3.1.1. Diabetic mice display lower BM-KSL Cells
Diabetic mice were used after as early of 4 weeks induction, which determined by constant hyperglycemia (>300 mg/dL or higher). We started our study by confirming the effect of diabetes model on functional bone marrow-derived endothelial progenitor cells (BM-EPC). Sufficient studies have demonstrated that diabetes caused decrease in number of endothelial progenitor cells [4,6,42,43]. Our findings demonstrated that diabetic BM-EPC, as sorted for Lin-/Sca-1+/c-Kit+ (KSL) cells by flow cytometry, significantly has lower KSL cell number (8,649+1,906) compare to control (19,729+4,649; p<0.05) (figure 1A). In addition, we also investigated whether the KSL percentage within bone marrow were also affected by diabetes. Here we found that the KSL cell percentage within bone marrow significantly decreased in diabetes (0.98+0.10) compare to control (1.58+0.08; p<0.05) (figure 1B). The results were indicated that 4 weeks of diabetic induction, which represent as early stage of diabetic, has caused quantity impairment in diabetic BM-KSL cells. The result was in line with previous study which demonstrated lower level of EPC under diabetic condition [4,5].
3.1.2. Vasculogenic Potential is decreased in diabetic BM-KSL cells
Since vascular impairment is a hallmark in diabetes and that BM-KSL which represent as bone marrow-EPC is we then investigated whether this impairment is started from the primitive endothelial progenitor cells reside within bone marrow. Previous studies have demonstrated that diabetes altered bone marrow niche and caused detrimental effect in bone marrow microenvironment, lead to perturb of microenvironment cells . However whether this result also impairs the vasculogenic function of BM-KSL cells reside in bone marrow, is still remain to be elucidated. We investigated in further for the vasculogenic potential of diabetic BM-KSL using colony formation assay (CFA) as previously described [8,10,45,46]. Using colony formation assay, enable us to distinguish two types of colonies which represent different EPC vasculogenic function. First is primitive DM-EPC showed significantly lowered in number of primitive endothelial progenitor cells colonies (pEPC) (diabetic 5.30 + 1.30; control 11.50 + 0.55, p < 0.05) and total endothelial progenitor cell (tEPC) colony (diabetic 5.90+1.65; control 12.17+0.80; p<0.05), however number of definitive endothelial progenitor cell (dEPC) colonies were similar between diabetic and control mice (diabetic 0.67 + 0.33; control 0.67 +0.49; p > 0.05) (Figure 2). Decreased in bone marrow number of total colony forming unit (BM-CFU) in further will affect vasculogenic potential of the EPC when involve in neovascularization .
3.2. Diabetic BM-KSL exhibit comparable oxidative stress level as control
It is purposed that within bone marrow, the stem/progenitor cells are surrounding with unique microenvironment (niche) which has low tension of Oxygen and hypoxic...