Objectives The chemokine stromal cell-derived factor-1 (SDF-1) plays a critical role in mobilising CXCR4-positive precursor cells in the bone marrow and may be essential for efficient vascular regeneration and repair. We recently reported that CXCR4 and the angiogenic potential of bone marrow derived cells (BMCs) were regulated positively by calcium and negatively by ageing. We hypothesise that atherosclerosis may also affect CXCR4 surface expression and functions. This study is to determine that atherosclerosis defects BMCs both on the decreased population of CXCR4+ progenitor cells and on the impaired mobility as well as to explore possible mechanisms.
Methods We established atherosclerosis model on ApoE-/- mice under at least 6-month High Fat Diet (group Ath), and used 3∼4-week aged ApoE-/- mice as control (group Ctrl). In order to evaluate impact of the disease on CXCR4 expression from gene transcription to protein synthesis, surface CXCR4 expression on BMCs from the two groups of mice were analysed with FCM; the mRNA of CXCR4 was detected by real-time PCR. To compare the inducibility of CXCR4 expression in response to environmental change, 1 mM CaCl2 was applied as an extracellular stimuli to treat BMCs for 4 h. To examine how BMCAth are defect on calcium-induced CXCR4 surface expression, calcium influx was analysed by measuring the increase of intracellular calcium after BMCs were mixed with CaCl2. Cell-cell adhesion assays were used to detect the SDF-1 mediated adhesion of BMCs to the HUVEC monolayer and vertical invasion assays were used respectively to quantify trans-endothelial migration of BMCs in response to a gradient of SDF-1.
Results BMCAth have less number of cells expressing CXCR4 as compared to BMCCtrl (8.54±1.96% vs 13.75±3.94%; n=10, p<0.01). CXCR4 expression on BMCCtrl could be enhanced by calcium, but CXCR4 surface expression in BMCAth increased significantly lesser then BMCCtrl (11.24±1.31% vs 26.59±4.92%; n=10, p<0.01). It is partly because of the defective calcium influx in BMCAth which reduced the CXCR4 gene transcription, consequently leading to impaired responses on calcium-induced CXCR4 surface expression. BMCAth showed weaker lower mobility and lower trans-endothelial migration (0.80±0.11 mm vs 1.17±0.15 mm; n=4, p<0.05), and this was not enhanced by calcium pretreatment.
Conclusions Atherosclerosis impairs CXCR4 surface expression on BMCs and related cell functions.