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Background: Residual kidney function (RKF) is associated with improved survival and quality of life in dialysis patients. Previous studies have suggested that initiation of peritoneal dialysis (PD) may slow RKF decline compared to the pre-dialysis period. We sought to evaluate the association between PD initiation and RKF decline in the Initiating Dialysis Early And Late (IDEAL) trial., Methods: In this post hoc analysis of the IDEAL randomized controlled trial, PD participants were included if results from 24-hour urine collections had been recorded within 30 days of dialysis initiation, and at least one value pre- and one value post-dialysis commencement were available. The primary outcome was slope of RKF decline, calculated as mean of urinary creatinine and urea clearances. Secondary outcomes included slope of urine volume decline and time from PD initiation to anuria., Results: The study included 151 participants (79 early start, 72 late start). The slope of RKF decline was slower after PD initiation (-2.69±0.18mL/min/1.73m2/yr) compared to before PD (-4.09±0.33mL/min/1.73m2/yr; change in slope +1.19 mL/min/1.73m2/yr, 95%CI 0.48-1.90, p<0.001). In contrast, urine volume decline was faster after PD commencement (-0.74±0.05 L/yr) compared to beforehand (-0.57±0.06L/yr; change in slope -0.18L/yr, 95%CI -0.34--0.01, p = 0.04). No differences were observed between the early- and late-start groups with respect to RKF decline, urine volume decline or time to anuria., Conclusions: Initiation of PD was associated with a slower decline of RKF compared to the pre-dialysis period., Competing Interests: I have read the journal’s policy and the authors of this manuscript have the following competing interests: YC reports personal fees from Baxter, outside the submitted work. CH reports personal fees from Janssen, GlaxoSmithKline and Osuka, and grant support from Baxter, Fresenius, Shire, PKD Aus and NHMRC, outside the submitted work. MGW has received honorarium for scientific lectures from AstraZeneca, Retrophin, Amgen and Baxter, outside the submitted work. His employer, the George Institute for Global Health, holds research contracts for trials in cardiovascular and/or kidney disease with a range of commercial organizations. DJ reports personal fees from AWAK, Astra-Zeneca, Baxter Healthcare and Fresenius Medical Care, and grant support from Baxter Extramural Grant and Baxter CEC Grant, outside the submitted work. The other authors declare no conflicts of interest. This does not alter our adherence to PLOS ONE policies on sharing data and materials.

Transforming growth factor β (TGF-β) is the key cytokine involved in causing fibrosis through cross-talk with major profibrotic pathways. However, inhibition of TGF-β to prevent fibrosis would also abrogate its anti-inflammatory and wound-healing effects. β-catenin is a common co-factor in most TGF-β signaling pathways. β-catenin binds to T-cell factor (TCF) to activate profibrotic genes and binds to Forkhead box O (Foxo) to promote cell survival under oxidative stress. Using a proximity ligation assay in human kidney biopsies, we found that β-catenin/Foxo interactions were higher in kidney with little fibrosis, whereas β-catenin/TCF interactions were upregulated in the kidney of patients with fibrosis. We hypothesised that β-catenin/Foxo is protective against kidney fibrosis. We found that Foxo1 protected against rhTGF-β1-induced profibrotic protein expression using a CRISPR/cas9 knockout of Foxo1 or TCF1 in murine kidney tubular epithelial C1.1 cells. Co-administration of TGF-β with a small molecule inhibitor of β-catenin/TCF (ICG-001), protected against kidney fibrosis in unilateral ureteral obstruction. Collectively, our human, animal and in vitro findings suggest β-catenin/Foxo as a therapeutic target in kidney fibrosis.

Regulatory T cells (Tregs) have been recognized as having a major role in maintaining peripheral tolerance and preventing and limiting autoimmune and chronic inflammatory diseases. Tregs derive from the thymus and also develop peripherally. In this review, we discuss recent progress in our understanding of the basic mechanisms involved in Treg development and function in protecting against autoimmunity in the periphery, including thymic selection, peripheral induction and the many mechanisms of Treg suppression. Specifically in kidney disease, Tregs have been shown to play a role in limiting injury and may potentially have a therapeutic role., (© 2016 Asian Pacific Society of Nephrology.)

Macrophages are found in normal kidney and in increased numbers in diseased kidney, where they act as key players in renal injury, inflammation, and fibrosis. Macrophages are highly heterogeneous cells and exhibit distinct phenotypic and functional characteristics in response to various stimuli in the local microenvironment in different types of kidney disease. In kidney tissue necrosis and/or infection, damage- and/or pathogen-associated molecular patterns induce pro-inflammatory macrophages, which contribute to further tissue injury, inflammation, and subsequent fibrosis. Apoptotic cells and anti-inflammatory factors in post-inflammatory tissues induced anti-inflammatory macrophages, which can mediate kidney repair and regeneration. This review summarizes the role of macrophages with different phenotypes in kidney injury, inflammation, and fibrosis in various acute and chronic kidney diseases. Understanding alterations of kidney microenvironment and the factors that control the phenotype and functions of macrophages may offer an avenue for the development of new cellular and cytokine/growth factor-based therapies as alternative treatment options for patients with kidney disease., (©2015 Int. Union Physiol. Sci./Am. Physiol. Soc.)

Conventional markers of macrophages (Mфs) and dendritic cells (DCs) lack specificity and often overlap, leading to confusion and controversy regarding the precise function of these cells in kidney and other diseases. This study aimed to identify the phenotype and function of renal mononuclear phagocytes (rMPs) expressing key markers of both Mфs and DCs. F4/80(+)CD11c(+) cells accounted for 45% of total rMPs in normal kidneys and in those from mice with Adriamycin nephropathy (AN). Despite expression of the DC marker CD11c, these double-positive rMPs displayed the features of Mфs, including Mф-like morphology, high expression of CD68, CD204, and CD206, and high phagocytic ability but low antigen-presenting ability. F4/80(+)CD11c(+) cells were found in the cortex but not in the medulla of the kidney. In AN, F4/80(+)CD11c(+) cells displayed an M1 Mф phenotype with high expression of inflammatory mediators and costimulatory factors. Adoptive transfer of F4/80(+)CD11c(+) cells separated from diseased kidney aggravated renal injury in AN mice. Furthermore, adoptive transfer of common progenitors revealed that kidney F4/80(+)CD11c(+) cells were derived predominantly from monocytes, but not from pre-DCs. In conclusion, renal F4/80(+)CD11c(+) cells are a major subset of rMPs and display Mф-like phenotypic and functional characteristics in health and in AN., (Copyright © 2015 by the American Society of Nephrology.)

Background and Objectives: The Initiating Dialysis Early and Late study showed that planned early or late initiation of dialysis, based on the Cockcroft and Gault estimation of GFR, was associated with identical clinical outcomes. This study examined the association of all-cause mortality with estimated GFR at dialysis commencement, which was determined using multiple formulas., Design, Setting, Participants, & Measurements: Initiating Dialysis Early and Late trial participants were stratified into tertiles according to the estimated GFR measured by Cockcroft and Gault, Modification of Diet in Renal Disease, or Chronic Kidney Disease-Epidemiology Collaboration formula at dialysis commencement. Patient survival was determined using multivariable Cox proportional hazards model regression., Results: Only Initiating Dialysis Early and Late trial participants who commenced on dialysis were included in this study (n=768). A total of 275 patients died during the study. After adjustment for age, sex, racial origin, body mass index, diabetes, and cardiovascular disease, no significant differences in survival were observed between estimated GFR tertiles determined by Cockcroft and Gault (lowest tertile adjusted hazard ratio, 1.11; 95% confidence interval, 0.82 to 1.49; middle tertile hazard ratio, 1.29; 95% confidence interval, 0.96 to 1.74; highest tertile reference), Modification of Diet in Renal Disease (lowest tertile hazard ratio, 0.88; 95% confidence interval, 0.63 to 1.24; middle tertile hazard ratio, 1.20; 95% confidence interval, 0.90 to 1.61; highest tertile reference), and Chronic Kidney Disease-Epidemiology Collaboration equations (lowest tertile hazard ratio, 0.93; 95% confidence interval, 0.67 to 1.27; middle tertile hazard ratio, 1.15; 95% confidence interval, 0.86 to 1.54; highest tertile reference)., Conclusion: Estimated GFR at dialysis commencement was not significantly associated with patient survival, regardless of the formula used. However, a clinically important association cannot be excluded, because observed confidence intervals were wide.

Background and Aims: Studies of diet in relation to chronic kidney disease (CKD) have focused on individual nutrients. The relationship between overall patterns of food intake and renal function has not been well explored. We aimed to investigate the associations between diet quality with the prevalence, incidence and progression of CKD in a cohort of older adults., Methods and Results: 1952 participants aged ≥50 years at baseline were examined between 1992-1994 and 2002-2004. Dietary data were collected using a semi-quantitative food frequency questionnaire. A modified version of the Healthy Eating Index for Australians was developed to determine total diet scores (TDS). Baseline biochemistry including serum creatinine was measured. CKD was defined as MDRD estimated glomerular filtration rate (eGFR) <60 mL min⁻¹·1.73 m⁻². Participants in the highest quartile of mean TDS compared to those in the first quartile (reference), had a 41% reduced likelihood of having eGFR <60 mL min⁻¹·1.73 m⁻², [multivariable-adjusted odds ratio, OR, 0.59 (95% confidence intervals, CI, 0.41-0.85), P-trend = 0.005]. Each unit increase in TDS was associated with a 15% decrease in the odds of having prevalent CKD, multivariable-adjusted OR 0.85 (95% CI 0.74-0.97). A non-significant, inverse association between TDS and CKD incidence was observed (P-trend = 0.10)., Conclusion: Older adults with better diet quality had a reduced likelihood of having eGFR <60 mL min⁻¹·1.73 m⁻². Adherence to dietary guidelines were not prospectively associated with CKD incidence or progression. Further studies with adequate power are warranted to assess the longitudinal association between diet quality and CKD., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Increasing evidence indicates that vitamin D deficiency exacerbates chronic kidney injury, but its effects on renal enlargement in polycystic kidney disease (PKD) are not known. In this study, male Lewis polycystic kidney disease (LPK) rats received a normal diet (ND; AIN-93G) supplemented with or without cholecalciferol (vitamin D-deficient diet, VDD; both 0.5% calcium), commenced at either postnatal week 3 (until weeks 10-20; study 1) or from week 10 (until week 20; study 2). Levels of 25-hydroxy vitamin D were reduced in groups receiving the VDD (12 ± 1 nmol/l vs. 116 ± 5 in ND; P < 0.001). In study 1, food intake and weight gain increased by ∼25% in LPK rats receiving the VDD ad libitum, and at week 20 this was associated with a mild reduction in the corrected serum calcium (SCa(2+), 7.4%) and TKW:BW ratio (8.8%), and exacerbation of proteinuria (87%) and hypertension (19%; all P < 0.05 vs. ND). When LPK rats were pair-fed for weeks 3-10, there was a further reduction in the SCa(2+) (25%) and TKW:BW ratio (22%) in the VDD group (P < 0.05 vs. ND). In study 2, the VDD did not alter food intake and body weight, reduced SCa(2+) (7.7%), worsened proteinuria (41.9%), interstitial monocyte accumulation (26.4%), renal dysfunction (21.4%), and cardiac enlargement (13.2%, all P < 0.05), but there was a trend for a reduction in the TKW:BW ratio (13%, P = 0.09). These data suggest that chronic vitamin D deficiency has adverse long-term actions on proteinuria, interstitial inflammation, renal function, and cardiovascular disease in PKD, and these negate its mild inhibitory effect on kidney enlargement.

Macrophages (MΦ) are located throughout kidney tissue, where they play important roles in homeostasis, surveillance, tolerance, and cytoprotection. MΦ are highly heterogeneous cells and exhibit distinct phenotypic and functional characteristics depending on their microenvironment and the disease type and stage. Recent studies have identified a dual role for MΦ in several murine models of kidney disease. In this review, we discuss the pathogenic and protective roles of the various MΦ subsets in experimental and human kidney diseases and summarize current progress toward the therapeutic use of MΦ in kidney diseases.

Objectives: Renal function has been shown to be influenced by thyroid status in animal models and human studies. We aimed to assess the cross-sectional association between thyroid hormones and function with prevalence of chronic kidney diseases (CKD) in older adults., Study Design: 1571 Blue Mountains Eye Study participants aged ≥ 60 years were analyzed in 2002-4. Thyroid dysfunction was defined using serum thyrotropin (TSH) screen, followed by serum free T4 (FT4) assessment. Baseline biochemistry including serum creatinine was measured. Moderate CKD was defined as estimated glomerular filtration rate (eGFR) <60 ml/min/1.73 m(2)., Results: After adjusting for age, sex, receipt of pension payment, body mass index, smoking, hypertension and diabetes, persons with any thyroid dysfunction (hyperthyroidism or hypothyroidism) had 84% higher likelihood of having CKD, odds ratio, OR, 1.84 (95% confidence intervals, CI, 1.03-3.31). Participants in the highest versus lowest quartile (reference) of serum TSH and FT4 had a significantly greater odds of prevalent CKD, OR 1.82 (95% CI 1.22-2.71), and OR 1.64 (95% CI 1.10-2.45), respectively. Similarly, among participants not receiving treatment for their thyroid dysfunction (n=1329), those in the third and fourth quartiles of serum TSH had significantly greater odds of having prevalent CKD, OR 1.83 (95% CI 1.15-2.92) and OR 1.96 (95% CI 1.23-3.13), respectively, Ptrend=0.001. Significant associations were not observed between type of thyroid dysfunction (hyperthyroidism or hypothyroidism) and prevalent CKD., Conclusions: Increasing serum TSH was associated with a greater likelihood of prevalent CKD among older adults, independent of the influence of age, diabetes and hypertension., (Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.)

A pro-fibrotic role of matrix metalloproteinase-9 (MMP-9) in tubular cell epithelial-mesenchymal transition (EMT) is well established in renal fibrosis; however studies from our group and others have demonstrated some previously unrecognized complexity of MMP-9 that has been overlooked in renal fibrosis. Therefore, the aim of this study was to determine the expression pattern, origin and the exact mechanism underlying the contribution of MMP-9 to unilateral ureteral obstruction (UUO), a well-established model of renal fibrosis via MMP-9 inhibition. Renal MMP-9 expression in BALB/c mice with UUO was examined on day 1, 3, 5, 7, 9, 11 and 14. To inhibit MMP-9 activity, MMP-2/9 inhibitor or MMP-9-neutralizing antibody was administered daily for 4 consecutive days from day 0-3, 6-9 or 10-13 and tissues harvested at day 14. In UUO, there was a bi-phasic early- and late-stage upregulation of MMP-9 activity. Interestingly, tubular epithelial cells (TECs) were the predominant source of MMP-9 during early stage, whereas TECs, macrophages and myofibroblasts produced MMP-9 during late-stage UUO. Early- and late-stage inhibition of MMP-9 in UUO mice significantly reduced tubular cell EMT and renal fibrosis. Moreover, MMP-9 inhibition caused a significant reduction in MMP-9-cleaved osteopontin and macrophage infiltration in UUO kidney. Our in vitro study showed MMP-9-cleaved osteopontin enhanced macrophage transwell migration and MMP-9 of both primary TEC and macrophage induced tubular cell EMT. In summary, our result suggests that MMP-9 of both TEC and macrophage origin may directly or indirectly contribute to the pathogenesis of renal fibrosis via osteopontin cleavage, which, in turn further recruit macrophage and induce tubular cell EMT. Our study also highlights the time dependency of its expression and the potential of stage-specific inhibition strategy against renal fibrosis.

Plasmacytoid dendritic cells play important roles in inducing immune tolerance, preventing allograft rejection, and regulating immune responses in both autoimmune disease and graft-versus-host disease. In order to evaluate a possible protective effect of plasmacytoid dendritic cells against renal inflammation and injury, we purified these cells from mouse spleens and adoptively transferred lipopolysaccharide (LPS)-treated cells, modified ex vivo, into mice with adriamycin nephropathy. These LPS-treated cells localized to the kidney cortex and the lymph nodes draining the kidney, and protected the kidney from injury during adriamycin nephropathy. Glomerulosclerosis, tubular atrophy, interstitial expansion, proteinuria, and creatinine clearance were significantly reduced in mice with adriamycin nephropathy subsequently treated with LPS-activated plasmacytoid dendritic cells as compared to the kidney injury in mice given naive plasmacytoid dendritic cells. In addition, LPS-pretreated cells, but not naive plasmacytoid dendritic cells, convert CD4+CD25- T cells into Foxp3+ regulatory T cells and suppress the proinflammatory cytokine production of endogenous renal macrophages. This may explain their ability to protect against renal injury in adriamycin nephropathy.

Macrophages are important mediators of injury in most types of human kidney diseases; however, the pathogenic importance of both macrophage number and activation status is unknown. To examine this question, severe-combined immunodeficient mice with adriamycin nephrosis, an experimental model of human focal segmental glomerulosclerosis, were treated intravenously with either resting (1 x 10(6) to 5 x 10(6)) or activated (1 x 10(3) to 1 x 10(6)) macrophages on day 6 postadriamycin administration, and the effects on kidney injury were examined. On day 28, renal injury was worse in the group that received activated macrophages at doses as low as 1 x 10(4) macrophages per mouse compared with control adriamycin nephrotic mice. However, treatment with resting macrophages at doses as high as 5 x 10(6) macrophages per mouse had no significant effect on either renal histology or function. The transferred activated macrophages homed to inflamed kidneys during the middle-to-late stages of the disease, but such homing was not observed for resting macrophages. This study of in vivo cell adoptive transfer supports the importance of macrophage activation status over macrophage number in causing renal injury. These data suggest that therapeutic strategies for treating progressive kidney diseases should target activated macrophages.

In adriamycin nephropathy (AN), a model of chronic proteinuric renal injury, the absence of functional B and T cells with residual natural killer (NK) cells, and macrophages in severe combined immunodeficient (SCID) mice results in more severe disease than in immunocompetent mice. We have recently shown expression of the stimulatory NK cell molecule NKG2D and its ligand RAE-1 in the adriamycin (ADR) kidney. Therefore, we sought to determine the role of NK cells in AN. We used anti-asialo GM1 NK cell depletion in immunocompetent BALB/c mice with AN, and also compared AN in immunodeficient SCID mice and immunodeficient nonobese diabetic (NOD)-SCID mice (that have impaired NK cell function). The number of NK cells was increased in AN in BALB/c mice compared with normal controls. NK cell depletion or reduction of NK function in NOD-SCID mice did not affect the severity of disease. In both wild type and immunodeficient models, ADR upregulated RAE-1 in the kidney. High levels of Class I major histocompatibility complex molecules were found in both models of AN. In conclusion, NK cells do not play a significant role in AN.

Background: We have previously shown that monocyte chemoattractant protein-1 (MCP-1) and regulated upon activation, normal T-cell expressed and secreted (RANTES) are significantly increased in renal cortex in adriamycin nephropathy. In this study, we tested the effect of DNA vaccination encoding the C-C chemokines MCP-1 and RANTES in a rat model of adriamycin nephropathy., Methods: Both reverse transcription-polymerase chain reaction (RT-PCR) products of MCP-1 and RANTES used as constructs were cloned into a pTarget vector for naked DNA vaccination. Two hundred micrograms of DNA was injected into the tibialis anterior muscle four times at weekly intervals. One week after the last DNA vaccination, rats received adriamycin. All animals were sacrificed 4 weeks after adriamycin administration. Changes in renal function and histologic features were assessed. Enzyme-linked immunosorbent assay (ELISA) and Western blot were used for autoantibody determination. Antibody specificity was assessed in in vitro transmigration assays., Results: Chemokine DNA vaccination significantly reduced proteinuria (P < 0.05) and ameliorated creatinine clearance (P < 0.05) at 2, 3, and 4 weeks after adriamycin administration. Morphometric analysis showed less glomerular sclerosis (P < 0.001) and interstitial infiltrates (P < 0.005) in chemokine DNA vaccination group compared with control groups. Anti-MCP-1 and RANTES autoantibodies were detected in higher concentrations in chemokine DNA vaccinated rats than in control rats (P < 0.001) and serum from vaccinated rats blocked T-cell transmigration to MCP-1 and RANTES., Conclusion: In this study, we have shown that naked DNA vaccination against MCP-1 and RANTES ameliorates the progression of renal disease in the rat adriamycin nephropathy model of chronic proteinuric renal disease. The protective mechanism may involve the production of autoantibodies against MCP-1 and RANTES.

Cellular hypoxia has been proposed as a major factor in the pathogenesis of chronic renal injury, yet to date there has been no direct evidence to support its importance. Therefore, we examined cortical hypoxia in an animal model of chronic renal injury (murine adriamycin nephrosis; AN) by assessing nuclear localization of the oxygen-dependent alpha-subunit of hypoxia-inducible factor-1 (HIF-1alpha) in animals 7, 14, and 28 days after adriamycin. Results were assessed in conjunction with quantitation of the cortical microvasculature (by CD34 immunostaining) and cortical expression of VEGF. Cortical apoptosis was also examined by terminal deoxynucleotidyl transferase dUTP nick-end labeling staining. A dramatic and significant increase in nuclear localization of HIF-1alpha was seen 28 days after adriamycin in the context of severe glomerular and tubulointerstitial damage. Areas of nuclear HIF-1alpha staining did not colocalize with areas of cellular apoptosis. AN was also associated with a significant attenuation of the peritubular capillaries that was significant at 14 and 28 days after adriamycin. Cortical VEGF expression fell in a stepwise manner from day 7 until day 28 after adriamycin. In conclusion, these data are consistent with a significant increase in cellular hypoxia occurring in the advanced stages of murine AN. Increased cortical hypoxia was preceded by significant reductions in both the number of peritubular capillaries (i.e., oxygen supply) and the angiogenic cytokine VEGF. Apart from providing the first direct evidence for cellular hypoxia in a model of chronic renal disease, these results suggest that a primary dysregulation of angiogenesis may be the cause of increased hypoxia in this model.

Background: Interaction between CD40 and CD40 ligand (CD40L) is involved in both cognate and innate immune responses. Blockade of CD40-CD40L interactions reduces severity of renal injury in murine lupus nephritis and membranous nephropathy. We hypothesized that CD40-CD40L could contribute to renal injury in models that are not antibody-dependent, and that anti-CD40L could diminish inflammation and fibrosis in murine adriamycin nephropathy., Methods: Male BALB/c mice were divided into three groups (N = 6 per group): (1). saline-treated, age-matched control; (2). adriamycin only; and (3). MR1 + adriamycin. In group 3, mice were treated with intraperitoneal injections of anti-CD40L antibody (clone MR1, 0.4 mg per mouse) after the onset of proteinuria at days 5, 7, 9, and 11 after adriamycin treatment. Animal subgroups were compared at 14 and 42 days after induction of adriamycin nephropathy. Functional and pathologic markers of disease severity, cellular components of interstitial inflammation, and the degree of CD40 expression were assessed. Relative cortical RNA expression of the chemokine monocyte-chemoattractant protein-1 (MCP-1) and regulated on activation normal T cell expressed and secreted (RANTES) was also compared between animal groups., Results: CD40 was weakly expressed in tubules of normal mice but was expressed in tubules, interstitium, and glomeruli of mice with adriamycin nephropathy in a time-dependent manner. MR1 treatment resulted in a significant attenuation of the severity of adriamycin nephropathy at day 42 [e.g., glomerular sclerosis (%), group 3, 20.1 +/- 4.7 vs. group 2, 30.2 +/- 7.2, P < 0.001]. CD40L blockade significantly reduced tubulointerstitial injury as well [tubular diameter microm), group 3, 42.5 +/- 6.9 vs. group 2, 66.3 +/- 13.7, P < 0.001; and group 1, 37.3 +/- 5.7, P < 0.01; tubular cell height microm), group 3, 16.3 +/- 1.7 vs. group 2, 11 +/- 1.8, P < 0.01; and group 1, 18.2 +/- 1.9, P < 0.01; interstitial volume (%), group 3, 13.9 +/- 5.1 vs. group 2, 26.2 +/- 4.9, P < 0.001; and group 1, 1.3 +/- 0.7, P < 0.001; proteinuria (mg/24 hours), group 3, 1.8 +/- 0.6 vs. group 2, 4.3 +/- 0.8, P < 0.001; and group 1, 0.7 +/- 0.2, P < 0.05; and creatinine clearance microL/min), group 3, 75 +/- 4 vs. group 2, 35 +/- 2, P < 0.001; and group 1, 82 +/- 4, P < 0.01] were also improved by MR1. MR1 treatment also resulted in a significant reduction in the number of cortical macrophages at both 14 and 42 days after adriamycin (P < 0.01). Cortical expression of MCP-1 and RANTES was significantly reduced by MR1 treatment at 42 days after adriamycin (P < 0.01 and P < 0.05, respectively)., Conclusion: Blockade of CD40-CD40L interaction protects against renal structural and functional injury in this murine model of chronic proteinuric renal disease.

Background: CD4(+) T cells play an important role in various types of immunologic renal disease, including lupus nephritis, IgA nephropathy, and crescentic glomerulonephritis. CD4(+) T cells are also major infiltrating lymphocytes in chronic tubulointerstitial inflammation associated with nonimmunological renal diseases. We suspected that CD4(+) T cells might contribute to disease progression and loss of renal function in chronic proteinuric renal disease (CPRD). To investigate this possibility, the effect of monoclonal antibody against CD4(+) lymphocytes (anti-CD4) was studied in a murine model (adriamycin nephropathy) of CPRD., Methods: Adriamycin nephropathy was produced in male BALB/c mice by a single intravenous injection of adriamycin (11 mg/kg). Anti-CD4 was given by intraperitoneal injection following the development of proteinuria at days 5, 6, 7, 21, and 37 after adriamycin. After six weeks, renal function and histology were studied by histomorphometry, immunohistochemistry, and flow cytometry., Results: Flow cytometric analysis showed a marked decrease in the number of CD4(+) T cells in blood and spleen of the antibody-treated animals (N = 7, P < 0.01). Adriamycin plus CD4(+) depletion mice had significantly greater mesangial expansion, glomerular sclerosis, and interstitial expansion than the mice on adriamycin alone. Interstitial infiltration with macrophages and CD8(+) cells was significantly increased in adriamycin plus CD4(+) depletion mice. Creatinine clearance (17.5 +/- 0.54 vs. 29.2 +/- 0.89 microL/min, P < 0.001) was significantly worse in the adriamycin plus CD4(+) depletion mice than in adriamycin alone mice and correlated with histologic change in glomeruli and interstitium., Conclusions: Depletion of CD4(+) T cells promotes glomerular and interstitial injury in mice with established adriamycin nephropathy. These findings suggest that CD4(+) T cells have a protective role against the progression of adriamycin nephropathy.

Urinary and proximal tubular iron are increased after subtotal nephrectomy, and iron depletion has been shown to be beneficial in proteinuric models of chronic renal disease in rats. In this study, iron depletion by low iron pair-fed diet and periodic phlebotomy was induced for 6 months in rats with partial (5/6) nephrectomy, resulting in a reduction in hematocrit and serum iron in all iron-deficiency subgroups. Tubular iron, assessed by energy dispersive analysis and electron microscopy, was reduced in quantity but not number of iron-containing lysosomes only within 1 subgroup of severe iron deficiency (p < 0.05). There was no improvement in serial isotopic glomerular filtration rate measurements, urinary protein and transferrin excretion, tubular damage scores, serum creatinine, or measures of reactive oxygen species (ROS) generation. In a subgroup of rats with no supplementation of sulfhydryl amino acids (cysteine and methionine) which can act as ROS scavengers, iron deficiency increased urinary protein excretion (213.3 +/- 23.0 mg/24 h, mean +/- SEM, vs. 87.4 +/- 16.1, p < 0.001), urinary transferrin excretion (p < 0.05), kidney weight (p < 0.05) and tissue malondialdehyde, a lipid peroxidation product (0.78 +/- 0.16 nmol/mg protein vs. 0.57 +/- 0.19, p < 0.05), consistent with increased ROS generation. Hence, no beneficial effect of iron-deficiency was demonstrated by any measure of structure of function in the remnant kidney, and it may enhance damage if sulfhydryl repletion is not provided.

Reactive oxygen species (ROS) have been implicated in progression of disease in the rat remnant kidney (RK) model of chronic renal failure. Substantial amounts of iron accumulate in proximal tubular lysosomes of RK and could damage tubules by ROS generation. The effect of dietary protein intake on ROS, tubular damage and iron accumulation assessed by energy dispersive analysis was determined in RK (5/6 nephrectomy, N = 12) and sham-operated kidneys (SO, N = 10). In RK, mean lysosomal iron concentration, urinary iron and protein excretion and morphological damage were increased and GFR decreased. Dietary protein loading (40% vs. 12%) increased the number of iron-containing lysosomes (P < 0.05) and the mean lysosomal iron (P < 0.02) in proximal tubular cells after four weeks. In RK, high protein diet increased renal weight (P < 0.01), numerical density of iron-containing lysosomes and tubular damage (both P < 0.05). ROS generation, assessed by tissue and plasma malondialdehyde (MDA), was also increased (both P < 0.05). Plasma MDA correlated with tubular iron accumulation (r = 0.75). In RK fed a high protein diet (N = 18) treatment with the iron-chelator desferrioxamine reduced serum iron, urinary volume, and tubular iron accumulation and damage compared to controls (P < 0.01). In summary, in RK dietary protein manipulation altered urinary iron and protein excretion, proximal tubular iron accumulation, renal cortical ROS generation and ultrastructural damage. Desferrioxamine treatment reduced tubular lysosomal iron and ultrastructural damage. These results suggest a role for tubular iron as a determinant of tubular injury associated with dietary protein loading in rats with partial nephrectomy.

Iron has been implicated in the pathophysiology of several models of acute and chronic renal disease. In this study, energy-dispersive x-ray spectrometry was used to quantify and localize iron in rat remnant kidneys (RK) and normal kidneys (NK) and to determine its pathophysiologic significance. Substantial iron accumulation occurred in proximal tubular cell secondary lysosomes of RK (P < 0.001 versus NK) and reached a plateau at 8 wk after partial nephrectomy. In NK, minor increases of iron also occurred with aging (P < 0.02). Proximal tubular iron accumulation correlated independently with protein excretion (r = 0.90) and impairment of GFR (r = 0.70) and was associated with tubular damage and phosphate accumulation (both P < 0.001). Iron nitrilotriacetate (1 mg/kg ip) increased tubular lysosomal iron accumulation and tubular damage (P < 0.001 versus nitrilotriacetate) in NK, comparable to levels seen in untreated RK, and increased cortical cytosolic malondialdehyde, consistent with reactive oxygen species generation. The iron chelator deferoxamine (30 mg/kg per day ip) significantly reduced iron accumulation and tubular damage in RK at 4 wk, compared with deferoxamine chelated to iron and untreated RK. These results suggest that filtered iron enters the remnant tubular lysosomes across the brush border membrane by endocytosis and may produce tubular damage in chronic renal disease by the generation of reactive oxygen species.

To understand further the contribution of heightened net sodium reabsorption (TNa+) and other nontransport processes to increased and altered metabolic activity of the rat remnant kidney (RK), isolated RK and normal kidneys (NK) were perfused with and without metabolic inhibitors and following 4 weeks of low (12%)- or high (40%)-protein diet. 3-Mercaptopicolinate (200 microM) and 2-deoxyglucose (200 microM) reduced glucose production and consumption, respectively, without altering oxygen consumption (QO2). Ouabain reduced TNa+ and QO2 in NK but not RK, and increased glucose consumption in RK (by 67%, p < 0.02, n = 12) and NK (by 54%, p < 0.05, n = 12). Raising perfusate pH by 0.3 U to 7.65 increased glucose production in RK but not NK and reduced TNa+ in NK but not RK. Dietary protein restriction reduced QO2 (2.18 +/- 0.29 vs. 4.13 +/- 0.20 mumol-1 x min-1 x g-1, p < 0.001), TNa+ (8.74 +/- 4.39 vs. 38.83 +/- 8.32, p < 0.01) and ammoniagenesis (0-30 min, 0.16 +/- 0.05 vs. 0.32 +/- 0.10, p < 0.05) in RK, but not NK. In summary: (1) responses to ouabain and alkalosis, but not other metabolic inhibitors, were quite distinct in remnant versus normal kidneys, and (2) protein restriction limited sodium-transport-dependent and -independent hypermetabolism in RK.

Oxygen consumption is increased in the rat remnant kidney (RK), but the basis of this enhanced metabolic activity has not been defined fully. To characterize the hypermetabolism further, isolated RK and normal kidneys (NK) were perfused using a range of perfusion pressures and substrates, and at varying times after 5/6 nephrectomy. Altered and increased RK metabolism was found as early as 1 week after 5/6 nephrectomy. The rate of net glucose consumption was higher in RK than NK (e.g., 0-30 min, 2.42 +/- 0.28 vs. 1.21 +/- 0.33 mumol.min-1 x g-1), a difference not explained by changes in oxygen delivery or glycosuria. Ammonia production was significantly greater early in perfusion in RK than NK. Calculated nephron oxygen consumption (QO2) was greater in RK than NK for all substrates tested (e.g., pyruvate 5 mM: 552.3 +/- 43.1 vs. 173.3 +/- 25.6 pmol.min-1, p < 0.001). When considered together, site I mitochondrial substrates supported QO2 and net sodium reabsorption (TNa+) better than site II substrates. In RK, QO2 did not change with increasing perfusion pressure, and TNa+ rose only slightly for pressures > or = 120 mm Hg. In summary, RK hypermetabolism is characterized by increased oxygen consumption, glycolysis and ammoniagenesis, is not substrate-specific and does not appear to be an artefact of the altered physiology of remnant perfusion.

In vivo energy metabolism in remnant and normal kidney nephrons and the effect of dietary phosphate restriction on energy metabolism were studied. Tissue concentrations of ATP did not differ between remnant and normal kidneys; however, the tissue concentration of inorganic phosphate (Pi) was higher in remnant kidneys (1.17 +/- 0.10 vs. 0.82 +/- 0.09 mumol/g, P less than 0.05) as assessed by 31P-nuclear magnetic resonance (NMR). Intracellular pH was also higher in remnant kidneys (7.48 +/- 0.04 vs. 7.20 +/- 0.05, P less than 0.01), as was oxygen consumption (QO2) when normalized for rate of net tubular sodium reabsorption (Tna; 0.032 +/- 0.005 vs. 0.018 +/- 0.002 mumol.min-1.g-1, P less than 0.01). Compared with glycine supplemented controls, dietary phosphate restriction induced by the phosphate binder diaminodihydroacetoacetate, a maneuver which independently of protein restriction has been shown to ameliorate the progression of chronic renal failure, resulted in no major change in tissue levels of ATP, intracellular pH, or TNa+ but decreased Pi (0.62 +/- 0.07 vs. 1.40 +/- 0.11 mumol/g, P less than 0.01) and QO2 (4.0 +/- 0.4 vs. 7.7 +/- 1.0 mumol.min-1.g-1, P less than 0.01) in remnant kidneys. For the amount of sodium reabsorption performed, remnant kidneys are hypermetabolic, and this hypermetabolism is attenuated by dietary phosphate restriction. These data suggest that hypermetabolism may be involved in the tubulointerstitial mechanisms operant in the progression of chronic renal failure that occurs in the rat remnant kidney, and that attenuation of this abnormality may be the mechanism by which dietary phosphate restriction ameliorates this process.

Phosphate depletion may prevent progression of experimental chronic renal failure. The present study was designed to examine the effect of phosphate depletion on the severity and progression of ischemic acute renal failure. Two groups of rats, phosphate-depleted (PD) and phosphate-repleted (PR), were pair-fed a regular diet for 10 days; the diet of the PD rats also contained 15 g% dihydroxyaluminum aminoacetate. After 10 days, serum phosphate in PD rats averaged 3.9 +/- 0.7 mg%, compared to 6.1 +/- 0.6 mg% in the PR rats (p less than 0.001). The weight gain during the equilibration period was comparable in both groups (24.2 +/- 9.3 vs. 25.7 +/- 10.4 g; NS). Creatinine clearance, hematocrit, renal cortical adenine nucleotides, tissue calcium, mitochondrial calcium and mitochondrial respiration were similar in both groups after 10 days of the diet. A 50-min bilateral clamping of both renal arteries and veins was then undertaken in both groups. The PD rats demonstrated more severe acute renal failure at 24 h after the ischemic insult. Serum creatinine and creatinine clearance for the PD and PR rats were 4.0 +/- 0.6 versus 3.1 +/- 0.9 mg% (p less than 0.005) and 7.8 +/- 8.1 versus 85.3 +/- 70.7 microliter/min (p less than 0.001), respectively. Tissue calcium (131.3 +/- 103.0 vs. 63.5 +/- 36.0 mmol/kg dry weight; p less than 0.02) and mitochondrial calcium (49.4 +/- 24.0 vs. 29.5 +/- 14.6 nmol/mg prot.; p less than 0.02) were significantly higher in the PD rats.(ABSTRACT TRUNCATED AT 250 WORDS)

To investigate the mechanisms whereby verapamil and dietary phosphate restriction slow progression of nephron loss after renal ablation, the effects of these maneuvers on inulin clearance (CIn), net sodium reabsorption (TNa+), oxygen consumption (QO2), and net glucose production (GP) were examined in isolated perfused normal and remnant kidneys. Preliminary studies characterized the isolated perfused remnant kidney perfusion. Adaptation to renal ablation was greater for QO2 (102% of normal), GP (138%), and kidney weight (79%) than CIn (51%) or TNa+ (40%). Verapamil (50 microM) lowered QO2 in remnant kidneys (1.64 +/- 0.24 vs. control 2.86 +/- 0.16 mumol.min-1.g-1, P less than 0.005), as did phosphate restriction (1.81 +/- 0.22 vs. control 3.05 +/- 0.40 mumol.min-1.g-1, P less than 0.05). These effects could not be accounted for by changes in CIn, TNa+, or GP and were not observed in normal kidneys. In summary 1) remnant kidneys are hypermetabolic compared with normal kidneys when assessed by QO2 and GP; 2) verapamil and phosphate restriction diminish the enhanced metabolic activity of remnant kidneys, an effect that is independent of TNa+; and thus 3) verapamil and phosphate restriction may slow progression of renal disease, at least in part by reducing renal metabolic demands.