Effect of long non-coding RNA np-5318 on intestinal ischemia-reperfusion injury in rats

CHEN Jing-yu; WANG Xin-yu; ZHANG Cheng

doi:10.16048/j.issn.2095-5561.2023.05.04

Abstract：Objective To investigate the expression and role of long non-coding RNA(lncRNA)np-5318 in intestinal ischemia-reperfusion(I/R)injury,and to analyze interaction between np-5318 and epidermal growth factor receptor(EGFR)signaling pathway.Methods Twenty male SD rats were randomly divided into sham operation group(n=10)and I/R group(n=10).After the anesthesia,1.0-1.5 cm incision was made along the midline of the abdomen in the I/R group to expose the superior mesenteric artery(SMA),and the SMA was clamp with arteriolar clamp to block the blood flow.Forty-five minutes later,the artery clamp was removed through the incision to restore the blood supply,and the rat intestinal I/R injury model was established.In the sham operation group,the SMA was isolated but not clamped.The rats in the I/R group were killed 6 hours after the reperfusion,meanwhile the rats in the sham operation group were killed,and the intestinal tissues were removed immediately.The dry-wet ratio(W/D)of intestinal tract in sham operation group and I/R group was measured.Real-time quantitative polymerase chain reaction(qRT-PCR)was used to detect the expression of np-5318 in the intestinal tissues of the sham operation group and the I/R group.Enzyme linked immunosorbent assay(ELISA)was used to detect intestinal transforming growth factor-β(TGF-β),interferon-γ(INF-γ),interleukin-4(IL-4),interleukin-6(IL-6),interleukin-17A(IL-17A)and cyclooxidase-2(Cox-2)levels.The intestinal epithelial cells of rats were divid-ed into control group,I/R model group and transfected si-np-5318 group.In-vitro I/R model was constructed in the intestinal epithelial cells of transfecting si-np-5318 group and I/R model group,and si-np-5318 was transfected to inhibit the expression of lncRNA np-5318 in transtecting si-np-5318 group and corresponding control(si-NC)was transfecting in I/R model group,respectively.The cell activity was detected by MTT colorimetry,and the expression levels of np-5318 and EGFR signaling pathway related genes were de-tected by qRT-PCR.Results The W/D and expression levels of inflammatory factors TGF-β,INF-γ,IL-4,IL-6,IL-17A and Cox-2 in intestinal tissue of the I/R group were higher than those of the sham operation group,and the differences were statistically sig-nificant(P<0.05).The results of qRT-PCR showed that the expression level of np-5318 in the I/R group was(2.07±0.14),which was higher than that in the sham operation group(1.02±0.11),and the difference was statistically significant(P<0.05).MTT colorimetric assay results showed that the cell activity in the I/R model group was lower than that in the control group,while the cell activity in the transfected si-np-5318 group was higher than that in the I/R model group,with statistical significance(P<0.05).There was no significant difference in cell activity between the control group and the transfected si-np-5318 group(P>0.05).The expression levels of EGFR signaling pathway genes HBEGF,CSK and HGS in the I/R model group were significantly lower than those in the control group,while the expression levels of EGFR signaling pathway genes HBEGF,CSK and HGS in the transfect-ed si-np-5318 group were significantly higher than those in the I/R model group,and the differences were statistically significant(P<0.05).Conclusion In intestinal I/R,lncRNA np-5318 can inhibit EGFR signaling pathway,that may be the potential mecha-nism of lncRNA np-5318 aggravating intestinal I/R injury.