Effect Of Colchicine On Human Keloid Fibroblasts

PART ONE CULTIVATION OF PRIMARY HUMAN KELOID-DERIVED FIBROBLAST AND THE INFLUENCE OF COLCHICINE ON PROLIFERATION AND CELL CYCLE OF KELOID FIBROBLASTSObjective: To culture the primary human keloid-derived fibroblast in vitro and investigate the regulating effects of colchicine on proliferation and cell cycle of keloid fibroblasts.Methods: The primary human keloid-derived fibroblast were cultured by tissue. The proliferative behavior of KFB was studied after pretreating with various concentrations of colchicines (1μg/ml, 2μg/ml, 4μg/ml, 8μg/ml, 16μg/ml, 32μg/ml) by 4 methyl thiazolyl tetrazolium method and the cell cycle was assessed by flow cytometry.Results: The primary human keloid-derived fibroblast was cultivated successfully; Not only was the proliferation of KFB inhibited,but also the prophase of cell division (G2) affected depending on the time and drug concentration.Conclusion: The primary human keloid-derived fibroblast should be cultured by tissue adherent culture method and colchicine may regulate the proliferation and prophase of cell division of KFB at a certain concentration.PART TWO EFFECT OF COLCHICINE ON HUMAN KELOID FIBROBLASTS MATRIX METALLOPROTEINASEObjective: To investigate the effects of colchicine on the matrix metalloproteinase-1 (MMP-1), matrix metalloproteinase-2 and matrix metalloproteinase-9 (MMP-9) in human keloid fibroblasts.Methods: Various concentrations of colchicine (2μg/ml, 4μg/ml, 8μg/ml, 16μg/ml) were used to pretreat human keloid fibroblasts. The expression of MMP-1, MMP-2 and MMP-9 mRNA in the cells was studied by RT-PCR, and their protein expression was assessed by Western blot.Results: The expression of mRNA and protein level were significantly increased in colchicine group as compared with control group. The expression of MMP-1 mRNA were 0.282±0.015, 0.369±0.022, 0.541±0.027, 0.672±0.020, compared with control group (0.174±0.016) were increased (P<0.05); The expression of MMP-2 mRNA were 0.371±0.026, 0.548±0.031, 0.653±0.019, 0.796±0.024, compared with control group (0.201±0.033) were increased (P<0.05), and depend on the drug concentration. The expression of MMP-1 protein were 0.205±0.013, 0.293±0.027, 0.349±0.034, 0.512±0.035, compared with control group (0.127±0.031) were increased (P<0.05); The expression of MMP-2 protein were 0.395±0.036, 0.526±0025, 0.673±0.014, 0.792±0.037, compared with control group (0.278±0.023) were increased (P<0.05), depended on the drug concentration. But there was no expression of MMP-9 in colchicine group.Conclusion: Colchicine increased the expression of MMP-1 and MMP-2, and also increased the degradation of collagen in keloid, may participate the process of the formation of keloid.