The Effect Of Fibrinogen On EPCs' Angiogenic Function

Background: It is reported that both endothelial progenitor cells(EPCs)and fibrinogen could be involved in the endothelium repair and angiogenesis.Bone marrow derived EPCs can migration and homing in damage position to take part in the endothelium repair and angiogenesis to response to the injury of vascular.Fibrinogen can participate in angiogenesis in tumour by promoting the migration and proliferation of endothelial cells(ECs).In addition,chronic exposure to fibrin and fibrinogen could affect migration and proliferation by differentially elevating intracellular Ca2+ in human pulmonary arterial smooth cells(PASMCs)and endothelial cells(PAECs).Objective: We sought to explore the possible effect of fibrinogen on angiogenesis by culturing EPCs on fibrinogen.Methods: Coverslips will be either coated with fibrinogen of two concentrations,4 and 40?g/ml diluted in PBS,or PBS alone.Plate EPCs derived from rat bone marrow on the coated coverslips for 72 hours after culture in vitro 9 days.Use computer program such as Image J to quantify the tube network by measuring the following parameters: numbers of loops/meshes,numbers of branch sites/nodes and length of tubes.Results: EPCs' morphological changes: One day later: cells adherent to the wall,most of the cell morphology were round;Four days later: part of the adherent cells were fusiform or polygon and visible cells arranged in line;Six days later: the number of fusiform or polygon cells increased and saw to be preferential growth;Eight days later: almost of the cells were fusiform or polygon and cells slabstone shaped.EPCs fluorescence staining:The rate of the cells which both FITC-UEA-1 and Dil-ac-LDL positive were 85.726%±2.568%(n=3).Angiogenesis: length of tubes is 9.782±0.666 mm,number of loops/meshes is 7.433±0.286,number of branch sites/nodes is 8.533±1.347(n=3).Control: length of tubes is 9.975±0.563 mm,number of loops/meshes is 8.967±0.260,number of branch sites/nodes is 11.800±0.436(n=3);The parameters of 4?g/ml group did not show significant statistically differences compared to control: length of tubes is12.604±1.354mm(P=0.231),number of loops/meshes is 10.767±0.649(P=0.064),number of branch sites/nodes is 14.400±1.179(P=0.071)(n=3);However,the parameters of 40?g/ml group showed significant statistically differences compared to control: length of tubes is 25.764±1.920mm(P=0.000),number of loops/meshes is 16.133±0.677(P=0.001),number of branch sites/nodes is 21.833±0.726(P=0.001)(n=3)? When compared to 4?g/ml group,the parameters of 40?g/ml group: length of tubes(P=0.001),number of loops/meshes(P=0.002),number of branch sites/nodes(P=0.001)showed significant statistically differences.Conclusion: Exposure to fibrinogen(40 ? g/ml)could increase the angiogenic function of EPCs,suggesting that fibrinogen may promote the angiogenic function of EPCs which is related to the concentration of fibrinogen.However,the mechanism still needs further study.