e., regeneration, repair or replacement of weakened or injured tissue) were not altered or induced using Matrase Reagent. The use of enzyme did not manipulate the cells as expression of embryonic genes and pluripotent differentiation capacity was equally present in cells recovered with or without enzyme.