| Donor after cardiac death(DCD)is increasingly important in transplantation,it plays a very important role to alleviate the shortage of organs.However,due to warm ischemia process of the donor organ(i.e.kidney)after cardiac death,the internal environment homeostasis was break and the quality of donor kidney decreased,which results poor kidney transplantation prognosis.therefore,how to reduce the renal warm ischemia injury has become a research hot spots.In this study,we improved the classic method of renal warm ischemia injury in rats,and successfully established a simple model of renal warm injury in rats,which can simulate renal warm ischemia injury on DCD donor.Then,liposomal prostaglandin E1(Lipo-PGE1)was added in hypertonic citrate adenine solution(HCA),and mixed liquid in low temperature was perfused in rat kidney for 2hours.We observed the pathomorphology and detected the molecular expression,to verify the protective effect of Lipo-PGE1 continuous hypothermic perfusion on renal warm ischemia injury in rats.finally,we detect the indicators related to oxidative stress to explore protection mechanism of Lipo-PGE1.This paper is divided into two parts.PART I Establishment of renal warm ischemic injury model of SD ratsOBJECTIVE: To establish a reliable model of kidney warm ischemia in rat with improved methods,paving way for the next stage of kidney transplantation after warm ischemia in rats,which simulate the phenomenon of warm ischemic injury of kidney after cardiac death in clinical.METHODS: The rats have the anatomical characteristics that right kidney in SD rats were higher than left.so the model of left renal warm ischemic injury was realized by occlusion of the abdominal aorta by clamping between the renal arteries using non-invasive micro-vascular clamp.Blood supply of right kidney is not affected.After detachment of left kidney,another micro-vascular clamp is placed in the abdominal aorta above right renal artery,which should occlude the blood supply of right kidney,achieving the purpose of warm ischemic injury of right kidney.After kidney warm ischemia model was established,intubation was perfused in the abdominal aorta of the rats,and the color of the kidney was observed by the naked eye.Results: In the first stage,after 30 cases of training,rat kidney warm ischemic injury mode could be established stably.In the second stage,The average operation time was 90 ± 20 minutes and the success rate was 78%(31/40),which was better than the first stage whose average operation time was 115 ± 25 min,success rate of operation was 43%(13/30).The appearance of the kidney in the second stage SD rats after renal thermal ischemic injury was purple.After perfusion,the rats could achieve full yellow-brown.Conclusion: Our improved method can achieve reliable warm ischemic injury in rats,which also has the advantages of simple and easy to practice,high success rates.It is a good warm ischemia injury model in SD rats.Part II The protective effect of Lipo-PGE1 on renal warm ischemia injury in SD ratsOBJECTIVE: To investigate the protective effect of continuous hypothermia perfusion of liposomal prostaglandin E1(Lipo-PGE1)in hyperosmotic citrate purine(HCA)on renal warm ischemia compared to HCA perfusion solution only.Methods: The left kidney and the right kidney of each rat were randomly divided into A and B groups.Group A: perfusion fluid was Lipo-PGE1 + HCA.Group B:perfusion solution was Na Cl + HCA only.The kidneys were hypothermia infused with the indicated solutions for 2 hours.Then kidneys were harvested and examined by: 1)HE staining followed by pathological examination;2)Malondialdehyde(MDA)and superoxide dismutase(SOD)were detected by spectrophotometry;3)expression of KIM-1,IL-8 were detected by Western blotting.Results: Compared with the control group(group B),pathologic results showed that the degree of necrosis was significantly reduced in experimental group(group A).In experiment group(group A),the brush border of the renal tubules was relatively intact,the epithelial cells were relatively intact,infiltration of inflammatory cells was significantly reduced,micro-thrombus was hardly seen.The activity of SOD in the experimental group was 652.59 ± 52.31 U / mg,and itwas nearly 3 times of control group,which is 237.86 ± 19.95 U / mg.This is statistically significant difference checked by paired t test.MDA concentration in the experimental group,the control group were 5.82 ± 0.83,12.53 ± 1.61 nmol/mg respectively,and the difference was statistically significant.The results of western blot showed that expression of KIM-1 and IL-8 were decreased in the experimental group compared with control group.Conclusion: 1.Low temperature perfusion with Lipo-PGE1 in HCA solution can effectively alleviate the renal injury caused by warm ischemia for a long time,and improve the quality of renal preservation.2.Low temperature perfusion with Lipo-PGE1 in HCA solution may relieve the renal injury induced by reactive oxygen species,and thus prevent in kidney from warm ischemia injury. |
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