The Effect Of Arachidonic Acid On Proliferation Of Human Hypertrophic Scar Fibroblasts

Objective:To observe the effects of arachidonic acid on human hypertrophic scar(HS) fibroblasts(FB) proliferation in vitro and to investigate the possible mechanism of arachidonic acid(AA)preventing hypertrophic scar fibroblasts proliferation. Methods:(1) HS tissue taken from hospital patients of burn surgery. Fibroblasts were cultured with modified trypsin digestion in vitro, the fourth generation cells were used for the experiment.(2)The fibroblasts were treated with 0μmol/L、5μmol/L、10μmol/L、20μmol/L、40μmol/L、80μmol/L concentrations of AA in 10% FBS culture medium for a certain time, then to observe the morphological changes of the fibroblasts.(3)The fibroblasts were treated with 0μmol/L、5μmol/L、10μmol/L、20μmol/L、40μmol/L、80μmol/L concentrations of AA in 10% FBS culture medium for 24h、48h and 72 h respectively, then the cell toxicity of arachidonic acid was assessed by CCK.(4)The cultured cells were treated with AA at the above concentrations for 24 h, the expression of Ki-67 protein was detected by immunocytochemical method. Results:(1) From the HS tissue blocks inoculated sixth days the cells begin to swim out of. Cells were single scattered and long fusiform.The 10 th day tissue surrounding cells increased significantly, about 15 th day cells around the tissue blocks growth intensive, begin to integrate. About 7 days after cell passage, cells covered the bottom of the culture bottle. The cells showed the brush, the swirling or radial growth,cells were long fusiform, surrounding cells showing different lengths of neurites, and the cells arranged in neat rules.(2) In the AA effect, the morphology of the fibroblasts was changed. When the concentration were 5μmol/L and 10μmol/L, An increase in cell number, cell morphology and growth trend was not obvious affected by AA. When the concentrations of AA were 20μmol/L,40μmol/Land 80μmol/L,the shape of the FB began to exhibit irregular, cell size is not uniform, the shape changed,part of the cells were round blunt, the cells number reduced and nucleus condensation. The AA concentration is high, change the FB of a more obvious.(3) FB at 5μmol/L and 10μmol/L of AA under the action, the survival rate of FB was not affected(p>0.05); In the 20-80μmol/L AA concentration range, the inhibition rate of fibroblasts gradually increased with increasing concentration and duration of AA(P<0.05,and comparison between groups P < 0.01).(4) When the concentrations of AA were 20μmol/L、40μmol/L and 80μmol/L,the percentages of the fibroblasts that had the positive Ki-67 antigen were significantly smaller than that of the control(P<0.05),and the concentration more higher,the percentage of the fibroblasts that had the positive Ki-67 antigen more smaller(comparison between groups P<0.01).When the concentration were 5μmol/L and 10μmol/L, the percentage of Ki-67 protein positive cells less than AA concentration of 0μmol/L, but not significantly different from that of the control(P>0.05). Conclusion:A certain concentration of AA can inhibit the proliferation of human HSFb, then the number of mitotic cells decreased, which may be one of the mechanisms for the inhibition of hypertrophic scar.