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Biomedical subjects

Y Hara

Publications and source records attributed to Y Hara.

At least 109 records · Page 6Linked to original sources
PubMed2000

Postnatal development of orexin/hypocretin in rats.

We examined developmental changes of orexins/hypocretins and their receptors (OX1R and OX2R) in the rat hypothalamus from postnatal day 0 to 10 weeks, using in situ hybridization histochemistry for the prepro-orexin, OX1R and OX2R mRNAs and immunohistochemistry for orexin-A and orexin-B. The prepro-orexin mRNA was weakly detected in the lateral hypothalamic area (LHA) from days 0 to 15. Orexin-A- and -B-like immunopositive cells and fibers were not detected from days 0 to 10, but they were observed after day 15. The prepro-orexin mRNA in the LHA markedly increased between days 15 and 20. The OX1R mRNA was detected in the ventromedial hypothalamic area (VMH) at day 0. The OX2R mRNA was not detected in the paraventricular nucleus (PVN) at days 0 and 1, but weakly observed on day 5. The OX1R mRNA in the VMH and OX2R mRNA in the PVN gradually increased throughout the postnatal period. Next, we examined the effects of milk deprivation and intraperitoneal (i.p.) administration of leptin on the hypothalamic prepro-orexin mRNA in pups. Although 24-h milk deprivation did not affect the level of the prepro-orexin mRNA at days 5 and 10, i.p. administration of leptin from days 0 to 3 caused a significant increase in the prepro-orexin mRNA on days 5 and 10. These results suggest that the development of orexins may be associated with developmental changes such as increase of leptin, weaning, feeding and sleep/wakefulness states.

Animals↗
PubMed2000

Spontaneous single-channel activity of neuronal TRP5 channel recombinantly expressed in HEK293 cells.

Mammalian homologues of the Drosophila transient receptor potential (trp) protein (TRP) form Ca(2+) permeable cation channels activated in response to stimulation of G-protein-coupled receptors. Establishing biophysical characteristics of basal TRP activity is of great importance in understanding modulatory processes, which underlie enhancement of TRP activity via receptor stimulation. We have examined spontaneous activity of the TRP5 channel recombinantly expressed in human embryonic kidney cells, using the conventional whole-cell mode of the patch-clamp technique in a low-Ca(2+) external solution. The unitary Na(+) conductance of the TRP5 channel was linear, being 47.6 pS. By contrast, the open probability of the TRP5 channel showed a voltage-dependent decrease below -50 mV. These biophysical properties are important hallmarks in distinguishing the TRP5 channel in native neuronal preparations, whose spontaneous activity may contribute to control of resting membrane potentials and generation of action potentials.

Calcium Channels↗
PubMed2000

Simultaneous analysis of tea catechins, caffeine, gallic acid, theanine and ascorbic acid by micellar electrokinetic capillary chromatography.

A micellar electrokinetic capillary chromatography (MEKC) method for the simultaneous analysis of five tea catechins, theanine, caffeine, gallic acid and ascorbic acid has been developed. The catechins are (-)-epicatechin, (+)-catechin, (-)-epigallocatechin, (-)-epicatechin gallate and (-)-epigallocatechin gallate. p-Nitrophenol serves as both reference and internal standard. All the components are separated within 13 min with a 57 cm uncoated fused-silica column. On-column detection was carried out at 200 nm. This method has been used to measure these compounds in fresh tea leaves and tea liquor. The limit of detection for all analytes ranged from 1 to 20 microg/ml.

Ascorbic Acid↗
PubMed2000

Purification and structural determination of SCB1, a gamma-butyrolactone that elicits antibiotic production in Streptomyces coelicolor A3(2).

Early stationary phase culture supernatants of Streptomyces coelicolor A3(2) contained at least four small diffusible signaling molecules that could elicit precocious antibiotic synthesis in the producing strain. The compounds were not detected in exponentially growing cultures. One of these compounds, SCB1, was purified to homogeneity and shown to be a gamma-butyrolactone of structure (2R, 3R,1'R)-2-(1'-hydroxy-6-methylheptyl)-3-hydroxymethylbutanolide . Bioassays of chemically synthesized SCB1, and of its purified stereoisomers, suggest that SCB1 acts in a highly specific manner to elicit the production of both actinorhodin and undecylprodigiosin, the two pigmented antibiotics made by S. coelicolor.

4-Butyrolactone↗
PubMed2000

The B cell-restricted adaptor BASH is required for normal development and antigen receptor-mediated activation of B cells.

B cell antigen receptor signals development, activation, proliferation, or apoptosis of B cells depending on their condition, and its proper signaling is critical for activation and homeostasis of the immune system. The B cell-restricted adaptor protein BASH (also termed BLNK/SLP-65) is rapidly phosphorylated by the tyrosine kinase Syk after BCR ligation and binds to various signaling proteins. BASH structurally resembles SLP-76, which is essential for T cell development and T cell receptor signaling. To evaluate the role for BASH in B cell development and function in vivo, we disrupted BASH alleles in embryonic stem cells by means of homologous recombination and used these cells to complement lymphocyte-incompetent blastocysts from RAG2-deficient mice. In the resultant chimeric mice, T cell development was apparently normal, but B cell development was impaired, and a normally rare population of large preB cells expressing preB cell receptor dominated in the bone marrow in place of small preB cells, although they were mostly noncycling. In addition, the mature B cell populations in the periphery and the bone marrow profoundly decreased in size, as did B-1 cells in the peritoneal cavity, and serum Ig was severely reduced. The BASH-deficient B cells scarcely proliferated or up-regulated B7-2 in response to BCR ligation and poorly proliferated upon CD40 ligation or lipopolysaccharide stimulation. This phenotype indicates that BASH is critical for preB cell receptor signaling inducing proliferation of large preB cells and the following differentiation, for peripheral B cell maturation, and for BCR signaling inducing activation/proliferation of B cells.

Adaptor Proteins, Signal Transducing↗
PubMed2000

Increase of urocortin-like immunoreactivity in the supraoptic nucleus of Dahl rats given a high salt diet.

Urocortin-like immunoreactivity (Ucn-LI) in the supraoptic nucleus (SON) of Dahl rats was examined. Dahl salt-sensitive (S) rats fed with a high salt diet developed hypertension. Numbers of Ucn-LI neurons in the SON in Dahl S on a high salt diet were markedly increased, compared with those in Dahl salt-resistant (R) rats on the same. Sporadic Ucn-LI neurons were found in the SON of both Dahl S and R on a normal diet. Numbers of Ucn-LI neurons in the SON of spontaneously hypertensive rat (SHR) and stroke-prone SHR, genetic models of hypertension, and control rats (Sprague-Dawley and Wistar-Kyoto) were similar. These results suggest that Ucn in the SON is associated with salt loading-induced hypertension rather than spontaneous hypertension.

Animals↗
PubMed2000

Expression of estrogen receptor alpha exon 5 and 7 deletion variant in human breast cancers.

BACKGROUND: An exon deletion variant of estrogen receptor (ER) mRNA has been reported, as one of the possible mechanisms of loss of ER function. METHODS: We examined the expression of exons 3, 5, and 7 in ER alpha mRNA and the frequency of exon deletion variant expression in 64 cases of human breast cancers and in 8 non-cancerous breast tissues using reverse transcriptase polymerase chain reaction (RT-PCR). RESULTS: Approximately the same amount of wild-type (wt) mRNA was detected in all the non-cancerous breast tissues. In cancers, expression of wild-type exon 3 (w3), exon 5 (w5), and exon 7 (w7) was detected in 93.5%, 93.5%, and 91.3% of ER alpha protein (pER) positive cases, respectively, and 27.8%, 38.9%, and 44.4% in negative cases, respectively (p < 0.0001, p = 0.0035, and p = 0.0002). Although the variants for exon 5 (d5) and 7 (d7) were detected in both non-cancerous and cancerous tissues respectively, the variant for exon 3 was not detected at all. Comparatively, the ratio of d5/w5 was significantly higher in pER positive and progesterone receptor protein (pPgR) negative cases. CONCLUSIONS: We suspect that the exon 5 deletion does not work as a dominant positive.

Breast↗
PubMed2000

Sandhoff disease in Cyprus: population screening by biochemical and DNA analysis indicates a high frequency of carriers in the Maronite community.

In the last 15 years, four patients with the infantile form of Sandhoff disease were diagnosed in four different families in Cyprus (population 703,000, birth rate 1.7%). Three of these cases came from the Christian Maronite community (less than 1% of the population) and one from the Greek community (84% of the population). This relatively large number of patients prompted us to initiate an epidemiological study in order to establish the frequency of the mutant allele in Cyprus. Carrier detection was initially based on the measurement of beta-hexosaminidase A and B in both leucocytes and serum. Using the enzyme test, 35 carriers were identified among 244 random Maronite samples and 15 among 28 Maronites with a family history of Sandhoff disease, but only one carrier was found out of 115 random samples from the Greek community. In parallel to the biochemical screening, DNA studies were undertaken in one of the three Maronite patients and in a Greek carrier related to the Greek patient. These studies resulted in the identification of two novel mutations, a deletion of A at nt76 and a G to C transversion at position 5 of the 5'-splice site of intron 8, which have been published. We subsequently screened the carriers detected in the biochemical study for these two mutations using PCR-based tests. Of 50 Maronite carriers examined, 42 were found to have the nt76 deletion. Eight Maronite samples, designated carriers from the biochemical results, were negative for both mutations. It is possible that these individuals were incorrectly classified as carriers since their enzyme values are equivocal, although the presence of another mutation has not been excluded. Two Greek Cypriot carriers and two obligate Lebanese carriers were negative for both mutations. We conclude that there is a high frequency of Sandhoff disease carriers in the Maronite community of Cyprus, approximately 1 in 7, and that a single mutation predominates in this population.

Cyprus↗
PubMed2000

Adenocarcinoma of the ileum producing carbohydrate antigen 19-9: report of a case.

We report herein the case of an 81-year-old woman found to have small intestinal carcinoma producing carbohydrate antigen (CA)19-9, in whom recurrence on the abdominal wall was strongly suspected 4 months after resection. She presented to our hospital with acute abdominal pain with severe anemia. Marked serum elevation of CA19-9 to 164.8 U/ml suggested a progression to malignancy. A fluorography using an ileus tube revealed an abnormal mucosal pattern. An exploratory laparotomy showed an incomplete annular constrictive Borrmann type 2 tumor, located approximately 190 cm from Treitz's ligament, without any signs of peritoneal or hepatic metastases. Histological examination confirmed a diagnosis of papillotubular adenocarcinoma without metastases of the regional lymph nodes. CA19-9 antigenicity was detected in the cytoplasm and on the surface of the cancer cells, using the monoclonal CA19-9 antibody, NS19-9. In this report, we demonstrate the CA19-9 productivity and distribution of the cancer tissues in relation to their prognosis.

Adenocarcinoma, Papillary↗
PubMed2000

Analysis of the addition products of alpha-tocopherol with phosphatidylcholine-peroxyl radicals by high-performance liquid chromatography with chemiluminescent detection.

A chemiluminescence-based high-performance liquid chromatographic method was developed for the analysis of the addition products of alpha-tocopherol with phosphatidylcholine-peroxyl radicals (TOO-PC). The TOO-PC eluted from a reversed-phase column was reacted with a chemiluminescent reagent consisting of a Cypridina luciferin analog and a lipid-soluble iron chelate in acidic methanol at 50 degrees C, and the generated chemiluminescence was monitored. The detection limit for TOO-PC by this method was about 1 pmol. This method was applied to the detection of TOO-PC in the peroxidized membranes prepared from rabbit erythrocyte ghosts. When the erythrocyte ghosts were peroxidized by the addition of a water-soluble free radical initiator, a peak corresponding to TOO-PC was detected on the chromatogram with chemiluminescent detection. The amount of TOO-PC in the erythrocyte membranes increased with the depletion of endogenous alpha-tocopherol. The results indicate that this method proved useful for the detection of the TOO-PC formed by the peroxyl-radical scavenging reactions of alpha-tocopherol in biological systems.

Animals↗
PubMed2000

The configuration of methyl jasmonate affects paclitaxel and baccatin III production in Taxus cells.

All stereoisomers of methyl jasmonate (MJA) were prepared, and their effects on cell yield and promotion of paclitaxel (Taxol) and baccatin III production investigated in cell suspension cultures of Taxus media. (3R,7S)-MJA showed the strongest cell growth inhibition, followed by (3R,7R)-MJA. In contrast, (3S,7R)- and (3S,7S)-MJA had very low inhibitory effects, indicating that this inhibition depends largely on the (3R)-configuration. In terms of the promotion of paclitaxel and baccatin III production, (3R,7R)-MJA had the highest activity. Although it showed considerable activity at low concentration, at higher concentrations the activity was decreased due to strong inhibition of cell growth. Interestingly, paclitaxel and baccatin III contents increased even at a high (3S,7R)-MJA concentration, whereas the other isomers had the opposite effects. These findings are interpreted to suggest that the optimum configuration is (3R,7R), the (3R)-configuration not being indispensable, and that the (7R)-configuration is suitable for the promotion of paclitaxel and baccatin III production.

Acetates↗
PubMed2000

Antioxidative activity of green tea treated with radical initiator 2, 2'-azobis(2-amidinopropane) dihydrochloride.

This study investigated the antioxidative activity of green tea extract, and a green tea tannin mixture and its components, under conditions of radical generation using the hydrophilic azo compound, 2,2'-azobis(2-amidinopropane) dihydrochloride (AAPH) to generate peroxyl radicals at a constant and measurable rate in the cultured renal epithelial cell line, LLC-PK(1), which is susceptible to oxidative damage. Treatment with AAPH decreased cell viability and increased the formation of thiobarbituric acid-reactive substances. However, green tea extract, and the tannin mixture and its components, comprising (-)-epigallocatechin 3-O-gallate (EGCg), (-)-gallocatechin 3-O-gallate (GCg), (-)-epicatechin 3-O-gallate (ECg), (-)-epigallocatechin (EGC), (+)-gallocatechin (GC), (-)-epicatechin (EC), and (+)-catechin (C), showed protective activity against AAPH-induced cellular damage. The tannin mixture and its components exhibited higher antioxidative activity than the green tea extract. Furthermore, EGCg and GCg had higher activity than EGC and GC, respectively. In particular, EGCg exerted the most significant cellular protective activity against AAPH. These results indicate that green tea tannin may inhibit cellular loss and lipid peroxidation resulting from the peroxyl radical generated by AAPH, and that the chemical structure of tannin is also involved in the activity, suggesting that the O-dihydroxy structure in the B ring and the galloyl groups are important determinants for radical scavenging and antioxidative potential.

Amidines↗
PubMed2000

Identification of biliary metabolites of (-)-epigallocatechin gallate in rats.

After oral administration of (-)-epigallocatechin gallate (EGCg) to rats, its biliary metabolites were examined. Although a large part of the biliary metabolites was found to exist in conjugated forms, it was difficult to separate the conjugated forms. Thus the free form of biliary metabolites was prepared by beta-glucuronidase/sulfatase treatment and was purified by HPLC. Six compounds purified were subjected to FABeta-MS and NMR analyses. The six metabolites thus obtained were shown to be EGCg, 3'-O-methyl-EGCg, 4'-O-methyl-EGCg, 3' '-O-methyl-EGCg, 4' '-O-methyl-EGCg, and 4',4' '-di-O-methyl-EGCg, respectively. The six EGCg metabolites and their conjugates excreted during a 4-h period were estimated to be roughly 0.1% and 3.3% of the administered EGCg, respectively. In addition, 4' '-O-methyl-EGCg and 4',4' '-di-O-methyl-EGCg were estimated to exist only in the sulfate form, but the other four metabolites existed in both glucuronide (and/or sulfoglucuronide) and sulfate forms.

Animals↗
PubMed2000

Green tea polyphenols inhibit the sodium-dependent glucose transporter of intestinal epithelial cells by a competitive mechanism.

Intestinal glucose uptake is mainly performed by the sodium-dependent glucose transporter, SGLT1. The transport activity of SGLT1 was markedly inhibited by green tea polyphenols, this inhibitory activity being most pronounced in polyphenols having galloyl residues such as epicatechin gallate (ECg) and epigallocatechin gallate (EGCg). Experiments using brush-border membrane vesicles obtained from the rabbit small intestine demonstrated that ECg inhibited SGLT1 in a competitive manner, although ECg itself was not transported via SGLT1. The present results suggest that tea polyphenols such as ECg interact with SGLT1 as antagonist-like molecules, possibly playing a role in controlling the dietary glucose uptake in the intestinal tract.

Animals↗
PubMed2000

Inhibition of radical reaction of apolipoprotein B-100 and alpha-tocopherol in human plasma by green tea catechins.

(-)-epicatechin (EC), (-)-epigallocatechin (EGC), (-)-epicatechin gallate (ECg), (-)-epigallocatechin gallate (EGCg), and Trolox inhibited the decreases of apolipoprotein B-100 (apoB) and alpha-tocopherol in a radical reaction of human plasma initiated by Cu(2+). The concentrations of EC, EGC, ECg, EGCg, and Trolox for 50% inhibition (IC50) of apoB fragmentation were 39.1, 42.2, 14.6, 21.3, and 36.2 microM, respectively. Similar IC50 values were observed for alpha-tocopherol consumption, indicating the close relationship between apoB fragmentation and alpha-tocopherol consumption. These results demonstrate that tea catechins serve as an effective antioxidant in plasma and that the gallate group has a strong antioxidative activity.

Antioxidants↗
PubMed2000

Biosynthesis of beta1,4- and beta1,beta1-galactopyranosyl xylopyranosides in the mammary gland of lactating cow.

Lactose is a principal carbohydrate in nearly all species of mammalian milk. In order to examine the acceptor substrate specificity of lactose synthase in vivo, D-xylose as an acceptor substrate was injected into the jugular vein of a Holstein cow during lactation, then a milk sample obtained by milking. beta1, beta1-galactopyranosyl xylopyranoside, a nonreducing disaccharide, was separated from the bovine milk sample after elimination of reducing sugars, identified by fast-atom bombardment (FAB)-MS and 1H-NMR analysis. A mixture of beta1,beta1- and beta1, 4-galactopyranosyl xylopyranoside fractions was also obtained by thin layer chromatography from the milk sample and elucidated by electrospray ionization (ESI)-MS and 1H NMR analysis. Comparison of the integrated intensity of the products shows that the beta1,beta1 and beta1,4 isomers are present in a ratio of 1.0 : 1.4, suggesting that D-xylose, transported from capillary blood across the plasma membrane of the mammary gland, was recognized by lactose synthase in its normal and reverse orientation owing to high symmetry of its structure. While the beta1,4-isomer is known as a fragment of the linkage region between the protein and the polysaccharide chain of proteoglycans, the beta1,beta1-isomer has not been identified in vivo. Here, we demonstrate that galactosylation of D-xylose transported from the capillary blood can occur by lactose synthase catalysis in the mammary gland while the usual galactosylation of D-glucose proceeds. In addition, these results suggest that the possibility of endogenous occurrence of the beta,beta-trehalose type disaccharide in the mammary gland of lactating mammals may not be ruled out.

Animals↗
PubMed2000

Preperitoneal fat thickness in childhood obesity: association with serum insulin concentration.

BACKGROUND: The clinical significance of visceral fat accumulation in childhood obesity has remained controversial. METHODS: In 35 subjects (18 boys and 17 girls) aged 9-15 years, visceral and subcutaneous fat were estimated by measuring the maximum thickness of preperitoneal fat (Pmax) and the minimum thickness of subcutaneous fat (Smin) in the upper median abdomen, respectively, by ultrasonography. The associations with fasting serum insulin levels converted to logarithm (log(IRI)) were studied for six obesity-related indices, including the obesity index, body fat percentage, waist-to-hip circumference ratio (WHR), Pmax, Smin and the ratio of Pmax to Smin (abdominal wall fat index, AFI). RESULTS: In girls, the log(IRI) was positively correlated with the Pmax, obesity index, body fat percentage and Smin. In contrast, the log(IRI) was positively correlated only with the Pmax and Smin in boys. There were no significant correlations between the log(IRI) and the WHR or AFI in either sex. The Pmax revealed the highest correlation with the log(IRI) in both boys (r = 0.703, P < 0.001) and girls (r = 0.921, P < 0.0001). CONCLUSION: These results indicate the important contribution of visceral fat accumulation to the development of hyperinsulinemia, regardless of sex. Accordingly, it is suggested that evaluating Pmax by ultrasonography may be the most sensitive and reliable method of predicting insulin resistance-associated metabolic derangements in children, compared with other conventional indices of obesity.

Adipose Tissue↗