Biomedical subjects

H Hirano

Publications and source records attributed to H Hirano.

At least 181 records · Page 10Linked to original sources

PubMed1996

Experimental liver fibrosis induced in rats receiving high doses of alcohol and alternating between regular and vitamin-depleted diets.

Liver fibrosis was induced in rats by simulating human alcoholic eating and drinking patterns. Alcohol addiction was established by gradually increasing the ethanol concentration in the drinking water; salts were added at the terminal stage. The hepatocytes of rats receiving alcohol concentrations exceeding 50% (v/v) (similar to vodka) exhibited alcoholic hyaline (Mallory bodies). Alcoholic liver fibrosis was induced by alternating between regular and autoclaved (vitamin-depleted) diets, simulating the irregular eating habits of human alcoholics. In the livers of rats receiving 70% (v/v) ethanol (comparable to absinthe) with 25% saline and fed the alternating diets, pericellular fibrosis was induced. No significant difference in calorie intake between control and alcohol rats was detected except when rats underwent drinking bouts (heavy drinking phase). This indicates that neither a high-fat diet nor a choline-depleted diet is necessary to induce the alcoholic fibrosis seen in human alcoholics.

Animals↗

PubMed1996

Enhanced glomerular profilin gene and protein expression in experimental mesangial proliferative glomerulonephritis.

Profilin is a cytoplasmic protein that binds to actin monomer and regulates actin polymerization. In a number of experimental and human glomerular diseases, the mesangial cell expresses alpha-smooth muscle actin and undergoes a phenotypic change to myofibroblast. We used a rat model of mesangial proliferative nephritis induced with antibody to the Thy 1 antigen present on mesangial cells to investigate whether profilin is upregulated. We amplified and sequenced rat profilin cDNA by the reverse-transcribed-polymerase chain reaction (RT-PCR). The nucleotide and amino acid sequences were highly conserved across the mammalian profilins. We raised affinity purified antibody to rat profilin in rabbits immunized with a synthetic profilin peptide (EFTMDLRTKS). At 7 days after disease induction, enhanced expression in both profilin mRNA and protein was demonstrated in the isolated glomeruli by RT-PCR and Western blot analysis. These results suggest that profilin may be involved in the pathogenesis of glomerulonephritis by reorganizing actin cytoskeleton.

Amino Acid Sequence↗

PubMed1996

Rom1p and Rom2p are GDP/GTP exchange proteins (GEPs) for the Rho1p small GTP binding protein in Saccharomyces cerevisiae.

The RHO1 gene encodes a homolog of the mammalian RhoA small GTP binding protein in the yeast Saccharomyces cerevisiae. Rho1p is localized at the growth site and is required for bud formation. Multicopy suppressors of a temperature-sensitive, dominant negative mutant allele of RHO1, RHO1(G22S, D125N), were isolated and named ROM (RHO1 multicopy suppressor). Rom1p and Rom2p were found to contain a DH (Dbl homologous) domain and a PH (pleckstrin homologous) domain, both of which are conserved among the GDP/GTP exchange proteins (GEPs) for the Rho family small GTP binding proteins. Disruption of ROM2 resulted in a temperature-sensitive growth phenotype, whereas disruption of both ROM1 and ROM2 resulted in lethality. The phenotypes of deltarom1deltarom2 cells were similar to those of deltarho1 cells, including growth arrest with a small bud and cell lysis. Moreover, the temperature-sensitive growth phenotype of deltarom2 was suppressed by overexpression of RHO1 or RHO2, but not of CDC42. The glutathione-S-transferase (GST) fusion protein containing the DH domain of Rom2p showed the lipid-modified Rholp-specific GDP/GTP exchange activity which was sensitive to Rho GDP dissociation inhibitor. These results indicate that Rom1p and Rom2p are GEPs that activate Rho1p in S.cerevisiae.

Amino Acid Sequence↗

PubMed1996

Rice gibberellin-binding phosphoprotein structurally related to ribulose-1,5-bisphosphate carboxylase/oxygenase activase.

A gibberellin A (GA)-binding protein was identified from rice (Oryza sativa L.) leaves by a ligand-binding assay. The dissociation constant of GA-binding protein and GA complex was about 100 nM. This protein has a relative molecular mass of 47 000 and an isoelectric point of 5.1. The partial amino acid sequence of the protein was determined for 54 residues from both the N-terminal and internal regions. A sequence homology search indicated that the amino acid sequence of GA-binding protein was homologous to that of the ribulose-1,5-biphosphate carboxylase/oxygenase activase from barley, Arabidopsis, spinach and Chlamydomonas. The GA-binding protein was immunologically detected in two polypeptides in the protein extract from leaves. The GA-binding protein identified was phosphorylated with Ca2+, Mg2+ and ATP in the leaf protein extracts of rice grown in the presence of exogenous GA.

Amino Acid Sequence↗

PubMed1996

Self-incompatibility (S) alleles of the Rosaceae encode members of a distinct class of the T2/S ribonuclease superfamily.

Stylar ribonucleases (RNases) are associated with gametophytic self-incompatibility in two plant families, the Solanaceae and the Rosaceae. The self-incompatibility-associated RNases (S-RNases) of both the Solanaceae and the Rosaceae were recently reported to belong to the T2 RNase gene family, based on the presence of two well-conserved sequence motifs. Here, the cloning and characterization of S-RNase genes from two species of Rosaceae, apple (Malus x domestica) and Japanese pear (Pyrus serotina) is described and these sequences are compared with those of other T2-type RNases. The S-RNases of apple specifically accumulated in styles following maturation of the flower bud. Two cDNA clones for S-RNases from apple, and PCR clones encoding a further two apple S-RNases as well as two Japanese pear S-RNases were isolated and sequenced. The deduced amino acid sequences of the rosaceous S-RNases contained two conserved regions characteristic of the T2/S-type RNases. The sequences showed a high degree of diversity, with similarities ranging from 60.4% to 69.2%. Interestingly, some interspecific sequence similarities were higher than those within a species, possibly indicating that diversification of S-RNase alleles predated speciation in the Rosaceae. A phylogenetic tree of members of the T2/S-RNase superfamily in plants was obtained. The rosaceous S-RNases formed a new lineage in the tree that was distinct from those of the solanaceous S-RNases and the S-like RNases. The findings suggested that self-incompatibility mechanisms in Rosaceae and Solanaceae are similar but arose independently in the course of evolution.

Alleles↗

PubMed1996

Immunohistochemical localization of androgen receptor in mouse testicular germ cells during fetal and postnatal development.

BACKGROUND: Determination of the cellular distribution of the androgen receptor (AR) in testicular cells is necessary for understanding the mode of AR action in the testis. We here investigated immunohistochemically the localization of AR by use of anti-human AR polyclonal antibody NH27, with special reference to the AR in germ cells in the developing mouse testis. METHODS: ICR mouse testes taken from day 14 post coitum (p.c.) to day 56 post partum (p.p) were used for AR immunohistochemistry by the routine immunoperoxidase method at the light microscopic level and the pre-embedding method at the electron microscopic level. RESULTS: On day 14 p.c., AR immunoreactivity was present in nuclei of prospermatogonia but not in those of Sertoli cells or interstitial cells. On day 14 p.p., the AR was detected in the nuclei of spermatogonia, Sertoli cells, and myoid cells. AR immunoreactivity in nuclei of Leydig cells appeared on day 21 p.p. In the mature mouse testis, the AR was present in the nuclei of spermatogonia, Sertoli cells, myoid cells, and Leydig cells. CONCLUSIONS: AR was present both in germ cells and in somatic cells during fetal and postnatal development of the mouse testis. In the fetal testis, AR was localized exclusively in prospermatogonia and spermatogonia, suggesting that androgen may act directly on germ cells during prespermatogenesis and the early stage of spermatogenesis. Based on the fact that AR is expressed in Sertoli cells, myoid cells, and Leydig cells around the onset of spermatogenesis, the regulation of AR expression in the germ cells seems to be different from that in the somatic cells. Furthermore, our present data suggest the ultrastructural localization in nuclei of mouse testicular cells is similar to that of some other steroid receptors, both in germ cells and somatic cells.

Age Factors↗

PubMed1996

Inhibition by tunicamycin of mucin synthesis, not morphological changes, in epidermis during retinol-induced mucous metaplasia of chick embryonic cultured skin.

BACKGROUND: Our previous studies have shown that epidermal mucous metaplasia of chick embryonic skin can be induced by culture in medium containing 20 microM retinol for only 8 hr and then in a chemically defined medium without retinol for 2 days and that retinol primarily affects the dermal cells, which then transform the epithelial cells into mucus-secreting cells. METHODS: Tarsometatarsal skin of 13-day-old chick embryo was cultured with 20 microM retinol for 1 day and then without the vitamin but with 0.1 microgram/ml tunicamycin for 5 days. Effect of tunicamycin on epidermal mucous metaplasia was studied biochemically and morphologically. RESULTS: Tunicamycin, which prevents the formation of N-glycans and inhibits maturation or morphological organization of various epithelial cells, irreversibly inhibited the synthesis of sulfated glycoproteins (O-glycans, mucin) in the epidermis only when applied to retinol-pretreated skin. Microvilli on the surface of the cells were well developed, but mucous granules surrounded by a limiting membrane were not observed in the upper cell layer of the epidermis, and many vesicles without electron-dense materials (mucin) and dilated rough endoplasmic reticulum were seen in the intermediate cell layers of the epidermis. When recombinants of 13-day-old normal epidermis and cultured dermis, which had been treated with retinol for 24 hr and with only tunicamycin for 2 days, were cultured without the antibiotic for 5 days, epidermal mucous metaplasia was induced. CONCLUSION: These results suggest that tunicamycin did not prevent morphological changes induced by retinol but inhibited mucin synthesis by a direct action on the epidermis of retinol-pretreated skin. Because in some cell-line mucin precursors contain high mannose N-linked oligosaccharides side chains, tunicamycin may have inhibited mucin synthesis. Interaction between epidermal basal cells and retinol-pretreated dermal fibroblasts is prerequisite for epidermal mucous metaplasia. Thus, the present study suggests that N-linked protein glycosylation is not required for this interaction.

Animals↗

PubMed1996

Adherence of synovial cells on EDA-containing fibronectin.

OBJECTIVE: To investigate the role of EDA-containing fibronectin (EDA+ FN), a splice variant of FN detectable in association with cellular transformation, in the adherence of synovial cells (SC) on rheumatoid cartilage surface. METHODS: The number of SC adherent on cartilage slices or on culture plates containing either EDA+ FN or plasma FN (pFN) was enumerated under a phase-contrast microscope. The portion of the FN molecule responsible for adherence of SC onto EDA+ FN was investigated by inhibition studies using antibodies or peptide fragments. RESULTS: SC adhered more strongly on the surfaces containing EDA+ FN than on those containing pFN (P < 0.01). When monoclonal antibodies against the EDA or the carboxyl-terminal heparin-binding (Hep2) domains were used, adhesion of SC onto EDA+ FN was reduced to a level comparable with that onto pFN. FN fragments containing Hep2 or heparan sulfate inhibited the adhesion of SC onto EDA+ FN. Treatment of SC with heparitinase, but not heparinase, reduced the adhesion of SC onto EDA+ FN. CONCLUSION: EDA+ FN enhances adherence of SC on the matrix via the Hep2 region of EDA+ FN.

Alternative Splicing↗

PubMed1996

Extracellular matrices in peritendinous connective tissue after surgical injury to the chicken flexor tendon.

Gene expression of fibronectin, type I collagen and type III collagen and the presence of fibronectin in peritendinous connective tissue were investigated, following flexor tendon injury, using slot-blot hybridization and immunohistochemistry. Fibrous adhesion around the chicken flexor tendon was induced by a partial laceration using a surgical knife. Total RNA was purified from the peritendinous connective tissue surrounding a tendon by combining the guanidine thiocyanate procedure and isoelectric precipitation. Slot-blot hybridization revealed that gene expression of fibronectin and collagens type I and III increased gradually after tendon injury, and levels were maintained high throughout the experimental period of 16 weeks. Indirect immunofluorescence revealed fibronectin to be preferentially located in peritendinous connective tissue around the injured tendon. The prolonged increase of fibronectin in the peritendinous connective tissue may relate to the development of histological adhesion after flexor tendon injury.

Animals↗

PubMed1996

Effects of ethanol on meal-stimulated secretion of pancreatic polypeptide and cholecystokinin: comparison of healthy volunteers, heavy drinkers, and patients with chronic pancreatitis.

Tiscornia and Dreiling (Physiopathogenic Hypothesis of Alcoholic Pancreatitis: Supranormal Ecbolic Stimulation of the "Pancreon" Units Secondary to the Loss of the Negative Component of Pancreas Innervation. Pancreas 1987;2:604-612.) proposed that hypertonicity of intrapancreatic cholinergic neurons provoked by chronic alcoholism may contribute to the pathogenesis of chronic pancreatitis (CP). In the present study, the validity of this hypothesis was investigated in humans by studying the effects of atropine, cisapride, and ethanol on the meal-stimulated secretion of pancreatic polypeptide (PP) and cholecystokinin (CCK) in healthy volunteers, heavy drinkers, and CP patients. In healthy volunteers, the early phase PP response (0-40 min) to a test meal was completely blocked by atropine, whereas it was augmented by cisapride, an enhancer of acetylcholine release from cholinergic nerves. The early phase PP response to a test meal was inhibited by ethanol in healthy volunteers, whereas, in heavy drinkers, the response was augmented and the inhibition by ethanol was abrogated. In CP patients, ethanol tended to enhance the early phase PP response. Ethanol did not affect the early phase CCK response to a test meal in any group, but it significantly enhanced the late phase CCK response (40-120 min) in CP patients. These results suggest that: (i) oral ethanol may inhibit the postprandial activation of the cholinergic neural pathway to the pancreas in healthy subjects, (ii) in heavy drinkers, postprandial cholinergic tone may be augmented and become resistant to the inhibition by ethanol, and (iii) the ethanol-induced increase in the postprandial CCK response in CP patients may play some role in the pathophysiology of this disease.

Adult↗

PubMed1996

An immunohistochemical study of the 32-kDa galectin (beta-galactoside-binding lectin) in the nematode Caenorhabditis elegans.

The localization of the 32-kDa galectin (beta-galactoside-binding lectin) of the nematode Caenorhabditis elegans, which is the first lectin to be found in a nematode, was examined immunohistochemically using an anti-lectin antiserum. The lectin was found to be localized most abundantly in the adult cuticle and also in the terminal bulb of the pharynx. However, it was difficult to locate the galectin in larval animals, though immunochemical experiments suggested its presence. These results suggest that one of the fundamental roles of the galectin may be as a component of the durable outer barrier, as in the case of the morphogenesis of chick embryonic skin.

Animals↗

PubMed1996

Detection of alpha 1-adrenoceptor subtypes in human hypertrophied prostate by in situ hybridization.

Adrenergic stimulation induces contraction of hypertrophied prostatic tissue via the alpha 1 adrenoceptor, and the results of pharmacological studies suggested the existence of adrenoceptor subtypes. Recently three subtypes (alpha 1a, alpha 1b, and alpha 1d) were cloned. Using probes for these subtypes, we demonstrated their expression in the tissues of ten cases of benign prostatic hypertrophy, using in situ hybridization. To determine the ratio between these subtypes, an RNase protection assay was also performed in three cases. Expression of the alpha 1a and alpha 1d adrenoceptors was diffuse in the smooth muscles of the interstitium, but was absent in glandular epithelial cells. On the contrary, the alpha 1b adrenoceptor was hardly detectable. The RNase protection assay confirmed the absence of the alpha 1b adrenoceptor, the ratio of alpha 1a and alpha 1d being 4:1. These results supported the idea that the differences in prostatic contractile response to several adrenergic drugs are based on the affinities of these drugs for the different subtypes.

Aged↗

PubMed1996

Changes in cellular proteins of Deinococcus radiodurans following gamma-irradiation.

In order to examine radiation-induced proteins in an extremely radioresistant bacterium, Deinococcus radiodurans R1, changes in cellular proteins after gamma-irradiation were analysed by two-dimensional gel electrophoresis and silver staining. Nine proteins (190, 120, 87,60, 58, 52, 46, 41 and 41 kDa) were increased (or appeared) and more than 13 proteins diminished after gamma-irradiation at 6 kGy. Increase of eight proteins (except for 190-kDa protein) was prevented when the cells were irradiated in the presence of chloramphenicol. Three proteins, 87, 60 and 46 kDa, continued to be synthesized during post-irradiation incubation, and the amounts of these proteins increased with higher doses in a range of 1-12 kGy. Changes in the amount of proteins after irradiation in the R1 strain were compared with those in a moderately radioresistant mutant (rec I) and in a highly radiosensitive mutant (rec30). These three proteins were increased in both R1 and recI, but not in rec 30, suggesting that they are characteristic for radioresistant strains. In addition, from the microsequence analysis, the 46-kDa protein was found to be homologous to the EF-Tu protein of Escherichia coli, whereas the remarkable homologous sequence to the N-terminal of the 60-kDa protein was not found among the known proteins.

Amino Acid Sequence↗

PubMed1996

Down-regulation of S-adenosylhomocysteine hydrolase in the active methyl transfer system in the brain of genetically epileptic El mice.

To study the role of cerebral methylation in epileptogenesis, we investigated the active methyl transfer pathway in the brain of genetically epileptic El mice. We examined S-adenosylhomocysteine (AdoHcy) hydrolase activity (by high performance liquid chromatography), the corresponding mRNA levels (by competitive reverse transcription-polymerase chain reaction), as well as S-adenosylmethionine (AdoMet) and AdoHcy levels in epileptic El and non-epileptic control ddY mice. The level of AdoHcy, a potent feedback inhibitor in the methyl transfer system, was relatively high throughout growth, particularly in 15 week old El mice where it was increased approximately 20% compared to control ddY mice. AdoHcy hydrolase activity in the El mice brain did not increase during growth, which was confirmed by the finding that mRNA synthesis from the hydrolase gene behaved likewise. In contrast, ddY mice exhibited a gradual increase in the mRNA synthesis up to three fold and 20% increase in the enzyme activity at 15 weeks of age compared to those of El mice in which the seizure frequency was 100%. The levels of AdoMet, a versatile methyl donor, did not change throughout growth. We concluded that the down-regulation of AdoHcy hydrolase results in the accumulation of AdoHcy, possibly inducing an unstable state including seizures in the El mouse brain. El mice predisposed to epilepsy may be characterized by disordered feedback regulation of the AdoMet-dependent methyl transfer pathway.

Adenosylhomocysteinase↗

PubMed1996

Technegas ventilation SPECT for evaluating silicosis in comparison with computed tomography.

To compare the subtle pulmonary parenchymal morphologic changes with ventilation function in patients with silicosis, the conventional CT, high resolution CT and technegas ventilation SPECT were performed. In 25 silicotic patients and six controls, the pulmonary ventilation state was evaluated by an index called the coefficient of variation (CV), which expresses the subliminal heterogeneous distribution of technegas in the lungs. The results showed that with silicosis the CV value is significantly higher than that without silicosis. The CV value was proved by multifactorial analysis to independently reflect the extent of the appearance of small scattered interstitial findings such as nodules, septal thickening and bulla, which were typical findings for silicosis. The CV value calculated from the technegas SPECT correlated well with the severity of silicosis. It is considered that the CV value can also express the functional state of the silicotic lung.

Adult↗

PubMed1996

Infantile choriocarcinoma: a case report with MRI, angiography and bone scintigraphy.

Infantile and maternal choriocarcinoma is a very rare disease. We report a case with the characteristic clinical features of infantile choriocarcinoma: developing anemia, hemorrhagic liver tumors, rapid progression to death and maternal choriocarcinoma. Bone scintigraphy showed increased uptake by the liver tumors. In this case there were two possible primary sites: the placenta of this pregnancy and a hydatidiform mole that had been present 2 years previously.

Angiography↗

PubMed1996

Partial purification and characterization of a calcium-dependent protein kinase in rice leaves.

A protein from rice leaves, which was partially purified by sequential chromatography on DE52, MONO-Q and Superose 12, presented calcium-dependent protein kinase (CDPK) activity. This protein kinase phosphorylated the substrate, histone III-S, in a Ca(2+)-dependent manner and the half-maximum concentration of Ca2+ to protein kinase activity (EC50) was 1 microM. This phosphorylation was independent of phosphatidylserine and a phorbol ester. The apparent M(r) of the protein kinase, as determined by phosphorylation in SDS-polyacrylamide gel containing histone III-S, was 45 k. This kinase was found to react differently from other protein kinases, such as protein kinase C from rat brain or CDPK from soybean leaves, owing to the absence of a phospholipid or phorbol ester dependency. CDPK phosphorylated three endogenous proteins as detected by in vitro phosphorylation on two-dimensional PAGE.

Animals↗

PubMed1996

Isolation and primary culture of rat cerebral microvascular endothelial cells for studying drug transport in vitro.

To establish the cerebral microvascular endothelial cell (CMEC) culture system for animals commonly utilized in in vivo studies, we developed a method for isolation and culture of rat CMECs, and the model system was used in preliminary in vitro transport experiments. The isolated rat brains were minced. After an incubation with dispase, a fraction of microvessels was obtained by the dextran gradient. The tissue was filtered and dissociated using collagenase/dispase. After the enzyme treatment, the microvessels were layered onto the top of Percoll gradient and centrifuged for purification. Specific enzyme activities of alkaline phosphatase and gamma-glutamyltransferase in the preparations gradually increased as the isolation process progressed. The isolated cells reached confluence after 5-7 days in culture. The cultured cells had Factor VIII-related antigen and an uptake of acetylated-low density lipoprotein labeled with 1,1'-dioctadecyl-3,3, 3',3'-tetramethyl-indocarbocyamine perchlorate (Dil-Ac-LDL). In the transport studies, thawed cells after several months under -80 degrees C were used. The cultured cells after the freezing preservation also had CMEC characteristics, the same as the cells seeded immediately after isolation. The permeability of [14C]-mannitol, an impermeable marker for blood-brain barrier, was considerably reduced when the cell monolayer was present. The transport of [3H]3-O-methyl-D-glucose was significantly inhibited by the unlabeled compound. Furthermore, 2,4-dinitrophenol, an uncoupler of oxidative phosphorylation, significantly diminished the transport of [3H]L-proline. These results indicated that the cell monolayer obtained in the present study could be applicable to drug transport studies through the blood-brain barrier in vitro.

3-O-Methylglucose↗