The purpose of this study was evaluated the osteogenic differentiation in vitro of mesenchymal stem cells (MSC) of SHR (Spontaneously Hypertensive Rats) and Wistar (Normotensive) rats.Therefore, MSC from femoral bone marrow of Wistar and SHR rats 4 weeks old were utilized. The experimental protocols were approved by the Animal Experiments Local Ethics Committee (Process 00716-2012). The osteogenic medium (OM) consist of the proliferation medium (MEM) supplemented with 50 μg/ml ascorbic acid, 10 mM β-glicerophosphato and 10−8M dexamethasone. We evaluated the cell proliferation, alkaline phosphatase (ALP) activity and total protein by colorimetric methods; mineralization of nodules was analyzed using alizarin red staining; and osteoblasts-associated protein expression by qPCR. Statistical analysis we used two-way ANOVA test, followed by Bonferroni test. Differences were regarded as significant if P<0.05. The results demonstrated reduced proliferation rate, total protein content and mineralization in osteoblast of SHR plus OM (SHROM) than Wistar plus OM (WOM). However, ALP activity has no difference between both groups. The transcription factors Osterix and β-catenin were low in SHROM, suggesting reduced differentiation in this group. The decreased expression of osteoblast-associated proteins as such OCN, BSP, COL I and OPN in SHROM, revealed the low quality of extracellular matrix (ECM). All this results described above evidence reduced osteoblast differentiation in SHROM. IL-6 and CXCL1/CINC2 production was increased in SHRC (proliferation medium) compared to WC (proliferation medium), suggesting a proinflammatory stage in SHR. In conclusion, SHR osteoblast differentiation is reduced than compared to Wistar. Lower mineralization and poor ECM quality was also observed. These data suggest that the influence of hypertensive genotype on bone metabolism of young SHR, before the hypertension development should be considered. These differences may be the reason of the bone alterations in the SHR strain. Financial support: FAPESP (Process 2012/01924-9; 2011/06070-5; 2011/19458-1).
17 May 2014 - 20 May 2014