New genotypes of hepatitis C virus.
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Genes, Viral↗
Biomedical subjects
A Takada
Publications and source records attributed to A Takada.
Correlation between serotonergic measures in cerebrospinal fluid and blood of subhuman primate.
The relationship between the concentration of serotonin (5-hydroxytryptamine; 5-HT), its precursor; tryptophan (Trp) and the main metabolite 5-hydroxyindoleacetic acid (5-HIAA) in cerebrospinal fluid (CSF) and blood of monkey have been studied. 5-HT, Trp and 5-HIAA underwent circadian changes in both CSF and blood. Significant correlations were found between 5-HT, 5-HIAA and Trp in CSF and blood. The significance of these findings and their relationship to the use of peripheral serotonergic system as a functional model of the central nervous system are discussed.
Animals↗
Rapid steady-state analysis of blood-brain transfer of L-Trp in rat, with special reference to the plasma protein binding.
We estimated constants for the binding of tryptophan (Trp) to plasma proteins, and for the transfer of Trp from plasma to brain in rat. The measurements were made under conditions in which the plasma and brain concentrations of Trp were raised to new steady-states for at least 10 min before being measured. The concentration of other competing amino acids were also at a steady-state. The plasma Trp concentration was elevated by i.p. injection of different doses of L-tryptophan methyl ester 60 min before the measurement of the plasma-brain transfer. We simultaneously measured blood flow with [14C]-butanol, and the brain tissue Trp uptake with [3H]Trp. The maximal velocity (Vmax), apparent half-saturation Michaelis-Menten constant (Km(app)), and diffusion constant (PdS) for Trp transport from plasma into brain were found to be 7.0 +/- 2.1 nmol g-1 min-1, 36 +/- 17 microM, and 0.065 +/- 0.006 ml g-1 min-1, respectively. The maximum plasma protein binding (Bmax) and dissociation constant (KD) for Trp were estimated at 360 +/- 16 nmol/ml-plasma and 81 +/- 10 microM, respectively. We conclude that the plasma protein binding of Trp inhibits the blood-brain transfer in inverse proportion to the plasma free Trp concentration.
Animals↗
Differences in the hepatitis C virus genotypes in different countries.
We recently classified the hepatitis C virus (HCV) into 4 types (HCV-PT, -K1, -K2a and -K2b) according to differences in nucleotide sequences. It was found that HCV-PT, the prototype reported from the U.S.A., was rare in Japan, suggesting that distribution of HCV genotypes may be different in various countries. The prevalence of HCV genotypes was therefore compared in different countries. Genotyping of HCV was performed by slot-blot hybridization analysis using cDNA probes specific to each type of HCV or by restriction fragment length polymorphism analysis. In 121 Japanese non-cancer patients, the prevalence of HCV genotypes was 77.7% for HCV-K1, 16.5% for HCV-K2a and 5.0% for HCV-K2b. HCV-PT was detected in only 1 patient (0.8%). The prevalence in 43 Japanese hepatocellular carcinoma (HCC) patients was 74.4% for HCV-K1, 18.6% for HCV-K2a and 4.7% for HCV-K2b. HCV-PT was found in only 1 sample. In 19 European non-cancer patients, HCV-PT was found in 42.1% and HCV-K1 was found in 52.6%. HCV-K2 was not found. All 7 samples from European HCC patients were HCV-K1, indicating a significantly higher prevalence than in non-cancer patients. In 13 Brazilian non-cancer patients, the distribution pattern was similar to that of the Europeans. In 10 samples from the U.S.A., HCV-PT was found in 70% and HCV-K2 was found in 1 sample. In 18 Chinese non-cancer patients, HCV-K1 was found in 44.4%, HCV-K2a in 50.0% and HCV-K2b in 5.6% HCV-PT was not found. Two samples from Chinese HCC patients were HCV-K1.(ABSTRACT TRUNCATED AT 250 WORDS)
Brazil↗
Analysis of the characteristics of microheterogeneity of various serum glycoproteins in chronic alcoholics.
It has been reported that microheterogeneity of serum glycoproteins including transferrin is found in alcoholic liver disease. In the present study, microheterogeneity of serum glycoproteins in alcoholic liver disease patients was analysed using the Western blotting technique after isoelectric focusing. Microheterogeneity was found for serum alpha 1-antitrypsin, alpha 2-macroglobulin, caeruloplasmin, alpha 1-acid glycoprotein and hemopexin as well as transferrin. Microheterogeneity disappeared following treatment with sialidase in some but not all glycoproteins. In hemopexin, microheterogeneity was recognized only after treatment with sialidase. These results suggest that mechanisms of microheterogeneity of serum glycoproteins in alcoholic liver disease may vary. One mechanism may be the interference of glycosylation of glycoproteins in the Golgi apparatus, and another may be the decrease of asialo-protein receptors in hepatocytes.
Blotting, Western↗
Effects of ethanol and acetaldehyde on the maturation of hepatic secretory glycoproteins.
The effects of ethanol and acetaldehyde on the maturation and secretion of hepatic glycoproteins in cultured rat hepatocytes were analysed using a pulse-chase labelling method and autoradiography following two-dimensional gel electrophoresis. A spot for transferrin was detected at mol. wt 77,000 and pI 5.2-5.4 on the autoradiogram in the control culture. The spot for transferrin shifted to the more basic isoforms by treatment for 3 hr with monensin or tunicamycin. The spot for transferrin also shifted to the more basic side by treatment with acetaldehyde for 3 hr or by additional treatment for 3 hr following pretreatment with ethanol for 6 hr. However, the shift of transferrin was not observed following treatment with ethanol for 3 hr. These results suggest that glycosylation of secretory glycoproteins in the Golgi apparatus is inhibited by acetaldehyde, but not by ethanol itself, and that this inhibition may play an important role in the development of alcoholic liver disease.
Acetaldehyde↗
Ethanol and acetaldehyde metabolism in cultured rat hepatocytes.
In the present study, ethanol (Et-OH) and acetaldehyde (Ac-CHO) metabolism in primary cultured hepatocytes isolated from chronically alcohol-fed rats were analysed to elucidate the characteristics of the metabolism of Ac-CHO produced through the non-alcohol dehydrogenase (non-ADH) pathway. Dimethyl sulfoxide was added to the culture medium to prevent a decrease in P450 IIE1 activity. Ac-CHO formed in the culture medium was trapped by semicarbazide to prevent a reutilization of Ac-CHO by hepatocytes. The degradation rate of Ac-CHO in the liver was similar in alcohol and non-alcohol-treated rats. Therefore, the characteristic of the metabolism of Ac-CHO in the chronically alcohol-fed rats was not clarified in this study. The possibility that the Ac-CHO trapping by semicarbazide was only 70%, and that the remaining Ac-CHO was reutilized for oxidation in the hepatocytes, should be considered. Some method to trap Ac-CHO completely in the culture medium is needed to clarify the characteristics of Ac-CHO metabolism.
Acetaldehyde↗
Type IV collagen and laminin contents of livers from patients with alcoholic liver disease.
Characteristic histological features of alcoholic liver disease (ALD) are pericellular and perivenular fibrosis. It has been emphasized from immunohistochemical studies that pericellular and perivenular fibrosis may be caused by the increase of type IV collagen (IV-C) and/or laminin (LM). However, quantitative changes of hepatic IV-C and LM contents in ALD are not well known. Recently, we have developed assay systems for IV-C and LM contents in liver biopsy specimens. In the present study, hepatic IV-C and LM contents in ALD and non-ALD patients were measured. Liver biopsy specimens were obtained from 36 patients with ALD, 24 patients with non-ALD and five patients without liver disease. IV-C and LM contents in liver biopsy specimens were measured using the one-step sandwich enzyme immunoassay system for human serum IV-C and LM levels. Total collagen (T-C) content was also measured by the method of Leon and Rojkind. Hepatic IV-C, LM and T-C contents were significantly higher in all types of liver disease than in controls, and tended to increase with the progression of fibrosis. Especially in ALD, both IV-C and LM contents increased from the early stage, and the values in each type of ALD were significantly higher than those in the corresponding type of non-ALD. The ratio of IV-C or LM to T-C was also significantly higher in ALD than in the corresponding non-ALD. The prominent increases of IV-C and LM at the early stage of fibrosis may be one of the characteristics of collagen metabolism in ALD.(ABSTRACT TRUNCATED AT 250 WORDS)
Biomarkers↗
Relationship between alcoholic liver disease and HCV infection.
The high prevalence of hepatitis C virus (HCV) markers in alcoholic liver cirrhosis (AL-LC) and hepatocellular carcinoma (HCC) suggests a close aetiopathogenic relationship between alcoholic liver disease (ALD) and HCV infection. In the present study, HCV markers in ALD were measured by the highly sensitive methods, and the changes of sequential HCV markers after abstinence in ALD patients were analysed in order to elucidate the effect of alcohol on HCV. Antibodies to HCV-related antigen were determined using the first or second generation test kit. HCV-RNA genomes encoding the NS-5 region were detected using the RT-PCR method. In the HCV-NS5 negative serum, HCV genomes of the 5'-noncoding region were detected using the two-stage PCR method. Titres of HCV-RNA were measured by multiple cyclic PCR and cDNA dot blotting. Typing of HCV genomes was carried out on the PCR product from the NS-5 region by slot blot hybridization using type-specific cDNA probes, or by restriction fragment length polymorphisms analysis. In alcoholic fibrosis and alcoholic hepatitis, the prevalence of HCV markers was low, suggesting that the main aetiological factor is alcohol but not HCV in these types of ALD. HCV markers were positive in the half of the patients with AL-LC, and in more than 80% of patients with AL-CH and AL-HCC, indicating that HCV infection closely relates to these types of ALD. The ratio of the K1 type to the K2 type of HCV genomes was 4:1 in all types of NANB liver disease.(ABSTRACT TRUNCATED AT 250 WORDS)
Adult↗
Role of the aldehyde dehydrogenase-1 isozyme in the metabolism of acetaldehyde.
At least four types of aldehyde dehydrogenase (ALDH) isozymes exist in human liver: acetaldehyde (Ac-CHO) is metabolized mainly by ALDH2. However, the role of ALDH1 in Ac-CHO metabolism is not well known, because ALDH1 deficiency is rare. In the present study, changes in blood levels of Ac-CHO were analyzed in a person with ALDH1 deficiency who is also heterozygous for the ALDH2 alleles, and subjects with normal ALDH1, in order to clarify the roles of ALDH1 in the metabolism of Ac-CHO. By isoelectric focusing analysis of ALDH in red blood cells (RBC), the ALDH1 band was not detectable in a male subject, indicating ALDH1 deficiency. In this subject, ALDH1 activity was nearly null, but the activities in his wife and two children were normal. Serial changes of blood Ac-CHO levels following a small amount of ethanol were analyzed. The peak levels of Ac-CHO were not different between the subject with ALDH1 deficiency and the person with normal ALDH1. However, the decrease from the peak values of Ac-CHO was clearly slower in the deficient subject than in ALDH1 normal subjects. Changes in Ac-CHO in blood were also analyzed in vitro. Ac-CHO metabolism by RBC from the ALDH1 deficient subjects was clearly slower than in those from controls. These results indicate that ALDH1 plays an important role in the metabolism of Ac-CHO in blood.
Acetaldehyde↗
Genetic factors related to the development of carcinoma in digestive organs in alcoholics.
Heavy drinkers do not always develop alcoholic liver disease or cancer of the digestive organs, indicating that genetic factors may be involved. Recently, the presence of genetic polymorphisms of cytochrome P4502E1 (2E1) was confirmed. The genotype of 2E1 was separated into three types; type A which is homozygous for the c1 gene, type B which is heterozygous for the c1 and c2 genes, and type C which is homozygous for the c2 genes. In the present study the relationship between genotypes of 2E1 and digestive diseases, including alcoholic liver disease, was analyzed. In healthy controls, the prevalence of types A, B and C was 69%, 31% and 0%, respectively. In patients with esophageal cancer, four out of five patients (80%) were type B, and the prevalence of type B was significantly higher than that in normal controls. These results suggest that the c2 gene of 2E1 may be related to the development of esophageal cancer. In patients with gastric disease, the prevalence of types A and B was 72% and 28%, respectively. In patients with large bowel disease, the prevalence of type A was 83-100%, significantly higher than that in healthy controls. These results suggest that the c2 gene of 2E1 may not be related to the development of large bowel disease. However, the possibility should be considered that the c1 gene of 2E1 may link to its development. In this study, the relationship between the genotype of 2E1 and alcoholic liver disease was also analyzed. All patients with alcoholic liver disease, including one hepatoma, had the c2 gene but not the c1 gene. The frequency of the c2 gene was quite different from that in non-alcoholic liver disease.(ABSTRACT TRUNCATED AT 250 WORDS)
Alcoholism↗
The relationship between the development of alcoholic liver and pancreatic diseases and the induction of gamma glutamyl transferase.
It is well known that the responses of serum gamma glutamyl transferase (GGT) to chronic alcohol drinking are different depending on the individual. In order to clarify the genetic backgrounds in the development of alcoholic liver and pancreatic diseases, the relationships between serum GGT response and alcoholic liver and pancreatic diseases in heavy drinkers were studied. The responses of GGT to alcohol drinking were classified into three groups: non-response, mild-response and hyperresponse. In alcoholic liver disease, non-responders were scarcely found and the response of GGT tended to increase in parallel with the progression of liver disease, when the hepatitis C virus (HCV) marker-positive patients were excluded. The differences in GGT levels between just after and at 4 weeks after abstinence in the HCV marker-negative patients were significantly higher than those in the HCV marker-positive patients. The rate of decrease in GGT activities during 4 weeks following abstinence was significantly higher in the HCV marker-negative patients than in the HCV marker-positive patients, indicating higher GGT induction in the HCV marker-positive patients. All patients with alcoholic pancreatitis, but without liver disease, were non-responders. All patients, except one, with severe pancreatitis were also non-responders. In alcoholic pancreatic disease, GGT induction correlated negatively with the development of pancreatitis. These results suggested that genetic polymorphism of GGT may link with the induction of GGT by alcohol drinking, and consequently link with the development of alcoholic liver and pancreatic diseases.
Adult↗
Studies on capillarization of the hepatic sinusoids in alcoholic liver disease.
It has been reported that basement membranes were found around the sinusoidal walls in cirrhotic livers, indicating the development of capillarization of the sinusoids. It has been also emphasized that capillarization of the sinusoids is more prominent in alcoholic liver disease (ALD). In the present study, factor VIII related antigen (VIII-Ag) and UEA-1 were identified immunohistochemically in order to analyze capillarization of the sinusoids in chronic liver diseases. Electron microscopic studies on the endothelial cells and sinusoids were also performed. Electron microscopic studies revealed that the number of fenestra in the endothelial cells decreased and basement membranes were clearly observed in the space of Disse from an early stage of ALD. However, these changes were not observed in the early stage of non-ALD. VIII-Ag or UEA-1 was not stainable in the sinusoidal cells of normal livers or at an early stage of non-ALD. However, in ALD, both VIII-Ag and UEA-1 were clearly demonstrated in the sinusoidal cells from the early stage of fibrosis. These results suggest that the sinusoidal endothelial cells may transform to vascular endothelial cells from an early stage of ALD. The alterations in the sinusoidal endothelium and the basement membrane formation in the Disse space indicate that capillarization of the sinusoid may occur. Capillarization of the sinusoid may cause a disturbance in exchanges of many bioactive substances between the sinusoidal blood and hepatocytes across the Disse space and may thereby contribute to the pathogenesis of ALD.
Basement Membrane↗
Effects of alcohol on the replication of hepatitis C virus.
We have reported that hepatitis C virus (HCV) RNA encoding the NS5 region (HCV-NS5) became negative following abstinence in some patients with alcoholic liver disease (ALD). In order to clarify the relationship between alcohol misuse (abuse) and the replication of HCV, serial changes of HCV-NS5 and the titer of HCV (HCV-titer) in blood after abstinence were studied in 11 male patients with ALD. Disappearance of HCV-NS5 after abstinence was found in five out of 11 patients (45%) and HCV-titer in blood decreased in all of these cases. In one patient, HCV-NS5 became positive and HCV-titers increased again following resumption of alcohol misuse. The etiology of liver disease in heavy drinkers was subdivided into three categories: alcohol alone, alcohol and HCV, and HCV alone, according to the diagnostic criteria of the Japanese research group for ALD. The etiology in four patients was a combination of alcohol and HCV, and was HCV alone in the remaining seven patients. In all four patients of the alcohol and HCV group, and one out of seven patients in the HCV alone group, HCV-NS5 became negative and HCV-titer decreased after abstinence. The decrease in HCV-titer was not related to the HCV genotypes. These results suggest that alcohol misuse may enhance the replication of HCV.
Adult↗
Effects of ethanol and acetaldehyde on the contraction of cultured Ito cells.
The effects of ethanol (Et-OH) and acetaldehyde (Ac-CHO) on the contraction of cultured Ito cells were studied using a time-interval videotape recorder (VTR). Ito cells were isolated from Wistar rats and cultured for 4 days. The motion of the cells was recorded by VTR and the sequential changes of the surface area of the each cell were evaluated by an image analyzer. No contraction of Ito cells was observed after incubation with Et-OH (100 mM) or Ac-CHO (500 microM) alone. Ten nanomoles endothelin-1 (ET-1) caused Ito cells to contract gradually. Pre-incubation of ET-1 with Ac-CHO at 37 degrees C for 1 hr inhibited the contraction of Ito cells completely. This inhibition was not seen in pre-incubation of ET-1 with Et-OH. In addition, the Ito cells which were pre-cultured with Ac-CHO for 6 hr did not contract at all after incubation with ET-1. In the isoelectric focusing of ET-1, a band was clearly seen at pI 4.6 after the treatment with Et-OH as well as non-treated ET-1. Two extra bands of ET-1 appeared in the anodic side of the gel after treatment with Ac-CHO, indicating the conformational changes of ET-1. These results indicated that the contraction of Ito cells induced by ET-1 was inhibited completely by Ac-CHO. This inhibitory effect of Ac-CHO was thought to be caused by binding Ac-CHO to ET-1 itself or to ET-1 receptors.
Acetaldehyde↗
Correlation between tumor proliferation and tumor tissue level of urokinase-type plasminogen activator.
We established a xenotransplanted human colon cancer strain, TK-3, which produces urokinase-type plasminogen activator (U-PA) and carcinoembryonic antigen (CEA) simultaneously. Immunohistochemical staining of U-PA revealed that U-PA was located in the cytoplasm of cancer cells. Using TK-3, we investigated whether the tissue level of U-PA changed when the tumor proliferated locally. The mice were divided into three groups: mice of group A, B and C were sacrificed at 4, 5 and 6 weeks after tumor inoculation, respectively. The tissue level of U-PA was 0.78 +/- 0.183 ng/mg protein in group A, 0.95 +/- 0.189 in group B and 1.13 +/- 0.311 in group C. The values of groups B and C increased significantly compared with those of group A, and the tumor weight in each group showed a similar increase. The level of plasminogen activator inhibitor type 2 also increased (0.14 +/- 0.078 ng/mg protein in group A, 0.17 +/- 0.096 in group B, 0.24 +/- 0.172 in group C). On the other hand, the tissue level of CEA did not change significantly (78 micrograms/g tissue in group A, 88 in group B, 76 tissue in group C), and no correlation was observed between the tissue levels of U-PA and CEA. These results suggest that U-PA plays an important role not only in metastasis, but also in local tumor proliferation, and that its biological action in the autocrine system is independent of CEA.
Animals↗
The physiology of the fibrinolytic system.
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Aging↗
Impaired fibrinolysis in hypertension and obesity due to high plasminogen activator inhibitor-1 level in plasma.
In order to elucidate the influence of the risk factors of coronary heart disease on the fibrinolytic activity, relationships between blood pressure, body mass index (BMI), plasma lipoprotein (a) (Lp(a)) level and the plasma levels of tissue plasminogen activator (tPA) and plasminogen activator inhibitor-1 (PAI-1) were analyzed in the subjects with mild hypertension. Systolic blood pressure showed a positive correlation with total PAI-1 and free PAI-1. Diastolic blood pressure showed no correlation with these proteins involved in the fibrinolytic system. BMI had a positive correlation with total PAI-1, free PAI-1 and euglobulin clot lysis time (ECLT). Plasma Lp(a) level showed correlation with neither blood pressure nor fibrinolytic parameters, but it showed weak negative correlation with body mass index (BMI). These results suggest that high blood pressure and obesity tend to increase free PAI-1 which reduces fibrinolytic activity. Lp(a), however, seems not to influence directly the fibrinolytic system but may work to decrease fibrinolytic activity only in conjunction with other risk factors. The effects of daily drinking of alcohol and smoking on the fibrinolytic system were also investigated in the present study and we obtained the results that habitual drinking increased plasma levels of both tPA and PAI-1 whereas smoking did not affect fibrinolytic activity. These results suggest that risk factors for coronary heart disease such as hypertension and obesity are closely related to the impaired fibrinolysis.
Adult↗