Biomedical subjects
Kenichiro Kitamura
Publications and source records attributed to Kenichiro Kitamura.
Successful treatment of a patient with severe calcific uremic arteriolopathy (calciphylaxis) by etidronate disodium.
A 59-year-old woman with a 10-year history of hemodialysis was admitted to our hospital for painful skin ulcers on her right thigh, right calf, and left upper arm. A whole-body plain computed tomographic scan showed diffuse calcification of the uterus and marked calcification of the mitral valve. Skin biopsy specimens from the left thigh showed calcium deposition in numerous small blood vessels in the dermis and fat, leading to a diagnosis of calcific uremic arteriolopathy (CUA). Despite antibiotic therapy and aggressive wound care for 2 months, the skin ulcers enlarged and the patient's general condition worsened. Surprisingly, oral administration of etidronate disodium (200 mg/d) strikingly improved the focal infection and decreased the size of the skin ulcers within several days. She was discharged from the hospital 2 months later, when epithelialization of the ulcers was almost complete. We report a case of CUA that was improved dramatically by treatment with etidronate. Etidronate therapy should be considered for refractory CUA.
Fibrocystin interacts with CAML, a protein involved in Ca2+ signaling.
The predicted structure of the autosomal recessive polycystic kidney disease protein, fibrocystin, suggests that it may function as a receptor, but its function remains unknown. To understand its function, we searched for proteins that interact with the intracellular C-terminus of fibrocystin using the yeast two-hybrid system. From the screening, we found calcium modulating cyclophilin ligand (CAML), a protein involved in Ca(2+) signaling. Immunofluorescent analysis showed that both proteins are co-localized in the apical membrane, primary cilia, and the basal body of cells derived from the distal nephron Epitope-tagged expression constructs of both proteins were co-immunoprecipitated from COS7 cells. The intracellular C-terminus of fibrocystin interacts with CAML, a protein with an intracellular distribution that is similar to that of PKD2. Fibrocystin may participate in regulation of intracellular Ca(2+) in the distal nephron in a manner similar to PKD1 and PKD2 that are involved in autosomal dominant polycystic kidney disease.
The structure and function of oxidized albumin in hemodialysis patients: Its role in elevated oxidative stress via neutrophil burst.
Oxidized albumin is a reliable marker of oxidative stress in hemodialysis (HD) patients. However, oxidized albumin in vivo and its possible clinical significance has been rarely investigated. In the present study, the qualitative modification of albumin in HD patients (n = 20) was examined and their results were compared with healthy age-matched controls (n = 10). The increase in plasma protein carbonyl levels in HD patients was largely due to an increase in oxidized albumin. Human serum albumin (HSA) of HD patients, HSA of HD patients (HD-HSA) and normal subjects (Normal-HSA) were purified on a blue Sepharose CL-6B column. Spectroscopic analysis confirmed that the HD-HSA samples contained higher levels of carbonyls than Normal-HSA. An HPLC analysis also suggested that the state of the purified HSA used throughout the experiments accurately reflects the redox state of albumin in blood. HD-HSA was found to have a decreased the antioxidant activity, and was able to trigger the oxidative burst of human neutrophils, compared to Normal-HSA. HD-HSA was conformationally altered, with its hydrophobic regions more exposed and to have a negative charge. In binding experiments, HD-HSA showed impaired Site II-ligand binding capabilities. Collectively, the oxidation of plasma proteins, especially HSA, might enhance oxidative stress in HD patients.
[Roll of aldosterone in pathogenesis of kidney disorders].
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Mutations in human urate transporter 1 gene in presecretory reabsorption defect type of familial renal hypouricemia.
To date, 11 loss of function mutations in the human urate transporter 1 (hURAT1) gene have been identified in subjects with idiopathic renal hypouricemia. In the present studies we investigated the clinical features and the mutations in the hURAT1 gene in seven families with presecretory reabsorption defect-type renal hypouricemia and in one family with the postsecretory reabsorption defect type. Twelve affected subjects and 26 family members were investigated. Mutations were analyzed by PCR and the direct sequencing method. Urate-transporting activities of wild-type and mutant hURAT1 were determined by [14C]urate uptake in Xenopus oocytes. Mutational analysis revealed three previously reported mutations (G774A, A1145T, and 1639-1643 del-GTCCT) and a novel mutation (T1253G) in families with the presecretory reabsorption defect type. Neither mutations in the coding region of hURAT1 gene nor significant segregation patterns of the hURAT1 locus were detected in the postsecretory reabsorption defect type. All hURAT1 mutants had significantly reduced urate-transporting activities compared with wild type (P < 0.05; n = 12), suggesting that T1253G is a loss of function mutation, and hURAT1 is responsible for the presecretory reabsorption defect-type familial renal hypouricemia. Future studies are needed to identify a responsible gene for the postsecretory reabsorption defect-type familial renal hypouricemia.
Inhibition of prostasin expression by TGF-beta1 in renal epithelial cells.
BACKGROUND: Prostasin has been shown to be involved in the regulation of sodium handling in the kidney. TGF-beta1 has been demonstrated to suppress alphaENaC expression and sodium uptake. Therefore, we hypothesized that TGF-beta1 may regulate prostasin expression to modulate sodium reabsorption in the kidney. METHODS: To determine if TGF-beta1 has an effect on prostasin expression, we isolated 2.9 kb of the rat prostasin promoter, and measured its transcriptional activity with a luciferase assay in mouse cortical collecting duct cell line (M-1). The effect of TGF-beta1 on the mRNA and protein abundance of prostasin, and amiloride-sensitive (22)Na uptake was determined. RESULTS: Treatment of M-1 cells with 20 ng/mL of TGF-beta1 for 24 hours significantly decreased the promoter activity by 50 +/- 1%, and the inhibitory effect was dose dependent over the range of 0.1 to 20 ng/mL. We identified a 50 bp region (-410 to -360) containing c-Rel-like sequence in prostasin promoter that is responsible for the TGF-beta1-mediated inhibition, and found that TGF-beta1 increases IkappaBalpha expression in M-1 cells. TGF-beta1 reduced endogenous prostasin mRNA and protein expression in M-1 cells by 50 +/- 12% and 44 +/- 12%, respectively, and the amiloride-sensitive (22)Na uptake by 35.9 +/- 4.8%. CONCLUSION: Our findings indicate the possibility that TGF-beta1 transcriptionally inhibits prostasin expression by the induction of IkappaBalpha and the subsequent inhibition of NF-kappaB/Rel activity in M-1 cells, and also suggest the possibility that TGF-beta1 might inhibit sodium reabsorption through a reduction in prostasin expression and subsequent inhibition of ENaC activity.
Oxidation and carboxy methyl lysine-modification of albumin: possible involvement in the progression of oxidative stress in hemodialysis patients.
Hemodialysis (HD) patients are frequently in a state of increased oxidative stress, and hyperglycemia appears to be a major factor. We recently found that oxidized human serum albumin (HSA) is a reliable marker of oxidative stress in HD patients. However, the issue of whether oxidized HSA is associated with the progression of oxidative stress in HD patients with or without diabetes is not clear. In the present study, we examined the effect of a qualitative modification of HSA in HD patients with or without diabetes. Blood samples from 10 HD patients with diabetes, 7 HD patients without diabetes, and 10 healthy age-matched controls were examined. The increase in plasma protein carbonyl content and advanced glycation endproducts (AGEs) in HD patients was largely due to an increase in the levels of oxidized HSA. Furthermore, these increases were greatest in HD patients with diabetes. Purified HSA from HD patients (non-DM-HSA) was carbonylated and AGE-modified. The amount of modified HSA was the highest in HD patients with diabetes (DM-HSA). Carboxy methyl lysine (CML)-modified HSA triggered a neutrophil respiratory burst, and this activity was closely correlated with the increase in the CML/HSA ratio. These findings indicate that uremia plays an important role in the progression of oxidative stress in HD patients via an increase in CML-modified HSA. They also indicate that diabetic complications further exacerbate the progression of oxidative stress by further increasing the amount of these modified HSA molecules.
Effect of telmisartan on ambulatory blood pressure monitoring, plasma brain natriuretic peptide, and oxidative status of serum albumin in hemodialysis patients.
The effect of telmisartan on ambulatory blood pressure, plasma neurohormonal parameters, and oxidation of serum albumin has not been investigated in hemodialysis (HD) patients. Thirteen hypertensive HD patients were treated with 40 mg telmisartan once daily, and 24-h ambulatory blood pressure monitoring was performed after 0, 4, and 8 weeks of treatment. Plasma renin activity, plasma aldosterone concentration (PAC), brain natriuretic peptide (BNP) level, and serum oxidized albumin level were determined at the same time points. Serum telmisartan concentration was also measured at 4 and 8 weeks. Telmisartan significantly reduced systolic blood pressure and diastolic blood pressure (both awake and sleeping) after 4 weeks, and these pressures showed a further significant decrease after 8 weeks. Plasma levels of aldosterone, BNP, and serum oxidized albumin were markedly decreased after 4 weeks and these lower levels were maintained at 8 weeks. The trough serum telmisartan concentration was not significantly different at 8 weeks compared with 4 weeks. Throughout the treatment period, there were no significant adverse effects. Telmisartan effectively lowers blood pressure and reduces PAC, BNP, and oxidative stress and is safe and well-tolerated by HD patients. A long-term study in a large population is required to determine the influence of telmisartan therapy on cardiovascular mortality and morbidity in HD patients.
Familial pseudohyperkalemia: a rare syndrome, but diverse genetic heterogeneity.
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[Regulation of sodium and water balance by the kidney].
Maintenance of consistent internal environment is essential for mammalians to survive in waterless circumstances. Extracellular fluid (ECF) volume is maintained within a narrow range in normal humans, despite day-to-day variations in the dietary intake of salt and water. In general, kidneys are the major organs that play a predominant role in the control of the fluid and electrolyte balance. Recent advances in the molecular biology have provided significant contribution to the identification of kidney specific channels and transporters for sodium and water, leading to better understandings of the regulation of sodium and water balance. Generalized edema formation, a clinical hallmark of ECF volume expansion, represents the accumulation of excessive fluid volumes in the interstitial compartment and is invariably associated with renal sodium retention. In this article, we review the regulation of sodium and water balance in the kidney to understand the renal mechanisms of edema formation.
[Renal action of aldosterone].
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Liddle's syndrome caused by a novel mutation in the proline-rich PY motif of the epithelial sodium channel beta-subunit.
Liddle's syndrome is an autosomal dominant form of salt-sensitive hypertension and has been shown to be caused by missense or frameshift mutations in the amiloride-sensitive epithelial sodium channel (ENaC), which is composed of three subunits: alpha, beta, and gamma. All disease mutations either remove or alter amino acids of the target proline-rich PPPxY sequence (PY motif) of beta- or gamma-ENaC and result in increased channel activity. In this report, we present a family with Liddle's syndrome whose abnormality is caused by a novel missense mutation, P616R, in the PY motif of the betaENaC. Functional studies using the P616R mutant expressed in Xenopus oocytes showed an approximately 6-fold increase in the amiloride-sensitive sodium channel activity compared with that of the wild type. These findings provide additional clinical evidence that a conserved PY motif is critically important for the regulation of ENaC activity.
Intravenous iron administration induces oxidation of serum albumin in hemodialysis patients.
BACKGROUND: Intravenous iron administration (IVIR) is effective for correcting anemia in hemodialysis (HD) patients. However, it may also enhance the generation of hydroxyl radicals. Recently, plasma proteins have been demonstrated to be extremely susceptible to oxidative stress. Therefore, we investigated the effect of IVIR on the oxidative status of albumin, a major plasma protein, in HD patients. METHODS: Eleven hemodialysis (HD) patients were treated with 40 mg of saccharated ferric oxide intravenously after every dialysis session for four weeks, and 11 age-/gender-matched HD patients were treated with vehicle. We performed high performance liquid chromatography (HPLC) analysis of serum albumin and determined the levels of reduced and oxidized albumin. Carbonyl formation of plasma proteins were also measured using an anti-2,4 dinitrophenylhydrazine antibody in patients with or without IVIR. RESULTS: IVIR resulted in an increase in both disulfide form (f(HNA-1)) and oxidized form (f(HNA-2)) of albumin in HD patients (36.0 +/- 6.03 vs. 41.7 +/- 6.27; 5.46 +/- 1.50 vs. 8.7 +/- 2.22, respectively, P < 0.05). The findings here also show that IVIR substantially increased plasma protein carbonyl content by oxidizing albumin. In addition, we found a strong correlation between plasma carbonyl content and the levels of oxidized albumin (f(HNA-1) and f(HNA-2)) in HD patients (R= 0.674 and R= 0.724, respectively, P < 0.01). CONCLUSION: The results of this study indicate that the HPLC analysis of serum albumin represents a potentially useful method for the quantitative and qualitative evaluation of oxidative stress in HD patients, and strongly suggest the possibility that oxidative stress, generated by IVIR, enhances the oxidation of albumin in those patients.
Increased endothelin-1 expression in the kidney in hypercalcemic rats.
BACKGROUND: Although hypercalcemia causes diuresis and natriuresis, the molecular mechanisms of these effects are not well established. Recently, the important role of the calcium-sensing receptor (CaR) in hypercalcemia-induced polyuria was reported. Endothelin-1 (ET-1) that is locally produced in the nephron has been suggested to have the natriuretic and/or diuretic effects in the kidney. Therefore, we hypothesized that ET-1 expression could be increased through the activation of CaR in the kidney in hypercalcemia. METHODS: Rats were made hypercalcemic by dihydrotachysterol (DHT) treatment. The urinary concentration of ET-1 and the mRNA expression of ET-1 in the kidney were determined. Immunohistochemistry was performed to determine types of the cells that produce ET-1. CaR and ET-1 promoter luciferase constructs were co-expressed in COS-7 cells and the ET-1 promoter activity following the addition of extracellular calcium was measured by the luciferase assay. RESULTS: In hypercalcemic rat, urinary ET-1 excretion was increased by twofold, and ET-1 mRNA expression was increased in the kidney cortex by threefold. In cortical collecting duct (CCD), both principal cells and intercalated cells synthesized ET-1. In cells that express CaR, ET-1 promoter was activated in a dose-dependent manner by extracellular calcium over the range of 0.5 to 3.0 mmol/L. CONCLUSIONS: First, activation of CaR increases ET-1 transcription in a dose-dependent manner. Second, hypercalcemia increases ET-1 production in the kidney cortex. These data suggest the possibility that CaR might play an important role in hypercalcemia-induced increase in ET-1 production.
Two male siblings with hereditary renal hypouricemia and exercise-induced ARF.
Familial renal hypouricemia with exercise-induced acute renal failure (ARF) is rare. A 45-year-old man presented with abdominal pain, vomiting, and oliguria after severe exercise. The diagnosis was ARF based on high serum creatinine (SCr) level (5.1 mg/dL [451 micromol/L]). Renal function recovered completely within 2 weeks of conservative treatment (creatinine clearance [Ccr], 100.4 mL/min [1.67 mL/s]). After remission, laboratory results showed serum urate (SUA) of 0.8 mg/dL (48 micromol/L), and fractional excretion of uric acid (FE(UA)) of 46%. The final diagnosis was ARF associated with idiopathic renal hypouricemia. Other diseases that could increase the excretion of urate were excluded. Because only mild responses were observed both in pyradinamide and benzbromarone loading tests, he was considered to be a presecretory reabsorption disorder type. The younger brother (42 years old) also had episodes of low and middle back pain after severe exercise and experienced similar attacks at least 5 times since the age of 29. SCr level was elevated in every attack. Hypouricemia (SUA, 1.0 mg/dL [59 micromol/L]) and high urinary urate excretion (FE(UA), 65.7%) also were detected. Renal function recovered almost completely without any specific treatment. Radiologic examination of the 2 cases showed bilateral urolithiasis probably caused by the high urinary urate excretion. Sequence analysis of a urate anion exchanger known to regulate blood urate level (URAT1 gene) in both brothers showed homozygous mutation in exon 4 (W258Stop), resulting in a premature truncated URAT1 protein. Both their parents and their children showed heterozygous mutation of the URAT1 gene. This is the first report of the 2 male siblings of familial renal hypouricemia complicated with exercise-induced ARF, with definite demonstration of genetic abnormality in the responsible gene (URAT1).
Inhibition of prostasin secretion by serine protease inhibitors in the kidney.
A serine protease, prostasin, has been shown to stimulate the activity of amiloride-sensitive sodium channels (ENaC). Prostasin is a glycosylphosphatidylinositol-anchored protein that is found free in physiologic fluids and tissue culture medium, but the mechanism by which prostasin is secreted from the cells has not been elucidated. The current studies found that serine protease inhibitor aprotinin blocked the secretion of prostasin in a mouse cortical collecting duct (CCD) cell line (M-1 cells). A synthetic serine protease inhibitor, nafamostat mesilate (NM), which is commonly used for the treatment of pancreatitis and disseminated intravascular coagulation in Japan, also inhibited the secretion of prostasin in M-1 cells. Continuous infusion of NM into rats resulted in a substantial decrease in urinary prostasin and urinary sodium excretion. p-guanidinobenzoic acid and 6-amidino-2-naphtol, catalytically inactive metabolites of NM, had no effect on prostasin secretion both in M-1 cells and in rats. These findings suggest that a serine protease-sensitive mechanism is involved in the secretion of prostasin in vitro as well as in vivo. Potassium secretion in the CCD is tightly linked to sodium reabsorption through EnaC; therefore, NM-induced decrease in prostasin secretion and subsequent inhibition of ENaC activity could account for the side effects of hyponatremia and/or hyperkalemia that are found sometimes in patients treated with NM. The results indicate an important role for prostasin in sodium reabsorption in the kidney under pathophysiologic conditions.