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Kidney fibrosis constitutes the shared final pathway of nearly all chronic nephropathies, but biomarkers for the non-invasive assessment of kidney fibrosis are currently not available. To address this, we characterize five candidate biomarkers of kidney fibrosis: Cadherin-11 (CDH11), Sparc-related modular calcium binding protein-2 (SMOC2), Pigment epithelium-derived factor (PEDF), Matrix-Gla protein, and Thrombospondin-2. Gene expression profiles in single-cell and single-nucleus RNA-sequencing (sc/snRNA-seq) datasets from rodent models of fibrosis and human chronic kidney disease (CKD) were explored, and Luminex-based assays for each biomarker were developed. Plasma and urine biomarker levels were measured using independent prospective cohorts of CKD: the Boston Kidney Biopsy Cohort, a cohort of individuals with biopsy-confirmed semiquantitative assessment of kidney fibrosis, and the Seattle Kidney Study, a cohort of patients with common forms of CKD. Ordinal logistic regression and Cox proportional hazards regression models were used to test associations of biomarkers with interstitial fibrosis and tubular atrophy and progression to end-stage kidney disease and death, respectively. Sc/snRNA-seq data confirmed cell-specific expression of biomarker genes in fibroblasts. After multivariable adjustment, higher levels of plasma CDH11, SMOC2, and PEDF and urinary CDH11 and PEDF were significantly associated with increasing severity of interstitial fibrosis and tubular atrophy in the Boston Kidney Biopsy Cohort. In both cohorts, higher levels of plasma and urinary SMOC2 and urinary CDH11 were independently associated with progression to end-stage kidney disease. Higher levels of urinary PEDF associated with end-stage kidney disease in the Seattle Kidney Study, with a similar signal in the Boston Kidney Biopsy Cohort, although the latter narrowly missed statistical significance. Thus, we identified CDH11, SMOC2, and PEDF as promising non-invasive biomarkers of kidney fibrosis.

An understanding of the ethical underpinnings of human subjects research that involves some risk to participants without anticipated direct clinical benefit-such as the kidney biopsy procedure as part of the Kidney Precision Medicine Project (KPMP)-requires a critical examination of the risks as well as the diverse set of countervailing potential benefits to participants. This kind of deliberation has been foundational to the development and conduct of the KPMP. Herein, we use illustrative features of this research paradigm to develop a more comprehensive conceptualization of the types of benefits that may be important to research participants, including respecting pluralistic values, supporting the opportunity to act altruistically, and enhancing benefits to a participant's community. This approach may serve as a model to help researchers, ethicists, and regulators to identify opportunities to better respect and support participants in future research that entails some risk to these participants as well as to improve the quality of research for people with kidney disease.

BACKGROUND: Kidney transplant recipients with coronavirus disease 2019 (COVID-19) are at increased risk for adverse outcomes, such as acute kidney injury (AKI), intensive care unit (ICU) admission, and death. The association of inflammatory biomarkers with outcomes and the impact of changes in immunosuppression on biomarker levels are unknown. METHODS: We investigated factors associated with a composite of AKI, ICU admission, or death, and whether immunosuppression changes correlated with changes in inflammatory biomarkers and outcomes in kidney transplant recipients with a positive severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) polymerase chain reaction. RESULTS: Of 59 patients, 50% had estimated glomerular filtration rate (eGFR)

Sepsis is a systemic inflammatory state in response to infection, and concomitant acute kidney injury (AKI) increases mortality significantly. Endoplasmic reticulum stress is activated in many cell types upon microbial infection and modulates inflammation. The role of endoplasmic reticulum signaling in the kidney during septic AKI is unknown. Here we tested the role of the spliced X-box binding protein 1 (Xbp1s), a key component of the endoplasmic reticulum stress- activated pathways, in the renal response to sepsis in the lipopolysaccharide (LPS) model. Xbp1s was increased in the kidneys of mice treated with LPS but not in other models of AKI, or several chronic kidney disease models. The functional significance of Xbp1s induction was examined by genetic manipulation in renal tubules. Renal tubule-specific overexpression of Xbp1s caused severe tubule dilation and vacuolation with expression of the injury markers Kim1 and Ngal, the pro-inflammatory molecules interleukin-6 (Il6) and Toll-like receptor 4 (Tlr4), decreased kidney function and 50% mortality in five days. Renal tubule-specific genetic ablation of Xbp1 had no phenotype at baseline. However, after LPS, Xbp1 knockdown mice displayed lower renal NGAL, pro-apoptotic factor CHOP, serum creatinine levels, and a tendency towards lower Tlr4 compared to LPS-treated mice with intact Xbp1s. LPS treatment in Xbp1s-overexpressing mice caused a mild increase in NGAL and CHOP compared to LPS-treated mice without genetic Xbp1s overexpression. Thus, increased Xbp1s signaling in renal tubules is unique to sepsis- induced AKI and contributes to renal inflammation and injury. Inhibition of this pathway may be a potential portal to alleviate injury.

Background and objectives Induction therapy with IL-2 receptor antagonist (IL2-RA) is recommended as a first line agent in living donor renal transplantation (LRT). However, use of IL2-RA remains controversial in LRT with tacrolimus (TAC)/mycophenolic acid (MPA) with or without steroids. Design, setting, participants, & measurements The Organ Procurement and Transplantation Network registry was studied for patients receiving LRT from 2000 to 2012 maintained on TAC/MPA at discharge ( n =36,153) to compare effectiveness of IL2-RA to other induction options. The cohort was initially divided into two groups based on use of maintenance steroid at time of hospital discharge: steroid ( n =25,996) versus no-steroid ( n =10,157). Each group was further stratified into three categories according to commonly used antibody induction approach: IL2-RA, rabbit anti-thymocyte globulin (r-ATG), and no-induction in the steroid group versus IL2-RA, r-ATG and alemtuzumab in the no-steroid group. The main outcomes were the risk of acute rejection at 1 year and overall allograft failure (graft failure or death) post-transplantation through the end of follow-up. Propensity score-weighted regression analysis was used to minimize selection bias due to non-random assignment of induction therapies. Results Multivariable logistic and Cox analysis adjusted for propensity score showed that outcomes in the steroid group were similar between no-induction (odds ratio [OR], 0.96; 95% confidence interval [95% CI], 0.86 to 1.08 for acute rejection; and hazard ratio [HR], 0.99; 95% CI, 0.90 to 1.08 for overall allograft failure) and IL2-RA categories. In the no-steroid group, odds of acute rejection with r-ATG (OR, 0.73; 95% CI, 0.59 to 0.90) and alemtuzumab (OR, 0.53; 95% CI, 0.42 to 0.67) were lower; however, overall allograft failure risk was higher with alemtuzumab (HR, 1.27; 95% CI, 1.03 to 1.56) but not with r-ATG (HR, 1.19; 95% CI, 0.97 to 1.45), compared with IL2-RA induction. Conclusions Compared with no-induction therapy, IL2-RA induction was not associated with better outcomes when TAC/MPA/steroids were used in LRT recipients. r-ATG appears to be an acceptable and possibly the preferred induction alternative for IL2-RA in steroid-avoidance protocols.

We previously reported that expression of the transcription factor interferon regulatory factor 1 (IRF1) is an early, critical maladaptive signal expressed by renal tubules during murine ischemic acute kidney injury (AKI). We now show that IRF1 mediates signals from reactive oxygen species (ROS) generated during ischemic AKI and that these signals ultimately result in production of α-subtypes of type I interferons (IFNαs). We found that genetic knockout of the common type I IFN receptor (IFNARI−/−) improved kidney function and histology during AKI. There are major differences in the spatial-temporal production of the two major IFN subtypes, IFNβ and IFNαs: IFNβ expression peaks at 4 h, earlier than IFNαs, and continues at the same level at 24 h; expression of IFNαs also increases at 4 h but continues to increase through 24 h. The magnitude of the increase in IFNαs relative to baseline is much greater than that of IFNβ. We show by immunohistology and study of isolated cells that IFNβ is produced by renal leukocytes and IFNαs are produced by renal tubules. IRF1, IFNαs, and IFNARI were found on the same renal tubules during ischemic AKI. Furthermore, we found that ROS induced IFNα expression by renal tubules in vitro. This expression was inhibited by small interfering RNA knockdown of IRF1. Overexpression of IRF1 resulted in the production of IFNαs. Furthermore, we found that IFNα stimulated production of maladaptive proinflammatory CXCL2 by renal tubular cells. Altogether our data support the following autocrine pathway in renal tubular cells: ROS > IRF1 > IFNα > IFNARI > CXCL2.

The elastic properties of renal glomeruli and their capillaries permit them to maintain structural integrity in the presence of variable hemodynamic forces. Measured by micro-indentation, glomeruli have an elastic modulus (E, Young’s modulus) of 2.1 kPa, and estimates from glomerular perfusion studies suggest that the E of glomeruli is between 2 and 4 kPa. F-actin depolymerization by latrunculin, inhibition of acto-myosin contractility by blebbistatin, reduction in ATP synthesis, and reduction of the affinity of adhesion proteins by EDTA reduced the glomerular E to 1.26, 1.7, 1.5, and 1.43 kPa, respectively. Actin filament stabilization with jasplakinolide and increasing integrin affinity with Mg2+ increased E to 2.65 and 2.87 kPa, respectively. Alterations in glomerular E are reflected in commensurate changes in F/G actin ratios. Disruption of vimentin intermediate filaments by withaferin A reduced E to 0.92 kPa. The E of decellularized glomeruli was 0.74 kPa, indicating that cellular components of glomeruli have dominant effects on their elasticity. The E of glomerular basement membranes measured by magnetic bead displacement was 2.4 kPa. Podocytes and mesangial cells grown on substrates with E values between 3 and 5 kPa had actin fibers and focal adhesions resembling those of podocytes in vivo. Renal ischemia and ischemia-reperfusion reduced the E of glomeruli to 1.58 kPa. These results show that the E of glomeruli is between 2 and 4 kPa. E of the GBM, 2.4 kPa, is consistent with this value, and is supported by the behavior of podocytes and mesangial cells grown on variable stiffness matrices. The podocyte cytoskeleton contributes the major component to the overall E of glomeruli, and a normal E requires ATP synthesis. The reduction in glomerular E following ischemia and in other diseases indicates that reduced glomerular E is a common feature of many forms of glomerular injury and indicative of an abnormal podocyte cytoskeleton.

There has been enormous progress in the understanding of acute kidney injury (AKI) over the last five years. This article reviews some of the salient new findings, the challenges revealed by these findings, and new insights into the pathogenesis of ischemic AKI. Clinical studies have demonstrated that even a small, transient rise in serum creatinine increases the risk of mortality in hospitalized patients and that a single event of AKI increases the risk for developing chronic kidney disease. Although the overall mortality rate from AKI has improved over the last two decades, it continues to be significant. Current treatment is focused on maintaining renal perfusion and avoiding volume overload. However, new therapeutic targets are emerging for the treatment of AKI as our understanding of the pathogenesis of ischemic injury and inflammation increases. Early diagnosis, however, continues to be challenging as the search continues for sensitive and specific biomarkers.

Toll-like receptor 4 (TLR4), a receptor forDamage Associated Molecular Pattern Molecules and also the lipopolysaccharide receptor, is required for early endothelial activation leading to maximal inflammation and injury during murine ischemic acute kidney injury. DNA microarray analysis of ischemic kidneys from TLR4-sufficient and deficient mice showed that pentraxin 3 (PTX3) was upregulated only on the former while transgenic knockout of PTX3 ameliorated acute kidney injury. PTX3 was expressed predominantly on peritubular endothelia of the outer medulla of the kidney in control mice. Acute kidney injury increased PTX3 protein in the kidney and the plasma where it may be a biomarker of the injury. Stimulation by hydrogen peroxide, or the TLR4 ligands recombinant human High-Mobility Group protein B1 or lipopolysaccharide, induced PTX3 expression in the Mile Sven 1 endothelial cell line and in primary renal endothelial cells suggesting that endothelial PTX3 was induced by pathways involving TLR4 and reactive oxygen species. This increase was inhibited by conditional endothelial knockout of Myeloid differentiation primary response gene 88, a mediator of a TLR4 intracellular signaling pathway. Compared to wild type mice, PTX3 knockout mice had decreased endothelial expression of cell adhesion molecules at 4 hours of reperfusion possibly contributing to a decreased early maladaptive inflammation in the kidneys of knockout mice. At 24 hours of reperfusion, PTX3 knockout increased expression of endothelial adhesion molecules when regulatory and reparative leukocytes enter the kidney. Thus, endothelial PTX3 plays a pivotal role in the pathogenesis of ischemic acute kidney injury.

IL-2 receptor antagonist (IL2-RA) is recommended as a first-line agent for induction therapy in renal transplantation. However, this remains controversial in deceased donor renal transplantation (DDRT) maintained on tacrolimus (TAC)/mycophenolic acid (MPA) with or without steroids.We studied the United Network for Organ Sharing Registry for patients receiving DDRT from 2000 to 2012 maintained on TAC/MPA at transplantation hospital discharge (n=74,627) to compare outcomes of IL2-RA and other induction agents. We initially divided the cohort into two groups on the basis of steroid use at the time of discharge: steroid (n=59,010) versus no steroid (n=15,617). Each group was stratified into induction categories: IL2-RA, rabbit antithymocyte globulin (r-ATG), alemtuzumab, and no induction. The main outcomes were incidence of acute rejection within the first year and overall graft failure (defined as graft failure and/or death) post-transplantation. Propensity score (PS), specifically inverse probability of treatment weight, analysis was used to minimize selection bias caused by nonrandom assignment of induction therapies.Median (25th, 75th percentiles) follow-up times were 3.9 (1.1, 5.9) and 3.2 (1.1, 4.9) years for steroid and no steroid groups, respectively. Acute rejection within the first year and overall graft failure within 5 years of transplantation were more common in the no induction category (13.3%; P0.001 and 28%; P=0.01, respectively) in the steroid group and the IL2-RA category (11.1%; P=0.16 and 27.4%; P0.001, respectively) in the no steroid group. Compared with IL2-RA, PS-weighted and covariate-adjusted multivariable logistic and Cox analyses showed that outcomes in the steroid group were similar among induction categories, except that acute rejection was significantly lower with r-ATG (odds ratio [OR], 0.68; 95% confidence interval [95% CI], 0.62 to 0.74). In the no steroid group, compared with IL2-RA, odds of acute rejection with r-ATG (OR, 0.80; 95% CI, 0.60 to 1.00) and alemtuzumab (OR, 0.68; 95% CI, 0.53 to 0.88) were lower, and r-ATG was associated with better graft survival (hazard ratio, 0.86; 95% CI, 0.75 to 0.99).In DDRT, compared with IL2-RA induction, no induction was associated with similar outcomes when TAC/MPA/steroids were used. r-ATG seems to offer better graft survival over IL2-RA in steroid avoidance protocols.

Although leukocytes infiltrate the kidney during ischemic acute kidney injury (AKI) and release interleukin 6 (IL6), their mechanism of activation is unknown. Here, we tested whether Toll-like receptor 4 (TLR4) on leukocytes mediated this activation by interacting with high-mobility group protein B1 (HMGB1) released by renal cells as a consequence of ischemic kidney injury. We constructed radiation-induced bone marrow chimeras using C3H/HeJ and C57BL/10ScNJ strains of TLR4 (−/−) mice and their respective TLR4 (+/+) wild-type counterparts and studied them at 4h after an ischemic insult. Leukocytes adopted from TLR4 (+/+) mice infiltrated the kidneys of TLR4 (−/−) mice, and TLR4 (−/−) leukocytes infiltrated the kidneys of TLR4 (+/+) mice but caused little functional renal impairment in each case. Maximal ischemic AKI required both radiosensitive leukocytes and radioresistant renal parenchymal and endothelial cells from TLR4 (+/+) mice. Only TLR4 (+/+) leukocytes produced IL6 in vivo and in response to HMGB1 in vitro . Thus, following infiltration of the injured kidney, leukocytes produce IL6 when their TLR4 receptors interact with HMGB1 released by injured renal cells. This underscores the importance of TLR4 in the pathogenesis of ischemic AKI.

Ischemic acute kidney injury (AKI) triggers expression of adaptive (protective) and maladaptive genes. Agents that increase expression of protective genes should provide a therapeutic benefit. We now report that bardoxolone methyl (BARD) ameliorates ischemic murine AKI as assessed by both renal function and pathology. BARD may exert its beneficial effect by increasing expression of genes previously shown to protect against ischemic AKI, NF-E2-related factor 2 (Nrf2), peroxisome proliferator-activated receptor-γ (PPARγ), and heme oxygenase 1 (HO-1). Although we found that BARD alone or ischemia-reperfusion alone increased expression of these genes, the greatest increase occurred after the combination of both ischemia-reperfusion and BARD. BARD had a different mode of action than other agents that regulate PPARγ and Nrf2. Thus we report that BARD regulates PPARγ, not by acting as a ligand but by increasing the amount of PPARγ mRNA and protein. This should increase ligand-independent effects of PPARγ. Similarly, BARD increased Nrf2 mRNA; this increased Nrf2 protein by mechanisms in addition to the prolongation of Nrf2 protein half-life previously reported. Finally, we localized expression of these protective genes after ischemia and BARD treatment. Using double-immunofluorescence staining for CD31 and Nrf2 or PPARγ, we found increased Nrf2 and PPARγ on glomerular endothelia in the cortex; Nrf2 was also present on cortical peritubular capillaries. In contrast, HO-1 was localized to different cells, i.e., tubules and interstitial leukocytes. Although Nrf2-dependent increases in HO-1 have been described, our data suggest that BARD's effects on tubular and leukocyte HO-1 during ischemic AKI may be Nrf2 independent. We also found that BARD ameliorated cisplatin nephrotoxicity.

Ischemic acute kidney injury (AKI) triggers an inflammatory response which exacerbates injury that requires increased expression of endothelial adhesion molecules. To study this further, we used in situ hybridization, immunohistology, and isolated endothelial cells, and found increased Toll-like receptor 4 (TLR4) expression on endothelial cells of the vasa rectae of the inner stripe of the outer medulla of the kidney 4h after reperfusion. This increase was probably due to reactive oxygen species, known to be generated early during ischemic AKI, because the addition of hydrogen peroxide increased TLR4 expression in MS1 microvascular endothelial cells in vitro . Endothelial TLR4 may regulate adhesion molecule (CD54 and CD62E) expression as they were increased on endothelia of wild-type but not TLR4 knockout mice in vivo . Further, the addition of high-mobility group protein B1, a TLR4 ligand released by injured cells, increased adhesion molecule expression on endothelia isolated from wild-type but not TLR4 knockout mice. TLR4 was localized to proximal tubules in the cortex and outer medulla after 24h of reperfusion. Thus, at least two different cell types express TLR4, each of which contributes to renal injury by temporally different mechanisms during ischemic AKI.

The clinical utility of cisplatin is limited by nephrotoxicity. Oh et al. report that β-lapachone prevents this nephrotoxicity but not cisplatin's cytotoxicity for cancers. In addition to its potential clinical importance, the beneficial effect of β-lapachone on cisplatin acute kidney injury may illustrate fundamental processes that ordinarily link alterations in nutrient availability and intracellular reactive oxygen species on the one hand, with inflammation and cell death on the other hand.

Antibodies against HLA antigens offer a window to the response of transplant recipients to the challenge of allografts. HLA alleles produced by recombinant technology and attached to color-coded polystyrene microspheres are now in widespread use. They offer antibody detection by flow cytometry at a high level of sensitivity. Methods used in the author's laboratory for the interpretation of these tests are described. They include a correction for variable antigen density, and the binding of normal immunoglobulins to the conjugated beads. The 'normalized ratio' provides a good method of scoring results. Antibodies specific for the antigens of the donors were associated with kidney and heart allograft loss. Antibodies against crossreactive antigens, without reactivity against donor HLA, did not appear to increase the risk of developing chronic rejection. Antibody production by peripheral blood B cells was analyzed. The idea of sequestration of HLA antibodies in the graft as a major cause for their absence in the blood after transplantation is discussed. IgM antibodies against donor HLA antigens were found before transplantation and also were produced both early and late in the post-transplant course. In addition, IgM antibodies against donor HLA appeared to be associated with decreased survival of kidney and heart allografts.

We report a renal transplant patient with a functioning allograft who had an inferior vena cava (IVC) filter placed above the renal allograft vein. The patient occluded the IVC filter and developed extensive distal thrombosis. This included complete occlusion of the renal allograft vein (RAV). However, this complication did not lead to a loss of kidney transplant function. Alternate allograft venous drainage via capsular collaterals and subsequent recanalization of transplant vein were demonstrated.

The actin cytoskeleton is dynamically remodeled during Fcgamma receptor (FcgammaR)-mediated phagocytosis in a phosphatidylinositol (4,5)-bisphosphate (PIP(2))-dependent manner. We investigated the role of type I phosphatidylinositol 4-phosphate 5-kinase (PIP5K) gamma and alpha isoforms, which synthesize PIP(2), during phagocytosis. PIP5K-gamma-/- bone marrow-derived macrophages (BMM) have a highly polymerized actin cytoskeleton and are defective in attachment to IgG-opsonized particles and FcgammaR clustering. Delivery of exogenous PIP(2) rescued these defects. PIP5K-gamma knockout BMM also have more RhoA and less Rac1 activation, and pharmacological manipulations establish that they contribute to the abnormal phenotype. Likewise, depletion of PIP5K-gamma by RNA interference inhibits particle attachment. In contrast, PIP5K-alpha knockout or silencing has no effect on attachment but inhibits ingestion by decreasing Wiskott-Aldrich syndrome protein activation, and hence actin polymerization, in the nascent phagocytic cup. In addition, PIP5K-gamma but not PIP5K-alpha is transiently activated by spleen tyrosine kinase-mediated phosphorylation. We propose that PIP5K-gamma acts upstream of Rac/Rho and that the differential regulation of PIP5K-gamma and -alpha allows them to work in tandem to modulate the actin cytoskeleton during the attachment and ingestion phases of phagocytosis.

Purpose of review Ischemic acute kidney injury may be exacerbated by an inflammatory response. How injury elicits inflammation remains a major question in understanding acute kidney injury. The present review examines the hypothesis that molecules released by injured cells elicit inflammation. Recent findings After necrotic death, intracellular molecules find their way into the extracellular space. These molecules include heat shock proteins and HMGB1. Receptors for these proteins include TLR4, TLR2, CD91 and RAGE. These proinflammatory mechanisms may be so useful that nature has evolved mechanisms for programming necrotic death via poly(ADP-ribose) polymerase and cyclophilin D. In addition, apoptosis may also elicit inflammation. Summary The concepts discussed in this review are important for clinical medicine. Drugs and genetic manipulation may ameliorate ischemic kidney injury by regulating the inflammatory response to cell injury.

T lymphocytes are established mediators of ischemia reperfusion (IR)–induced AKI, but traditional immune principles do not explain their mechanism of early action in the absence of alloantigen. Nrf2 is a transcription factor that is crucial for cytoprotective gene expression and is generally thought to have a key role in dampening IR-induced AKI through protective effects on epithelial cells. We proposed an alternative hypothesis that augmentation of Nrf2 in T cells is essential to mitigate oxidative stress during IR-induced AKI. We therefore generated mice with genetically amplified levels of Nrf2 specifically in T cells and examined the effect on antioxidant gene expression, T cell activation, cytokine production, and IR-induced AKI. T cell–specific augmentation of Nrf2 significantly increased baseline antioxidant gene expression. These mice had a high frequency of intrarenal CD25+Foxp3+ regulatory T cells and decreased frequencies of CD11b+CD11c+ and F4/80+ cells. Intracellular levels of TNF-α, IFN-γ, and IL-17 were significantly lower in CD4+ T cells with high Nrf2 expression. Mice with increased T cell expression of Nrf2 were significantly protected from functional and histologic consequences of AKI. Furthermore, adoptive transfer of high-Nrf2 T cells protected wild-type mice from IR injury and significantly improved their survival. These data demonstrate that T cell–specific activation of Nrf2 protects from IR-induced AKI, revealing a novel mechanism of tissue protection during acute injury responses.

The role of IL-6 was investigated in murine ischemic acute renal failure. The renal pedicles were clamped for 17 min, and the mice were studied at various times after reperfusion. We found that serum IL-6 increased after murine ischemic renal injury. This increase was associated with increased IL-6 mRNA in the ischemic kidney but not in the contralateral kidney or the liver. Maximal IL-6 production occurred at 4 to 8 h and decreased to baseline by 24 h. Reperfusion of the kidney was required for IL-6 production. In situ hybridization and immunohistochemistry showed that macrophages infiltrated areas adjacent to the vascular bundles in the outer medulla within hours of reperfusion and showed that these macrophages produced IL-6 mRNA. For understanding how macrophages were stimulated to produce IL-6, an in vitro model in which S3 proximal tubular cells were injured by reactive oxygen species was set up. These injured cells released molecules that activated macrophages to produce IL-6 in vitro. IL-6 that was produced in response to renal ischemia was maladaptive because transgenic knockout of IL-6 ameliorated renal injury as measured by serum creatinine and histology. IL-6 transgenic knockout mice were lethally irradiated, and their bone marrow was reconstituted with wild-type IL-6 cells. Such bone marrow transfers abolished the protective effects of transgenic IL-6 knockout. It is concluded that macrophages infiltrate the area of the vascular bundles of the outer medulla, these macrophages produce IL-6, and this IL-6 exacerbates ischemic murine acute renal failure.

The lipopolysaccharide-triggered mesangial transcriptome: Evaluating the role of interferon regulatory factor-1. Background Presently, we do not have a clear picture of how the mesangial transcriptome evolves following stimulation. The present study was designed to address this, using an innate trigger to stimulate murine mesangial cells. Methods Three independent mesangial cell lines derived from C57BL/6 mice were stimulated with lipopolysaccharide (LPS). The mesangial cell transcriptomes were defined 1, 6, 24, and 60 hours poststimulation with LPS, using a 17,000 gene oligonucleotide array. Results Interferon regulatory factor-1 ( IRF-1 ), ScyA2/MCP1 , ScyA20/MIP3 α ( ScyB1/Gro1 , and ScyB2/MIP2α/Gro2 were the earliest genes to be hyperexpressed after LPS stimulation. Later-appearing genes included ScyA7/MCP3 , ScyD1/fractalkine , GM-CSF/CSF-2 , PDGF, epiregulin, NfKb , C/EBP , TIMP-1 , MMP11 , MMP13 , PTGS2/COX2 , SpI2-1 , Spp1 , PAI-1 , VCAM-1, C3 , and defensin-β1 , among others. Several of these changes were validated by real-time polymerase chain reaction (PCR) or enzyme-linked immunosorbent assay (ELISA). Rapid IRF-1 hyperexpression was also noted following stimulation of mesangial cells with peptidoglycan, poly I:poly C, interferon-γ?(IFN-γ), and heat-aggregated IgG. However, the blocking of IRF-1 using RNA interference and the use of mesangial cells isolated from IRF-1 –deficient mice could not substantiate an obligatory role for IRF-1 in LPS-induced mesangial cell activation. Likewise, IRF-1 deficiency did not impact the development of anti-glomerular basement membrane (GBM)-induced immune nephritis. Conclusion Innate stimuli such as LPS appear to trigger successive waves of mesangial cell gene expression. Although IRF-1 surfaces as an "early-on, early-off" transcription factor following several different triggers, it does not appear to be an essential molecule for mesangial cell activation by innate triggers or for anti-GBM disease.

STAT3 is a ubiquitous transcription factor that is indispensable during early embryogenesis. To study the functions of STAT3 postnatally, we generated conditional STAT3-deficient mice. To that end, STAT3(lox/lox) mice were crossed with mice expressing Cre under the control of rat insulin II gene promoter (RIP-Cre mice). Immunohistochemical and Western blot analyses showed that STAT3 is deleted from beta cells in the islets of Langerhans. Genomic DNA PCR revealed that STAT3 deletion also occurred in the hypothalamus. Hypothalamic Cre expression was further confirmed by crossing RIP-Cre/STAT3(lox/lox) mice with the ROSA26 Cre reporter strain and staining for lacZ activity. Double immunohistochemical staining confirmed that deletion of STAT3 occurred in leptin receptor (OB-Rb isoform)-positive neurons. RIP-Cre/STAT3(lox/lox) mice are mildly hyperglycemic and hyperinsulinemic at the time of weaning, become hyperphagic immediately after weaning, and exhibit impaired glucose tolerance. Body weight, body fat, and mRNA and protein levels of leptin are all significantly increased in RIP-Cre/STAT3(lox/lox) mice. Administration of recombinant leptin by intracerebroventricular infusion failed to cause complete loss of body fat in RIP-Cre/STAT3(lox/lox) mice. Transplantation of wild-type islets into RIP-Cre/STAT3(lox/lox) mice also failed to decrease adiposity or to correct other abnormalities in these mice. These data thus suggest that loss of STAT3 in the hypothalamus caused by RIP-Cre action likely interferes with normal body weight homeostasis and glucose metabolism.

Background: Biliary obstruction affects the renal response to ischemia and also elicits a hepatic cytokine response. Using a murine model, we now test the hypothesis that these hepatic cytokines help determine the outcome of ischemic acute renal failure. Methods: C3H/HEN mice were subjected to bile duct ligation 24 h (ABDL) or 7 days (CBDL) prior to induction of acute ischemic renal failure (ARF). Serum creatinine (Scr), cytokine mRNA abundance, and renal histology were studied 24 h after renal ischemia. Results: ABDL prior to ARF resulted in amelioration of renal injury (Scr 0.7 ± 0.1 mg/dl compared to 2.5 ± 0.1 mg/dl in sham/ARF group, (mean ± SE, n = 11/group). CBDL exacerbated renal injury. Increased hepatic mRNA for interleukin-10 (IL10) and interleukin-1 receptor antagonist (IL1RA) was detected in the ABDL/ARF group but not in the CBDL/ARF group. These data suggest that hepatic production of IL10 and IL1RA in response to ABDL ameliorates ischemic ARF, an effect that is lost after several days of BDL. Conclusion: Our data support the concept that hepatic cytokines modulate renal injury. This adds a new dimension in our understanding of renal injury in the setting of hepatic disease.

This study demonstrates that intraperitoneal injections of DHA (all cis 4,7,10,13,16,19 docosahexaenoic acid C22: n-3) bound to bovine serum albumin ameliorate murine acute renal failure (ARF) induced by temporary occlusion of the renal artery. Three micromoles of DHA decreased serum creatinine (Scr) from 2.3 mg/dl to 1.1 mg/dl 24 h after reperfusion (n = 15; P < 0.05). Scr of the treated animals were significantly lower than controls throughout a 7-d time course. Although lower doses of DHA were less effective, higher doses were not more effective. Ribonuclease (RNase) protection assays showed that ischemia increased mRNA abundance for TNF-alpha and inducible nitric oxide synthase (iNOS) at 24 h. This increase was prevented by DHA administration. Because TNF-alpha and iNOS contribute to renal ischemic injury, their inhibition may contribute to DHA's salutary effect. In addition, the data may have therapeutic implications, because the DHA improves ARF even when administered at 4 h after reperfusion.

Acute pancreatitis results in many deaths each year. Our understanding of the pathophysiology is limited. To better understand the impact of apoptosis versus necrosis, we compared cerulein-induced pancreatitis in Fas-deficient (MRL lpr/lpr) versus Fas-sufficient (MRL +/+) mice. Average amylase values in Fas-deficient mice were substantially greatr than in the Fas-sufficient mouse. Histology graded on edema, inflammation, vacuolization, and necrosis showed greater injury in the Fas-deficient mouse. This finding suggests that the Fas pathway is important in controlling cerulein-induced pancreatitis.

Face transplants have been clinically established, and early acute rejections have been reported. Late acute rejections have been less common. Immediate and accurate diagnosis along with successful treatment is critical to prevent graft damage. This case report describes the successful treatment of a severe, steroid-resistant rejection 2 years after a full face transplant.

Recent data suggest that long-term allograft survival might be affected by two factors. The first is the endowment of the allograft, which consists of two elements: the nephron mass and the ability of these nephrons to repair injuries sustained during the transplant process. The second factor is renal inflammation. Although inflammation is traditionally ascribed to alloreactivity, recent data have shown that there is also a renal inflammatory response to early injury after transplantation, to brain death in the donor, and as part of the maladaptive response to nephron loss. These two factors contribute to the detrimental effects of delayed graft function or acute rejection on the long-term survival seen in most studies, and the beneficial effects of anti-inflammatory agents on the maladaptive response to nephron loss.

Background Nitric oxide produced by inducible nitric oxide synthase (iNOS) may be cytotoxic during cardiac, hepatic, and renal acute allograft rejection. Because the incidence of rejection is decreased by fish oils that contain docosahexaenoic acid (DHA), we investigated the effects of DHA on iNOS. Using nuclear run-on assays and iNOS-promoter constructs, we previously showed that docosahexaenoic acid (DHA) inhibits activation of the iNOS gene by murine macrophages that had been stimulated in vitro by IFNgamma plus lipopolysaccharide. Methods In our current investigation, our purpose has been to determine how DHA inhibits iNOS gene activation in murine macrophages, by using gel retardation and Northern blotting techniques. We studied the effects of DHA on the formation nuclear protein complexes that interact with the critical iNOS promoter's response elements for IRF-1 (IRF-E -923 to -913 bp) and NF-kappaB (NFkappa d -85 to -75 bp). Results We now show that DHA inhibited increases of IRF-1 mRNA abundance in response to IFNgamma plus lipopolysaccharide. As expected, we found that this prevented formation of the nuclear protein complex that binds to the IRF-E DNA response element. We also found that inhibition of IRF-1 inhibited formation of the nuclear protein complex that binds to the NFkappa d DNA response element. Conclusions DHA decreases the abundance of IRF-1 mRNA in stimulated cells. That, in turn, results in the decreased nuclear protein binding to the major iNOS promoter response elements (IRF-E and NF-kappaB). We found that this occurred because IRF-1 is a component of both the nuclear protein complex that binds to IRF-E and the nuclear protein complex that binds to NFkappa d.

Dendritic cells participate in the regulation of CD4 and CD8 T cells during transplant rejection. Understanding what causes increased numbers of dendritic cells to appear in the renal transplant is therefore important. We performed syngeneic renal transplants between rats. We used the monoclonal antibody OX62 to detect dendritic cells, and OX6 to detect major histocompatability complex (MHC) Class II in the renal transplant. One week after transplant, dendritic cells appeared. This indicates that the injury of transplantation itself is sufficient to increase the number of dendritic cells in the kidney in a model where there is no alloreactivity.

Murine macrophage activation is deficient in the fetus and the neonate, and in areas of the placenta perfused by the fetal circulation. Fetal and neonatal serum concentrations of docosahexaenoic acid (DHA) are 150 microM, or approximately 50-fold higher than in the adult. We previously showed that DHA inhibits activation of the gene for inducible nitric oxide synthase (iNOS) in murine macrophages stimulated in vitro with interferon gamma (IFN gamma) plus lipopolysaccharide (LPS). We have now pursued these observations in greater depth. An assay system was developed which separated the stimulation of macrophages by IFN gamma plus LPS, and the actual production of nitric oxide (NO). It was found that macrophages do not produce NO until they have been stimulated by IFN gamma plus LPS for a period of 10 h. NO is produced during the subsequent 10 h, even though IFN gamma plus LPS are not longer present. DHA, if present, inhibited only during the initial 10 h stimulation; DHA did not inhibit the production of NO by macrophages which had previously been stimulated by IFN gamma plus LPS, and were already producing NO. It was also found that DHA was less inhibitory if given prior to the IFN gamma plus LPS stimulation. In a dose-responsive manner, DHA inhibited the increased abundance of iNOS mRNA by macrophages stimulated by IFN gamma plus LPS. NO contributes to the host defense against Listeria monocytogenes and other intracellular pathogens. We therefore investigated the ability of DHA to inhibit NO production by macrophages stimulated by IFN gamma plus Listeria monocytogenes in vitro; DHA inhibited transcription of the iNOS gene and also the listeriocidal activity of activated macrophages. Inhibition of NO production by DHA may contribute to the increased susceptibility of the fetoplacental unit and neonate to intracellular infections.

Acute ischemia reperfusion injury (IRI) is a major cause of acute kidney injury (AKI). A major conceptual insight is that the total amount of renal injury is the result of not only the initial ischemic insult, by the ensuing responses. This simple insight has profound clinical implications: it suggests that therapy after the initial insult has already occurred may ameliorate the course of AKI. Some of these responses are maladaptive. Examples include inappropriate intrarenal hemodynamics, altered mitochondrial and other metabolic functions, endothelial dysfunction, and tubular obstruction and back-leak. Other responses are reparative. Examples include the production of growth factors such as Wnt7.

PROBLEM: We previously demonstrated profound inhibition of macrophage activation in the murine placenta in vivo. Given the importance of macrophages both in initiating cellular immunity by presenting antigen in the context of Ia to CD4+ T cells, and in killing cellular targets by producing nitric oxide (NO), inhibition of macrophage functions in the placenta may account for the increased susceptibility of the placenta to infection. We have also showed that docosahexaenoic acid (DHA), at concentrations present in the fetal circulation, has a major role in inhibiting macrophage Ia-expression and NO production in the placenta. The concentration of DHA in fetal serum perfusing the placenta is 50x higher than in the adult. DHA has previously been reported to profoundly affect prostanoid production, to be metabolized by lipoxygenases, and to affect lipoxygenases. We now determine if these activities of DHA account for its inhibition of macrophage NO production and Ia-expression. METHODS: Murine macrophages were cultured in vitro, exposed to IFNγ, endotoxin, DHA, and various eicosanoids, and their ability to produce NO or express Ia determined. RESULTS: Although the cyclooxygenase inhibitor, indomethacin, did inhibit NO production, DHA inhibited by a different mechanism. DHA further inhibited NO production by macrophages exposed to doses of indomethacin known to maximally inhibit prostanoid production. Stable, biologically active prostanoids did not reverse the inhibitory effect of DHA. Although DHA is metabolized by lipoxygenases, the lipoxygenase inhibitor NDGA did not reverse the inhibition of either NO production nor Ia expression. This indicates that lipoxygenase products of DHA did not mediate inhibition. NDGA itself inhibited NO production and Ia expression. However, DHA did not inhibit by inhibiting lipoxygenase activity because DHA further inhibited macrophages exposed to doses of DHA known to maximally inhibit lipoxygenases. Furthermore, stable biologically active analogs of lipoxygenase products did not reverse DHA inhibition. DHA also did not inhibit by preventing PAF production because PAF did not reverse inhibition of NO production. CONCLUSION: DHA did not inhibit Ia-expression or NO production via its known effects on eicosanoid or PAF metabolism, nor by being metabolized by lipoxygenases.

listeria monocytogenes is an intracellular bacterial pathogen. A single gene product, listeriolysin (LLO), is critical for the induction of protective immunity. We now show that listeria that produce functional LLO augment Ia expression by macrophages and are better presented to a Th1, CD4+ anti-listeria T cell line. We used two genetically engineered strains of listeria which differed only in their ability (Ly+) or inability (Ly-) to produce functional LLO. Ia-negative murine macrophages ingested either Ly+ or Ly-, and then were stimulated by interferon-γ(IFN-γ). Increasing numbers of live Ly+, but not Ly-, augmented IFN-γ-induced Ia expression. Ly+ by itself did not induce Ia expression. Heat-killed Ly+ and Ly- did not augment IFN-γ-induced Ia expression. The abundance of Ia on the macrophage cell surface is one major determinant of antigen presentation to CD4+ T cells. Consistent with their ability to augment Ia expression, Ly+ were better presented than Ly- to a CD4+, Th1, anti-listeria T cell line. When macrophages and T cells were from different inbred mouse strains, antigen presentation required identity at the Class II region of the MHC gene complex. This indicated that antigen presentation occurred via Ia molecules. The increased ability of macrophages to present Ly+ is a product of the macrophage-listeria interaction, not a property of the T cell line 86. If Ia-negative macrophages ingested listeria and were then stimulated by IFN-γ, Ly+ was presented more efficiently than Ly-. On the other hand, if Ia-positive macrophages ingested Listeria, then Ly+ and Ly- were presented equally well to T cells. Altogether our data is consistent with the hypothesis that macrophages interact differently with Ly+, and that this contributes to the ability of only live Ly+-to induce protective immunity.

Macrophage activation is deficient in the fetus and neonate when the serum concentrations of docosahexaenoic acid (DHA) are 150 microM, or 10-50-fold higher than in the adult. We now show that DHA inhibits production of nitric oxide (NO) by macrophages stimulated in vitro by IFNgamma plus LPS, or by IFNgamma plus TNFalpha. The half-maximal inhibitory activity of DHA was approximately 25 microM. There were strict biochemical requirements of the fatty acid for inhibition. Polyenoic fatty acids with 22 carbons were more inhibitory than those with 20 carbons. Among 22-carbon fatty acids, those with a greater number of double bonds and a double bond in the n-3 position were more inhibitory. DHA was the most inhibitory of the polyenoic acids we tested. Inducible nitric oxide synthase (iNOS) is the enzyme responsible for the production of NO by macrophages. NO production is initiated after new iNOS enzyme is synthesized following transcription of the iNOS gene. In macrophages stimulated by IFNgamma plus LPS, DHA inhibited accumulation of iNOS mRNA, as measured by Northern blotting, and iNOS transcription, as measured by nuclear run-on assays. We transfected RAW 264.7 macrophages with a construct containing the iNOS promoter fused to the chloramphenicol acetyl transferase gene. DHA inhibited activation of this promoter by IFN gamma plus LPS. By inhibiting iNOS transcription in the fetus and neonate, DHA may contribute to their increased susceptibility to infection.

To understand the increased susceptibility of preterm neonates to infection.A murine listeriosis model using immunohistochemistry, microarray technology, and real-time polymerase chain reaction (PCR).We report that recombinant serum amyloid A (SAA) administered prophylactically 18 h before intraperitoneal (i.p.) inoculation with Listeria monocytogenes conferred a dramatic survival benefit compared with administration of only vehicle in neonatal mice. Neonates that received the recombinant SAA protein had significantly fewer Listeria colony counts on plating of infected liver and showed significantly more activated macrophages, but SAA did not affect postnatal growth. Real-time PCR was used to confirm the microarray findings that gene expression levels for the SAA proteins 1 (Saa1) and 2 (Saa2), in addition to that for orosomucoid-2 (Orm2), were strikingly elevated in the adult compared with those in the neonate. Real-time PCR analysis showed that of the acute phase cytokines, tumor necrosis factor (TNF) gene expression increased exponentially with time in the infected adult, whereas neonates did not show similar increases.The increased susceptibility of neonatal mice to listeriosis is in part mediated by a deficiency in the acute phase response, specifically expression of SAA, and that prophylactic SAA protein before neonatal murine listeriosis results in more macrophage activation, lower Listeria counts, and greater survival.

Ischemic acute kidney injury (AKI) contributes to considerable morbidity and mortality in hospitalized patients and can contribute to rejection during kidney transplantation. Maladaptive immune responses can exacerbate injury, and targeting these responses holds promise as therapy for AKI. In the last decade, a number of molecules and receptors were identified in the innate immune response to ischemia–reperfusion injury. This review primarily focuses on one pathway that leads to maladaptive inflammation: toll-like receptor 4 (TLR4) and one of its ligands, high mobility group box protein 1 (HMGB1). The temporal–spatial roles and potential therapeutics targeting this particular receptor–ligand interaction are also explored.

Renal transplantation is the preferred treatment modality for patients with ESRD who are good surgical risks and able to comply with chronic immunosuppressive medications. Clinical transplantation has advanced significantly, with most transplant centers reporting 1-yr renal allograft survival rates of better than 80%. Nevertheless, rejection and a progressive loss of allografts over time continue to occur. The immunosuppressive agents currently used may lead to the development of life-threatening infections, malignancies, and advanced atherosclerosis as a consequence of some of their side effects. This review examines the mechanisms involved in allograft rejection as currently understood. The recent knowledge into the mechanism of action of cyclosporine, FK506, and rapamycin on T cell activation is presented. Information recently available on some of the established therapies such as steroids, antimetabolites and monoclonal antibodies as well as the newer agents is also discussed. The interaction between clinical transplantation and basic research in immunology continues to result in exciting advances in both fields.

Because of their paternal antigens, the fetus and placenta may be considered an allograft in the maternal host. Understanding the mechanisms which prevent maternal immunological rejection of the fetus remains a fundamental unsolved problem in immunology. We have previously reported that macrophages are inhibited by maternal decidual stromal cells residing at the maternal-fetal interface. In view of the central role of macrophages in cell-mediated immunity, this inhibition may contribute to preventing maternal antifetal responses. We now report that it was the solid phase signals embedded in the extracellular matrix (ECM) made by decidual cells which are responsible for inhibiting macrophage-mediated lysis of TNF-alpha-resistant P815 mastocytoma cells. The latter macrophage function is acquired after stimulation by interferon gamma and endotoxin. All these macrophage functions were also inhibited by ECM isolated from the Engelberth-Holm-Swarme (EHS) tumor. This tumor ECM has a similar biochemical composition to decidual ECM. This ECM inhibited the effector, as opposed to the stimulator, phase of macrophage-mediated tumor lysis. Laminin, type IV collagen, and heparan sulfate proteoglycans, the major known components of decidual and EHS ECMs, did not inhibit the above macrophage functions. Altogether these data indicate that macrophages were inhibited by solid phase signals embedded in decidual and EHS ECMs. Whether the solid phase signals in these two ECMs are biochemically identical remains to be determined. To our knowledge, such signals are a novel pathway of inhibiting macrophage functions which may be important in understanding the maternal-fetal immunologic relationship, and the pathogenesis of perinatal infections. Furthermore, the ability of EHS tumor ECM to inhibit macrophage functions may indicate that some tumors may defend themselves against host macrophage responses using solid phase signals. This may be important in understanding some host-tumor relationships.

Acute renal ischemia elicits an inflammatory response that may exacerbate acute kidney injury, but the regulation of the initial signals that recruit leukocytes is not well understood. Here, we found that IFN regulatory factor 1 (IRF-1) was a critical, early proinflammatory signal released during ischemic injury in vitro and in vivo. Within 15 min of reperfusion, proximal tubular cells of the S3 segment produced IRF-1, which is a transcription factor that activates proinflammatory genes. Transgenic knockout of IRF-1 ameliorated the impairment of renal function, morphologic injury, and inflammation after acute ischemia. Bone marrow chimera experiments determined that maximal ischemic injury required IRF-1 expression by both leukocytes and radioresistant renal cells, the latter identified as S3 proximal tubule cells in the outer medulla by in situ hybridization and immunohistochemistry. In vitro, reactive oxygen species, generated during ischemia/reperfusion injury, stimulated expression of IRF-1 in an S3 proximal tubular cell line. Taken together, these data suggest that IRF-1 gene activation by reactive oxygen species is an early signal that promotes inflammation after ischemic renal injury.

Because of their favorable effects on renal hemodynamics, calcium antagonists may have a major role in the prevention and management of certain types of acute renal dysfunction. In fact, verapamil (VP) was shown to prevent cyclosporin A (CsA)-induced decreases in RBF in mice and in cadaver renal transplant (CRT) recipients. The study presented here of 59 cadaver renal transplant patients evaluates the outcome from perioperative treatment with VP (N = 30) administered intraoperatively into the renal artery (10 mg) followed by oral administration of 120 mg every 8 to 12 h for 14 days versus no drug (N = 29). Early immunosuppression included azathioprine, corticosteroids, and antilymphocyte globulin with subsequent overlapping with CsA on days 5 and 6. Actuarial graft survival at 1 yr was different when the two groups were compared (P less than 0.05). Estimated graft survival at 1 yr for VP patients was 93.3 compared with 72.4% in control patients. The improved graft survival was most striking in repeat transplants with 90% graft survival at 1 yr for VP recipients versus 37.5% for controls. Compared with controls, VP recipients had significantly improved renal parenchymal diastolic blood flow velocities on the first day after surgery (7.8 versus 5.8 cm/s). By day 7, GFR were greater with VP (44 +/- 29 mL/min) versus controls (28 +/- 22 mL/min). Of VP patients, 67% (18 of 24) had GFR greater than 30 mL/min versus 33% (9 of 26) for control patients. Similarly, on the seventh day, 77% (21 of 30) of VP patients had serum creatinines less than 2.0 mg% versus 34% (10 of 29) for controls.(ABSTRACT TRUNCATED AT 250 WORDS)

Elderly patients are increasingly being considered for kidney transplantation due to a global explosion of the aging population with end-stage renal disease (ESRD). However, mounting scarcity of available organs for transplant has led to a wider disparity between organ supply and demand. Consequently, the criteria for accepting kidneys for transplantation have been extended in an attempt to allow the use of organs from elderly donors or those with significant co-morbidities, so-called “expanded criteria donor” (ECD) kidneys. Excellent outcomes have been achieved from ECD kidneys with appropriate donor and recipient profiling and selection. With increasing recovery efforts directed at older donors, the concept of age-matching is becoming more accepted as a method of optimizing utilization of organs in elderly donors and recipients. Utilization of pulsatile perfusion has further improved ECD outcomes and helped the decision-making process for the UNOS (United Network for Organ Sharing) offer. However, age-related immune dysfunction and associated co-morbidities make the elderly transplant recipients ever more susceptible to complications associated with immunosuppressive agents. Consequently, the elderly population is at a higher risk to develop infections and malignancy in the post-transplant period notwithstanding improved transplant outcomes. Appropriate immunosuppressive agents and dosages should be selected to minimize adverse events while reducing the risk of acute rejections and maximizing patient and renal allograft survival.

Biliary obstruction in the setting of hepatic bacterial infection has great morbidity and mortality. We developed a novel murine model to examine the effect of biliary obstruction on the clearance of hepatic Escherichia coli infection. This model may allow us to test the hypothesis that biliary obstruction itself adversely affects clearance of hepatic infections even if the bacteria are introduced into the liver by a nonbiliary route. We ligated the bile ducts of C57BL/6 mice on days − 1, 0, or + 1, relative to a day 0 portal venous injection of E. coli. We monitored survival, hepatic bacterial growth, pathology, and IL-10 protein levels. The role of IL-10 in this model was further examined using IL-10 knockout mice. Mice with bile duct ligation at day +1 or 0, relative to portal venous infection at day 0, had decreased survival compared with mice with only portal venous infection. The impaired survival was associated with greater hepatic bacterial growth, hepatic necrosis, and increased production of IL-10. Interestingly, the transgenic knockout of IL-10 resulted in impaired survival in mice with bile duct ligation and portal venous infection. Biliary obstruction had a dramatic detrimental effect on hepatic clearance of portal venous E. coli infection. This impaired clearance is associated with increased IL-10 production. However, transgenic knockout of IL-10 increased mortality after hepatic infection.

Ischemic acute renal failure involves not only the kidney but also extrarenal organs such as the bone marrow that produces inflammatory cells. By ELISA and RNase protection assays, we now show that renal ischemia-reperfusion increases serum concentrations of granulocyte macrophage colony-stimulating factor (G-CSF) protein and increases both G-CSF mRNA and protein in the ischemic kidney. In situ hybridization localized the increased G-CSF mRNA to tubule cells, including medullary thick ascending limb cells (mTAL), in the outer medulla. We also show that mTAL produce G-CSF protein and increase G-CSF mRNA after stimulation by reactive oxygen species in vitro. The production of G-CSF by the kidney after ischemia-reperfusion provides a means of communication from the injured kidney to the bone marrow. This supports the known inflammatory response to ischemia.

Lymphoproliferative disorders occur with a greater frequency in recipients of solid organ transplants. Unlike in the general population, these tumors tend to arise in extranodal sites and are often confined to a single organ. We report on a patient with lymphoproliferative disease confined to the hilum of the renal allograft. The only clinical clue to the presence of the tumor was an increase in the serum creatinine secondary to ureteral obstruction by the mass.

Cholangitis requires bile duct obstruction and infection. Patients with cholangitis are often more affected than those with infections that reach the liver through the portal vein. We will attempt to study the influences of (i) route of entry and (ii) presence of bile duct obstruction on hepatic infection. C57BL/6 mice received injections of Escherichia coli or lipopolysaccharide into the obstructed bile duct or portal vein and were monitored for survival. Livers were assayed for bacteria, and cytokine mRNA was measured. In order to examine the effect of biliary obstruction on hepatic infection, animals were subjected to bile duct ligation 1 day prior to portal vein injection and were monitored for survival. The 50% lethal dose (LD 50 ) for E. coli injected into the bile duct was 50 CFU/animal; the LD 50 for E. coli injected into the portal vein was 5 × 10 7 CFU/animal. Initial hepatic delivery of bacteria was equivalent 1 h after injection into the bile duct or portal vein. However, by 24 h, a significantly greater amount of bacteria was recovered from the livers of the bile duct-injected group. Interleukin 10 (IL-10) and IL-1RA mRNA was expressed at greater levels in the bile duct-injected group. Prior bile duct ligation followed by portal vein injection resulted in a higher incidence of death than when sham operation was performed prior to portal vein injection. Our data suggest that the increased mortality from cholangitis, compared with that from other hepatic infections, is related to the different route of delivery of pathogen and the maladaptive response (possibly involving IL-10 and IL-1RA) to biliary obstruction itself.