T cells as mediators in renal ischemia/reperfusion injury

Inflammation has been established to contribute substantially to the pathogenesis of ischemia/reperfusion (I/R) with a central role for particular cells, adhesion molecules, and cytokines. Until recently, most of the research trying to unravel the pathogenesis of I/R injury has been focused on the role of neutrophils. However, recent studies have brought evidence that T cells and macrophages are also important leukocyte mediators of renal and extrarenal (liver) I/R injury. In vivo depletion of CD4+ cells but not CD8+ cells in wild-type mice was protective in I/R of the kidney. A marked preservation of liver function was also found after I/R in T-cell deficient athymic mice. Blocking the b130/CD28 costimulatory pathway by CTLA-4 Ig (recombinant fusion protein) ameliorated renal dysfunction and decreased mononuclear cell infiltration in I/R of the kidney. b130-1 expression was found limited to the membrane of the endothelial cells of the ascending vasa recta, resulting in trapping of CD28-expressing CD4 T cells. This trapping of leukocytes results in the upstream congestion in the ascending arterial vasa recta, generating the since more than 150 years described medullary vascular congestion of the kidney soon after ischemic injury. It seems worthwhile to study a combination therapy using anti-inflammatory/anti-adhesion molecules in the early phase of I/R.     

共刺激分子B7 mRNA在大鼠热缺血再灌注损伤肾组织中的表达

目的：进一步验证Ｂ７－ＣＤ２８共刺激通路在肾缺血再灌注损伤中的作用。方法：在大鼠单肾热缺血再灌注损伤模型的基础上,将６０只雄性ＳＤ大鼠均分为假手术组,缺血３０ｍｉｎ再灌注组和缺血６０ｍｉｎ再灌注组,利用多聚酶链反应（ＲＴ－ＰＣＲ）半定量技术检测不同损伤程度的肾组织中共刺激分子Ｂ７的ｍＲＮＡ表达水平。结果：正常和缺血肾组织中Ｂ７ｍＲＮＡ的表达处于极低水平,再灌注后肾组织中Ｂ７ｍＲＮＡ的表达开始逐渐升     

阻断B7/CD28共刺激通路抑制大鼠肝移植急性排斥反应的研究

目的:建立大鼠原位肝移植(ROLT)急性排斥反应模型,观察细胞毒淋巴细胞抗原4免疫球蛋白(CTLA-4Ig)在大鼠肝移植术后对共刺激分子B7-1/B7-2的影响及其抗排斥反应的作用。方法:采用"二袖套管"法先行建立DA-Lewis大鼠组合肝移植急性排斥反应模型,随机分为对照组(A组)与实验组(B组)两组,于肝移植术后48h每只受体大鼠腹腔内一次性注射CTLA-4Ig 75μg,分别于术后3、5、7和10d采用RT-PCR检测B7-1和B7-2 mRNA在两组肝脏组织中的表达情况,并同时观察其肝功能变化。结果:1)B7-1和B7-2 mRNA在A组高水平表达,而在B组表达明显降低(P<0.01);2)B组动物术后未见明显排斥反应,血清ALT、TBIL和DBIL水平明显低于对照组(P<0.01)。结论:移植术后应用CTLA-4Ig可以降低肝组织中B7-1和B7-2的表达;动态检测B7分子的表达有助于观察肝移植排斥反应的进程。     

Induction therapy with monoclonal antibodies specific for CD80 and CD86 delays the onset of acute renal allograft rejection in non-human primates

CD80 and CD86 (also known as B7-1 and B7-2, respectively) are both ligands for the T cell costimulatory receptors CD28 and CD152. Both CD80 and CD86 mediate T cell costimulation, and as such, have been studied for their role in promoting allograft rejection. In this study we demonstrate that administering monoclonal antibodies specific for these B7 ligands can delay the onset of acute renal allograft rejection in rhesus monkeys. The most durable effect results from simultaneous administration of both anti-B7 antibodies. The mechanism of action does not involve global depletion of T or B cells. Despite in vitro and in vivo evidence demonstrating the effectiveness of the anti-B7 antibodies in suppressing T cell responsiveness to alloantigen, their use does not result in durable tolerance. Prolonged therapy with murine anti-B7 antibodies is limited by the development of neutralizing antibodies, but that problem was avoided when humanized anti-B7 reagents are used. Most animals develop rejection and an alloantibody response although still on antibody therapy and before the development of a neutralizing antibody response. Anti-B7 antibody therapy may have use as an adjunctive agent for clinical allotransplantation, but using the dosing regimens we used, is not a tolerizing therapy in this non-human primate model.     

Recombinant erythropoietin rapidly treats anemia in ischemic acute renal failure

BACKGROUND: The anemia associated with acute renal failure (ARF) is currently treated with blood transfusions, while the anemia of chronic renal failure is treated with recombinant erythropoietin (EPO). We hypothesized that EPO treatment during ARF could rapidly improve hemoglobin levels and be a useful therapeutic approach. In addition, as tubular epithelial cells have EPO receptors that can mediate proliferation, enhanced recovery of renal function may occur with EPO use. METHODS: An established rat model of ischemic ARF was studied, using either moderate or severe ischemia. EPO was administered in a dose of 500 or 3000 U/kg starting at time of ischemia. Hematocrit (Hct), serum creatinine, reticulocyte count, and mortality rate were measured. RESULTS: EPO treatment led to a rapid and significant increase in Hct at 48 and 72 hours after moderate ischemic renal reperfusion injury (IRI) in EPO (500 U/kg)-treated rats compared with control (saline treated) rats (mean +/- SE; 45.6 +/- 0.3% vs. 42.0 +/- 1.0%, P < 0.01) and (46.6 +/- 0.3 vs. 41.0 +/- 1.0, P < 0.01, N = 3 per group). In severe renal IRI, EPO treatment also led to significantly increased Hct at 48 (40.0 +/- 4.4% vs. 36.8 +/- 0.3%, P < 0.01, N = 3 per group) and 72 hours (43.5 +/- 1.5% vs. 34.7 +/- 2.3%, P < 0.01, N = 3 per group). Higher dose (3000 U/kg) EPO led to a more pronounced Hct increase after severe IRI at 48 hours compared with the 500 U/kg dose (43.5 +/- 0.3 vs. 40.3 +/- 0.3, P < 0.01, N = 3 per group). EPO treatment during moderate or severe renal IRI did not change the course of the renal dysfunction. EPO treatment (N = 19) had a significant protective effect on mortality during severe IRI. In addition, loss of body weight during ARF was not affected by EPO therapy. CONCLUSIONS: Recombinant EPO can rapidly increase Hct and improve mortality during ARF. Human studies are warranted to evaluate the clinical applicability of this important finding.     

共刺激分子B7-1、B7-2在大鼠热缺血再灌注肝脏表达意义的研究

目的:探讨共刺激分子B71和B72在大鼠肝脏热缺血伤再灌注损伤模型中的表达情况及其免疫学意义。方法:将雄性Wistar大鼠随机分为3组:A组(缺血30min再灌注24h)、B组(缺血60min再灌注24h)、C组(假手术组)。模型制备参考Ohmorid的方法。分别取模型之肝左叶,采用实时反转录聚合酶链反应(PTPCR)检测B71和B72在3组肝左叶组织中mRNA表达情况。结果:B71和B72mRNA在C组以极低水平表达,而在A组、B组表达却明显增多(P<0.01),且B组高于A组(P<0.05)。结论:热缺血再灌注之肝脏通过上调B71和B72的表达增加了肝脏的免疫原性。     

Effects of Combined T- and B-Cell Deficiency on Murine Ischemia Reperfusion Injury

B and T cells have been implicated in the pathogenesis of renal ischemia reperfusion injury (IRI); however, it is unknown if B and T cells interact in early injury responses, as seen in adaptive immune responses. Recent evidence has shown that B-cell deficient and T-cell deficient mice are partially protected from renal IRI. Renal IRI was induced in recombinase activating gene (RAG)-1 deficient mice, which lack both B and T cells. RAG-1 deficient mice from two different background strains were not protected from renal IRI. Adoptive transfer of either B or T cells into RAG-1 deficient mice led to a significant protection of renal injury, which was independent of effects on neutrophil trafficking. Neutrophil depletion in RAG-1 deficient mice did not protect from IRI. While deficiency of either B or T cells reduced IRI, combined lack of both is not protective. These results demonstrate that complex interactions between B and T cells are likely occurring in kidney IRI.     

Prolongation of renal allograft survival in rats by replication-defective recombinant adenovirus-mediated coexpression of CD40L and CTLA4Ig

BACKGROUND: Blockade of costimulatory signals has been shown to prolong allograft survival. The aim of the present study was to investigate the effect of simultaneous blockade of CD40/CD40L and CD28/B7 costimulatory pathways by replication-defective adenovirus-mediated expression of secretable extracellular domain of human CD40L (shCD40L) and CTLA4Ig to prolong rats renal allograft survival. METHODS: We constructed Adv-shCD40L-IRES2-CTLA4Ig, a replication-defective adenovirus carrying genes encoding human CD40L and CTLA4Ig. Coexpression of shCD40L and CTLA4Ig was evaluated by confocal laser scanning microscopy. The function of these two molecules was examined in human mixed lymphocyte reactions (MLRs) in vitro and in experimental BN-to-LEWIS rat renal transplantation in vivo. RESULTS: Successful construction of Adv-shCD40L-IRES2-CTLA4Ig was confirmed by polymerase chain reaction. Coexpression of shCD40L and CTLA4Ig on human kidney cell line HK-2 cells after transfection was detected by direct immunofluorescence staining. Human MLR was inhibited to 52.2%+/-0.6% and 42.1%+/-0.2% of the vehicle control by Adv-shCD40L and Adv-CTLA4Ig, respectively. Adv-shCD40L-IRES2-CTLA4Ig resulted in further inhibition of MLR to 22.0%+/-0.2% of vehicle control. Transfection with Adv-shCD40L or Adv-CTLA4Ig alone prolonged renal graft survival to 24.8+/-2.5 days and 27.3+/-3.6 days, respectively, as compared to vehicle-treated controls (7.8+/-0.3 days). Cotransfection of both genes extended graft survival to 41.8+/-3.7 days. CONCLUSIONS: Adv-shCD40L-IRES2-CTLA4Ig, a replication-defective adenovirus carrying genes encoding human CD40L and CTLA4Ig, achieved simultaneous blockade of CD40/CD40L and CD28/B7 costimulatory pathways, Adv-shCD40L-IRES2-CTLA4 by Ig synergistically inhibited human T-cell proliferation in MLR, and prolonged rats renal allograft survival.     

Expression of inducible lymphocyte costimulatory molecules in human renal allograft

Background: CTLA-4/CD28-B7 and CD40-CD40L interactions constitute two key costimulatory pathways in lymphocyte signalling during experimental allograft rejection. Studies on the expression of these molecules in human transplant rejection are still lacking. Methods: The immunohistochemical study was performed on renal biopsies obtained for various clinical complications from 25 renal transplant patients. Expression of B7-1 and B7-2 and their counter-receptor CTLA-4, and of CD40 and its counter-receptor CD40L was examined. Results: In acute rejection a focal intense infiltration of B7-1⁺ and B7-2⁺ cells (mainly CD20^- CD14⁺) and of CTLA-4⁺ T lymphocytes (mainly CD8⁺) was present. In contrast, CD40 and CD40L were rarely expressed. Accumulations of T lymphocytes were found in the interstitium in the same area containing B7-1⁺ and B7-2⁺ cells. The scattered CD40L⁺ cells found in the T-cell infiltrate exhibited the CD4⁺ phenotype. In chronic rejection only a few B7-1⁺, B7-2⁺ or CTLA-4⁺ cells were detectable. In contrast, several CD40L⁺CD4⁺ cells were present both in the interstitium and in glomeruli. Moreover, an intense expression of CD40 on the endothelium was observed. In patients with cyclosporin nephrotoxicity cells positive for B7-1, B7-2, CTLA-4, CD40, or CD40L were absent. Conclusions: These results demonstrate a differential expression of costimulatory molecules in renal biopsies of allograft recipients undergoing acute or chronic rejection. Moreover, their detection may prove useful to discriminate rejection from cyclosporin nephrotoxicity.     

Inhibiting the complement system does not reduce injury in renal ischemia reperfusion.

The complex pathogenesis of ischemia reperfusion injury (IRI) includes endothelial expression of adhesion molecules, leukocyte recruitment and activation, reactive oxygen species production, and apoptotic and necrotic cell death. A role for complement in IRI of different organs, including kidney, has been proposed on the basis of results of experiments that used pharmacologic inhibitors as well as animals that were deficient in individual complement proteins. Here, renal IRI in mice was examined. Animals that were deficient in C3 had partial protection from IRI induced by 27.5 min of bilateral renal ischemia, followed by 20 h of reperfusion (blood urea nitrogen [BUN] values, 46.6 +/- 6.9 and 68.4 +/- 7.9 mg/dl in C3 -/- and C3 +/+ mice; n = 7 and 8, respectively; P = 0.033). Given the reduction in IRI in C3 -/- mice, it was investigated, by use of the rodent C3 convertase inhibitor CR1-related gene/protein y-Ig (Crry-Ig), whether exogenous administration of a complement inhibitor could lessen renal injury. Despite the use of Crry-Ig in high doses, there was no significant reduction of injury induced by 20 to 30 min of ischemia followed by up to 30 h of reperfusion. Histologic examination revealed acute tubular necrosis and neutrophilic infiltration, both of which correlated significantly with BUN values (P < 0.001). Of interest, C3 deposition around renal tubules was significantly less in animals with IRI, compared with that in unmanipulated controls (P < 0.001). In Crry-Ig-treated animals, C3 deposition was inversely proportional to BUN values (r = -0.63; P < 0.001), which presumably indicates that severe vascular IRI allowed access of the 160 kD Crry-Ig to the interstitium. Thus, renal IRI in mice may have a partial complement dependence, yet pharmacologic inhibition of the complement system does not seem to be effective, likely because of the presence of other mediator systems that operate in parallel.     

Treg expansion with trichostatin A ameliorates kidney ischemia/reperfusion injury in mice by suppressing the expression of costimulatory molecules

Innate immune reactions are believed to be associated with ischemia/reperfusion injury (IRI), and IRI might be treatable by expanding regulatory T cells (Tregs), which can suppress the excessive responses of the immune system. Organ IRI is known to be closely involved in the expression of costimulatory molecules. The present study aimed to assess whether Tregs endogenously expanded by the administration of trichostatin A (TsA), a histone deacetylase inhibitor, could reduce renal IRI and to clarify their association with the expression of costimulatory molecules in a murine model.In this study, the wild-type mice used for an IRI model were randomly divided into the following four treatment groups: TsA group, DMSO group (control), DMSO+PC61 group, and TsA + PC61 group. Renal injury in the early phase after IRI was ameliorated in the TsA group (increased Tregs) when compared with the other groups. After renal IRI, both the mRNA and the protein levels of anti-inflammatory cytokines, IL-10 and TGF-β in the kidney and spleen were significantly higher in the TsA group than in the other groups, whereas the IL-6 levels were significantly lower in the TsA group than in the other groups. These results were offset by the administration of PC61, supporting that the renoprotective effect of TsA in this study is Treg dependent. mRNA expression levels of CD80, CD86, and ICAM-1 were lower in the TsA group, consistent with Treg control of injury through costimulatory molecules.Our findings suggest that endogenously expanded Tregs coordinate postischemic immune responses and decrease the expression of costimulatory molecules after renal IRI, and thus, they might ameliorate renal IRI. TsA administration for expanding Tregs is a promising therapeutic strategy for renal IRI.     

Inflammatory cells in ischemic acute renal failure

Ischemic acute renal failure (ARF) is increasingly recognized as involving a complex cascade of mechanisms with both acute and chronic consequences. Attention to nontraditional mediators of ARF such as inflammatory pathways and microvascular events has yielded new paradigms and avenues of research. The initiation phase of renal ischemia/reperfusion (I/R) injury damage involves microvascular hemodynamic changes characterized by red blood cell sludging with platelets and leukocytes. Blocking leukocyte-endothelial interactions has yielded significant protection from renal I/R injury in experimental models. However, experiments focusing on the role of the neutrophil have led to a modest expectation of its role in ARF. Recent studies have found that T cells directly mediate renal injury in experimental I/R injury. The CD4+ T cell, working both via interferon-gamma (IFN-gamma) and costimulatory molecules appears to be an important modulator of ARF. The B cell has recently been implicated in ARF. Little is known about the role for the macrophage. Finally, resident kidney cells likely contribute to the inflammatory pathogenesis of I/R damage and protection/repair, but how, and to what extent they are involved is not known. New tools to modulate inflammatory cells, particularly mononuclear leukocytes, hold promise for clinical trials in ARF.     

Renal ischemia/reperfusion leads to macrophage-mediated increase in pulmonary vascular permeability.

BACKGROUND: Despite the advent of dialysis, survival with acute renal failure when associated with multiorgan failure is poor. The development of lung injury after shock or visceral ischemia has been shown; however, the effects of isolated renal ischemia/reperfusion injury (IRI) on the lungs are unclear. We hypothesized that isolated renal IRI could alter pulmonary vascular permeability (PVP) and that macrophages could be important mediators in this response. METHODS: Rats (N = 5 per group) underwent renal ischemia for 30 minutes, followed by reperfusion. Lung vascular permeability was evaluated by quantitation of Evans blue dye extravasation from vascular space to lung parenchyma at 1, 24, 48, or 96 hours after reperfusion. Serum was collected for blood urea nitrogen and creatinine at each time point. To examine the role of the macrophage, the macrophage pacifant CNI-1493, which inhibits the release of macrophage-derived inflammatory products, was administered in a blinded fashion during renal IRI. RESULTS: PVP was significantly (P < 0.05) increased at 24 hours and peaked at 48 hours after IRI compared with shams as well as baseline levels. PVP after IRI became similar to shams after 96 hours. This correlated with increases in blood urea nitrogen and creatinine at similar time points. At 48 hours, CNI-1493 significantly abrogated the increase in PVP compared with IRI alone. However, CNI-1493 did not alter the course of the acute renal failure. Pulmonary histology demonstrated interstitial edema, alveolar hemorrhage, and red blood cell sludging after renal IRI, which was partially attenuated by CNI-1493. CONCLUSIONS: Increased PVP develops after isolated renal IRI, and macrophage-derived products are mediators in this response. These findings have implications for understanding the mechanisms underlying respiratory dysfunction associated with acute renal failure.     

Cholesterol ester accumulation: an immediate consequence of acute in vivo ischemic renal injury

BACKGROUND: Cholesterol is a major constituent of plasma membranes, and recent evidence indicates that it is up-regulated during the maintenance phase of acute renal failure (ARF). However, cholesterol's fate and that of the cholesterol ester (CE) cycle [shuttling between free cholesterol (FC) and CEs] during the induction phase of ARF have not been well defined. The present studies sought to provide initial insights into these issues. METHODS: FC and CE were measured in mouse renal cortex after in vivo ischemia (15 and 45 minutes)/reperfusion (0 to 120 minutes) and glycerol-induced myoglobinuria (1 to 2 hours). FC/CE were also measured in (1) cultured human proximal tubule (HK-2) cells three hours after ATP depletion and in (2) isolated mouse proximal tubule segments (PTSs) subjected to plasma membrane damage (with cholesterol oxidase, sphingomyelinase, phospholipase A2, or cytoskeletal disruption with cytochalasin B). The impact of cholesterol synthesis inhibition (with mevastatin) and FC traffic blockade (with progesterone) on injury-evoked FC/CE changes was also assessed. RESULTS: In vivo ischemia caused approximately threefold to fourfold CE elevations, but not FC elevations, that persisted for at least two hours of reperfusion. Conversely, myoglobinuria had no effect. Isolated CE increments were observed in ATP-depleted HK-2 cells. Neither mevastatin nor progesterone blocked this CE accumulation. Plasma membrane injury induced with sphingomyelinase or cholesterol oxidase, but not with phospholipase A(2) or cytochalasin B, increased tubule CE content. High CE levels, induced with cholesterol oxidase, partially blocked hypoxic PTS attack. CONCLUSIONS: In vivo ischemia/reperfusion acutely increases renal cortical CE, but not FC, content, indicating perturbed CE/FC cycling. The available data suggest that this could stem from specific types of plasma membrane damage, which then increase FC flux via aberrant pathways to the endoplasmic reticulum, where CE formation occurs. That CE levels are known to inversely correlate with both renal and nonrenal cell injury suggests the potential relevance of these observations to the induction phase of ischemic ARF.     

PDE5 inhibition against acute renal ischemia reperfusion injury in rats: does vardenafil offer protection?

Purpose

To evaluate the effect of vardenafil on renal function after renal ischemia–reperfusion (IR) injury (IRI) in a rat model.

Materials and methods

Seventy-one Wistar rats were divided into 7 groups including (1) a vehicle-treated group, (2) a vehicle pretreated-IR group, (3–6) vardenafil pretreated-IR groups in doses of 0.02, 0.2, 2 and 20 μg/kg, respectively, (7) a group of IR followed by treatment with 2 μg/kg of vardenafil. Vardenafil or vehicle solution was administered one hour before unilateral nephrectomy and the induction of 45 min of ischemia on the contralateral kidney by clamping of renal pedicle. Four hours of reperfusion were allowed after renal ischemia. Studied parameters were serum creatinine, fractional excretion of sodium (FENa), and histological evaluation of renal specimens. In addition, renal tissue cGMP levels, ERK1/2 phosphorylation as well as renal function by renal scintigraphy were also evaluated.

Results

Administration of vardenafil before the induction of ischemia resulted in a significant reduction in creatinine and FENa levels as well as in less histological lesions observed in treated kidneys in comparison with the vehicle-treated group. The underlying mechanism of cytoprotection was cGMP depended and involved the phosphorylation of ERK proteins. Renal scintigraphy confirmed that PDE5 inhibition attenuates renal IRI.

Conclusions

Vardenafil attenuates renal IRI. Based on similar results from relevant studies on other PDE-5 inhibitors in renal and cardiac IRI, it can be assumed that all PDE-5 inhibitors share a common mechanism of cytoprotection.     

Ang-1基因转染的内皮祖细胞移植对大鼠肾脏缺血再灌注后的保护作用

目的 评估Ang-1基因转染的内源性内皮祖细胞(endothelial progenitor cells,EPCs)移植治疗肾脏缺血再灌注损伤的作用,进一步完善EPCs对缺血肾脏产生保护作用的旁分泌机制.方法利用重组腺病毒载体Ad-Ang-1感染EPCs,并进行转染效率鉴定,进一步移植治疗大鼠缺血肾脏,术后72 h给予肾功能评估,并检测大鼠缺血肾脏中VEGF、Ang-1以及Ang-2表达情况.结果EPCs移植治疗后,大鼠肾脏功能明显改善,术后72 h检测大鼠患肾,其内VEGF、Ang-1以及Ang-2表达均明显提升.结论 Ang-1基因转染的EPCs移植治疗可以有效地治疗大鼠肾脏IRI,其作用机制可能与EPC归巢后旁分泌过量Ang-1相关,并且促使肾脏血管新生.     

Notch2/Hes-1 pathway plays an important role in renal ischemia and reperfusion injury-associated inflammation and apoptosis and the γ-secretase inhibitor DAPT has a nephroprotective effect

This study aims to investigate the role of Notch pathway in the renal ischemia/reperfusion injury (IRI)-associated inflammation and apoptosis. Materials and methods: Male Sprague-Dawley rats were divided into three groups: normal saline (NS)-treated sham rats, NS-treated ischemia/reperfusion (I/R) rats, and N-[N-(3,5-difluorophenacetyl-l-alanyl)]-S-phenylglycine t-butyl ester (DAPT) (a γ-secretase inhibitor) treated I/R rats. I/R rat model underwent nephrectomy of the right kidney and was subjected to 60 min of left renal pedicle occlusion followed by 24 h, 48 h, and 72 h of reperfusion, respectively. The levels of creatinine, urea nitrogen (BUN), interleukin (IL)-6, tumor necrosis factor (TNF)-α in serum samples and urinary N-acety-β-d-glucosaminidase (NAG) were assayed. Histological examinations were performed. The protein expression of Notch2, hairy/enhancer of split 1 (hes-1), NF-κB2, monocyte chemoattractant protein (MCP)-1, B-cell lymphoma 2 (bcl-2), and bcl-2-associated X (bax) were detected and the degree of apoptosis of tubular cells was evaluated. Results: Renal IR induced severe tubular damage, caused significant increases in the Scr, BUN, IL-6, TNF-α, urinary NAG, Notch2, hes-1, NF-κB2, MCP-1, ratio of tubule cells apoptosis, and reduction in the ratio of bcl-2 to bax. However, DAPT treatment significantly reduced the level of Scr, BUN, IL-6, TNF-α, and NAG. Thus, I/R activates Notch2/hes-1 signaling and DAPT treatment can ameliorate the severity of tubular damage after renal IRI, lower the expression of NF-κB2, MCP-1, and bax protein, increase the expression of bcl-2 protein, and reduce the ratio of terminal 2-deoxyuridine 5-triphosphate nick end-labeling-positive cells. Conclusion: Notch signaling plays an important role in the renal IRI-associated inflammation and apoptosis. DAPT can protect against IRI through partly suppressing inflammation and apoptosis, which could constitute a new target for AKI.     

Erythropoietin attenuates apoptosis after ischemia-reperfusion-induced renal injury in transiently hyperglycemic Wister rats

Background

Hyperglycemia is associated with a decreased tolerance to ischemia and an increased severity of renal ischemia reperfusion (I/R) injury. It has been suggested that erythropoietin (EPO) attenuates this effect in normoglycemic animals. This study sought to examine the effects of EPO on treatment renal I/R injury (IRI) in transiently hyperglycemic rats.

Material and Methods

Twenty-eight male Wister rats anesthetized with isoflurane received glucose (2.5 g.kg⁻¹ intraperitoneally) before right nephrectomy. They were randomly assigned to four groups: sham operation (S); IRI (ISO); IRI+EPO, (600 UI kg⁻¹ low-dose EPO [EL]); and IRI+EPO 5000 UI kg⁻¹ (high-dose EPO [EH]). IRI was induced by a 25-minute period of left renal ischemia followed by reperfusion for 24 hours. Serum Creatinine and glucose levels were measure at baseline (M1), immediately after the ischemic period (M2), and at 24 hours after reperfusion (M3). After sacrificing the animals, left kidney specimens were submitted for histological analysis including flow cytometry to estimate tubular necrosis and the percentages of apoptotic, dead or intact cells.

Results

Scr in the ISO group was significantly higher at M3 than among the other groups. Percentages of early apoptotic cells in ISO group were significantly higher than the other groups. Percentages of late apoptotic cells in S and ISO groups were significantly greater than EL and EH groups. However, no significant intergroup differences were observed regarding the incidence of tubular necrosis.

Conclusions

Our results suggested that, although not preventing the occurrence of tubular necrosis, EPO attenuated apoptosis and glomerular functional impairment among transiently hyperglycemic rats undergoing an ischemia/reperfusion insult.     

Effects of Immunosuppressive Drugs on Rat Renal Ischemia Reperfusion Injury

Introduction

Recent evidence has demonstrated that the immune response and, more specifically, lymphocytes (T and B) and dendritic cells participate as mediators of renal ischemia reperfusion injury (IRI). The aim of this study was, therefore, to evaluate the effect of various immunosuppressive drugs with known activity to prevent IRI among rats undergoing a scheme that is potentially applicable in the clinic.

Methods

Male Sprague-Dawley rats (200-300 g) underwent 60 minutes of ischemia by renal artery clamping and contralateral nephrectomy. The experimental groups (n = 6-7) were as follows: I, Sham; II, Control; III, Rapamycin (R; 1 mg/kg); IV, Methylprednisolone (M; 15 mg/kg); V, Vitamin D3 (VD3; 2 μg/kg); VI, VD3 (1 μg/kg); and VII, M (15 mg/kg) + R (1 mg/kg). Each drug was administered in 2 doses at 6 hours and 1 hour before surgery. Creatinine (Cr) was determined on days 0.1, 2, 3, 5, and 7, and Cr clearance was determined on days 3 and 7. At 7 days nephrectomy was performed to obtain samples for histology to evaluate the degree of acute tubular necrosis.

Results

Mortality from renal insufficiency was between 0 and 33%, except in group V (66%; 4/6; P = .01). Kidney function was similar to controls in all groups except for creatinine at 7 days between group VI (VD3) and control (1.05 vs 0.65; P < .05) but no difference in Cr clearance. Histologically moderate to severe renal damage was greater in groups V and VI (VD3) than controls (P = .04).

Conclusion

We observed that none of the drugs conferred protection against IRI in a time setting relevant to kidney transplantation. Controversy exists regarding R, because some prior studies have shown a deleterious effect on IRI injury, although we did not observe any deleterious effect.     

肾脏缺血再灌注损伤过程中天然免疫分子高迁移率族蛋白B1释放变化

目的 观察高迁移率族蛋白B1(HMGB1)在小鼠肾脏缺血再灌注损伤(IRI)中的表达变化.方法 建立小鼠IRI模型,再灌注0、3、6 h或24 h后取外周血及左肾,测定血肌酐(Cr)和尿素氮(BUN)水平,免疫组织化学及Western blot检测肾组织全蛋白和胞质蛋白HMGB1的表达,并观察病理学变化(HE)及细胞凋亡.结果与假手术组比较,再灌注0 h时小鼠血清Cr和BUN无明显升高,而再灌注3、6、24 h后呈阶梯性显著性升高.假手术组肾组织HMGB1几乎均表达于细胞核内,而肾脏缺血损伤后HMGB1即开始在(主要是肾小管上皮细胞)胞质或胞外表达,且HMGB1在细胞核外的表达量在再灌注3 h时最强,之后缓慢减弱,而24 h内细胞总蛋白HMGB1表达量未见明显变化.再灌注0、24 h病理损伤渐进性加重,而凋亡细胞显著性增加.结论 HMGB1在肾IRI早期表达总量恒定而表达部位发生改变,且表达部位的改变先于肾功能的变化,HMGB1可能作为重要早期炎症因子在IRI启动中发挥作用.