Biomedical subjects
Publications and source records attributed to H Yoshida.
Baroreceptor sensitivity and carotid sinus distensibility were lower in patients with angina associated with significant coronary artery disease than in patients with vasospastic angina. Baroreceptor sensitivity was significantly correlated with carotid sinus distensibility in both groups of patients.
Murine T cell receptor (TCR) alpha beta intestinal intraepithelial lymphocytes (IEL), which express the CD8 molecule as a homodimer (CD8 alpha alpha), can be divided into two subsets: those which are CD4+ (CD4+CD8+alpha alpha) and those which are CD4- (CD4-CD8+alpha alpha). Here, we demonstrate that most TCR alpha beta CD4+CD8+alpha alpha IEL and TCR alpha beta CD4-CD8+alpha alpha IEL subsets appear to be of thymus origin, as neonatal thymectomy of BALB/c mice on Day 3 nearly eliminated both subsets. To further support this hypothesis, we demonstrate by grafting the thymus of CBF1 (BALB/c x C57BL/6) mice into nude mice that the thymus is capable of generating both TCR alpha beta CD4-CD8+alpha alpha IEL and TCR alpha beta CD4+CD8+alpha alpha IEL. However, which of the two TCR alpha beta IEL subsets is generated depends largely on the age of the thymus. The thymus from fetal up to 2 weeks of age generates predominantly TCR alpha beta CD4-CD8+alpha alpha IEL, but very scant amounts CD4+CD8+alpha alpha IEL. In contrast, the thymus after 2 weeks of age generates very little TCR alpha beta CD4-CD8+alpha alpha IEL, but generates an abundant amount of TCR alpha beta CD4+CD8+alpha alpha IEL. These results are consistent with the observation in euthymic mice that TCR alpha beta CD4-CD8+alpha alpha IEL precede the appearance of TCR alpha beta CD4+CD8+alpha alpha IEL by several weeks, thus further suggesting that the thymus is the major source of both TCR alpha beta IEL subsets.
The untranslatable, RNA polymerase II-dependent gene (dutA) of Dictyostelium discoideum is induced early in development. However, unlike other early genes, dutA induction was not affected by cAMP pulses and occurred normally in various cAMP-related mutant cells, the results indicating that this induction depended solely on factors other than cAMP. In the knockout strain of the catalytic subunit of protein kinase A, dutA expression was severely blocked and not recovered by cAMP pulses. This demonstrates that even the cAMP-independent gene, dutA, requires protein kinase A for its expression.
Acute promyelocytic leukemia (APL) is associated with a chromosomal translocation t(15;17) and successfully differentiated by all-trans-retinoic acid (ATRA) in vivo as well as in vitro. The PML-retinoic acid receptor alpha (RARA) oncoprotein, which is generated by the translocation, blocks the differentiation, and ATRA is thought to modulate the dominant negative function of PML-RARA. However, the molecular effect of ATRA on PML-RARA is unknown. In this study, we showed by means of immunoblotting that the expression of PML-RARA decreased within 12 h in APL cells treated with ATRA at concentrations greater than 0.1 microM. The decrease of PML-RARA was associated with restoration of the normal subcellular PML localization. PML-RARA transcripts were not down-regulated by ATRA. However, lactacystin, a specific inhibitor of the proteasome, almost completely inhibited the decrease of PML-RARA. These data indicate that the PML-RARA degradation is accelerated by pharmacological concentrations of ATRA, suggesting that ATRA allows APL cells to differentiate by relieving the differentiation block.
Deficiency of plasma platelet-activating factor (PAF) acetylhydrolase is an autosomal recessive syndrome that has been associated with severe asthma in Japanese children. Acquired deficiency has been described in several human diseases usually associated with severe inflammation. PAF acetylhydrolase catalyzes the degradation of PAF and related phospholipids, which have proinflammatory, allergic, and prothrombotic properties. Thus, a deficiency in the degradation of these lipids should increase the susceptibility to inflammatory and allergic disorders. Miwa et al. reported that PAF acetylhydrolase activity is absent in 4% of the Japanese population, which suggests that it could be a common factor in such disorders, but the molecular basis of the defect is unknown. We show that inherited deficiency of PAF acetylhydrolase is the result of a point mutation in exon 9 and that this mutation completely abolishes enzymatic activity. This mutation is the cause of the lack of enzymatic activity as expression in E. coli of a construct harboring the mutation results in an inactive protein. This mutation as a heterozygous trait is present in 27% in the Japanese population. This finding will allow rapid identification of subjects predisposed to severe asthma and other PAF-mediated disorders.
14-3-3 proteins have recently been implicated in the regulation of intracellular signaling pathways via their interaction with several oncogene and protooncogene products. We found recently that 14-3-3 associates with several tyrosine-phosphorylated proteins and phosphatidylinositol 3-kinase (PI3-K) in T cells. We report here the identification of the 120-kDa 14-3-3tau-binding phosphoprotein present in activated T cell lysates as Cbl, a protooncogene product of unknown function which was found recently to be a major protein-tyrosine kinase (PTK) substrate, and to interact with several signaling molecules including PI3-K, in T lymphocytes. The association between 14-3-3tau and Cbl was detected both in vitro and in intact T cells and, in contrast to Raf-1, was markedly increased following T cell activation. The use of truncated 14-3-3tau fusion proteins demonstrated that the 15 C-terminal residues are required for the association between 14-3-3 and three of its target proteins, namely, Cbl, Raf-1, and PI3-K. The findings that 14-3-3tau binds both PI3-K and Cbl, together with recent reports of an association between Cbl and PI3-K, suggest that 14-3-3 dimers play a critical role in signal transduction processes by promoting and coordinating protein-protein interactions of signaling proteins.
To understand the molecular mechanisms whereby normal human salivary gland cells become malignant and escape growth-inhibitory control by transforming growth factor (TGF)-betaI, we examined the effect of TGF-betaI on the proliferation and expression of TGF-beta receptors in cells and the expression of TGF-beta type II receptor (TbetaR-II) mRNA. An SV40-immortalized normal human salivary gland duct cell clone (NS-SV-DC) with no tumorigenic ability, originally obtained via s.c. implantation into nude mice, was partially resistant to the growth-inhibitory effect of TGF-betaI, while a neoplastic human salivary gland duct cell clone (HSGc) with tumorigenic, but not metastatic, potential in nude mice was more resistant to the growth-suppressive effect of TGF-betaI than NS-SV-DC. Metastatic cell clones derived from carcinogen-treated HSGc were completely refractory to the anti-proliferative effect of TGF-betaI. Affinity cross-linking revealed that NS-SV-DC possesses the types I, II (TbetaR-II) and III receptors. However, HSGc and metastatic cell clones lacked expression of detectable levels of the TbetaR-II protein. Moreover, we evaluated TbetaR-II mRNA expression in these cell clones by Northern blot analysis and observed that, although NS-SV-DC expressed a large amount of TbetaR-II mRNA, a small amount of TbetaR-II mRNA was detectable in HSGc. In contrast, no significant bands were detected in metastatic cell clones. Our results, therefore, suggest that one of the possible mechanisms of escape from autocrine or paracrine growth inhibition by TGF-betaI during human salivary gland carcinogenesis involves reduced expression or lack of TbetaR-II.
A method was developed for obtaining a homogeneous silkworm hemolymph protein (peptidoglycan recognition protein, PGRP) which has affinity for peptidoglycan and the ability to trigger the prophenoloxidase cascade upon its binding to peptidoglycan. The purified PGRP had a molecular mass of about 19 kDa and is composed of a single polypeptide with an isoelectric point of 6.5. It bound to peptidoglycan in the absence of divalent cation, whereas its binding to beta1,3-glucan and chitin was not detected. N-Acetyl-D-glucosaminyl-(beta1-4)-N-acetylmuramyl-L-alanyl-D-isogluta mine did not inhibit purified PGRP to bind insoluble peptidoglycan, but fragmented soluble peptidoglycan did. PGRP seemed to require peptidoglycan as a possible ligand to keep its glycan portion consisting of at least two or more of the repeating unit. PGRP did not have any detectable lysozyme activity, and its amino acid composition and amino-terminal sequence of 20 amino acid residues were shown to be different from those of silkworm lysozyme. PGRP seems to be a hitherto unknown protein. In the absence of PGRP, the prophenoloxidase cascade in the plasma fraction of hemolymph could not be triggered by peptidoglycan, indicating that some type of activity, capable of activating the cascade, is generated upon their binding. However, the exact nature of this activity is not yet known. The purified PGRP bound to peptidoglycan did not hydrolyze significantly any of the 26 commercially available peptidyl-7-amino-4-methylcoumarins, substrates for various proteases.
In pancreatic acini, the high affinity cholecystokinin (CCK) receptor agonist, CCK-OPE which utilizes the phospholipase A2 (PLA2)/arachidonic acid (AA) pathway, dose-dependently increased intracellular Ca2+ spike frequency and amplitude. An uncoupler of proton gradients, FCCP, abolished Ca2+ oscillations and amylase secretion induced by CCK-OPE. Furthermore, FCCP or decreasing extravesicular pH inhibited ATP-dependent 45Ca2+ uptake into the endoplasmic reticulum (ER) fraction. On the other hand, cytosolic acidification induced by Na(+)-free medium led to Ca2+ oscillations. Depletion of intracellular ATP by antimycin resulted in an abolition of the response to CCK-OPE. Administration of the K+ ionophore, valinomycin, abolished the action of CCK-OPE. Decreasing K+ concentrations outside the ER vesicles inhibited ATP-dependent 45Ca2+ uptake and AA-induced 45Ca2+ release. Caffeine inhibited the actions of CCK-OPE, whereas ryanodine did not have any effects. These data suggest that IP3-independent Ca2+ oscillations mediated by the PLA2 cascades involve the release of intracellular Ca2+ by AA and reuptake of Ca2+ by an ATP-dependent Ca2+/H+ antiport. Furthermore, the presence of K+ membrane potential gradient across the ER membrane is required for normal Ca2+ oscillations.
PURPOSE: To evaluate the feasibility and efficacy of prophylactic entire hemithorax irradiation (EH) in addition to mediastinal irradiation (MRT) following a complete resection in Stage II-III invasive thymoma. METHODS AND MATERIALS: Forty-three patients with invasive thymoma treated with surgery and radiation therapy between 1978 and 1993 were analyzed retrospectively. All 43 patients underwent a complete surgical resection and were judged to have Masaoka's Stage II-III invasive thymoma. Of these, 23 patients received EH and MRT (EH-MRT) and the remaining 20 received MRT. Of the 23 patients with EH-MRT, 11 were Stage II and 12 Stage III. Of the 20 with MRT, 11 were Stage II and 9 Stage III. In most cases, EH was 15 Gy per 15 fractions over 3 weeks (without lung compensation calculation). In both the EH-MRT and MRT group, the total radiation doses to the mediastinum were similar with a median of 40 Gy. The median follow-up time after surgery was 63 months and no patients were lost to follow-up. RESULTS: Only one of the 23 patients with EH-MRT relapsed. On the other hand, eight of the 20 with MRT relapsed, six of whom died of disease. The pleura was the most common site of failure. At 5 years, the relapse-free rate was 100% for those receiving EH-MRT and 66% for those with MRT (p = 0.03); the overall survival rate was 96% for those with EH-MRT, and 74% for those with MRT (p: not significant). The only significant treatment-related complication was radiation pneumonitis requiring treatment, in one patient who received MRT and three who received EH-MRT, including one death of a 72-year-old man and one 68-year-old woman with severe lung fibrosis. CONCLUSION: Except for elderly patients, EH-MRT following a macroscopically complete resection appears to be safe and feasible, and can reduce intrathoracic relapses.
Tham Khuyen Cave (Lang Son Province, northern Vietnam) is one of the more significant sites to yield fossil vertebrates in east Asia. During the mid-1960s, excavation in a suite of deposits produced important hominoid dental remains of middle Pleistocene age. We undertake more rigorous analyses of these sediments to understand the fluvial dynamics of Pleistocene cave infilling as they determine how skeletal elements accumulate within Tham Khuyen and other east Asian sites. Uranium/thorium series analysis of speleothems brackets the Pleistocene chronology for breaching, infilling, and exhuming the regional paleokarst. Clast analysis indicates sedimentary constituents, including hominoid teeth and cranial fragments accumulated from very short distances and under low fluvial energy. Electron spin resonance analysis of vertebrate tooth enamel and sediments shows that the main fossil-bearing suite (S1-S3) was deposited about 475 thousand years ago. Among the hominoid teeth excavated from S1-S3, some represent Homo erectus and Gigantopithecus blacki. Criteria are defined to differentiate these teeth from more numerous Pongo pygmaeus elements. The dated co-occurrence of Homo erectus and Gigantopithecus blacki at Tham Khuyen helps to establish the long co-existence of these two species throughout east Asia during the Early and Middle Pleistocene.
Between 1986 and 1990, among a female population of about 0.95 million, 1184 women living in Kagoshima Prefecture were diagnosed pathologically as having mammary carcinoma. The number of patients with mammary carcinoma, the crude incidence rate per 100000 women and age-standardized mammary carcinoma incidence rates were 205, 21.29 and 19.42 in 1986, 212, 22.04 and 19.48 in 1987, 238, 24.74 and 21.8 in 1988, 245, 25.47 and 22.33 in 1989 and 284, 29.57 and 25.69 in 1990, respectively. The number of patients with this disease increased annually. The increase in the age-specific incidence rate of this type of cancer between 1986 and 1990 was 1.38-fold in women between 40 and 49 years of age, 1.46-fold in women between 50 and 59, 1.37-fold in the 60-69 age group, 1.48-fold in the 70-79 age group and 1.91-fold in women over 80. In both the urban and rural areas, the incidence increased annually during the course of this study. In 1990, the age-standardized mammary carcinoma incidence rate in urban area was 1.17 times that in rural area.
Cyclic ADP-ribose (cADPR), a well-known stimulator of ca(2+) release from the intracellular Ca(2+) pool, has recently emerged as a potential regulator of insulin secretion in pancreatic beta cells. As recently described, BST-1 is a glycosyl-phosphatidylinositol (GPI)-anchored surface molecule that exhibits homology with CD38 and Aplysia ADP-ribosyl cyclase. Like CD38, BST-1 has both ADP-ribosyl cyclase and cADPR hydrolase activities. As a step toward elucidating the physiological role of cADPR in insulin secretion we examined whether BST-1 is expressed in pancreatic islet cells. Sensitive reverse transcription-polymerase chain reaction detected almost as abundant expression of BST-1 mRNA in pancreatic islets as CD38 mRNA. Immunohistochemical analyses utilizing mirror image sections revealed that BST-1 protein is expressed in a majority of the cells in pancreatic islets and that at least beta cells and, to an even greater extent, alpha cells express BST-1. These observations suggest the involvement of multiple enzymes in the regulation of cADPR concentrations in pancreatic islet cells.
We have established a highly sensitive semi-nested PCR assay for the detection of H. pylori infection using gastric juice samples, which can be aspirated with disposable nasogastric tubes. The primers targeting H. pylori urease A gene were designed based on the sequence conservation analysis of sixteen H. pylori strains isolated from Japanese patients. The efficacy of the PCR assay, designated as the URA-PCR, was confirmed by in vitro and in vivo assessments. Its sensitivity was 97.5% in the gastric juice samples aspirated from forty patients with proven H. pylori infection, and was significantly higher than that obtained with previously described PCR assays.
The glucocorticoid dexamethasone inhibited the production of the rat cytokine-induced neutrophil chemoattractant CINC/gro, a counterpart of human melanoma growth-stimulating activity that belongs to the interleukin-8 (IL-8) family, in the normal rat kidney epithelial cell line NRK-52E stimulated with interleukin-1 beta (IL-1 beta), lipopolysaccharide, or tumor necrosis factor alpha. The accumulation of CINC/gro mRNA induced by these activators was also decreased comparably by dexamethasone. A nuclear run-on assay revealed that dexamethasone decreased the IL-1 beta-induced transcription of the CINC/gro gene. The half-life of CINC/gro mRNA transcripts did not change significantly after exposure to dexamethasone, suggesting that this glucocorticoid acts mainly at the transcriptional level. Transfection with luciferase expression vectors containing 5'-deleted and mutated CINC/gro gene sequences demonstrated that the 5'-flanking region containing the NF-kappa B binding site is involved in the IL-1 beta- and dexamethasone-induced activation and repression of the CINC/gro gene expression, respectively. Furthermore, a tandem repeat of the NF-kappa B sequence in the CINC/gro gene conferred the inducibility by IL-1 beta and suppression of luciferase activity by dexamethasone. In an electrophoretic mobility shift assay, dexamethasone diminished the IL-1 beta-induced formation of NF-kappa B complexes, which consisted of p65 and p50. Western blotting revealed that dexamethasone inhibited the IL-1 beta-induced translocation of p65 from the cytoplasm into the nucleus, while the nuclear level of NF-kappa B p50 remained almost unchanged. In addition, the degradation of I kappa B-alpha induced by IL-1 beta was not inhibited by dexamethasone. These results indicated that the suppression of the CINC/gro gene transcription by glucocorticoid occurs through the impairment of NF-kappa B activation, possibly by interference with the translocation of NF-kappa B p65 from the cytoplasm into the nucleus, thereby suppressing transactivation of the CINC/gro gene.
We have been interested in the effects of some popular peptides on tracheal smooth muscle. Previously, we reported that atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP) and C-type natriuretic peptide (CNP) had dose-dependent relaxant effects on guinea pig tracheal smooth muscle. In this study, we compared the effects of ANP, BNP and CNP on histamine release from rat peritoneal mast cells. ANP and BNP were more potent than CNP, dose-dependently increasing histamine release at a concentration of 10(-7) M or higher. CNP induced histamine release at a concentration of 10(-6) M or higher. Extracellular calcium inhibited the histamine release induced by all 3 peptides. In conclusion, the effects of these 3 peptides in rat peritoneal mast cells demonstrated adverse reactions for respiratory diseases, although our previous results showed that these peptides caused relaxation of guinea pig tracheal smooth muscle. We should note that the drugs have different actions in each organ.
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Platelet-activating factor (PAF), a potent mediator of inflammation and circulatory shock, is inactivated by the enzyme PAF acetylhydrolase. Plasma PAF acetylhydrolase deficiency occurs even in healthy subjects. We hypothesized that erythrocyte PAF acetylhydrolase could play a supplementary role in this plasma acetylhydrolase deficiency. We examined 1,030 subjects who participated in mass checkups, and assayed plasma and erythrocyte PAF acetylhydrolase. We also investigated the degradation of exogenous PAF by erythrocytes or other blood cells obtained from subjects who exhibited the plasma enzyme deficiency. The incidence of the plasma enzyme deficiency in this general Japanese population was 4.7% (48/1,030). None of the subjects with the deficiency had a history of allergy, circulatory shock, or chronic inflammatory diseases. The mean values for erythrocyte cytosolic PAF acetylhydrolase activity in the normal and deficient subjects were 0.51 +/- 0.15 (SD) and 0.71 +/- 0.28 nkat (nmol/s)/g protein, respectively, and the difference was significant (P < 0.001, Mann-Whitney U-test). The half-life of 10 nmol/l [3H]PAF in plasma from normal subjects was about 5 min, and the half-life in whole blood or erythrocyte suspension in autologous plasma was almost the same as that in plasma. In plasma from deficient subjects, unchanged PAF virtually remained and the degradation in whole blood or erythrocyte suspension was a little faster than in plasma. We conclude that erythrocytes contribute only little to PAF metabolism in normal blood but they account for almost all of the slow PAF degradation in blood from subjects deficient in plasma PAF acetylhydrolase.