| Acute kidney injury (AKI) is a common clinical syndrome as well as a kind ofcomplication in critically-ill patients, if the AKI develops into severe acute renal failure(ARF), the mortality rate will reach up to50%, and10%-14%of patients with acute renalfailure need for lifelong renal replacement therapy since the renal function cannot berestored. Ischemia-reperfusion injury is the one of the most important causes of acutekidney failure, which often occurs after the kidney transplantation, cardiopulmonary bypasssurgery and the context of trauma, hemorrhagic shock and sepsis etc. Currently, there is noother effective method except renal replacement treatment. So the illustration of ischemicAKI mechanism and the exploration of effective preventive and treatment measures havescientific significance in reducing the incidence and mortality of AKI.It is so sensitive to ischemia, toxic and other injury factors for renal tubular epithelialcells (RTECs). After kidney injury, RTECs have to be de-differentiation, the cytoskeletonand polarity have been destroyed, the epithelial cell markers, such as E-cadherin, α-Catenin,APQ1and THP, have been disappeared, the mesenchymal cell marker appeared; withoutinjury factors, the de-differentiated cells proliferate, re-differentiate, tubules reconstruct,and renal function has been recovered. It is known that some key factors for embryonicmetanephros development, such as NCAM1, Pax2, play important roles during thereparation of AKI.Trps1is one of atypical members of GATA transcription factors. It plays important rolefor embryonic metanephros development. The size of Trps1-/-mice kidney is normal, butdifferentiation markers (Pax2and Wt1) were lost. Gai's research showed that, on day7and14after UUO or sham-operation (sham) in wild-type (WT) and heterozygous Trps1-null(HT) mice, the obstructed kidneys showed tubular dilation, atrophy, a widenedinterstitial space, and a greater number of interstitial cells in WT and HT mice. And the degree of these changes was greater in HT mice than in WT mice. A new research alsoshowed that, Trps1could suppress EMT through suppressing ZEB2gene.Therefore, we hypothesize that Trps1also accelerate the repair of acute kidney injuryby promoting impaired RTECs (mesenchymal-like cells) re-differentiation (MET).I. Methods1. Injury servity decides kidney repair and prognosis.Kidney I/R model was established by clipping bilateral renal pedicles by artery clampafter isolating them from surrounding tissues for45min or60min. On1,3,7,14and21days after kidney I/R, SCr and BUN were detected, renal injury was observed under a lightmicroscope.2. The relationship between Trsp1and kidney repair after acute ischemia andreperfusion injury.Kidney I/R model was established by clipping bilateral renal pedicles by artery clampafter isolating them from surrounding tissues for45min or60min. On1,3,7,14and21 days after kidney I/R, Trps1was measured by immunohistochemistry. By using correlationanalysis, the relationship between Trps1expression and injury score were showed. Theactivity of Na+-K+ATPase was tested by Na+-K+ATPase assay kits.3. The effect of over-expression-Trps1in kidney repair of I/R injury.A Trps1-over-expression rat model was contructed, in which theultrasound-microbubble-mediated adenovirus transfer technique has been applied. And thenkidney I/R models were established on over-expression-Trps1rats. The rats divided intothree groups, control group, GFP-vector group, GFP-Trps1group. On1,3,7,14and21days after kidney I/R, SCr and BUN were detected, renal injury was observed under a lightmicroscope, and Trps1was measured by immunohistochemistry.4. The relationship between Trps1expression and RTEC re-differentiation.The expression of Trps1, α-Catenin and vimentin was measured byimmunohistochemistry and western blotting.II. Results1. Injury servity decides kidney repair and prognosis.1.1The changes of renal function.Renal function: compared with the control group, in I/R45min group, BUN and SCrconcentrations gradually increased, reaching a peak at1day after ischemic-reperfusion, andthen decayed. Both the two indexes in I/R60min group were higher than I/R45min groupon day1,3and7, reaching a peak at3days after ischemic-reperfusion, and then decayed.1.2The changes of renal pathology.Pathological changes: under the light microscope, tissue damages induced by renalischemia-reperfusion are mainly embodied through necrosis of renal tubular epithelial cellsand the cell tube formation, exposed basement membrane, interstitial edema, mononuclearcell infiltration. As for the lesion, the medulla is heavier than the cortex; the junction part isthe most important leather cord. The changes in injury severity and is related withischemia-reperfusion time. In I/R45min group, the worst time point was day1. Thejunction of renal tubular epithelial cell swelled, vacuolar degenerated and occasionallyrenal tubular epithelial cell abscised. Then the renal tissue began to be repaired on day3. InI/R60min group, the renal tissue had been destroyed on day1, but the worst time point was day3. Then the renal tissue began to be repaired on day7.2. The relationship between Trsp1and kidney repair of acute ischemia and reperfusioninjury.2.1The expression of Trps1In control group, trps1located on cortex tubules, the score was338±27. In I/R45mingroup, Trps1expression reach a peak on day1, and the score was109±42; on day3, theTrps1expression began to increase. In I/R60min group, Trps1expression decreased on day1and reach a peak on day3, the score was20±4; then the Trps1expression began toincrease on day7.2.2The relationship between Trps1expression and injury score.Immunohistochemistry analysis showed that in I/R45min group, Trps1expressiondecreased during the phase of injury, reaching a valley at day1, then rose during the phaseof repair. But in I/R60min group, Trps1expression was lower than I/R45min group at eachtime point, reaching a valley at day3. The expression trend of Trps1was opposite withinjury severity. Even in the same renal tissue, the tissue with high Trps1expression waswell repaired, while the tissue with low Trps1expression was not.2.3The relationship between Trps1expression and the activity of Na+-K+ATPase.The activity of Na+-K+-ATPase also decreased reaching a valley firstly, then increasedaccordingly with the reparation of injury both in I/R45min and I/R60min. But in I/R45min, the activity reach a valley on day1while in I/R60min on day3. Meanwhile, theactivity of Na+-K+-ATPase was lower in I/R60min group than I/R45min group on day3,7,14and21.3. The effect of over-expression-Trsp1in kidney repair of I/R injuryCompared with GFP-vector group, on1,3,7days, the levels of SCr and BUN weresignificantly lower, the PAS stain showed the lesions were lighter, and the Trps1expressionwere significantly higher.4. The relationship between Trps1expression and RTEC re-differentiation.It is observed that the α-Catenin was decreased during the phase of injury, andincreased again along with Trps1during the phase of repair in I/R60min group, whilevimentin was the opposite. On day3, Trps1and α-Catenin were disappeared and vimentincame to appear; then the expression of Trps1and α-Catenin incrased and vimentin expression decreased. On day21, the expression of Trps1and α-Catenin almost was equalto it in control group, while vimentin expression reaching a valley than in other time point.After Trps1over expressing, western blotting also showed the expression of Trps1andα-Catenin were higher in GFP-Trps1group than in GFP-vector group.III. ConclusionThese data show that, in I/R model, the Trps1is down-regulated during the phase ofkidney injury, and up-regulated during the phase of repair. The expression of Trps1iscorrelated with epithelial cell phenotype, indicate that Trps1may plays a positive role in thekidney repair.
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