Joris van Dongen

72 Chapter 3 Statistical analysis Immunohistochemistry images and colony formation assay images were analyzed using ImageJ, version 1.4.3.67 (NIH, USA). Descriptive statistics were used to evaluate the cell numbers, α -SMA and vWF expression, CD surface expression scores and colony area and intensity. Data were expressed as mean ± standard deviation (SD). A t-test was performed using Graphpad Prism, version 5.01 (Graph Pad Software Inc., Los Angeles). RESULTS Only living cells in the Stromal Vascular Fraction In the FAT-SVF as well as in the control only living cells (green) were detected (Fig. 2) in contrast to the dead cell (red) controls that contained only dead cells (Fig 3, Supplemental Content). FAT-SVF Control Figure 2. Immunofluorescent images of FAT-SVF and control viability. Living cells turned green, while dead cells turned red after staining with CFDA-SE/PI. Nuclei turned blue after staining with DAPI. Immunohistochemistry & Masson’s Trichrome The fraction of small vessels visible in FAT-SVF (determined as mean α -SMApositivity of 6.2%, [+/- 5.5] and mean vWF positivity of 7.0%, [+/- 4.2]) was higher than in the controls (mean α -SMA positivity of 0.6%, [+/-0.6] and mean vWF positivity of 0.3%, [+/-0.3]), respectively p<0.05 and p<0.01 (Fig. 3). Interestingly, in both groups, vessels were present that did not express vWF. Likely, the procedure to generate the FAT-SVF had activated part of the vessels and caused these to release their vWF. VonWillebrand Factor and other ‘damage-response’ proteins, are stored in the Weibel Palade bodies within endothelial cells, the contents of which are secreted after stress-induced