Effect of miR-34a-5p overexpression on ulcerative colitis and the intervention of berberine based on IL-6/STAT3 signaling pathway
Objective To explore the influence of microRNA-34a-5p(miR-34a-5p)overexpression on ulcerative colitis(UC)and the intervention mechanism of berberine(BBR)based on interleukin-6(IL-6)/signal transduction and activator of transcription 3(STAT3)signaling pathway.Methods HT-29 cells were cultured in vitro.HT-29 cells in the third passage,which were in logarithmic growth phase and had good growth status,were taken and randomly divided into the blank control group,NC mimics group and miR-34a-5p mimics group,respectively.Cells in the NC mimics group and miR-34a-5p mimics group were transfected with NC mimics and miR-34a-5p mimics,respectively,while cells in the blank control group were not transfected.After transfection for 6 h,we replaced the medium with the fresh medium and contin-ued to culture for 48 h.The transfection efficiency was verified by RT-qPCR.The transfected cells of the three groups were collected and cell viability was detected by CCK-8.The culture supernatants of the three groups were separated and the content of IL-6,IL-1β and TNF-α was detected by ELISA.The transfected cells of the three groups were collected,and the mRNA and protein expression levels of IL-6 and STAT3 were detected by RT-qPCR and Western blotting.The UC cell model was established by taking HT-29 cells and adding LPS for simulation of 48 h.UC cells were taken and treated with 0,2.5,5,10,20,40,80,160,320 μg/mL BBR for 24 h,and cell viability was detected by CCK-8.UC cells were ran-domly divided into the model group and BBR group.The BBR group was treated with BBR for 24 h,while the model group was not treated with BBR.The culture supernatant of the two groups was separated and the content of IL-6,IL-1β and TNF-α was detected by ELISA.The cells of the two groups were intervened for 24 h and the mRNA expression levels of miR-34a-5p,IL-6 and STAT3 were detected by RT-qPCR.Results The relative expression of miR-34a-5p was higher in the mimics group than in the NC mimics group and the blank control group(both P<0.05),but there was no significant dif-ference between the NC mimics group and the blank control group(P>0.05).The cell viability was lower in the miR-34a-5p mimics group than in the NC mimics group and blank control group(both P<0.05),but there was no significant difference between NC mimics group and blank control group(P>0.05).The levels of IL-6,IL-1β and TNF-α in culture supernatant of miR-34a-5p mimics group were lower than those of the NC mimics group and blank control group(all P<0.05),but there were no significant differences between NC mimics group and blank control group(all P>0.05).The relative expres-sion levels of IL-6,STAT3 mRNA and IL-6 protein in the miR-34a-5p mimics group were lower than those in the NC mim-ics group and blank control group(all P<0.05),but there were no statistically significant differences between the NC mim-ics group and blank control group(all P>0.05).The cell viability of UC cells treated with 40,80,160,320 μg/mL BBR for 24 h was higher than that treated with 0,2.5,5,10,20 μg/mL BBR for 24 h(all P<0.05),so 20 μg/mL BBR was selected for follow-up experiments.The levels of IL-6,IL-1β and TNF-α in the culture supernatant of BBR group were lower than those of the model group(all P<0.05).The relative expression of miR-34a-5p in BBR group was higher than that in the model group,and the relative expression levels of IL-6 and STAT3 mRNA in the BBR group were lower than those in the model group(all P<0.05).Conclusions Overexpression of miR-34a-5p can reduce the inflammatory response of UC by inhibiting IL-6/STAT3 signaling pathway.BBR can reduce UC inflammation by up-regulating miR-34a-5p expres-sion and inhibiting IL-6/STAT3 signaling pathway,delaying ulcerative colitis associated carcinogenesis.
ulcerative colitisberberinemicroRNA-34a-5pinterleukin-6/signal transduction and activator of transcription 3 signaling pathway
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