Abstracts Category : Other

Semi-automated extraction of stromal vascular fraction for autologous cell therapy

by Alexander Hanke

Introduction: The stem cell rich Stromal Vascular Fraction (SVF) can be obtained by enzymatic digestion with a collagenase followed by centrifugation from patients lipoaspirate or fat tissue. To date neither a standardized extraction method nor a generally accepted application procedure exists for common use on patient. A novel commercially available semi-automated device for the extraction of SVF promises sterility, consistent results and usability in the clinical routine. The aim of this work was to investigate the quantity and quality of the SVF obtained by a semi-automated process in comparison to an established manual laboratory method.Material and Methods: SVF was extracted from lipoaspirate by a prototype of the semi-automated UNiStation (NeoGenesis, Seoul, Korea) as well as by hand preparation with common laboratory equipment. The SVF was measured by multi-parametric flow-cytometry (FACSCanto-II, BD Biosciences) following the lysis of the remaining erythrocytes. The primary interest was the total cell number (quantity) of the extracted cells. In addition, the quality of the SVFs was investigated using the stem cell marker CD34, the leucocyte marker CD45 and the marker CD271 for highly proliferative stem cells. Furthermore, the distribution of these markers, double positive cells and the stain index were investigated.Results: Lipoaspirate obtained from six patients was processed with both the novel device (d) as the hand preparation using laboratory equipment (h), always resulting in a macroscopically visible SVF. However, there was a tendency of a fewer cell yield per gram of used lipoaspirate with the device (d: 1.1*1051.1*105 vs. h: 2.0*1051.7*105; p=0.06). Regarding the composition of the SVF, the percentage of CD34+ cells was significantly reduced with the device (d: 57.323.8% vs. h: 74.113.4%; p=0.02). On the contrary there was a tendency to a higher percentage of CD45+ leukocytes (d: 20.715.8% vs. h: 9.87.1%; p=0.07). The percentage of highly proliferative CD271+ cells was comparable for both methods (d: 13.411.6% vs. h: 12.99.6%; p=0.74). No significant difference was identified regarding the double positive cell fraction for CD34+/CD45+ (d: 0.50.6% vs. h: 0.30.2%; p=0.21) and CD34+/CD271+ (d: 1.92.3% vs. h: 2.42.0%; p=0.42). Double positive cells for CD45+/CD271+ were not detected in any sample. The stain index did not show a significant difference between the two extraction methods (p>0.12).Discussion: The semi-automated system was able to provide considerable amounts of sterile SVF without requiring much space. The SVF extracted by the semi-automated process showed only little difference in its composition compared with the SVF obtained by the hand preparation. Taken together both methods showed comparable extraction results which are in accordance with the data from literature. This semi-automated system offers an opportunity to take research and application of the SVF one step further to the clinic. Einleitung: Die stammzellreiche Stromal Vascular Fraction (SVF) kann ausAdvisors/Committee Members: Prantl, Lukas (advisor).