Effect Of Hyperbaric Oxygenation On The Expression Of IRE1α-ASK1-JNK Pathway Proteins Of Ischemia-reperfused Renal Tissue In Mice

Objective: A mouse model of renal ischemia-reperfusion injury(IRI)was constructed,and the renal function and renal tissue of the mice in the sham-operated group,the ischemia-reperfusion(IR)group,and the hyperbaric oxygenation(HBO)treatment group were observed.Pathology,endoplasmic reticulum stress IRE1α-ASK1-JNK pathway protein expression,and renal tubular epithelial cell apoptosis rate changes,to explore the effect of HBO treatment on endoplasmic reticulum stress IRE1α-ASK1-JNK pathway-related protein expression in renal IRI influence and regulation.Methods: The C57BL/6 mice were randomly separated into three groups,namely the sham operation group,the renal IRI group,and the renal IRI+HBO treatment group(HBO treatment group).Each group was divided into four subgroups according to reperfusion time: 1h,3h,6h,and 12 h,with 6 mice in each subgroup.In the sham operation group,only an abdominal incision was performed,bilateral renal pedicles were not blocked,and blood was collected from bilateral kidneys and heart at four points.In the renal IRI group:the abdominal cavity was incised,and the bilateral renal pedicles were blocked for 45 minutes and then opened.The bilateral kidneys and heart were collected for blood collection at four points after reperfusion.In the renal IRI+HBO treatment group: an abdominal incision was performed,and the bilateral renal pedicles of mice were stopped for 45 minutes before blood flow was perfused,and then the bilateral kidneys and heart were collected for blood collection.The serum creatinine(SCr)value of mice was detected through the instrument,and the pathological changes in mouse kidney tissue were observed by hematoxylin-eosin staining.The expression changes of IRE1α,ASK1,JNK m RNA,and protein were detected by immunohistochemistry,RT-q PCR,and Western blot.The number of apoptosis cells and apoptosis rate of renal tubular epithelial cells were detected by TUNEL assay.Results: 1.Changes in renal function: The SCr value at four points in the renal IRI group was significantly higher than that in the sham operation group(P<0.05),and with the increase in reperfusion time,the SCr value also increased.Compared with the renal IRI group,the SCr value at four points after HBO treatment was lower(P<0.05).2.Histopathological changes in the kidney: There were no pathological changes in renal histopathology at each time point in the sham operation group.In the renal IRI group,glomerular volume increased,tubular epithelial cell swelling and interstitial edema were observed at each time point,and the increase was increased with the prolongation of reperfusion time.Bleeding;after HBO treatment,the above injury was alleviated compared with the renal IRI group,and the effect of reperfusion for 1 h was the most obvious.3.Results of cell apoptosis in renal tissue: TUNEL-positive cells were rare in the sham-operated group;in the renal IRI group,the number of TUNEL-positive cells at each time point increased significantly and was higher than that in the sham-operated group(P<0.05);The number of TUNEL-positive cells at each time point was lower than that in the renal IRI group(P<0.05).4.Immunohistochemical results: The positive expression of IRE1α and ASK1 proteins were mainly located in the cytoplasm of renal tubular epithelial cells,while JNK protein was expressed in the cytoplasm and nucleus of renal tubular epithelial cells,and the positive expression showed yellow or brown granular changes.The expression changes of IRE1α,ASK1 and JNK proteins in renal tissue were the same.The results of the relative quantitative analysis showed that the expressions of IRE1α,ASK1 and JNK proteins at each time point were significantly higher than those in the sham-operated group(P<0.05).The expression levels of the three proteins were lower than those in the renal IRI group(P<0.05).5.RT-q PCR results: IRE1α,ASK1,and JNK m RNA expression changes in kidney tissue were the same: in the renal IRI group,the expression levels of the three protein m RNAs at each time point were higher than those in the sham operation group(P<0.05).The expression levels of the three proteins were significantly decreased after HBO treatment(P<0.05);in the renal IRI group and the HBO treatment group,the m RNA expression levels of the three proteins increased with the prolongation of reperfusion time,and there was a statistical difference between the two points.Scientific significance(P<0.05),reached the peak at 12 h after reperfusion.6.Western blot results: In the renal IRI group,the expression levels of IRE1α,ASK1,and JNK proteins at each time point were significantly higher than those in the sham operation group(P<0.05),and their expression levels decreased after HBO treatment(P<0.05).There were significant differences in the expression levels of ASK1 and JNK proteins at each time point in the renal IRI group and the HBO treatment group(P<0.05);in the renal IRI group,the protein expression levels of IRE1α at each time point were statistically significant(P<0.05),while there was no significant difference between 1h and 3h in the HBO treatment group.7.The results of Pearson correlation analysis showed that the increased protein expression levels of IRE1α,ASK1,and JNK in the endoplasmic reticulum stress pathway after renal IRI were significantly positively correlated with serum creatinine and apoptosis rate,and the correlation coefficient after HBO treatment was higher than that in the renal IRI group.decreased,but still showed a significant positive correlation.Conclusions: 1.Ischemia-reperfusion injury can induce the overexpression of related apoptosis proteins in the IRE1α-ASK1-JNK pathway in renal tissue,promote tissue cell apoptosis,and aggravate renal injury.2.Hyperbaric oxygen therapy can play a protective role in renal IRI by inhibiting the overexpression of IRE1α,ASK1,and JNK proteins in endoplasmic reticulum stress.