| AimsTo establish the rabbit model of urine-derived sepsis,exploring the effect of H2S expression on oxidative stress response and inflammatory factors of urosepsis with acute kidney injury,applying NaHS and PAG intervention,observing the expression of AOPP、ROS and SOD in renal tissues and the inflammatory factors changes of HMGB1,in order to reveals the possible mechanism of the function of H2S playing in oxidative stress and inflammatory reaction of urine-derived sepsis with acute kidney injury.Methods1 Injecting Escherichia coli（ATCC 25922）to the left ureter on rabbit and ligating the distal end of ureter with silk thread in order to copy themodel of the urogenic sepsis that is caused by acute obstruction of ureteral obstruction and infection.Divide randomly selected 40 of healthy male rabbits into five groups（eight for each group）.respectively:control group（normal control group）;sham group（sham operation group）;sepsis group（urosepsis group）;PAG group（intraperitoneal injection of DL-glycine glycine）;Na HS group（injected 8.4umol/kg NaHS）.2 Five groups of rabbits were splited into 5 observation points according to the time:preoperative,postoperative 12h,postoperative 24h,postoperative36h and postoperative 48h.The anal temperature（RT）,respiratory rate（RR）and heart rate（HR）of the five groups of rabbits were measured at each time point.3 Five groups of rabbits were splited into 5 observation points according to the time:preoperative,postoperative 12h,postoperative 24h,postoperative36h and postoperative 48h.The white blood cell count（WBC）,neutrophil count（neutrophil,N）,creatinine（Creatinine,Cr）and urea nitrogen（Blood Urea Nitrogen,BUN）of the peripheral venous blood of 5 groups of rabbits were measured at different time points.4 The renal tissues of rabbits were stained with HE after the operation of48h,and their morphology and structure were observed by light microscope.5 The expression of AOPP in renal tissues were detected by immunohistochemistry after the operation of 48h.6 The ROS、SOD and HMGB1 expression in the kidney tissues were detected by ELISA after the operation of 48h7 The expression of SOD mRNA in the kidney tissues were detected by PCR after the operation of 48h.Results:1,The vital signs:The RT,RR and HR of Sham group tended to be normal after operation 48h,as opposed to Control group the difference was not significant（P>0.05）;The RT,RR and HR of Sepsis group increased gradually with the passage of time,which was significantly higher than that of the Control group and Sham group at the same time point,the difference was statistically significant（P<0.05）;The RT、RR and HR of the PAG group at each time point after operation were significantly higher than those of the Sepsis group at the same time point after operation,the difference was statistically significant（P<0.05）;Compared with PAG group and Sepsis group,the RT,RR and HR of the NaHS group decreased after the operation,but still higher than the Control group at each time,and the difference was statistically significant（P<0.05）.2,Peripheral blood cell count:The levels of WBC and N of Sham group began to increase in 12h after operation,compared with the levels of WBC and N of Control group at the same time,the difference was statistically significant（P<0.05）;The levels of WBC and N of Sepsis group at each time point after operation were higher than those in the control group and the sham group at the same time point,the difference was statistically significant（P<0.05）;The levels of WBC and N at each time point after operation of PAG group were significantly higher than those in Sepsis group at the same time point（P<0.05）;The levels of WBC and N at each time point after operation of NaHS group is higher than that in control group and sham group at the same time point,while lower than in sepsis group and PAG group,and the difference was statistically significant（P<0.05）.3,There was no significant difference in renal function indexes（Cr,BUN）between the Control group and the Sham group（P>0.05）;The Cr and BUN of the Sepsis group at each time point after operation were significantly higher than those in the control group and the sham group at the same time points,the difference was statistically significant（P<0.05）;The Cr and BUN of the PAG group at each time point after operation were higher than that of the Sepsis group at the same time point,andincreased significantly（P<0.05）;while the Cr and BUN at each time point after operation of NaHS group is betterer than that in control group and sham group at the same time points,the difference was statistically significant（P<0.05）.4,The results of HE staining:there was no obvious abnormality of the structural form of kidney morphology in control group and sham group;The structural form of kidney morphology in Sepsis group arranged disorderly,glomerular deformation,renal capsule expansion,renal capsule dilated,renal interstitial hyperemia and swelling,and Surrounded by a large number of Inflammatory cells;The pathological changes of PAG group were more serious than that of Sepsis group;Compared with Sepsis group and PAG group,the pathological changes of NaHS group were alleviated.5,The expression of AOPP in renal tissue after the operation of 48h:The kidneys of Control group and Sham group had no obvious staining,renal glomerulus and interstitium of Sepsis group can be seen brown granulars;PAG group compared with Sepsis group,the brown granulars of glomerular and renal interstitial appeared significantly increased,while NaHS group compared with Sepsis group and PAG group,the brown granules decreased.6,The levels of ROS、SOD and HMGB1 in renal tissue after the operation of48h:Compared to the Control group,the expression levels of ROS、SOD and HMGB1 in the Sham group after the operation of 48h were statistically significant（P>0.05）.The expression levels of ROS and HMGB1 in the Sepsis group after the operation of 48h were increased compared with those in the Control group;The expression levels of ROS and HMGB1 in the PAG group after the operation of 48h compared to Sepsis group,its rises is more significant,while Compared with Sepsis group and PAG group,the expression level of ROS and HMGB1 in NaHS group after the operation of48hdecreased,andthedifferencewasstatisticallysignificant（P<0.05）.Compared with Control group and Sham group,the activity of SOD in Sepsis group after the operation of 48h was decreased,and the difference was statistically significant（P<0.05）;The activity of SOD in PAG group after the operation of 48h was lower than the Sepsis group,while the activity of SOD after the operation of 48h was increased in the NaHS group,and the difference was statistically significant（P<0.05）.7,The expression of SOD mRNA in renal tissue after the operation of48h:There was no significant difference in the expression of SODmRNA between Sham group and Control group after the operation of 48h（P<0.05）;The expression of SODmRNA in renal tissue of Sepsis group was lower than that in group Control;Compared with Sepsis group,the expression of SODmRNA in renal tissue of PAG group decreased more significantly,and the difference was statistically significant（P<0.05）;While the expression of SODmRNA in renal tissue of NaHS group was higher than that in Sepsis group and PAG group,but still lower than that in Control group,which was statistically significant（P<0.05）.Conclusions:1、The rabbit urospsis model can be copied successfully by ligaturing the left ureter and injecting the Escherichia coli.2、PAG can aggravate the urine-derived sepsis with acute kidney injury,the use of NaHS to increase exogenous H2S can reduce oxidative stress and inflammatory response in urine-derived sepsis with acute kidney injury3、H2S plays a protective role in oxidative stress and inflammation of urine-derived sepsis with acute kidney injury.The mechanisms of actions may be attributed to its modulating effects that reduced the level of AOPP,downregulated ROS activity,increased SOD activity,and inhibited the inflammatory factors of HMGB1. |
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