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Katsuhiro Kato,
Tsubasa Yazawa, Kentaro Taki,
Kazutaka Mori,
Shujie Wang,
Tomoki Nishioka,
Tomonari Hamaguchi,
Toshiki Itoh,
Tadaomi Takenawa,
Chikako Kataoka,
Yoshiharu Matsuura,
Mutsuki Amano,
Toyoaki Murohara,
Kozo Kaibuchi
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ABSTRACT: Cell migration is essential for various physiological and pathological processes. Polarization in motile cells requires the coordination of several key signaling molecules, including RhoA small GTPases and phosphoinositides. Although RhoA participates in a front-rear polarization in migrating cells, little is known about the functional interaction between RhoA and lipid turnover. We find here that src-homology 2-containing inositol-5-phosphatase 2 (SHIP2) interacts with RhoA in a GTP-dependent manner. The association between SHIP2 and RhoA is observed in spreading and migrating U251 glioma cells. The depletion of SHIP2 attenuates cell polarization and migration, which is rescued by wild-type SHIP2 but not by a mutant defective in RhoA binding. In addition, the depletion of SHIP2 impairs the proper localization of phosphatidylinositol 3,4,5-trisphosphate, which is not restored by a mutant defective in RhoA binding. These results suggest that RhoA associates with SHIP2 to regulate cell polarization and migration.
Molecular biology of the cell 05/2012; 23(13):2593-604. · 5.98 Impact Factor
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Mutsuki Amano,
Yuta Tsumura, Kentaro Taki,
Hidenori Harada,
Kazutaka Mori,
Tomoki Nishioka,
Katsuhiro Kato,
Takeshi Suzuki,
Yosuke Nishioka,
Akihiro Iwamatsu,
Kozo Kaibuchi
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ABSTRACT: Protein kinases are major components of signal transduction pathways in multiple cellular processes. Kinases directly interact with and phosphorylate downstream substrates, thus modulating their functions. Despite the importance of identifying substrates in order to more fully understand the signaling network of respective kinases, efficient methods to search for substrates remain poorly explored.
We combined mass spectrometry and affinity column chromatography of the catalytic domain of protein kinases to screen potential substrates. Using the active catalytic fragment of Rho-kinase/ROCK/ROK as the model bait, we obtained about 300 interacting proteins from the rat brain cytosol fraction, which included the proteins previously reported as Rho-kinase substrates. Several novel interacting proteins, including doublecortin, were phosphorylated by Rho-kinase both in vitro and in vivo.
This method would enable identification of novel specific substrates for kinases such as Rho-kinase with high sensitivity.
PLoS ONE 01/2010; 5(1):e8704. · 4.09 Impact Factor
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ABSTRACT: This study aimed to investigate the association of the KLOTHO gene single nucleotide polymorphisms (SNPs), G-395A and C1818T, with various laboratory data in 219 Japanese hemodialysis (HD) patients.
The genotyping of G-395A in the promoter region and C1818T in exon 4 was performed using polymerase chain reaction with confronting two-pair primers (PCR-CTPP) assay.
In HD patients, the allele frequencies of G-395A were 0.847 for the G allele and 0.153 for the A allele and those of the C1818T were 0.829 for the C allele and 0.171 for the T allele. There were no significant differences in allele frequencies of G-395A and those of the C1818T between HD patients and healthy subjects. Multivariate analysis adjusted for age and duration on HD demonstrated that uric acid was significantly high in A allele carriers of G-395A compared with GG genotype in all and female patients. Low-density lipoprotein cholesterol was significantly low in T allele carriers of C1818T compared with CC genotype in all and male patients.
KLOTHO gene SNPs G-395A and C1818T are associated with low-density lipoprotein cholesterol and uric acid in HD patients.
American Journal of Nephrology 09/2009; 30(4):383-8. · 2.54 Impact Factor
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ABSTRACT: Cardiac troponin T (cTnT) is used as a biomarker of myocardial damage for the diagnosis of acute myocardial infarction and acute coronary syndrome. The aim was to investigate the association between advanced glycation end products (AGEs) and cTnT in hemodialysis (HD) patients.
The plasma level of cTnT in 224 HD patients was measured using the electrochemiluminescence immunoassay. The plasma levels of N(epsilon)-(carboxymethyl)lysine (CML) and pentosidine were measured using an enzyme-linked immunosorbent assay.
The cTnT-positive group (>0.1 ng/ml) showed significantly high plasma levels of calcium, CML and pentosidine as compared with the cTnT-negative group. In multiple logistic regression analysis, the prevalence of patients with high plasma calcium (>median) was increased in the cTnT-positive group as compared with the cTnT-negative group (OR: 5.08, 95% CI: 1.62-15.92, p < 0.01). The prevalence of high plasma CML (>median) was increased in the cTnT-positive group (OR: 4.45, 95% CI: 1.41-14.03, p < 0.05). Further, the prevalence of high plasma pentosidine (>median) was also increased in the cTnT-positive group (OR: 4.94, 95% CI: 1.55-15.70, p < 0.01).
In addition to calcium, AGEs such as CML and pentosidine were associated with cTnT, a marker of myocardial damage, in HD patients.
American Journal of Nephrology 04/2008; 28(5):701-6. · 2.54 Impact Factor
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ABSTRACT: Metabolic syndrome (MetS) is characterized by the presence of atherogenic risk factors, and is associated with a marked increase in the risk of cardiovascular disease. Recently, the criteria of MetS were newly defined in Japan. We examined the relationship between MetS and the various metabolic parameters in Japanese subjects. This study included 458 Japanese subjects undergoing medical checkups at Nagoya University Hospital. New criteria developed by the joint committee of eight Japanese medical societies for the clinical recognition of MetS were adopted. We examined the association between MetS and various metabolic parameters, including liver enzymes (alanine aminotransferase, ALT; gamma glutamyltransferase, GGT) and highly sensitive C-reactive protein (hsCRP). The mean overall prevalence of MetS was 8.7% (male: 12.9%; female: 2.2%, p = 0.0001). MetS was significantly associated with elevated ALT (> 45 IU/L) (OR: 3.37, 95% CI: 1.19-9.52, p < 0.05) and GGT (> 64 IU/L in males, > 45 IU/L in females) (OR: 4.96, 95% CI: 2.31-10.66, p = 0.0001), respectively. MetS was also significantly associated with elevated hsCRP (> or = 0.1 ng/mL) (OR: 2.77, 95% CI: 1.20-6.41, p < 0.05). Thus, MetS was associated with elevated liver enzymes (especially, GGT), and inflammation (hsCRP).
Nagoya journal of medical science 03/2008; 70(1-2):1-9.
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Yoshiko Ishida,
Koji Suzuki, Kentaro Taki,
Toshimitsu Niwa,
Shozo Kurotsuchi,
Hisao Ando,
Akira Iwase,
Kazuko Nishio,
Kenji Wakai,
Yoshinori Ito,
Nobuyuki Hamajima
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ABSTRACT: Helicobacter pylori (H. pylori) infection in gastric mucosa may cause systemic inflammatory reaction. This study aimed to examine the association between the infection and serum high sensitivity C-reactive protein (hsCRP).
Subjects were comprised of three groups; 453 health checkup examinees from Yakumo town inhabitants in Hokkaido, Japan (YTI, 153 males and 300 females), 449 health checkup examinees (ENUH, 273 males and 176 females), and 255 female patients of an infertility clinic (PIC), Nagoya University Hospital. Twenty participants with hsCRP more than 1 mg/dl were excluded from the analysis. Those with hsCRP more than 0.1mg/dl were defined as high hsCRP individuals. H. pylori infection status was examined with a serum IgG antibody test.
When the three groups were combined, the geometric mean of hsCRP concentration was significantly higher among the seropositives (0.047 mg/dl) than among the seronegatives (0.035 mg/dl); p<0.0001 by a t-test. The percentage of high hsCRP individuals was also higher in the seropositives than in the seronegatives among any group; 23.3% and 20.1% in YTI, 22.0% and 16.0% in ENUH, and 32.7% and 18.7% in PIC, respectively, although the difference was significant only in ENUH. The summary odds ratio of the high hsCRP for the seropositives relative to the seronegatives was 1.38 (95% confidence interval, 1.01-1.89), when age, sex, body mass index, smoking, and subject group were adjusted by a logistic model.
In three groups, hsCRP was higher among the infected individuals. The summary odd ratio indicated that H. pylori infection could influence the serum hsCRP level.
International journal of medical sciences 01/2008; 5(4):224-9. · 2.24 Impact Factor
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Masato Kikuchi,
Makoto Nakamura,
Kohei Ishikawa,
Toshimitsu Suzuki,
Hiroaki Nishihara,
Tomomi Yamakoshi,
Kazuko Nishio, Kentaro Taki,
Toshimitsu Niwa,
Nobuyuki Hamajima,
Hiroko Terasaki
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ABSTRACT: To determine the relationship between systemic C-reactive protein (CRP) levels and polypoidal choroidal vasculopathy (PCV) and advanced neovascular age-related macular degeneration (AMD) in Japanese patients.
Case-control study.
Ninety-seven patients with PCV, 176 with advanced neovascular AMD, and 262 control subjects without any macular abnormality were studied.
Color fundus photographs of the macular area were taken from both eyes in all subjects. Indocyanine green angiography and fluorescein angiography were performed for diagnosis. The CRP level was measured by a high-sensitivity assay using a latex aggregation immunoassay, and the levels in patients with PCV and neovascular AMD were compared with that in the control group using the Kruskal-Wallis test. Associations between CRP and PCV or neovascular AMD were compared using logistic regression analysis by computing the odds ratios (ORs) and 95% confidence intervals (CIs) after the study populations were divided into quartiles.
The CRP levels in patients with PCV, patients with neovascular AMD, and control subjects. Standard univariate and multivariate analyses between groups.
Median CRP levels were significantly higher in cases with PCV (0.94 mg/l) or with advanced neovascular AMD (0.95 mg/l) than in control subjects (0.43 mg/l) (P<0.001 for Kruskal-Wallis test). After adjusting for baseline characteristics such as age, gender, smoking status, alcohol use, body mass index, history, and use of antiinflammatory drugs, the increase in risk was significant for the highest quartile of CRP for both PCV (OR, 3.53; 95% CI, 1.49-8.40) and neovascular AMD (OR, 4.08; 95% CI, 1.94-8.56), and for the third quartile of CRP for neovascular AMD (OR, 2.29; 95% CI, 1.07-4.91). The trends for an increase in risk of disease with increase in CRP were statistically significant for both PCV (P = 0.001) and neovascular AMD (P<0.001).
The significant associations between elevated serum CRP levels and PCV or neovascular AMD in the Japanese strongly suggest that inflammatory processes are involved in the pathogenesis of PCV and neovascular AMD.
Ophthalmology 09/2007; 114(9):1722-7. · 5.45 Impact Factor
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ABSTRACT: Indoxyl sulfate shows nephrotoxicity and is a stimulating factor for progression of chronic kidney disease (CKD). This study was conducted to determine (1) whether the indoxyl sulfate-lowering capacity of oral sorbents (Kremezin [AST-120], Kureha Corporation, Tokyo, Japan; Merckmezin, Merck Hoei Ltd., Osaka, Japan) affects the prognosis of kidney function in CKD, and (2) whether oral sorbents reduce the markers of oxidative stress.
Rats with CKD were produced by 4/5 nephrectomy and were randomized into 3 groups: control rats, Merckmezin-treated rats, and Kremezin-treated rats. Kremezin and Merckmezin were administered to rats at a dose of 4 g/kg with powder chow for 16 weeks, whereas powder chow alone was administered to control rats.
Administration of Kremezin significantly decreased serum and urine levels of indoxyl sulfate and serum creatinine and significantly increased creatinine clearance as compared with control values. The change in serum indoxyl sulfate noted from the initial to the final week showed a positive correlation with the change in serum creatinine and a negative correlation with the change in creatinine clearance. Kremezin significantly reduced urine levels of acrolein, a marker of oxidative stress, as compared with control levels.
The indoxyl sulfate-lowering capacity of oral adsorbents affects the prognosis of kidney function in CKD. The more serum indoxyl sulfate is reduced, the better kidney function is preserved. Kremezin alleviates oxidative stress in the kidneys by reducing serum levels of indoxyl sulfate.
Journal of Renal Nutrition 02/2007; 17(1):48-52. · 1.57 Impact Factor
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ABSTRACT: Indoxyl sulfate is a uremic toxin that accelerates the progression of chronic kidney disease (CKD). Serum levels of indoxyl sulfate are increased in dialysis patients. It was reported that indoxyl sulfate plays a role in endothelial dysfunction in uremic patients, and stimulates proliferation of rat vascular smooth muscle cells (VSMC). We examined associations between indoxyl sulfate and several markers related to atherosclerosis.
The association between indoxyl sulfate and atherosclerotic risk factors was studied in 224 hemodialysis (HD) patients (123 male, 101 female). Serum levels of indoxyl sulfate were measured by using high-performance liquid chromatography (HPLC).
There were significant differences in serum levels of creatinine, calcium x phosphate and pentosidine between high- and low-indoxyl sulfate level groups. Indoxyl sulfate showed significant positive correlations with pentosidine, creatinine, and protein catabolic rate, and a significant negative correlation with high-density lipoprotein (HDL) cholesterol. Further, pentosidine, creatinine, and HDL-cholesterol were independently associated with indoxyl sulfate by multiple linear regression analysis.
In addition to creatinine, pentosidine and HDL-cholesterol, the risk factors of atherosclerosis, were associated with indoxyl sulfate in HD patients. Indoxyl sulfate may be involved in the pathogenesis of atherosclerosis.
American Journal of Nephrology 02/2007; 27(1):30-5. · 2.54 Impact Factor
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ABSTRACT: Indoxyl sulfate shows nephrotoxicity and is a stimulating factor for progression of chronic renal failure (CRF). Indoxyl sulfate is taken up by renal proximal tubular cells through organic anion transporters 1 and 3 (OAT1/3), and is accumulated in the renal proximal tubular cells of uremic rats. To determine whether indoxyl sulfate is accumulated in human OAT1/3 (hOAT1/3)-positive renal proximal tubular cells, localization of indoxyl sulfate and hOAT1/3 in the kidneys of CRF patients was determined by immunohistochemistry. Kidney samples were obtained by autopsy from 9 CRF patients (mean serum creatinine 4.7 mg/dL, ranging from 2.0 to 14.5 mg/dL) and 9 patients with non-kidney disease (mean serum creatinine 0.6 mg/dL, ranging from 0.4 to 0.9 mg/dL). Immunohistochemistry was performed using antibodies against indoxyl sulfate, hOAT1, and hOAT3. Indoxyl sulfate was localized in the hOAT1- and hOAT3-positive renal tubular cells in the kidneys of CRF patients. The indoxyl sulfate-positive area in the kidneys was markedly increased in the kidneys of CRF patients compared with patients with non-kidney disease. The indoxyl sulfate-positive area was positively correlated with serum creatinine. In conclusion, in CRF patients, indoxyl sulfate is accumulated in the tubular cells with hOAT1 and/or hOAT3 localized at the basolateral membrane. The extent of indoxyl sulfate accumulation in the kidneys is more prominent in those patients with more severe CRF.
Journal of Renal Nutrition 08/2006; 16(3):199-203. · 1.57 Impact Factor
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ABSTRACT: Intestinal microflora is deranged in hemodialysis (HD) patients as an increase in aerobic bacteria such as Escherichia coli and a decrease in anaerobic bacteria such as Bifidobacterium . Bifidobacteria ferment carbohydrates to produce acetic acid and lactic acid, which inhibit the intestinal putrefaction. Thus, intake of Bifidobacteria effectively restores the disturbed microflora to normal. However, Bifidobacteria in most medical products and healthy foods cannot usually survive because of exposure to gastric juices before it reaches the intestines. A gastroresistant seamless capsule prevents Bifidobacteria from inactivation by acidic gastric juice and allows it to be active in the intestines. We showed that the oral administration of Bifidobacterium longum in a gastroresistant seamless capsule to HD patients is effective in decreasing the pre-HD serum levels of homocysteine, indoxyl sulfate, and triglyceride. The reduction in the serum level of homocysteine is mainly attributable to the supply of folate produced by Bifidobacterium longum in the human intestines.
Journal of Renal Nutrition 02/2005; 15(1):77-80. · 1.57 Impact Factor
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ABSTRACT: Intestinal microflora is deranged in hemodialysis (HD) patients as increased aerobacteria such as Escherichia coli, and decreased anaerobacteria such as bifidobacteria. Indole, a precursor of indoxyl sulfate, is produced by E coli but not by bifidobacteria. The serum levels of indoxyl sulfate are increased markedly in HD patients and cannot be reduced efficiently by HD because of its albumin binding.
To compare the effect of oral administration of Bifidobacterium longum in gastro-resistant seamless capsule (Bifina) on indoxyl sulfate levels with that of Bifidobacteria in powder formulation (Lac B), Bifina was administered to 11 HD patients for 5 weeks, and Lac B to another group of 11 HD patients for 5 weeks. The authors measured the serum level of indoxyl sulfate by using high-performance liquid chromatography.
The pre-HD serum levels of indoxyl sulfate significantly decreased in Bifina-treated patients (before, 4.9 +/- 1.7 mg/dL, 4.5 mg/dL, mean +/- SD, median, after 5 weeks, 3.5 +/- 1.3 mg/dL, 3.8 mg/dL; P < 0.005). However, they did not decrease in the Lac B-treated patients (before, 4.8 +/- 1.4 mg/dL, 4.5 mg/dL, after 5 weeks, 5.2 +/- 2.0 mg/dL, 5.1 mg/dL).
Oral administration of Bifina to HD patients is effective in reducing the serum levels of indoxyl sulfate by correcting the intestinal microflora. Gastro-resistant seamless capsule prevents bifidobacteria from its inactivation in acidic gastric juice, and allows it to be actived in the intestine.
American Journal of Kidney Diseases 03/2003; 41(3 Suppl 1):S142-5. · 5.43 Impact Factor
