Experimental Study On The Effect Of Luteolin On LPS-induced HaCaT Cells Of Proliferation,Apoptosis And Related Inflammatory Mediators

Psoriasis is an autoimmune skin disease characterized by abnormal proliferation of epidermal keratinocytes and infiltration of various inflammatory cells in the epidermis and dermis.Luteolin is a kind of bioflavonoids.This pure yellow crystal is widely found in fruits,vegetables and herbs.The known pharmacological effects of luteolin include inhibiting proliferation of cells,promoting death of cells,antiinflammation,antioxidant,anti-tumor,etc.It has been reported that the luteolin has therapeutic effect on psoriasis.Objective: This study established a psoriasis-like cell model by stimulating HaCaT cells with LPS,and explored the effects of luteolin on HaCaT cell proliferation,apoptosis regulation,and related inflammatory factors.Methods:1.The CCK-8 method was used to detect the survival rate of HaCaT cells after stimulation with different concentrations of luteolin(0,1,10,25,50,100?mol/L)at different times(24 h,48 h,72 h).Based on the above results,I formulate a reasonable luteolin stimulation protocal(10?mol/L,24 h).2.The CCK-8 method was used to detect the survival rate of HaCaT cells after stimulation with different concentrations of LPS(0,0.1,1,10,20?g/m L)for 24 h.The protein and m RNA expression of inflammatory mediators NF-?B P65 and IL-6 were detected by western blot and RT-PCR.Based on the above results,I choose a reasonable lipopolysaccharide stimulation concentration(20?g/m L)to establish the psoriasis cell model.3.LPS was used to stimulate HaCaT cells in the luteolin pre-stimulation group and control group,respectively,and the protein and m RNA expression of NF-?B P65 and IL-6 were detected by western blot and RT-PCR.4.Flow cytometry was used to detect the apoptosis of HaCaT cells in the luteolin pre-stimulation group and control group.5.The protein expressions of IL-17,IL-23 and TNF-? which associated with psoriatic in the medium of both pre-stimulation group and control group were detected by ELISA.Results: 1.With the increase of concentration and time of luteolin stimulation in HaCaT cells,luteolin showed different inhibitory effects on cells,and the differences were statistically significant(P<0.05);2.LPS stimulates the proliferation of HaCaT cells,and up-regulated expression of inflammatory mediators NF-?B P65 and IL-6,which is proportional to the concentration.Then induces psoriatic dermatitis-like changes in HaCaT cells;3.Compared with the control group,The protein and m RNA expression of inflammatory mediators NF-?B P65 and IL-6 decreased in the Pre-stimulation group,and the differences were statistically significant(P < 0.05);4.Compared with the blank group,luteolin Pre-stimulation group induced apoptosis of HaCaT cells,and the differences were statistically significant(P<0.05);5.Compared with the blank group,luteolin Pre-stimulation group inhibited the release of inflammatory mediators IL-17,IL-23 and TNF-?,which are related to LPS-induced psoriasis in HaCaT cells,and the differences were statistically significant(P<0.05).Conclusion:Luteolin inhibited the proliferation of HaCaT cells and can effectively down-regulate the expression of LPS-induced inflammatory mediators in HaCaT cells,including NF-?B P65,IL-6,TNF-?,IL-17,IL-23.In addition,luteolin also can induce apoptosis of HaCaT cells.