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Hyperbranched polyglycerol (HPG) is a biocompatible polyether polymer that is a potential colloid component in a preservation solution for suppressing interstitial edema during cold storage of a donor organ. This study evaluated the outcomes of kidney transplants after cold perfusion and storage with a HPG‐based preservation solution (HPGS) in a pig model of kidney autotransplantation. The left kidneys of farm pigs (weighing 35–45 kg) were perfused with and stored in either cold HPGS or standard UW solution (UWS), followed by transplantation to the right side after right nephrectomy. The survival and function of transplants were determined by the urine output, and serum creatinine (SCr) and blood urea nitrogen (BUN) of recipients. Transplant injury was examined by histological analysis. Here, we showed that there was no significant difference between HPGS and UWS in the prevention of tissue edema, but HPGS was more effective than UWS for initial blood washout of kidney perfusion and for the prevention of cold ischemia injury during cold storage. After autotransplantation, the kidneys preserved with HPGS (HPG group) had better functional recovery than those with UWS (UW group), indicated by significantly more urine output and lower levels of SCr and BUN. The survived grafts in HPG group had less tissue damage than those in UW group. In conclusion, as compared to the UWS the HPGS has less negative impact on kidney cold ischemia during cold storage, resulting in improving immediate functional recovery after transplantation, suggesting that HPG is a promising colloid for donor kidney preservation. [ABSTRACT FROM AUTHOR]

Background: Long-term outcomes after acute kidney injury (AKI) include incremental loss of function and progression towards chronic kidney disease (CKD); however, the pathogenesis of AKI to CKD remains largely unknown. Clusterin (CLU) is a chaperone-like protein that reduces ischemia-reperfusion injury (IRI) and enhances tissue repair after IRI in the kidney. This study investigated the role of CLU in the transition of IRI to renal fibrosis.Methods: IRI was induced in the left kidneys of wild type (WT) C57BL/6J (B6) versus CLU knockout (KO) B6 mice by clamping the renal pedicles for 28 min at the body temperature of 32 °C. Tissue damage was examined by histology, infiltrate phenotypes by flow cytometry analysis, and fibrosis-related gene expression by PCR array.Results: Reduction of kidney weight was induced by IRI, but was not affected by CLU KO. Both WT and KO kidneys had similar function with minimal cellular infiltration and fibrosis at day 14 of reperfusion. After 30 days, KO kidneys had greater loss in function than WT, indicated by the higher levels of both serum creatinine and BUN in KO mice, and exhibited more cellular infiltration (CD8 cells and macrophages), more tubular damage and more severe tissue fibrosis (glomerulopathy, interstitial fibrosis and vascular fibrosis). PCR array showed the association of CLU deficiency with up-regulation of CCL12, Col3a1, MMP9 and TIMP1 and down-regulation of EGF in these kidneys.Conclusion: Our data suggest that CLU deficiency worsens renal inflammation and tissue fibrosis after IRI in the kidney, which may be mediated through multiple pathways. [ABSTRACT FROM AUTHOR]

Cellular autophagy is a prosurvival mechanism in the kidney against ischemia-reperfusion injury (IRI), but the molecular pathways that activate the autophagy in ischemic kidneys are not fully understood. Clusterin (CLU) is a chaperone-like protein, and its expression is associated with kidney resistance to IRI. The present study investigated the role of CLU in prosurvival autophagy in the kidney. Renal IRI was induced in mice by clamping renal pedicles at 32°C for 45 min. Hypoxia in renal tubular epithelial cell (TEC) cultures was induced by exposure to a 1% O2 atmosphere. Autophagy was determined by either light chain 3-BII expression with Western blot analysis or light chain 3-green fluorescent protein aggregation with confocal microscopy. Cell apoptosis was determined by flow cytometric analysis. The unfolded protein response was determined by PCR array. Here, we showed that autophagy was significantly activated by IRI in wild-type (WT) but not CLU-deficient kidneys. Similarly, autophagy was activated by hypoxia in human proximal TECs (HKC-8) and WT mouse primary TECs but was impaired in CLU-null TECs. Hypoxia-activated autophagy was CLU dependent and positively correlated with cell survival, and inhibition of autophagy significantly promoted cell death in both HKC-8 and mouse WT/CLU-expressing TECs but not in CLU-null TECs. Further experiments showed that CLU-dependent prosurvival autophagy was associated with activation of the unfolded protein response in hypoxic kidney cells. In conclusion, these data suggest that activation of prosurvival autophagy by hypoxia in kidney cells requires CLU expression and may be a key cytoprotective mechanism of CLU in the protection of the kidney from hypoxia/ ischemia-mediated injury. [ABSTRACT FROM AUTHOR]

Renal repair begins soon after the kidney suffers ischemia-reperfusion injury (IRI); however, its molecular pathways are not fully understood. Clusterin (Clu) is a chaperone protein with cytoprotective functions in renal IRI. The aim of this study was to investigate the role of Clu in renal repair after IRI. IRI was induced in the left kidneys of wild-type (WT) C57BL/6J (B6) vs. Clu knockout (KO) B6 mice by clamping the renal pedicles for 28-45 min at the body temperature of 32°C. The renal repair was assessed by histology and confirmed by renal function. Gene expression was examined using PCR array. Here, we show that following IRI, renal tubular damage and Clu expression in WT kidneys were induced at day 1, reached the maximum at day 3, and significantly diminished at day 7 along with normal function, whereas the tubular damage in Clu KO kidneys steadily increased from initiation of insult to the end of the experiment, when renal failure occurred. Renal repair in WT kidneys was positively correlated with an increase in Ki67+ proliferative tubular cells and survival from IRI. The functions of Clu in renal repair and renal tubular cell proliferation in cultures were associated with upregulation of a panel of genes that could positively regulate cell cycle progression and DNA damage repair, which might promote cell proliferation but not involve cell migration. In conclusion, these data suggest that Clu is required for renal tissue regeneration in the kidney repair phase after IRI, which is associated with promotion of tubular cell proliferation. [ABSTRACT FROM AUTHOR]

There is no effective treatment for chronic rejection (CR) that largely limits long-term survival of kidney transplants. Transforming growth factor (TGF)-β is a fibrogenic factor for tissue fibrosis. This study was to test the efficacy of an anti-TGF-β antibody in preventing the CR of renal allografts in a preclinical model. Male Lewis rats (RT11) were orthotopically transplanted with donor kidneys from male Fischer 344 (RT11v1) rats and were treated with either anti-TGF-β or a control antibody. The CR of renal allografts was assessed by semiquantitative histological analyses, and intragraft cytokines and fibrosis-related genes ware examined by PCR arrays. Compared with the control antibody, anti-TGF-β antibody treatment significantly reduced recipients' proteinuria (P = 0.0002), and CR in renal transplants, which was indicated by the fewer injured renal tubules, glomeruli, and interlobular arterioles or arteries, and by less mononuclear cell infiltration and interstitial fibrosis in the anti-TGF-β antibody-treated group (P < 0.05), but not significantly attenuate the ratios of different infiltrating leukocytes. These pathological changes were associated with downregulation of TGF-β1, TGF-β2, and proinflammatory cytokines, or with upregulation of anti-fibrotic HGF, BMP5, and BMP7. The therapeutic effect of the anti-TGF-β antibody was further confirmed by its prevention of graft dysfunction, indicated by lower levels of serum creatinine and blood urea nitrogen or higher creatinine clearance in anti-TGF-β antibody-treated recipients compared with those in control recipients (P < 0.05). In conclusion, the anti-TGF-β antibody (1D11) treatment significantly reduces CR of renal allografts in rats, suggesting the therapeutic potential of this antibody therapy for treating CR of kidney transplants in patients. [ABSTRACT FROM AUTHOR]

Both humoral and cellular immune responses are involved in renal allograft rejection. Interleukin (IL)-6 is a regulatory cytokine for both B and Foxp3 (forkhead box P3)-expressing regulatory T (Treg) cells. This study was designed to investigate the impact of donor IL-6 production on renal allograft survival. Donor kidneys from IL-6 knockout (KO) vs. wild-type (WT) C57BL/6 mice (H-2b) were orthotopically transplanted to nephrotomized BALB/c mice (H-2d). Alloantibodies and Treg cells were examined by fluorescence-activated cell sorting analysis. Graft survival was determined by the time to graft failure. Here, we showed that a deficiency in IL-6 expression in donor kidneys significantly prolonged renal allograft survival compared with WT controls. IL-6 protein was upregulated in renal tubules and endothelium of renal allografts following rejection, which correlated with an increase in serum IL-6 compared with that in those receiving KO grafts or naive controls. The absence of graft-producing IL-6 or lower levels of serum IL-6 in the recipients receiving IL-6 KO allografts was associated with decreased circulating anti-graft alloantibodies and increased the percentage of intragraft CD4+CD25+Foxp3+ Treg cells compared with those with WT allografts. In conclusion, the lack of graft-producing IL-6 significantly prolongs renal allograft survival, which is associated with reduced alloantibody production and/or increased intragraft Treg cell population, implying that targeting donor IL-6 may effectively prevent both humoral and cellular rejection of kidney transplants. [ABSTRACT FROM AUTHOR]

Prevention of ischemia-reperfusion injury (IRI) is a challenge in clinical care of the patients with kidney transplants or acute kidney injury, and understanding of the intrinsic mechanisms of resistance to injury in the kidney will lead to a novel therapy. Clusterin, a secreted glycoprotein, is an antiapoptotic protein in cancer cells. Our study is to investigate the role of clusterin in renal IRI. Renal IRI in mice was induced by clamping renal vein and artery for 45 or 50 mm at 32°C. Apoptosis of renal tubular epithelial cells (TECs) was determined by FACS analysis. Clusterin expression was examined by Western blot or immunohistochemistry. Here, we showed that clusterin protein was induced in TECs following IRI, and more tubules expressed clusterin in the kidneys following ischemia at higher temperatures. In human proximal TEC HKC-8 cultures, clusterin was upregulated by removal of serum and growth factors in medium and was downregulated by TNF-α-IFN-γ mixture. The levels of clusterin were positively conelated with cell survival in these conditions. Knockdown or knockout of clusterin expression enhanced the sensitivity of TECs to apoptosis. In experimental models of renal IRI, deficiency in clusterin expression worsened the injury, as indicated by a significant increase in renal tissue damage with higher levels of serum creatinine and blood urea nitrogen and by a poorer recovery from the injury in cluster in deficient mice compared with wild-type mice. Our data indicate that the reduction of inducible expression of clusterin results in an increase in TEC apoptosis in the cultures and renders mice susceptibility to IRI, implying a protective role of clusterin in kidney injury. [ABSTRACT FROM AUTHOR]

Chronic allograft nephropathy (CAN), the most common cause of late kidney allograft failure, is not effectively prevented by immunosuppressive regimens. Activation of extracellular signal-regulated kinases 1 and 2 (ERK1/2) via MEK mediates actions of various growth factors, including transforming growth factor (TGF)-β1, which plays a key role in CAN. Hence, we tested the therapeutic potential of MEK-ERK1/2 signaling disruption to prevent CAN. Kidneys from C57BL/6J (H-2b) mice were transplanted to bilaterally nephrectomized BALB/c (H-2d) mice. At 14 days after transplantation, the recipients were subjected to 28 days of treatment with the MEK inhibitor CI-1040. All six CI-1040-treated allografts survived, while two of seven grafts in the vehicle-treated group were lost. At the end of the experiment, the function and structure of grafts in the CI- 1040-treated group were significantly preserved, as indicated by lower levels of serum creatinine or blood urea nitrogen than in the vehicle-treated group [30 ± 6 vs. 94 ± 39 μM creatinine (P = 0.0015) and 22 ± 8 vs. 56 ± 25 mM BUN (P = 0.0054)] and reduced CAN in the CI- 1040-treated group compared with vehicle controls (CAN score 4.2 vs. 10.3, P = 0.0119). The beneficial effects induced by CI-1040 were associated with reduction of ERK1/2 phosphorylation and TGFβ1 levels in grafts. Also, CI-1040 potently suppressed not only TGFβ biosynthesis in kidney cell cultures but also antiallograft immune responses in vitro and in vivo. Our data suggest that interference of MEK-ERK1/2 signaling with a pharmacological agent (e.g., CI-1040) has therapeutic potential to prevent CAN in kidney transplantation. [ABSTRACT FROM AUTHOR]