Renal Ischemia-reperfusion Injury In Mice Research Articles

Role of chemokine CXC-ligand 16 in renal fibrosis following renal ischemia-reperfusion injury in mice

Objective To evaluate the role of chemokine CXC-ligand 16(CXCL16) in renal fibrosis following renal ischemia-reperfusion(I/R) injury in mice. Methods Twenty-four healthy male C57BL/6 mice, aged 8-10 weeks, weighing 20-30 g, were divided into 4 groups(n=6 each) using a random number table method: sham operation group(group S), sham operation plus anti-CXCL16 antibody group (group S+ A), group I/R, and I/R plus anti-CXCL16 antibody group(group I/R+ A). Renal ischemia was induced by occlusion of the left renal artery for 30 min followed by 72-h reperfusion, and the right kidney was removed on 5th day in anesthetized mice. CXCL16 antibody 5 mg/kg and the equal volume of normal saline were intraperitoneally injected at 72 h after reperfusion twice a week for 2 consecutive weeks in I/R+ A and I/R groups, respectively. Sham operation was performed, and the equal volume of CXCL16 antibody 5 mg/kg and normal saline were intraperitoneally injected at 72 h after reperfusion twice a week for 2 consecutive weeks in S+ A and S groups, respectively. Orbital venous blood samples were collected at day 14 after reperfusion for determination of serum blood urea nitrogen(BUN) and creatinine(Cr) concentrations. Animals were then sacrificed and the left kidney was removed for measurement of the renal fibrosis size(using Sirius red staining), expression of α-smooth muscle actin(α-SMA), fibronectin(FN) and type Ⅰ collagen(Col-Ⅰ) in renal tissues by Western blot. Results Compared with group S, the serum BUN and Cr concentrations and renal fibrosis size were significantly increased, and the expression of α-SMA, FN and Col-Ⅰ was up-regulated in group I/R(P<0.05). Compared with group I/R, the serum BUN and Cr concentrations and renal fibrosis size were significantly decreased, and the expression of α-SMA, FN and Col-Ⅰ was down-regulated in group I/R+ A(P<0.05). Conclusion CXCL16 is involved in the process of renal fibrosis following renal I/R injury in mice. Key words: Chemokines, CXC; Kidney; Reperfusion injury; Fibrosis

Blockade of the G-CSF Receptor is Protective in a Mouse Model of Renal Ischemia Reperfusion Injury

Background Ischemia-reperfusion injury (IRI) is a complex inflammatory process that detrimentally affects the function of transplanted organs. Several pathways contribute to cell death and microvascular dysfunction in IRI, including activation of the innate and adaptive arms of the immune system. Inhibiting complement, a key component of innate immunity, by C5 blockade has been shown to protect against renal IRI in mice. In this study, we investigated whether targeting neutrophils with VR81, a monoclonal antibody recognising the granulocyte colony stimulating factor receptor (G-CSFR), would reduce inflammation and protect against injury in a mouse model of renal IRI. The anti-C5 antibody BB5.1 was used for comparison. Methods Male 10-12 week old C57BL/6 mice underwent right nephrectomy followed by clamping of the left renal pedicle for 22 min, or right nephrectomy only (Sham), at 37°C. Mice (n = 5-8 per group) were injected i.p. at 24 hr pre-ischemia with VR81 (100, 200 or 500 μg), BB5.1 (100 μg), or isotype-matched antibody (Ab) control (500 μg). Mice were euthanased 24 hr after reperfusion, and serum and plasma samples were collected to assess renal function (creatinine, urea) and complement activation (C5a ELISA). Kidneys were harvested to analyse complement C9 deposition and immune cell infiltration by immunofluorescence. Results Compared to Sham, the Ab control IRI group showed significantly increased creatinine, urea and C5a, deposition of C9, and infiltration of neutrophils and macrophages. As expected, C5 blockade with BB5.1 markedly reduced renal injury and inflammation, reflected in significantly lower creatinine, urea, C5a and C9 deposition versus Ab control. Surprisingly, VR81 was also strongly protective in a dose-dependent manner, with the 500 μg dose significantly reducing creatinine, urea and cellular infiltration. Conclusion Unexpectedly, G-CSFR blockade using VR81 significantly protected against renal IRI with an efficacy similar to that of anti-C5 treatment. Protection was associated with markedly reduced infiltration by neutrophils and macrophages. G-CSFR blockade is thus a promising new approach to reduce IRI and improve organ transplant function.

Methylene blue attenuates renal ischemia-reperfusion injury by negative regulation of NLRP3 signaling pathway.

To investigate the effect of methylene blue (MB) on renal ischemia-reperfusion (IR) injury in mice and its possible relevant mechanisms. A total of 30 male C57/BL6 mice aged 4 months old were randomly divided into the following three groups: Sham group (n=10), IR group (n=10), and MB group (n=10). Mice in MB group were treated with gavage continuously using methylene blue solution (dosage: 25 mg·kg-1·d-1) until they were 7 months old. Mice in the other two groups were administrated with the same amount of normal saline for gavage. After that, the abdomen of mice in Sham group was opened and closed, and bilateral renal pedicles of mice in IR group and MB group were occluded using a micro-artery clamp for 45 min for modeling. After modeling, the renal tissues and blood samples of mice were taken for detection. The levels of serum creatinine (Scr), blood urea nitrogen (BUN), and malondialdehyde (MDA), and activity of superoxide dismutase (SOD) in mice in each experimental group were detected and statistically analyzed, respectively. The degree of renal tubular necrosis in renal tissues of mice was observed under an optical microscope. Enzyme-linked immunosorbent assay (ELISA) was used to detect the levels of inflammatory factors [interleukin-1β (IL-1β), IL-18, IL-10 and transforming growth factor-β1 (TGF-β1)] in renal tissues of mice in each experimental group, followed by relevant statistical analyses. The expressions of relevant proteins [NACHT, LRR, and PYD domains-containing protein 3 (NLRP3), nuclear factor-κB (NF-κB), caspase-1, pro-IL-1β, and IL-1β] in renal tissues of mice in each experimental group were detected via Western blotting, and the gray value of each band was detected for statistical analysis. It was found in this experimental study that Scr and BUN levels in MB group were significantly lower than those in IR group, and the differences were statistically significant (p<0.05). Compared with that in IR group, the degree of renal tubular necrosis in MB group was significantly alleviated, and the difference was statistically significant (p<0.05). Compared with those in IR group, the levels of inflammatory factors (IL-1β and IL-18) in MB group were significantly decreased, but the levels of IL-10 and TGF-β1 in MB group were significantly increased, and the differences were statistically significant (p<0.05). Compared with those in IR group, the activity of SOD in MB group was increased significantly, but the level of MDA was decreased, and the differences were statistically significant (p<0.05). The protein expressions of NLRP3, NF-κB, caspase-1, and pro-IL-1β in MB group were decreased compared with those in IR group, but the expression of IL-1β in MB group was increased, and the differences were statistically significant (p<0.05). We showed that methylene blue can alleviate the apoptosis and inflammatory response induced by renal IR injury in mice, and its relevant mechanism may be related to the fact that methylene blue can negatively regulate NLRP3 signaling pathway.

Overexpression of Human CD55 and CD59 or Treatment with Human CD55 Protects against Renal Ischemia-Reperfusion Injury in Mice.

Deficiency in the membrane-bound complement regulators CD55 and CD59 exacerbates renal ischemia-reperfusion injury (IRI) in mouse models, but the effect of increasing CD55 and CD59 activity has not been examined. In this study, we investigated the impact of overexpression of human (h) CD55 ± hCD59 or treatment with soluble rhCD55 in a mouse model of renal IRI. Unilaterally nephrectomised mice were subjected to 18 (mild IRI) or 22 min (moderate IRI) warm renal ischemia, and analyzed 24 h after reperfusion for renal function (serum creatinine and urea), complement deposition (C3b/c and C9), and infiltration of neutrophils and macrophages. Transgenic mice expressing hCD55 alone were protected against mild renal IRI, with reduced creatinine and urea levels compared with wild type littermates. However, the renal function of the hCD55 mice was not preserved in the moderate IRI model, despite a reduction in C3b/c and C9 deposition and innate cell infiltration. Mice expressing both hCD55 and hCD59, on the other hand, were protected in the moderate IRI model, with significant reductions in all parameters measured. Wild type mice treated with rhCD55 immediately after reperfusion were also protected in the moderate IRI model. Thus, manipulation of CD55 activity to increase inhibition of the C3 and C5 convertases is protective against renal IRI, and the additional expression of hCD59, which regulates the terminal complement pathway, provides further protection. Therefore, anti-complement therapy using complement regulatory proteins may provide a potential clinical option for preventing tissue and organ damage in renal IRI.

Changes in expression of keratin genes in renal tissues during renal ischemia-reperfusion injury in mice

Objective To investigate the changes in the expression of keratin genes in renal tissues during renal ischemia-reperfusion(I/R)injury in mice. Methods Six wild type male C57/B6 mice, aged 50 days, weighing 20-30 g, were divided into 2 groups(n=3 each)using a random number table: sham operation group(Sham group)and I/R group.Right renal arteries and veins were clamped for 1 h followed by reperfusion, and the left kidneys were removed to establish the model of renal I/R injury.At 24 h of reperfusion, blood samples were collected from the left ventricle for determination of serum creatinine and urea nitrogen concentrations by colorimetric method.The right kidney specimens were obtained for pathologic examination and for determination of the expression of kidney injury molecule-1 and neutrophil gelatinase-associated lipocalin mRNA(by quantitative real-time polymerase chain reaction [qRT-PCR]) and keratin genes(by Affemetrixc DNA microarray). The differentially expressed genes identified were further confirmed by qRT-PCR. Results Compared with Sham group, the serum creatinine and urea nitrogen concentrations were significantly increased, the expression of kidney injury molecule-1 and neutrophil gelatinase-associated lipocalin mRNA was up-regulated(P<0.05), and the damage to the renal tubules was aggravated in I/R group.The results of microarray analysis showed that only keratin 20 gene(the expresion was up-regulated)was the differentially expressed gene(P<0.05), and the results measured by qRT-PCR were consistent with those measured by Affemetrixc DNA microarray. Conclusion Keratin 20 gene expression in renal tissues is up-regulated during renal I/R injury in mice, and the change may be involved in the endogenous protective mechanism during renal I/R injury. Key words: Keratins; Reperfusion injury; Kidney

Small interfering RNA targeting TNF-α gene significantly attenuates renal ischemia-reperfusion injury in mice.

Tumor necrosis factor-alpha (TNF-α) has been found to be centrally involved in the development of ischemia-reperfusion injury (IRI)-induced inflammation and apoptosis. Knockdown of TNF-α gene using small interfering RNA (siRNA) may protect renal IRI. Renal IRI was induced in mice by clamping the left renal pedicle for 25 or 35 min. TNF-α siRNA was administered intravenously to silence the expression of TNF-α. The therapeutic effects of siRNA were evaluated in terms of renal function, histological examination, and overall survival following lethal IRI. A single systemic injection of TNF-α siRNA resulted in significant knockdown of TNF-α expression in ischemia-reperfusion injured kidney. In comparison with control mice, levels of BUN and serum creatinine were significantly reduced in mice treated with siRNA. Pathological examination demonstrated that tissue damage caused by IRI was markedly reduced as a result of TNF-α siRNA treatment. Furthermore, survival experiments showed that nearly 90% of control mice died from lethal IRI, whereas more than 50% of siRNApretreated mice survived until the end of the eight-day observation period. We have demonstrated for the first time that silencing TNF-α by specific siRNA can significantly reduce renal IRI and protect mice against lethal kidney ischemia, highlighting the potential for siRNA-based clinical therapy.

Induction of microRNA-17-5p by p53 protects against renal ischemia-reperfusion injury by targeting death receptor 6

Renal ischemia-reperfusion injury is a leading cause of acute kidney injury; the pathogenesis of which remains poorly understood and effective therapies are still lacking. Here we tested whether microRNAs, identified as critical regulators of cell health and disease, are involved in this process. We found that miR-17-5p was significantly up-regulated during renal ischemia-reperfusion injury in mice and during hypoxia in cultured renal tubular cells. In cultured cells, miR-17-5p directly inhibited the expression of death receptor 6 (DR6) and attenuated apoptosis during hypoxia. Blockade of miR-17-5p abolished the suppression of DR6 and facilitated caspase activation and apoptosis. Invivo, an miR-17-5p mimic suppressed DR6 expression and protected against renal ischemia-reperfusion injury. We further verified that miR-17-5p induction during renal ischemia-reperfusion injury was dependent on p53. Inhibition of p53 with pifithrin-α or a dominant-negative mutant led to the repression of miR-17-5p expression under hypoxia invitro. Moreover, miR-17-5p induction during renal ischemia-reperfusion injury was attenuated in proximal tubule p53 knockout mice, supporting the role of p53 in miR-17-5p induction invivo. Thus, p53/miR-17-5p/DR6 is a new protective pathway in renal ischemia-reperfusion injury and may be targeted for the prevention and treatment of ischemic acute kidney injury.

Anti-LG3 Antibodies Aggravate Renal Ischemia-Reperfusion Injury and Long-Term Renal Allograft Dysfunction.

Pretransplant autoantibodies to LG3 and angiotensin II type 1 receptors (AT1R) are associated with acute rejection in kidney transplant recipients, whereas antivimentin autoantibodies participate in heart transplant rejection. Ischemia-reperfusion injury (IRI) can modify self-antigenic targets. We hypothesized that ischemia-reperfusion creates permissive conditions for autoantibodies to interact with their antigenic targets and leads to enhanced renal damage and dysfunction. In 172 kidney transplant recipients, we found that pretransplant anti-LG3 antibodies were associated with an increased risk of delayed graft function (DGF). Pretransplant anti-LG3 antibodies are inversely associated with graft function at 1 year after transplantation in patients who experienced DGF, independent of rejection. Pretransplant anti-AT1R and antivimentin were not associated with DGF or its functional outcome. In a model of renal IRI in mice, passive transfer of anti-LG3 IgG led to enhanced dysfunction and microvascular injury compared with passive transfer with control IgG. Passive transfer of anti-LG3 antibodies also favored intrarenal microvascular complement activation, microvascular rarefaction and fibrosis after IRI. Our results suggest that anti-LG3 antibodies are novel aggravating factors for renal IRI. These results provide novel insights into the pathways that modulate the severity of renal injury at the time of transplantation and their impact on long-term outcomes.

The Role of Activin A and B and the Benefit of Follistatin Treatment in Renal Ischemia-Reperfusion Injury in Mice.

BackgroundActivins, members of the TGF-β superfamily, are key drivers of inflammation and are thought to play a significant role in ischemia-reperfusion injury (IRI), a process inherent to renal transplantation that negatively impacts early and late allograft function. Follistatin (FS) is a protein that binds activin and inhibits its activity. This study examined the response of activin A and B in mice after renal IRI and the effect of exogenous FS in modulating the severity of renal injury.MethodsMice were treated with recombinant FS288 or vehicle before renal IRI surgery. Activin A, B, and FS levels in the serum and kidney, and renal injury parameters were measured at 3, 6, and 24 hours after reperfusion.ResultsSerum and kidney activin B levels were increased within 6 hours postrenal IRI, accompanied by renal injury—increased serum creatinine, messenger (m)RNA expression of kidney injury molecule-1 (KIM-1) and neutrophil gelatinase-associated lipocalin (NGAL); endothelial activation—increased E-selectin mRNA; and systemic inflammation—increased serum levels of IL-6, monocyte chemotactic protein-1 and TNF-α. Further injury was potentiated by an upsurge in activin A by 24 hours, with further increases in serum creatinine, KIM-1 and NGAL mRNA expression. Follistatin treatment significantly reduced the level of serum activin B and subsequently blunted the increase in activin A. Renoprotection was evident with the attenuated rise in serum creatinine, KIM-1 and NGAL expression, tubular injury score, renal cell apoptosis, and serum IL-6 and monocyte chemotactic protein-1 levels.ConclusionsWe propose that activin B initiates and activin A potentiates renal injury after IRI. Follistatin treatment, through binding and neutralizing the actions of activin B and subsequently activin A, reduced renal IRI by minimizing endothelial cell activation and dampening the systemic inflammatory response. These data support the potential clinical application of FS treatment to limit IRI during renal transplantation.

Role of chemokine CXC-ligand 16 in renal ischemia-reperfusion injury in mice

Objective To investigate the role of chemokine CXC-ligand 16 (CXCL16) in renal ischemia-reperfusion (I/R) injury in the mice. Methods Twelve healthy male C57BL/6 mice and 12 CXCL16-knockout (CXCL16-KO) mice, aged 8-10 weeks, weighing 20-30 g, were studied.The 12 C57BL/6 mice were randomly divided into 2 groups (n=6 each) using a random number table: C57BL/6 sham operation group (group C-S) and C57BL/6 I/R group (group C-I/R). The 12 CXCL16-KO mice were randomly divided into 2 groups (n=6 each) using a random number table: CXCL16-KO sham operation group (group KO-S) and CXCL16-KO I/R group (group KO-I/R). The right kidney was removed, and the left kidney was exposed, and the renal artery was then clamped for 45 min with atraumatic microclips followed by 24 h reperfusion to establish the model of renal I/R in anesthetized mice.Venous blood samples were taken at 24 h of reperfusion for determination of the serum blood urea nitrogen (BUN) and creatinine (Cr) concentrations.The renal specimens were obtained at 24 h of reperfusion for microscopic examination of the pathological changes, and the damage to the renal tubules was scored.The number of myeloperoxidase (MPO) positive cells (MPO+ cells), F4/80+ cells and CD3+ cells in renal tissues was counted by immunohistochemistry.The expression of tumor necrosis factor-alpha (TNF-α), interleukin-1beta (IL-1β), IL-6, and macrophage inflammatory protein-2 (MIP-2) mRNA in renal tissues was determined by real-time reverse transcriptase polymerase chain reaction. Results Compared with group C-S, the serum BUN and Cr concentrations, renal tubular damage score, and the number of MPO+ , F4/80+ , and CD3+ cells were significantly increased, and the expression of TNF-α, IL-1β, IL-6 and MIP-2 mRNA was significantly up-regulated in group C-I/R (P<0.05). Compared with group KO-S, the serum BUN and Cr concentrations, renal tubular damage score, and the number of MPO+ , F4/80+ , and CD3+ cells were significantly increased, and the expression of TNF-α, IL-1β, IL-6 and MIP-2 mRNA was significantly up-regulated in group KO-I/R (P<0.05). Compared with group C-I/R, the serum BUN and Cr concentrations, renal tubular damage score, and the number of MPO+ , F4/80+ , and CD3+ cells were significantly decreased, and the expression of TNF-α, IL-1β, IL-6 and MIP-2 mRNA was significantly down-regulated in group KO-I/R (P<0.05). Conclusion CXCL16 is involved in the pathophysiological process of renal I/R injury in the mice. Key words: Chemokines, CXC; Kidney; Reperfusion injury

Protective Effect of Polydeoxyribonucleotide Against Renal Ischemia-Reperfusion Injury in Mice

BackgroundPolydeoxyribonucleotide (PDRN) is an A2A receptor agonist that induces vascular endothelial growth factor (VEGF) production during the pathological condition of low tissue perfusion. Ischemia-reperfusion injury (IRI) is a major problem after renal transplantation. In the present study, we investigated whether PDRN exhibits reno-protective effects against ischemia-reperfusion–induced acute kidney injury in mice. MethodsRenal ischemia-reperfusion injury was induced in male C57BL/6 mice by bilateral renal pedicle occlusion for 30 minutes, followed by reperfusion for 48 hours. PDRN (8 mg/kg body weight intraperitoneally) was administered 30 minutes before IRI. ResultsTreatment with PDRN significantly decreased neutrophil gelatinase-associated lipocalin levels in the urine, blood urea nitrogen level, and serum creatinine levels as well as kidney tubular injury. Western blotting showed that PDRN significantly increased the levels of vascular endothelial growth factor and B-cell lymphoma protein and attenuated p38 mitogen-activated protein kinase, c-Jun N-terminal kinase, inducible nitric oxide synthase, and Bcl-2–associated X protein levels 48 hours after IRI. ConclusionsOur findings suggest that PDRN is a potential therapeutic agent for acute ischemia-induced renal damage.

CSCI hydrogel promote adipose derived mesenchymal stem cells in renal ischemia reperfusion injury mice

Objective To explored the therapeutic benefit and safety of acute kidney injury (AKI) mice induced by ischemia/reperfusion (I/R) treating by thermosensitive chitosan chloride (CSCI) hydrogel, an injectable scaffold for adipose-derived MSCs (ADMSCs) delivery. Methods After establishing the rat model of acute kidney injury, all rats were randomly divided into 4 groups: ADMSCs/PBS group, ADMSCs/CSCI group, PBS group and CSCI group.Live/Dead staining was used to evaluate the Cytocompatibility between CSCI hydrogel and ADMSCs, dihydroethidium (DHE) staining was used to detect the number of ROS in vivo, and bioluminescence imaging (BLI) was used to track the retention and survival of (fluc-mrfp) labeled ADMSCs in the rat kidney, respectively.The biocompatibility and biodegradability of CSCI hydrogel in kidney tissue were detected by HE staining.The improvement of renal function was evaluated by detecting the level of serum creatinine and urea nitrogen. Results Live/Dead staining manifested an satisfactory Cytocompatibility between CSCI hydrogel and ADMSCs.CSCI hydrogel can improve the micro-environment of AKI kidney tissue by lowering the level of ROS.In PBS and CSCI groups, the positive rate of DHE staining was 68.8%±8.5% and 38.5%±5.8%(P<0.05), respectively.These suggested that CSCI hydrogels could improve the retention and survival of grafted ADMSCs.In ADMSCs / CSCI hydrogel group, BLI fluorescent signal can be detected on the seventh day and was undetectable on the 21st day.ROI value of BLI signal changed from 38×105 p/(s·cm2·sr) on the first day to 8.5×105 p/(s·cm2·sr) on the 14th day.In ADMSCs / PBS group, BLI fluorescent signal only can be detected on the seventh day and was undetectable on the 14th day.ROI value of BLI signal changed from 17×105 p/(s·cm2·sr) on the first day to 3.3×105 p/(s·cm2·sr) on the 14th day.Results detected on the 3th and 28th day suggested that ADMSCs/CSCI hydrogel group , ADMSCs/PBS group and CSCI hydrogel group have a better self-repairing capability than the PBS injection group (P<0.05). In addition, compared with the ADMSCs / PBS group and CSCI hydrogel group, ADMSCs / CSCI hydrogel group got a lower tissue injury scores (P<0.05). At the 4th week, significant improvement of the renal function was observed in CSCI hydrogels with ADMSCs groups. Conclusions CSCI hydrogel is a very promising cell carrier for the treatment of AKI. Key words: Thermosensitive hydrogel; Chitosan; Adipose-derived mesenchymal stem cells; Acute kidney injury; Cell transplantation

Reduction in Renal Ischemia-Reperfusion Injury in Mice by a Phosphoinositide 3-Kinase p110gamma-Specific Inhibitor.

Although renal ischemia-reperfusion injury (IRI) can cause delayed graft function, a targeted therapy is not yet available. Because phosphoinositide 3-kinases (PI3K) p110γ and p110δ play important roles in immune cell migration and function, we investigated the effects of PI3K p110γ- and p110δ-specific inhibitors in a murine renal IRI model. Renal function was assessed by serum creatine and hematoxylin-eosin staining. Immune cell migration was assessed by flow cytometry and an in vitro cell migration assay using Transwell plates. Gene expression analysis and a multiplex cytokine/chemokine assay were performed to find cytokines/chemokines whose expression was upregulated in renal IRI and affected by p110γ-specific inhibitor. The PI3K p110γ-specific inhibitor, but not p110δ-specific inhibitor, significantly reduced serum creatine levels and acute tubular necrosis. These were accompanied by reduced infiltration of B cells and reduced expression of CXCL9, a CXCR3 ligand, suggesting that p110γ plays an important role in B-cell migration toward injured kidneys. An in vitro cell migration assay revealed for the first time that B-cell migration to injured kidney cells and to CXCL9 requires p110γ. p110γ-specific inhibitor ameliorates renal IRI by reducing necrosis and immune cell migration. This inhibitor may have the potential to reduce renal graft failure caused by renal IRI.

Abstract P600: Conditional Knockout of AT1a Receptors Selectively in The Proximal Tubules Attenuates Renal Ischemia-Reperfusion Injury in Mice.

The activation of the intrarenal renin-angiotensin system plays an important role in the pathogenesis of renal ischemia-reperfusion injury, but the underlying cellular and signaling mechanisms remain incompletely understood. In the present study, we tested the hypothesis that conditional knockout of AT1a receptors selectively in the proximal tubules attenuates renal ischemia-reperfusion injury in mice. Three groups (n=5-8/per group) of adult male C57BL/6J (WT), global AT1a receptor-deficient (AT1a-KO), and conditional proximal tubule-specific AT1a-KO mice (PT-AT1a-KO) were subjected to sham surgery or 30 min unilateral left kidney ischemia, followed by reperfusion for 24 h or 7 days, respectively. Under basal conditions, systolic blood pressure was 25 ± 3 mmHg lower in global AT1a-KO (p&lt;0.01) or 13 ± 3 mmHg lower in PT-AT1a-KO mice (p&lt;0.05), respectively. Systolic blood pressure, 24 h urine and urinary sodium excretion were not significantly altered in all strains 24 h or 7 days after renal ischemia-reperfusion. Kidney wt. to body wt. ratio, but not heart wt. to body wt. ratio, was significantly increased in both AT1a-KO and PT-AT1a-KO mice (p&lt;0.05). Renal ischemia-reperfusion injury was assessed by Masson Trichrome staining and compared between WT, AT1a-KO and PT-AT1a-KO mice. No significant glomerular, tubulointerstital, and peri-vascular fibrotic responses were observed in sham controls of all strains. However, significant fibrotic responses were observed in the glomeruli, cortical tubulointerstitial and peri-vascular tissues in WT mice 24 h or 7 days after renal ischemia-reperfusion (p&lt;0.01). Surprisingly, however, glomerular, tubulointerstital, and peri-vascular fibrotic responses were significantly worsen, rather than improved, in global AT1a-KO mice (p&lt;0.01). By comparison, conditional deletion of AT1a receptors selectively in the proximal tubules markedly attenuated glomerular, tubulointerstital, and peri-vascular fibrotic responses in PT-AT1a-KO mice 24 h or 7 days after renal ischemia-reperfusion (p&lt;0.01). Our results suggest that AT1a receptors in the proximal tubule play an important role in the pathogenesis of renal ischemia-reperfusion injury, and thus may represent a therapeutic target in the future.

Effects of kallistatin on oxidative stress and inflammation on renal ischemia-reperfusion injury in mice.

Kallistatin (KS) is a serine proteinase. The result of KS on 'Renal Ischemia- Reperfusion Injury' (IRI) has not clearly been researched. In this study, investigative research has been conducted to draw results on the administration of human KS on kidney response conducted within a mouse model of IRI. BALB/c mice were used and given 30 min ischemia that was injected into the kidney which was followed with 24 h reperfusion. The human KS gene contained within an adenoviral vector was injected intravenously 30 min before reperfusion and 12h after reperfusion. Analyses illustrated what impact KS had on renal IRI in realtion to tubular necrosis, apoptotic cell death, inflammatory cytokines, renal function, and inflammatory cell infiltration. KS gene transfer significantly had a positive impact on renal function (reduced blood urea-nitrogen: 73.5±13.6 vs. 195.4±14.6 mg/dL at day three, p < 0.05 and the serum creatinine levels: 0.23±0.02 vs. 0.71±0.14 mg/dL at day three, p < 0.05), reduced tubular necrosis and apoptosis of IRI kidneys. The permeation of cells that were inflamed and the manufacturing of pro-inflammatory cytokines (RANTES-is regulated through been activated with normal T-cell which are expressed and secreted, tumour necrosis interleukin-1β factor-α, and interferon-γ) resulted in significantly suppressing KS in mice with IRI. The efficacy to scavenge superoxide in tubule cells was also demonstrated by high-performance liquid chromatography. Our study suggests a novel role of KS in renal protection after 'Renal Ischemia-Reperfusion Injury' blocking of oxidative stress and renal inflammation.

Fenofibrate pre-treatment suppressed inflammation by activating phosphoinositide 3 kinase/protein kinase B (PI3K/Akt) signaling in renal ischemia-reperfusion injury.

The aim of this study was to investigate the possible beneficial effects of Fenofibrate on renal ischemia-reperfusion injury (IRI) in mice and its potential mechanism. IRI was induced by bilateral renal ischemia for 60 min followed by reperfusion for 24 h. Eighteen male C57BL/6 mice were randomly divided into three groups: sham-operated group (sham), IRI+saline group (IRI group), IRI+Fenofibrate (FEN) group. Normal saline or Fenofibrate (3 mg/kg) was intravenously injected 60 min before renal ischemia in IRI group and FEN group, respectively. Blood samples and renal tissues were collected at the end of reperfusion. The renal function, histopathologic changes, and the expression levels of pro-inflammatory cytokines [interleukin-8 (IL-8), tumor necrosis factor alpha (TNF-α) and IL-6] in serum and renal tissue homogenate were assessed. Moreover, the effects of Fenofibrate on activating phosphoinositide 3 kinase/protein kinase B (PI3K/Akt) signaling and peroxisome proliferator-activated receptor-α (PPAR-α) were also measured in renal IRI. The results showed that plasma levels of blood urea nitrogen and creatinine, histopathologic scores and the expression levels of TNF-α, IL-8 and IL-6 were significantly lower in FEN group than in IRI group. Moreover, Fenofibrate pretreatment could further induce PI3K/Akt signal pathway and PPAR-α activation following renal IRI. These findings indicated PPAR-α activation by Fenofibrate exerts protective effects on renal IRI in mice by suppressing inflammation via PI3K/Akt activation. Thus, Fenofibrate could be a novel therapeutic alternative in renal IRI.

Effect of preemptive treatment with human umbilical cord blood-derived mesenchymal stem cells on the development of renal ischemia-reperfusion injury in mice.

Human umbilical cord blood-derived mesenchymal stem cells (HUCB-MSCs) have been studied in several models of immune-mediated disease because of their unique immunomodulatory properties. We hypothesized that HUCB-MSCs could suppress the inflammatory response in postischemic kidneys and attenuate early renal injury. In 8- to 10-wk-old male C57BL/6 mice, bilateral ischemia-reperfusion injury (IRI) surgery was performed, and 1 × 10(6) HUCB-MSCs were injected intraperitoneally 24 h before surgery and during reperfusion. Renal functional and histological changes, HUCB-MSC trafficking, leukocyte infiltration, and cytokine expression were analyzed. Renal functional decline and tubular injury after IRI were attenuated by HUCB-MSC treatment. PKH-26-labeled HUCB-MSCs trafficked into the postischemic kidney. Although numbers of CD45-positive leukocytes in the postischemic kidney were comparable between groups, the expression of interferon-γ in the postischemic kidney was suppressed by HUCB-MSC treatment. The rapid decrease in intrarenal VEGF after IRI was markedly mitigated by HUCB-MSC treatment. In inflammatory conditions simulated in a cell culture experiment, VEGF secretion from HUCB-MSCs was substantially enhanced. VEGF inhibitor abolished the renoprotective effect of HUCB-MSCs after IRI. Flow cytometry analysis revealed the decreased infiltration of natural killer T cells and increased number of regulatory T cells in postischemic kidneys. In addition, these effects of HUCB-MSCs on kidney infiltrating mononuclear cells after IRI were attenuated by VEGF inhibitor. HUCB-MSCs attenuated renal injury in mice in the early injury phase after IRI, mainly by humoral effects and secretion of VEGF. Our results suggest a promising role for HUCB-MSCs in the treatment of renal IRI.

Protective effect of indomethacin in renal ischemia-reperfusion injury in mice.

To evaluate the renoprotection effects of non-steroidal anti-inflammatory drugs (NSAIDs) in renal ischemia-reperfusion injury (IRI) and the cyclooxygenase (COX)-1/2 blockade association by indomethacin (IMT) in the mice model. After the left renal pedicle of mice was clamped, IMT was administrated by intraperitoneal injection with four doses: 1, 3, 5, and 7 mg/kg. Blood and kidney samples were collected 24 h after IRI. The renal functions were assayed by the cytokines and serum creatinine (SCr) using enzyme-linked immunosorbent assay (ELISA) kits. Kidney samples were analyzed by hematoxylin and eosin (H&E) and immunohistochemistry stainings. The mice administered with 5 mg/kg IMT had a marked reduction in SCr and significantly less tubular damage. The tumor necrosis factor α (TNF-α) activity in renal homogenates and interleukin 6 (IL-6) activity in serum had a marked reduction at doses of 5 and 7 mg/kg IMT. The administration of 3 and 5 mg/kg IMT had a marked reduction in the ratio of thromboxane B2 to 6-keto-prostaglandin F1α. COX-1 and COX-2 stainings were weaker in 5 mg/kg IMT groups than that in the other groups. There was a dose response in the IMT function of renal IRI in mice, and IMT had a protective effect in a certain dose range. The effect of IMT on mice IRI was related to COX-1/2 blockades.

Follistatin Reduces Serum Activin and Attenuates Renal Ischemia-Reperfusion Injury in Mice.

Follistatin Reduces Serum Activin and Attenuates Renal Ischemia-Reperfusion Injury in Mice.

Protective Effect of Peptide GV1001 Against Renal Ischemia-Reperfusion Injury in Mice

BackgroundIschemia reperfusion injury (IRI) is a common complication after kidney transplantation. Peptide GV1001 is a peptide vaccine representing a 16−amino acid human telomerase reverse transcriptase sequence, which has been reported to possess potential antineoplastic and anti-inflammatory activity. This study aimed to investigate the potential effects of peptide GV1001 on renal IRI. MethodsPeptide GV1001 was subcutaneously administered to C57BL6/J mice 30 minutes before and 12 hours after bilateral IRI. Sham operation and phosphate-buffered saline (PBS) injection were used as controls. Blood and renal tissues were harvested at 1 day after IRI. ResultsPeptide GV1001 treatment significantly attenuated renal functional deterioration after IRI (peptide GV1001 group vs PBS group; blood urea nitrogen, P < .05; creatinine, P < .05). Peptide GV1001 treatment also attenuated renal tissue injury (tubular injury score; the peptide GV1001 group vs PBS group; P < .001). Renal apoptosis was also lower in the peptide GV1001 group. Immunohistochemical studies showed that IRI increased perirenal infiltration of both neutrophils and macrophages, and that peptide GV1001 significantly attenuated this process. Expression of interleukin-6 and monocyte chemotactic protein-1 was significantly reduced by peptide GV1001 treatment. ConclusionsPeptide GV1001 ameliorates acute renal IRI by reducing inflammation and apoptosis; therefore, it is promising as a potential therapeutic agent for renal IRI. The mechanisms of protection should be explored in further studies.