Experimental study on the protective effect of butyrate on inflammatory bowel disease
Objective:To explore the protective effect of butyrate(BT)in inflammatory bowel disease(IBD)cell model.Methods:Hu-man normal colon epithelial cells(FHC)were cultured and induced with lipopolysaccharide(LPS)to construct an inflammatory bowel disease cell model.The cells were divided into the following groups:control group,inflammation group,0.25 mmol/L BT+LPS group,0.5 mmol/L BT+LPS group,and 1 mmol/L BT+LPS group.The control group was cultured normally without the need for additional reagents,while the inflammation group was stimulated with LPS at a concentration of 100 g/mL for 24 hours to stimulate FHC cells.Dif-ferent BT+LPS groups were treated with corresponding concentrations(0.25,0.5,1 mmol/L)of BT to intervene in FHC cells for 24 hours,followed by stimulation with LPS at a concentration of 100 μg/mL for 24 hours.After treating FHC cells with BT at different concentra-tions of 0,0.5,1,2 and 5 mmol/L for 24 and 48 hours,the concentration range of BT was determined by cell viability detection.Then,0.25,0.5 and 1 mmol/L BT solutions were used to intervene in FHC cells for 24 hours.100 g/mL LPS was added and treated for 24 hours.RT-qPCR and ELISA were used to detect the levels of COX-2,IL-6 mRNA and protein in the control group,inflammation group,0.25 mmol/L BT+LPS group,0.5 mmol/L BT+LPS group,and 1 mmol/L BT+LPS group,respectively,to clarify the protective ef-fect of BT on FHC cells and its concentration relationship.Finally,the microstructural changes of cells in the control group,inflamma-tion group,and 1 mmol/L BT+LPS group were observed under electron microscopy.Results:When the concentration of BT was below 1 mmol/L,the viability of FHC cells was above 90％,and there was no significant change in cell viability after 24 and 48 hours of treat-ment.The RT-qPCR results showed that compared with the control group,COX-2 mRNA and IL-6 mRNA increased(both P<0.001)in the inflammation group.Compared with the inflammation group,the 1 mmol/L BT+LPS group showed a decrease in COX-2 mRNA and IL-6 mRNA(t=3.384,4.722,both P<0.05).The levels of COX-2 mRNA and IL-6 mRNA decreased in the 0.5 mmol/L BT+LPS group(t=2.817,3.753,both P<0.05).There was no statistically significant difference in the levels of COX-2 mRNA and IL-6 mRNA in the 0.25 mmol/LBT+LPS group(both P>0.05).Compared with the 0.5 mmol/L BT+LPS group,the 1 mmol/L BT+LPS group showed statistically significant differences in COX-2 mRNA and IL-6 mRNA(t=4.561,5.196,both P<0.05).The ELISA results showed that compared with the control group,the expression levels of COX-2 and IL-6 increased in the inflammatory group(both P<0.001).Compared with the inflammation group,the levels of COX-2 and IL-6 were reduced in the 1 mmol/L BT+LPS group(t=4.547,3.452,both P<0.001).The levels of COX-2 and IL-6 decreased in the 0.5 mmol/L BT+LPS group(t=2.927,3.265,both P<0.05).There was no statistically significant difference in the levels of COX-2 and IL-6 in the 0.25 mmol/L BT+LPS group(both P>0.05).Compared with the 0.5 mmol/L BT+LPS group,the 1 mmol/L BT+LPS group showed statistically significant differences in IL-6 and COX-2 levels(t=4.674,3.217,both P<0.05).Under electron microscopy,the control group of FHC cells showed normal and intact microstructures.In the experi-mental group,the microvilli of FHC cells became sparse,and the tight junction structure was disrupted.In the 1 mmol/L BT+LPS group,the microvilli of FHC cells showed partial shedding,and the damage to the tight junction structure was reduced.Conclusion:The pro-tective effect of BT on inflammatory bowel disease cell models increases with increasing concentration when the concentration is below 1 mmol/L.
万方数据
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