[Needs and importance of physical examination: discussion].
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Humans↗
Biomedical subjects
S Yamashiro
Publications and source records attributed to S Yamashiro.
Expression of CCR6 and CD83 by cytokine-activated human neutrophils.
Polymorphonuclear leukocytes (PMNLs) are thought to be terminally differentiated, short-lived, and unable to actively synthesize new proteins or to interact with T cells. In the current study, it was found that PMNLs incubated with supernatants of phytohemagglutinin (PHA)-stimulated peripheral blood mononuclear cells (PHA-sup) expressed high levels of CCR6 mRNA. Neutralization with IgG against several cytokines revealed that tumor necrosis factor (TNF)-alpha was largely responsible for the PHA-sup-induced CCR6 mRNA expression. Among recombinant cytokines, TNF-alpha induced high levels of CCR6 mRNA expression, whereas interferon (IFN)-gamma induced low levels. The 2 cytokines together exhibited a considerable synergy. Cytokine-activated PMNLs expressed functional CCR6, as detected by the binding of sodium iodide I 125-labeled liver and activation-regulated chemokine (LARC) and dose-dependent migration toward LARC. The induction of CCR6 suggested that these cytokine-activated PMNLs have more similarities with dendritic cells (DCs) that express CCR6 in an immature stage. In fact, the activation of PMNLs with TNF-alpha and IFN-gamma induced the expression of CD83, a dominant cell-surface marker of DCs. When PMNLs were activated with granulocyte macrophage-colony-stimulating factor, TNF-alpha, and IFN-gamma, these cells expressed CD40 and HLA-DR in addition to CD83. Taken together, PMNLs, under appropriate conditions, can undergo a differentiation process characterized by the acquisition of new phenotypes and functions, and such differentiated PMNLs may play more active roles in the adaptive immune response. (Blood. 2000;96:3958-3963)
Antigens, CD↗
Distinct roles of ROCK (Rho-kinase) and MLCK in spatial regulation of MLC phosphorylation for assembly of stress fibers and focal adhesions in 3T3 fibroblasts.
ROCK (Rho-kinase), an effector molecule of RhoA, phosphorylates the myosin binding subunit (MBS) of myosin phosphatase and inhibits the phosphatase activity. This inhibition increases phosphorylation of myosin light chain (MLC) of myosin II, which is suggested to induce RhoA-mediated assembly of stress fibers and focal adhesions. ROCK is also known to directly phosphorylate MLC in vitro; however, the physiological significance of this MLC kinase activity is unknown. It is also not clear whether MLC phosphorylation alone is sufficient for the assembly of stress fibers and focal adhesions. We have developed two reagents with opposing effects on myosin phosphatase. One is an antibody against MBS that is able to inhibit myosin phosphatase activity. The other is a truncation mutant of MBS that constitutively activates myosin phosphatase. Through microinjection of these two reagents followed by immunofluorescence with a specific antibody against phosphorylated MLC, we have found that MLC phosphorylation is both necessary and sufficient for the assembly of stress fibers and focal adhesions in 3T3 fibroblasts. The assembly of stress fibers in the center of cells requires ROCK activity in addition to the inhibition of myosin phosphatase, suggesting that ROCK not only inhibits myosin phosphatase but also phosphorylates MLC directly in the center of cells. At the cell periphery, on the other hand, MLCK but not ROCK appears to be the kinase responsible for phosphorylating MLC. These results suggest that ROCK and MLCK play distinct roles in spatial regulation of MLC phosphorylation.
3T3 Cells↗
Long-term results of root reconstruction using the Carrel patch.
OBJECTIVE: The treatment of annuloaortic ectasia in patients, including those with Marfan syndrome, ascending aortic dissection, and other disorders of the ascending aorta and aortic valve presents a surgical challenge that has, unfortunately, shown high hospital mortality up to now. Improvements in graft materials and advanced surgical techniques have, however, begun to reduce hospital mortality. SUBJECTS AND METHODS: We retrospectively analyzed the records of 47 consecutive patients who undergoing aortic root reconstruction using the Carrel patch between January 1991 and March 1999. RESULTS: Postoperative complications included myonephrotic metabolic syndrome caused by femoral artery cannulation in 2 patients. Reexploration was done to halt bleeding in 2 patients. In 1 of 4 acute dissection patients, retrograde filling of the false lumen was demonstrated postoperatively. Overall surgical mortality in this series was 2.1% (1 of 47). The cardiac-event free rate was 98% at 5 years and 88% at 8 years. Actual survival is 97.8% at 8 years. No anastomosis complications were seen during follow-up (average: 32.7 months) (about 2.73 years). CONCLUSION: Surgery is considered feasible in any anatomic variation of aortic root disease, even in coronary ostial minimal dislocation, and the Carrel patch holds hope in preventing of anastomotic pseudaneurysm and ensuring long-term survival. Our experience suggests that modified Bentall operation, or aortic root remodeling using the Carrel patch, has few late-term complications, even in Marfan patients.
Adult↗
Alteration in the responsiveness to tumour necrosis factor-alpha is crucial for maximal expression of monocyte chemoattractant protein-1 in human neutrophils.
We previously reported delayed expression of monocyte chemoattractant protein-1 (MCP-1) in human neutrophils cultured with a cytokine-rich crude supernatant of phytohaemagglutinin-stimulated peripheral blood mononuclear cells (PHA-sup). Tumour necrosis factor-alpha (TNF-alpha) contained in the PHA-sup played a key role in this event, but there appeared to be another factor(s) in the same supernatant that co-operated with TNF-alpha for maximal MCP-1 expression. In the present study, we reduced TNF-alpha concentrations in the PHA-sup to minimal levels using anti-TNF-alpha affinity columns (TNF-depleted-sup) and investigated the co-operation between TNF-alpha and TNF-depleted-sup. Nine hours of preincubation with TNF-depleted-sup altered the responsiveness of neutrophils to TNF-alpha and enabled TNF-alpha to increase the level of MCP-1 expression to a maximal level within 4 hr. The priming effect was not due to the increased expression of cell-surface TNF receptors. However, the activation of primed cells by TNF-alpha was clearly through TNF receptor-p55. Finally, the activity in the TNF-depleted-sup that co-operated with TNF-alpha was eluted at 60 000 MW on high-performance liquid chromatography-gel filtration. Thus, delayed neutrophil expression of MCP-1 is regulated by a cytokine-dependent mechanism that induces neutrophils to enter a 'mature' stage.
Antigens, CD↗
Abnormal illness behavior and psychiatric disorders: a study in an outpatient clinic in Japan.
Abnormal illness behavior, such as hypochondriacal attitude and inappropriate treatment-seeking, has been associated with various psychiatric disorders in which patients tend to abuse medical services and seek inappropriate treatment in general practice clinics rather than psychiatric clinics. However, the relationship between illness behavior and psychiatric disorders in Japan is yet to be elucidated. We examined the abnormal illness behavior of 243 patients who visited the outpatient department of psychiatry at Saga Medical School Hospital, Saga, Japan, using a Japanese version of the Illness Behavior Questionnaire (IBQ). Multivariate analysis indicated significant association between some of the IBQ scale scores and age, sex and employment status. Patients with anxiety disorder scored higher on five of the seven IBQ scales compared with patients with another major disorder (mood disorder, schizophrenia or somatoform disorder). When compared with the IBQ scale scores reported in Australian patients in a psychiatric hospital, most of the IBQ scale scores differed significantly in our patients; a higher score among Japanese patients on the general hypochondriasis scale was most prominent. A similar trend in IBQ scale scores was also noted among Japanese patients visiting the hospital's general medicine clinic in comparison with Australian patients visiting a general practice clinic. Japanese patients with anxiety disorder may display the most salient abnormal illness behaviors among patients with psychiatric disorders. Sociocultural background may contribute to the characteristic abnormal illness behaviors of Japanese patients.
Adolescent↗
Expression of the actin-bundling protein fascin in cultured human dendritic cells correlates with dendritic morphology and cell differentiation.
Dendritic cells are key players of the immune system as they efficiently induce primary immune responses by activating naive T cells. We generated human dendritic cells from CD14+ blood precursors and investigated expression of the actin-bundling protein fascin during maturation by western blotting, immunofluorescence, and cytofluorometry. Cells obtained by culture of CD14+ blood precursors in the presence of granulocyte-macrophage colony-stimulating factor and interleukin-4, which were only weakly positive for the maturation marker CD83, expressed low amounts of fascin. Addition of a cytokine cocktail including tumor necrosis factor alpha, interleukin-1beta, interleukin-6, and prostaglandin E2 induced maturation of the cells and enhanced fascin expression in parallel with CD83 expression. Isolated mature CD83+ cells displayed especially high fascin levels on western blots, as did gated CD83+ dendritic cells in cytofluorometry. Dendritic cells generated from CD34+ blood precursors expressed high levels of fascin as well. Confocal microscopy revealed that location of fascin within the cell was restricted to the area of the submembranous actin cytoskeleton and to the dendritic processes. Suppression experiments using antisense constructs of fascin hint at a retarded morphologic maturation of dendritic cells, supporting the view that fascin expression is pivotal for dendrite formation. Our data suggest that fascin could serve as a marker molecule to monitor the maturation state of in vitro generated dendritic cells for use in clinical trials.
Actins↗
Comparison of contractile responses relative to protein between isolated diabetic and non-diabetic rat aortae to KCl and alpha-adrenoceptor agonists in different extracellular calcium concentrations.
The influence of extracellular Ca2+ concentration on contractile responses of aortae isolated from diabetic rats to KCl and alpha-adrenoceptor agonists was compared with that of non-diabetic rat aortae. Diabetic rats 6 weeks after administration of streptozotocin showed significantly lower body weight and higher plasma glucose concentration, but the protein content per each aortic ring preparation was not significantly different from that of non-diabetic preparation. Both diabetic and non-diabetic aortae showed concentration-dependent contractile responses to norepinephrine, which were concentration-dependently inhibited by prazosin. On the other hand, clonidine induced small contractions in both aortae, which tended to be more inhibited by prazosin than yohimbine. In non-diabetic aortae, the contractile responses to KCl, norepinephrine, methoxamine and clonidine were significantly greater with 2.5 mmol/l of extracellular Ca2+ than 1.25 mmol/l. In diabetic aortae, however, the contractile responses were not significantly influenced by changes in extracellular Ca2+ concentration. Additionally, the contractile responses to each agonist were markedly greater in non-diabetic aortae than diabetic ones. The present results indicate that the contractions of diabetic vasculature do not respond to changes in extracellular Ca2+ concentration, suggesting that there may be some impairment of Ca2+ related mechanisms.
Adrenergic alpha-Agonists↗
Association of CTLA-4 gene A/G polymorphism in Japanese type 1 diabetic patients with younger age of onset and autoimmune thyroid disease.
OBJECTIVE: We studied the association between type 1 diabetes with autoimmune thyroid disease (AITD) and A/G allele polymorphism in exon 1 of the CTLA-4 gene in a Japanese population. RESEARCH DESIGN AND METHODS: We studied 74 Japanese type 1 diabetic patients with or without AITD and 107 normal subjects to identify the association between CTLA-4 polymorphism and type 1 diabetes using polymerase chain reaction-restriction fragment length polymorphism analysis. RESULTS: The frequency of the CTLA-4 G allele differed significantly between the type 1 diabetic patients (61%) and the normal control subjects (48%) (P = 0.016). The difference in the CTLA-4 G allele became greater between patients with a younger age of onset of type 1 diabetes (age at onset <30 years) and the normal control subjects (64% and 48%, respectively). However, the frequency of the CTLA-4 G allele did not differ between type 1 diabetic patients with younger and older age of onset (64% vs. 57%). The G allele frequencies in the patients with younger-onset type 1 diabetes and AITD increased more than in the control patients (P = 0.025). These differences reflected a significant increase in the frequency of G/G genotype--that is, 54% in those with younger-onset type 1 diabetes and AITD, 39% in those without AITD, and 28% in control subjects. CONCLUSIONS: An association was detected between the CTLA-4 gene polymorphism and younger-onset type 1 diabetes with AITD. The G variant was suggested to be genetically linked to AITD-associated type 1 diabetes of younger onset in this apanese population. The defect in these patients presumably lies in a T-cell-mediated autoimmune mechanism.
Abatacept↗
Alteration of tumor phenotypes of B16 melanoma after genetic remodeling of the ganglioside profile.
Gangliosides have been thought to be tumor markers of various neuroectoderm-derived cancers. However, their roles in malignant phenotypes of cancer cells are not well understood. We have performed the remodeling of ganglioside profiles of mouse melanoma B16 (B78 subline) by introducing GM2/GD2 synthase cDNA, and analyzed the phenotypic changes such as cell morphology, growth, adhesion and c-fos gene expression. Although newly expressed GM2 was clearly detected, GM2-positive transfectant cells showed rather reduced growth rates in vivo and in vitro, lower expression levels of c-fos gene and increased levels of cell adhesion to extracellular matrices compared to control cells. These results suggested that GM2 is involved in the regulation of cell-cell or cell-extracellular matrix interaction, and negatively control cell proliferation.
Animals↗
Cardiac operations in patients with severe pulmonary impairment.
Many reviews concerning pulmonary complications after cardiac surgical procedures in patients with serious pulmonary disease have been published. However, no strict pulmonary function guidelines were proposed to help the clinician identify the patients at greater risk. We considered whether a low pulmonary function became a risk factor of cardiac operations. We conducted a retrospective analysis of records of 32 patients with severely impaired preoperative pulmonary function who had undergone cardiac operations between July 1988 and March 1999. There was 1 hospital death. The over-all mortality rate was 3.1% (1 of 32). However, this death could not be directly attributed to postoperative pulmonary complications. Postoperative pulmonary complications were seen in 2 patients (6.3%) who required tracheostomy due to atelectasis and pneumonia. No late deaths due to pulmonary complications were observed during the follow-up period. The actual survival rate is 68% at 7 years. A low pulmonary function did not, by itself, become a risk factor of cardiac operations, although a pulmonary function test can be used to alert the clinician to possible postoperative complications, including the requirement of tracheostomy. Especially strict control of postoperative respiration is necessary in patients with forced expiratory volume (FEV) of 1.0 <= 800 ml and/or FEV1.0/BSA <= 600 ml/m2.
Aged↗
Long-term angiographic evaluation of free internal thoracic artery grafting for myocardial revascularization.
Between 1990 and 1998, 41 patients underwent free internal thoracic artery grafting for coronary artery bypass. To investigate usefulness of free internal thoracic artery grafting, we compared the postoperative graft patency of free internal thoracic artery grafts with that of in situ internal thoracic artery grafts, and compared the long-term results such as actual survival and cardiac-event free rate in patients receiving free internal thoracic artery grafts with those results in patients receiving in situ internal thoracic artery grafts. Postoperative changes in luminal diameter of free internal thoracic artery grafts were calculated as the difference between the first and secondary angiographic evaluation in 17 patients who were followed-up for more than 5 years. The early postoperative graft patency rate of free internal thoracic artery was 95.2%. All these early patent grafts remained patent at the time of the late study. At 9 years post-surgery, patients who received free internal thoracic artery grafts had a survival rate of 100% and a cardiac event-free rate of 77.5%; the luminal diameter enlargement was 0.57 mm, and the mean matching rate increased 27.8%. We conclude that the free internal thoracic artery provides long-term results comparable with those of in situ internal thoracic artery.
Aged↗
[Practice and application of evidence-based medicine].
Evidence-based medicine is an approach to clinical practice that aims to solve the patient's problems. EBM includes five steps: identifying patient's problems, searching medical literature efficiently, critically appraising the literature(evidence), applying it to patient care, and self-assessment of these steps. Since September 1999, we have practiced the EBM approach to patient care, in inpatient and outpatient settings at the department of General Medicine of Saga Medical School. We found ourselves more clearly differentiating between the EBM approach and past conventional practice. We found EBM to be more clinically useful and we have enjoyed reading medical literature because of the focus on patient's problems. To improve our use of the EBM approach, we need to study more about clinical epidemiology to evaluate and use medical literature in patient care.
Education, Medical, Continuing↗
Activation of myosin phosphatase targeting subunit by mitosis-specific phosphorylation.
It has been demonstrated previously that during mitosis the sites of myosin phosphorylation are switched between the inhibitory sites, Ser 1/2, and the activation sites, Ser 19/Thr 18 (Yamakita, Y., S. Yamashiro, and F. Matsumura. 1994. J. Cell Biol. 124:129- 137; Satterwhite, L.L., M.J. Lohka, K.L. Wilson, T.Y. Scherson, L.J. Cisek, J.L. Corden, and T.D. Pollard. 1992. J. Cell Biol. 118:595-605), suggesting a regulatory role of myosin phosphorylation in cell division. To explore the function of myosin phosphatase in cell division, the possibility that myosin phosphatase activity may be altered during cell division was examined. We have found that the myosin phosphatase targeting subunit (MYPT) undergoes mitosis-specific phosphorylation and that the phosphorylation is reversed during cytokinesis. MYPT phosphorylated either in vivo or in vitro in the mitosis-specific way showed higher binding to myosin II (two- to threefold) compared to MYPT from cells in interphase. Furthermore, the activity of myosin phosphatase was increased more than twice and it is suggested this reflected the increased affinity of myosin binding. These results indicate the presence of a unique positive regulatory mechanism for myosin phosphatase in cell division. The activation of myosin phosphatase during mitosis would enhance dephosphorylation of the myosin regulatory light chain, thereby leading to the disassembly of stress fibers during prophase. The mitosis-specific effect of phosphorylation is lost on exit from mitosis, and the resultant increase in myosin phosphorylation may act as a signal to activate cytokinesis.
Amino Acid Sequence↗
Dissociation of FAK/p130(CAS)/c-Src complex during mitosis: role of mitosis-specific serine phosphorylation of FAK.
At mitosis, focal adhesions disassemble and the signal transduction from focal adhesions is inactivated. We have found that components of focal adhesions including focal adhesion kinase (FAK), paxillin, and p130(CAS) (CAS) are serine/threonine phosphorylated during mitosis when all three proteins are tyrosine dephosphorylated. Mitosis-specific phosphorylation continues past cytokinesis and is reversed during post-mitotic cell spreading. We have found two significant alterations in FAK-mediated signal transduction during mitosis. First, the association of FAK with CAS or c-Src is greatly inhibited, with levels decreasing to 16 and 13% of the interphase levels, respectively. Second, mitotic FAK shows decreased binding to a peptide mimicking the cytoplasmic domain of beta-integrin when compared with FAK of interphase cells. Mitosis-specific phosphorylation is responsible for the disruption of FAK/CAS binding because dephosphorylation of mitotic FAK in vitro by protein serine/threonine phosphatase 1 restores the ability of FAK to associate with CAS, though not with c-Src. These results suggest that mitosis-specific modification of FAK uncouples signal transduction pathways involving integrin, CAS, and c-Src, and may maintain FAK in an inactive state until post-mitotic spreading.
Amino Acid Sequence↗
MCP-1 is selectively expressed in the late phase by cytokine-stimulated human neutrophils: TNF-alpha plays a role in maximal MCP-1 mRNA expression.
Culture supernatants of phytohemagglutinin (PHA)-stimulated human peripheral blood mononuclear cells (PHA-sup) induced monocyte chemoattractant protein-1 (MCP-1) mRNA expression in human neutrophils. MCP-1 mRNA was first detected by Northern analysis at 8 h, and the peak level was detected at 16 h and sustained until 72 h. Cycloheximide and genistein, but not pertussis toxin, inhibited the expression of MCP-1 mRNA. Recombinant tumor necrosis factor alpha (TNF-alpha) induced a low level MCP-1 mRNA accumulation in neutrophils, and addition of anti-TNF-alpha IgG blocked 30-70% of MCP-1 mRNA expression induced with PHA-sup. PHA-sup-stimulated PMN synthesized and secreted 3.1+/-1.3 ng/5 x 10(6) PMN MCP-1 within the first 24 h. Hybridization of 32P-labeled cDNA preparations to an array of human cytokine cDNAs further indicated that MCP-1 mRNA was selectively up-regulated in the late phase after stimulation with the PHA-sup.
Chemokine CCL2↗
Effects of glutamate receptor antagonists on lower urinary tract function in conscious unanesthetized rats.
OBJECTIVES: To study the effects of the intrathecal administered glutamate receptor antagonists on the bladder and urethral activities during isovolumetric bladder contraction in conscious normal and chronic spinal rats. METHODS: Twenty-eight female Wistar rats with and without previous spinal cord transection were used. Before and after intrathecal administration of glutamate receptor antagonist, urodynamic parameters under isovolmetric condition of the bladder were analyzed. RESULTS: In normal rats, MK-801 (noncompetitive N-methyl-D-aspartate [NMDA] receptor antagonist) and LY 293558 (competitive AMPA receptor antagonist) produced a decrease in bladder contraction pressure and urethral activity with dose dependent manner. In chronic spinal rats, detrusor-sphincter dyssynergia (DSD) was developed before drug administration. MK-801 and LY 293558 partially inhibited bladder contraction pressure, and markedly depressed urethral contraction concomitant with bladder contraction. LY 293558 produced urethral relaxation concomitant with bladder contraction. CONCLUSIONS: In both normal rats and chronic spinal rats, two subtypes of glutamate receptors (NMDA and AMPA receptors) in the spinal cord were involved in the control of bladder and urethral activities. The AMPA receptor in the spinal cord seems to take an important role in the development of DSD.
Animals↗
Cell-matrix adhesions differentially regulate fascin phosphorylation.
Cell adhesion to individual macromolecules of the extracellular matrix has dramatic effects on the subcellular localization of the actin-bundling protein fascin and on the ability of cells to form stable fascin microspikes. The actin-binding activity of fascin is down-regulated by phosphorylation, and we used two differentiated cell types, C2C12 skeletal myoblasts and LLC-PK1 kidney epithelial cells, to examine the hypothesis that cell adhesion to the matrix components fibronectin, laminin-1, and thrombospondin-1 differentially regulates fascin phosphorylation. In both cell types, treatment with the PKC activator 12-tetradecanoyl phorbol 13-acetate (TPA) or adhesion to fibronectin led to a diffuse distribution of fascin after 1 h. C2C12 cells contain the PKC family members alpha, gamma, and lambda, and PKCalpha localization was altered upon cell adhesion to fibronectin. Two-dimensional isoelectric focusing/SDS-polyacrylamide gels were used to determine that fascin became phosphorylated in cells adherent to fibronectin and was inhibited by the PKC inhibitors calphostin C and chelerythrine chloride. Phosphorylation of fascin was not detected in cells adherent to thrombospondin-1 or to laminin-1. LLC-PK1 cells expressing green fluorescent protein (GFP)-fascin also displayed similar regulation of fascin phosphorylation. LLC-PK1 cells expressing GFP-fascin S39A, a nonphosphorylatable mutant, did not undergo spreading and focal contact organization on fibronectin, whereas cells expressing a GFP-fascin S39D mutant with constitutive negative charge spread more extensively than wild-type cells. In contrast, C2C12 cells coexpressing S39A fascin with endogenous fascin remained competent to form microspikes on thrombospondin-1, and cells that expressed fascin S39D attached to thrombospondin-1 but did not form microspikes. Blockade of PKCalpha activity by TPA-induced down-regulation led to actin association of wild-type fascin in fibronectin-adherent C2C12 and LLC-PK1 cells but did not alter the distribution of S39A or S39D fascins. The association of fascin with actin in fibronectin-adherent cells was also evident in the presence of an inhibitory antibody to integrin alpha5 subunit. These novel results establish matrix-initiated PKC-dependent regulation of fascin phosphorylation at serine 39 as a mechanism whereby matrix adhesion is coupled to the organization of cytoskeletal structure.
Actins↗