Kosaku Nitta

Reduced klotho expression level in kidney aggravates renal interstitial fibrosis

Renal expression of the klotho gene is markedly suppressed in chronic kidney disease (CKD). Since renal fibrosis is the final common pathology of CKD, we tested whether decreased Klotho expression is a cause and/or a result of renal fibrosis in mice and cultured renal cell lines. We induced renal fibrosis by unilateral ureteral obstruction (UUO) in mice with reduced Klotho expression (kl/+ mice) and compared them with wild-type mice. The UUO kidneys from kl/+ mice expressed significantly higher levels of fibrosis markers such as α-smooth muscle actin (α-SMA), fibronectin, and transforming growth factor-β(1) (TGF-β(1)) than those from wild-type mice. In addition, in cultured renal fibroblast cells (NRK49F), the levels of α-SMA and PAI1 expression were significantly suppressed by addition of recombinant Klotho protein to the medium. The similar effects were observed by a TGF-β(1) receptor inhibitor (ALK5 inhibitor). These observations suggest that low renal Klotho expression enhances TGF-β(1) activity and is a cause of renal fibrosis. On the other hand, TGF-β(1) reduced Klotho expression in renal cultured epithelial cells (inner medullary collecting duct and human renal proximal tubular epithelium), suggesting that low renal Klotho expression is a result of renal fibrosis. Taken together, renal fibrosis can trigger a deterioration spiral of Klotho expression, which may be involved in the pathophysiology of CKD progression.

Vascular Calcification in Patients With Chronic Kidney Disease

Vascular calcification is very prevalent in patients with chronic kidney disease (CKD). In addition to having more traditional cardiovascular (CV) risk factors, CKD patients also have a number of non‐traditional CV risk factors that may play a prominent role in the pathogenesis of vascular calcification. The transformation of vascular smooth muscle cells into osteoblast‐like cells seems to be a key element in the pathogenesis of vascular calcification in the presence of calcium (Ca) and phosphorus (P) deposition due to abnormal bone metabolism and impaired renal excretion. Vascular calcification causes increased arterial stiffness, left ventricular hypertrophy, decreased coronary artery perfusion, myocardial ischemia, and increased cardiovascular morbidity and mortality. Although current treatment strategies focus on correcting abnormal Ca, P, parathyroid hormone, or vitamin D levels in CKD, a better understanding of the mechanisms of abnormal tissue calcification may lead to the development of new therapeutic agents that are capable of reducing vascular calcification and improving the CV outcome of CKD patients. This review article summarizes the following: (i) the pathophysiological mechanism responsible for vascular calcification; (ii) the methods of detecting vascular calcification in CKD patients; and (iii) the treatment of vascular calcification in CKD patients.u2003

Circulating levels of soluble α-Klotho in patients with chronic kidney disease.

To the Editor, The a-Klotho gene encodes a single-pass transmembrane protein, which was originally identified as an aging-related gene [1]. In adults, a-Klotho contributes to the regulation of calcium and phosphate homeostasis. a-Klotho participates in phosphate homeostasis by cooperating with fibroblast growth factor 23 (FGF23) and the FGF receptor [2]. FGF23 reduces the serum phosphate level both by suppressing phosphate reabsorption and activating vitamin D in the proximal tubules. In addition to the transmembrane form, a-Klotho also exists in a soluble form. The soluble form, which is produced by the shedding of the transmembrane protein, is detectable in serum, cerebrospinal fluid and urine. Although soluble a-Klotho is considered to be a humoral factor, its regulatory mechanisms and functions are largely unknown, and FGF23 signaling is thought to require the transmembrane form of the protein. Recently, a sandwich ELISA for soluble a-Klotho has been established [3]. In the present study, we used this assay to measure the serum levels of soluble a-Klotho in patients with chronic kidney disease (CKD) and compared them to the levels in healthy adult volunteers. We recruited CKD patients and adult volunteers, and obtained informed consent from all participants or their legal guardians. Thirty CKD patients (17 males and 13 females; mean age, 66.2 years) were enrolled. For comparison, 10 healthy adult volunteers aged from 20 to 44 years were also enrolled. The serum soluble a-Klotho levels were measured using an ELISA kit provided from Kyowa Hakko-Kirin (Tokyo, Japan) [3]. The serum levels of intact FGF23 were determined using a commercial sandwich ELISA kit (Kainos Laboratories, Inc., Tokyo, Japan). Serum calcium, phosphate, intact parathyroid hormone (PTH), albumin, and creatinine levels were also measured. The biochemical parameters of CKD patients are shown in Table 1. The mean values for soluble a-Klotho were 1413 ± 614 pg/ml (mean ± SD) in CKD patients and 404 ± 87 pg/ml in the adult volunteers (P 0.0001). There was no significant difference in serum FGF23 levels between the CKD patients (166 ± 509 pg/ml) and the adult volunteers (29 ± 10 pg/ml) (P = 0.2024). In addition, we found that soluble a-Klotho levels were not correlated with FGF23 levels (r = 0.3478; P = 0.0597). Since it is known that circulating soluble a-Klotho levels may be influenced by angiotensin-converting enzyme inhibitor, angiotensin receptor blocker, and statin, we compared the soluble a-Klotho levels in the presence or absence of these drugs. The use of these drugs did not affect the soluble a-Klotho levels. The reason why soluble a-Klotho levels were variable in our CKD patients may be due to the small sample size. In patients with early stage CKD with normophosphatemia, decreases in serum active vitamin D levels and increases in serum PTH levels have been reported previously. The fact that Klotho expression in the parathyroid is decreased in CKD patients may explain why high FGF23 levels fail to suppress PTH secretion [4]. High FGF23 and low vitamin D levels may further decrease Klotho expression in kidney and parathyroid tissue in CKD patients, suggesting that deranged phosphate metabolism in H. Sugiura K. Tsuchiya K. Nitta (&) Department of Medicine, Kidney Center, Tokyo Women’s Medical University, 8-1 Kawada-cho, Shinjuku-ku, Tokyo 162-8666, Japan e-mail: knitta@kc.twmu.ac.jp

Aortic arch calcification evaluated on chest X-ray is a strong independent predictor of cardiovascular events in chronic hemodialysis patients.

Vascular calcification is associated with cardiovascular disease in hemodialysis (HD) patients. Some reports have previously shown that simple assessment of aortic calcification using plain radiography is associated with cardiovascular (CV) events; however, these studies simply assessed whether aortic calcification was present or absent only, without considering its extent. Here, we evaluated the validity of grading aortic arch calcification (AoAC) to predict new CV events. We retrospectively reviewed chest X-rays in 212 asymptomatic HD patients who underwent measurement of pulse wave velocity (PWV) in 2006 without a past history of CV events. The extent of AoAC was divided into four grades (0–3). Among these subjects, the follow-up of CV events in 197 patients was completed. At baseline, AoAC grade was positively associated with age, dialysis vintage, PWV and parathyroid hormone levels, and negatively correlated with body weight and body mass index. Arterial stiffness, as determined by PWV, was also correlated with increasing AoAC grade. Eighty-nine CV events in total occurred during a mean follow-up period of 69xa0±xa045xa0months. With multivariate adjustment, Kaplan–Meier analysis showed that the incidence was significantly higher in patients with higher AoAC grade (grades 2 and 3) than in those with grade 0 or 1 (pxa0=xa00.013, log-rank test). Multivariate Cox proportional hazards analyses showed the predictive values of AoAC grade were significant (hazard ratio 1.512; pxa0=xa00.0351). AoAC detectable on chest X-ray is a strong independent predictor of CV events in accordance with PWV. Risk stratification by assessment of AoAC may provide important information for management of atherosclerotic disease in HD patients.

Kidney tubular damage in the absence of glomerular defects in HIV-infected patients on highly active antiretroviral therapy

BACKGROUNDnThe emergence of kidney disease as an important comorbidity among human immunodeficiency virus (HIV)-infected patients receiving highly active antiretroviral therapy (HAART) has emphasized the critical importance of early identification of patients at risk for kidney disease. Use of urine as a diagnostic medium may allow the noninvasive detection of incipient nephropathy in these patients.nnnMETHODSnHere, we conducted cross-sectional and 1-year prospective studies of 424 HIV-infected patients on HAART without proteinuria or significant impairment of glomerular function. N-acetyl-β-D-glucosaminidase, γ-glutamyl transpeptidase, β(2)-microglobulin and α(1)-microglobulin were measured as indices of tubular damage, which was diagnosed when urinary concentrations of at least three tubular biomarkers exceeded the reference range. Risk factors associated with tubular damage were examined using multivariate logistic regression analysis.nnnRESULTSnTubular damage was identified in 107 patients (25%), who were characterized by advanced age [odds ratio (OR), 1.04; 95% confidence interval (CI), 1.01-1.07], high C-reactive protein (OR, 1.96; 95% CI, 1.26-3.14) and coexisting diabetes mellitus (OR, 3.97; 95% CI, 1.44-12.2). The use of tenofovir, the most likely tubulotoxic agent, was not statistically involved in this subclinical tubular damage. The 1-year follow-up study showed that a decrease in estimated glomerular filtration rate (eGFR) and incidence of proteinuria during the period were significantly higher in patients with than without tubular damage.nnnCONCLUSIONSnA quarter of HIV-infected patients receiving HAART had subclinical tubular damage, which was associated with a near-term decline in eGFR and higher incidence of proteinuria. Periodic monitoring of urinary biomarkers might facilitate the early identification of HAART patients predisposed to significant kidney disease.

Clinical Characteristics of Kidney Disease in Japanese HIV-Infected Patients

Background: Kidney disease has become an important cause of morbidity and mortality in HIV-infected patients in Western countries. Japan is a country with an increasing number of newly infected HIV patients. However, only a few studies have investigated kidney disease in Asian populations infected with HIV. Methods: We studied the prevalence of kidney disease by reviewing the clinical data of 732 HIV-infected Japanese patients. Risk factors for proteinuria, albuminuria, and renal dysfunction were determined using multivariate logistic regression analysis. Results: Microalbuminuria, macroalbuminuria and proteinuria were present in 13.2, 4.55 and 9.52% of patients, respectively. The prevalence of chronic kidney disease of any stage and CKD ≧ stage 3 was 15.4 and 9.70%, respectively. Multivariate analysis showed significant associations between increasing levels of serum creatinine and cholesterol, and the coexistence of diabetes, hypertension and hepatitis C coinfection with either proteinuria or albuminuria, which was significantly related to the presence of renal dysfunction. Lower CD4 cell count was associated with the presence of renal dysfunction, but higher HIV-RNA level was not. Conclusions: Our study has shown the international dimension of the burden of kidney disease in HIV-infected patients. Either proteinuria or albuminuria is likely the most significant factor for renal dysfunction in these patients.

Overweight and obesity accelerate the progression of IgA nephropathy: prognostic utility of a combination of BMI and histopathological parameters

BackgroundAlthough more than 40xa0years have passed since IgA nephropathy (IgAN) was first reported, predicting the renal outcome of individual IgAN patients remains difficult. Emerging epidemiologic evidence indicates that overweight and obesity are risk factors for end-stage renal disease. We aimed to elucidate the outcome of overweight IgAN patients and improve our ability to predict the progression of IgAN based on a combination of body mass index (BMI) and histopathological parameters, including maximal glomerular area (Max GA).MethodsForty-three adult IgAN patients whose estimated glomerular filtration rate was ≥50xa0ml/min/1.73xa0m2 were enrolled in this study. Renal biopsy specimens were evaluated according to the Oxford classification of IgAN. A Kaplan–Meier analysis and the multivariate Cox proportional hazards method were used to evaluate 10-year kidney survival and the impact of covariates. The ability of factors to predict the progression of IgAN was evaluated by their diagnostic odds ratio (DOR).ResultsA BMI ≥25xa0kg/m2 was found to be an independent predictor of a ≥1.5-fold increase in serum creatinine value (DOR 7.4). The combination of BMI ≥25xa0kg/m2, Max GA ≥42,900xa0μm2, and presence of mesangial hypercellularity (Oxford M1) optimally raised predictive power for disease progression of IgAN (DOR 26.0).ConclusionA combination of BMI ≥25xa0kg/m2, the Oxford classification M1, and a Max GA ≥42,900xa0μm2 can serve as a predictor of long-term renal outcome of IgAN.

The significance of caveolae in the glomeruli in glomerular disease

Aims The aim of this study was to demonstrate expression of cell membrane invagination ‘caveolae’ in glomeruli and to correlate this with functional and structural characteristics of the human glomerular diseases. Methods The expression of caveolin-1 (Cav-1), which is the main component of caveolae, was examined in the glomeruli, and the relationship between Cav-1 expression and pathological and clinical findings was determined in 99 patients with glomerular disease and in 50 renal transplantation donors as controls. Results Cav-1 was expressed very weakly in the controls, and the area of Cav-1 expression relative to the total glomerular area was 0.57±0.65%. However, the area of Cav-1 expression was significantly larger in each glomerular disease (IgA nephropathy, 1.05±1.36%, p<0.05; crescent glomerulonephritis, 1.86±1.19%, p<0.001; minimal change disease, 2.38±1.24%, p<0.001; focal segmental glomerulosclerosis, 2.88±2.05%, p<0.01; membranous nephritis, 4.27±2.95%, p<0.001; membranoproliferative glomerulonephritis, 4.49±3.15%, p<0.001; and diabetic nephropathy, 2.45±1.52%, p<0.001; compared with the controls. Cav-1 expression was significantly decreased in glomerular disease treated with steroids. Co-localisation of Cav-1 and the endothelial marker ‘pathologische anatomie leiden-endothelium’ was prominent in an immunofluorescence study, and caveolae on the glomerular endothelial cells were observed in electron microscopy. Conclusions The expression of Cav-1 was significantly increased in the glomeruli of patients with glomerular disease, and it was related to urinary albumin excretion. Cav-1 expression and caveolae were observed in glomerular endothelial cells. It is hypothesised that they play a role in the recovery phase of capillary injury or endocytosis of albumin into endothelial cells. Basic research should be performed to elucidate the role played by Cav-1 and caveolae.

Sphingosine‐1‐phosphate acts as a key molecule in the direct mediation of renal fibrosis

The major sphingolipid metabolite, sphingosine‐1‐phosphate (S1P), has important biological functions. S1P serves as a ligand for a family of five G‐protein‐coupled receptors with distinct signaling pathways regulating important biological pathways. S1P induces renal fibrosis through an inflammatory pathway. However, its direct fibrosis‐inducing effect on the kidney has not been shown. The role of S1P as a direct mediator of renal fibrosis was investigated in normal rat kidney interstitial fibroblast (NRK‐49F) cells (in vitro) and kidneys of a unilateral ureteral obstruction (UUO) mouse model (in vivo). To clarify the role of S1P in renal fibrosis, we adopted nude UUO mice with immune response deficits. NRK‐49F cells were stimulated with various concentrations of exogenous S1P and FTY720 (a S1P receptor agonist) or N,N‐dimethylsphingosine (DMS; a sphingosine kinase inhibitor). C57BL6 and nude UUO mice were pretreated with FTY720, DMS, or saline. Expression levels of alpha‐smooth muscle actin (a‐SMA), E‐cadherin, collagen type 1 (COL1), collagen type 4 (COL4), tissue inhibitor of matrix metalloproteinase‐1 (TIMP1), and plasminogen activator inhibitor‐1 (PAI1) were examined. S1P stimulated fibrosis in NRK‐49F cells and UUO mice. Increased a‐SMA, COL1, COL4, TIMP1, and PAI1 and decreased E‐cadherin expression levels were observed in both the S1P‐stimulated cells and UUO mice. Nude UUO mouse kidneys expressed fibrotic markers. Fibrotic changes were successfully induced in both UUO and nude UUO mice, evident through prominent fibronectin and COL1 staining. These S1P‐induced fibrotic changes were suppressed by FTY720 and DMS both in vitro and in vivo. Thus, S1P essentially and directly mediates renal fibrosis.

Histological heterogeneity of glomerular segmental lesions in focal segmental glomerulosclerosis

Focal segmental glomerulosclerosis (FSGS) involves considerable histological heterogeneity in terms of location and quality of the glomerular segmental lesions. The present study investigated the heterogeneity of segmental lesions in each variant of FSGS, determined by the Columbia classification, and its clinical relevance. All glomerular segmental lesions of 80 cases of primary FSGS were evaluated histologically based on location [tip (TIP), perihilar (PH), or not otherwise specified (NOS)], and quality (cellular or fibrous). Among the 1,299 glomeruli of the 80 biopsy specimens, 210 glomeruli (16.2%) had segmental lesions, comprising 57 (27%) cellular TIP, 4 (2%) fibrous TIP, 42 (20%) cellular NOS, 86 (41%) fibrous NOS, and 21 (10%) fibrous PH lesions. Each case was also classified into one of the five histological variants of the Columbia classification: collapsing (COL), TIP, cellular (CEL), PH, or NOS. Overlap of segmental lesions in different location categories was seen in the COL, TIP, and PH variants, and heterogeneity of quality was apparent in the COL and CEL variants. Histological findings of the CEL variant (endocapillary hypercellularity) were observed in nine of the 13 COL variants. Both location and quality correlated with disease duration, degree of proteinuria, and histological severity of global glomerular sclerosis and tubulo-interstitial lesions. These results demonstrated the histological heterogeneity of glomerular segmental lesions in all variants of the Columbia classification, except NOS. However, the fidelity of location and dominance of histological features were generally conserved in the TIP and PH variants. The COL and CEL variants warrant further investigation because of their overlapping histological findings and apparent histological heterogeneity in the glomerular segmental lesions.