To Investigate The Mechanism Of MAPK Pathway In The Inhibition Of Psoriatic Cell Growth By Resveratrol

Objective: To observe the effect of resveratrol on the growth of psoriasis-HaCaT cell model and to explore its mechanism of action.Methods: 1.the establishment of psoriasis-HaCaT cell model: Stimulate human immortalized keratinocytes(HaCaT)cells with keratinocyte growth factor(KGF)at concentrations of 0,5,10,20,40 ng/ml,observe the growth of HaCaT cells with a microscope every day,record the period of good cell growth within a week,and use the thiazole blue colorimetric(MTT)method to detect the HaCaT stimulated by KGF at the concentration of 0,5,10,20,and 40ng/ml Cell proliferation in this period of time.Observe the influence of KGF on the proliferation ability of HaCaT cells,select the optimal stimulation concentration and time of KGF,and establish a psoriasis cell model.2.Determination of resveratrol IC50: Stimulate HaCaT cells with resveratrol at a concentration of 0,0.5,1,2,4 μmol/L,detect the OD value by the MTT method,calculate the IC50,and then select the resveratrol stimulation concentration for subsequent experiments.3.The effect of resveratrol on psoriasis-HaCaT cell model: After the psoriasis-HaCaT cell model is successfully established,set up a blank control group,a negative control group(normal saline),a low-concentration resveratrol group,a medium-concentration resveratrol group,a high-concentration resveratrol group,Positive control group(Tretinoin).MTT method and flow cytometry to detect the effects of different concentrations of resveratrol on the proliferation and apoptosis of the psoriasis-HaCaT cell model.And Western-Blot detects the protein expression of Caspase3,MAPK/P-ERK1/2,and MAPK/P-P38.4.Statistical processing: Graphad software and SPSS22.0 statistical software are used in data processing.The calculated data are expressed as mean±standard deviation((?)±s),and the difference between groups is compared by the method of one-way analysis of variance or repeated measures design of analysis of variance,the difference is statistically significant at P<0.05.Results: 1.Microscope results and MTT method test results show: Observed under a microscope,under the stimulation of KGF,the number and density of HaCaT cells increased on the 1st and 2nd d.The cells were round and long fusiform,densely distributed,and the cells grew well,while the cells grew well from the 3rd to the 7th day Poor growth.The MTT test results showed that the OD value of the KGF concentration 40ng/ml group was significantly greater than that of the other concentration groups from 24 h,and the difference between the groups was statistically significant(P<0.05).Therefore,starting with KGF stimulation at a concentration of 40ng/ml for 24 hours,HaCaT cells were used as a psoriasis cell model when the growth of HaCaT cells were vigorous,and the model was successfully established.2.The results of MTT method showed that resveratrol can inhibit the proliferation of HaCaT cells,and the calculated IC50 value is about 5.3μmol/L.Therefore,the stimulation concentration of resveratrol in subsequent experiments was set to 2.5 μmol/L,5 μmol/L,and7.5 μmol/L,respectively.3.After resveratrol intervenes in the psoriasis-HaCaT cell model,the results of MTT assay show: Starting from 48 h,compared with the negative control group,the resveratrol in the middle and high concentration groups has a certain inhibitory effect on the proliferation of the cells of the psoriasis-HaCaT cell model,and its inhibitory effect was consistent with the positive control group(P<0.05)).The results of flow cytometry assay showed: Compared with the negative control group,the middle and high concentration groups of resveratrol have a pro-apoptotic effect on the cells of the psoriasis-HaCaT cell model.The pro-apoptotic effect of the high concentration group of resveratrol is the same as that of the positive control group.(P<0.05).The Western-Blot test results showed that,compared with the negative control group,the expression of Caspase3 protein in the middle and high resveratrol groups increased,while the expression of P-ERK1/2 and P-P38 decreased(P<0.05).Conclusions: Resveratrol can inhibit the proliferation of psoriasis cells and promote their apoptosis.The possible mechanism is achieved by inhibiting the MAPK/ERK and/or MAPK/P38 pathways.