Biomedical subjects
Publications and source records attributed to M Shinohara.
We report on a 3-year-old boy in whom a complete occlusive thrombus in the right femoral artery was removed rapidly and successfully using a hydrodynamic thrombectomy catheter. There was no bleeding complication, reocclusion, vascular injury, or loss of motor or sensory function. Hydrodynamic thrombectomy can be an important part of the treatment of thrombosis in children.
UNLABELLED: Recent studies have shown that vitamin D receptor (VDR) gene polymorphism had regulatory effects on bone mineral density (BMD) and bone turnover. The VDR gene has also been indicated as a candidate gene for the susceptibility of osteoporosis. However, it is unclear whether VDR genotypes could be associated with alveolar bone loss of patients with periodontitis, or whether vitamin D receptor gene could be a candidate gene for susceptibility to periodontitis. The purpose of this study was to answer these two questions. METHODS: Twenty-four cases of adult periodontitis (AP), 37 cases of early onset periodontitis (EOP) and 39 healthy controls were recruited for the study. Individual samples of venous blood and DNA were obtained from each subject. Genotypes of the TaqI VDR gene were determined by PCR and TaqI restriction endonuclease digestion. RESULTS: One out of 24 AP patients, nine out of 37 EOP patients and two out of 39 healthy controls were detected with Tt genotype, while the rest had the TT genotype. The detected frequency of Tt genotype was significantly higher in EOP patients (24.3%) than in AP patients (4.2%) and healthy controls (5.1%). The frequency of t allele was also significantly higher in EOP patients. There was no statistical difference in the distribution of TaqI VDR genotypes between AP patients and healthy controls. The study suggests that Tt genotype might be a risk indicator for the susceptibility to EOP. Carriage of the allele (t) of the TaqI VDR gene may increase the risk of developing EOP.
Alterations of cell cycle-regulated genes play an important role in the process of carcinogenesis, and some of them are thought to be prognostic factors in esophageal cancer. The expressions of p53, p16, pRB and Cyclin D1 proteins were evaluated immunohistochemically in 144 patients who underwent curative esophagectomy without any adjuvant therapy before surgery. p53 overexpression was observed in 99 (69%) out of the 144 patients. No significant correlation was noted between p53 and any other gene expression. p16 expression was observed in 12 (8.3%) out of all cases. A negative correlation was recognized between p16 and Cyclin D1 expression (P=0.0004). pRB expression was observed in 130 (90.3%) out of all cases, whereas pRB expression was not observed in 11 out of the 12 patients with p16-positive tumors. A negative correlation was also found between p16 and pRB (P < 0.0001). A positive correlation was noted between pRB and Cyclin D1 expression (P=0.0009). The cumulative survival rate of patients without pRB expression was significantly lower than that of patients with positive expression (P=0.003). In the multivariate survival analysis, pRB expression was an independent prognostic factor. In 98% of all patients with esophageal cancer, impairment of the G1 checkpoint is due to a loss of function by p16, pRB or Cyclin D1, which showed a negative correlation in each factor. In addition, aberrant expression of pRB is useful as a prognostic factor in esophageal cancer.
BACKGROUND: A possible relation between maternal-fetal microchimerism and autoimmune diseases with some similarities to chronic graft versus host disease (cGVHD) has been reported. OBJECTIVE: To investigate whether cells with male DNA exist in female patients with Sjögren's syndrome (SS) as SS has clinical features similar to those of cGVHD. METHODS: DNA was extracted from 27 samples of peripheral blood mononuclear cells (PBMC), 42 biopsy samples of labial salivary glands (LSG), and nine samples of bronchoalveolar lavage fluid (BALF) cells from 56 female patients with SS. The presence of male DNA was determined by nested polymerase chain reaction (PCR) and by fluorescence in situ hybridisation (FISH). RESULTS: Among 56 female patients with SS, 42 patients had at least one male child. Among those 42 patients, none of the 22 PBMC but 10/28 (36%) LSG samples tested positive by PCR for the Y chromosome-specific sequence (p=0.0013). The Y chromosome-specific sequence was not detected in the samples of LSG in 10 patients without SS. In the BALF samples 2/9 (22%) patients with SS tested positive by PCR. Cells containing the Y chromosome were shown to exist in all the LSG specimens from three female patients with SS by FISH. CONCLUSIONS: Maternal-fetal microchimerism was shown for the first time to exist in the salivary glands and lungs of female patients with SS in this study. The presence of non-host cells in the inflammatory lesions but not in the peripheral blood suggests a possible role for maternal-fetal microchimerism in the pathogenesis of SS.
Porphyromonas gingivalis (P. gingivalis) is implicated in the initiation and progression of periodontitis. Human gingival fibroblasts (HGFs) are the major constituent of gingival connective tissue. P. gingivalis or its components such as lipopolysaccharide (LPS) upregulate the production of various inflammatory cytokines including interleukin (IL)-1 and IL-6 in HGFs. Recently, we demonstrated that the binding of P. gingivalis LPS to Toll-like receptor 4 (TLR4) on HGFs activates various second messenger systems (Biochem. Biophys. Res. Commun. 273, 1161-1167, 2000). In the present study, we examined the level of TLR4 expression on HGFs by flow cytometric analysis (FACS), and studied the levels of IL-1 and IL-6 in the culture medium upon LPS stimulation of HGFs by enzyme-linked immunosorbent assay (ELISA). Upon stimulation by P. gingivalis LPS for 24 h, HGFs that expressed a high level of TLR4 secreted significantly higher levels of IL-1 and IL-6 than HGFs that expressed a low level of TLR4. On the other hand, after stimulation with P. gingivalis LPS for 24 h, the level of TLR4 on the surface of HGFs decreased. These results suggest that the level of TLR4 expression on HGFs reflects the extent of inflammation in the gingival tissue, and that P. gingivalis LPS downregulates TLR4 expression on HGFs. These findings may be used to control inflammatory and immune responses in periodontal disease.
We developed a system by which antibodies, fused to fluorescent proteins with different wavelengths, can be prepared within a month against various antigens. An antibody library composed of a large number of single-chain Fv-CL fragment was constructed by means of a phage-display system. The constructs were designed to facilitate changing of the protein forms by simple enzyme manipulation. In the present study, we adopted a molecular form of antibody in which a single-chain Fv-CL fragment is fused with a green fluorescent protein (GFP) or red fluorescent protein (RFP). In addition, a His-tag was inserted between CL and GFP (or RFP). We describe the utility of this system using Caenorhabditis elegans embryo as a model.
We evaluate the role of endomorphins 1 and 2 on microglial functions. Endomorphins 1 and 2 blocked phagocytosis of Escherichia coli. In addition, both markedly inhibited chemotaxis toward zymosan-activated serum. In contrast, when microglia was preincubated with these endomorphins, followed by incubation with LPS before stimulation with phorbol 12-myristate 13-acetate (PMA) at 200 nM, they potentiated superoxide anion production. Furthermore, when microglia was preincubated with these endomorphins together with PMA at 20 nM, followed by stimulation with PMA at 200 nM, superoxide anion production was potentiated. These results suggest that endomorphins 1 and 2 modulate phagocytosis, chemotaxis and superoxide anion production by microglia.
The previously isolated cDNA encoding human adenylate kinase (AK) isozyme 3 was recently renamed AK4. Consequently, human AK3 cDNA remains to be identified and we have little information about the functional relationship between human AK3 and AK4. In pursuit of the physiological roles of both the AK3 and AK4 proteins, we first isolated an authentic human AK3 cDNA and compared their expression. Nucleotide sequencing revealed that the cDNA encoded a 227-amino-acid protein, with a deduced molecular mass of 25.6 kDa, that shares greater homology with the AK3 cDNAs isolated from bovine and rat than that from human. We named the isolated cDNA AK3. Northern-blot analysis revealed that AK3 mRNA was present in all tissues examined, and was highly expressed in heart, skeletal muscle and liver, moderately expressed in pancreas and kidney, and weakly expressed in placenta, brain and lung. On the other hand, we found that human AK4 mRNA was highly expressed in kidney, moderately expressed in heart and liver and weakly expressed in brain. Western-blot analysis demonstrated expression profiles of AK3 and AK4 that were similar to their mRNA expression patterns in each tissue. Over expression of AK3, but not AK4, in both Escherichia coli CV2, a temperature-sensitive AK mutant, and a human embryonic kidney-derived cell line, HEK-293, not only produced significant GTP:AMP phosphotransferase (AK3) activity, but also complemented the CV2 cells at 42 degrees C. Subcellular and submitochondrial fractionation analysis demonstrated that both AK3 and AK4 are localized in the mitochondrial matrix.
The glutathione redox cycle plays a major role in scavenging hydrogen peroxide (H2O2) under physiological conditions. Recently, we demonstrated that a high glucose concentration in the culture medium reduced the level of H2O2 scavenging activity of human vascular smooth muscle cells (hVSMCs). We also showed that a high glucose concentration reduced the intracellular glutathione (GSH) content and the rate of uptake of cystine, which itself is a rate-limiting factor that maintains the GSH level (FEBS Lett.421: 19-22,1998). In the present study, we investigated whether the hyperglycemic condition in diabetic rats impairs the glutathione content in the aortic tissue in vivo. Wistar rats were divided into the following three groups: streptozotocin-induced diabetic rats (STZ-D, n=7), insulin-treated STZ-D rats (I-STZ-D, n=8), and non-diabetic controls (C, n=7). Fourteen days after streptozotocin injection, the aortic tissue was extracted and the GSH content in the aortic tissue was measured. Furthermore, the relationship between the GSH content in the aortic tissue and blood glucose level in Otsuka Long-Evans Tokushima Fatty (OLETF) rats aged 30 weeks, which developed diabetes spontaneously, was investigated. The GSH content in the aortic tissue of the STZ-D group (0.99+/-0.14 nmol/mg protein) was significantly lower than that of the control group (1.68+/-0.15 nmol/mg protein). Insulin treatment to the diabetic rats restored the GSH content in the aortic tissue (I-STZ-D group; 1.45+/-0.11 nmol/mg protein). Among the 22 Wistar rats, the GSH content in the aortic tissue was negatively correlated with the blood glucose level (r=-0.69, p<0.01, n=22). Among the OLETF rats, a similar negative correlation between the GSH content in the aortic tissue and blood glucose level was seen (r=-0.64, p<0.05, n=10). We demonstrated in vivo that the hyperglycemic condition in STZ-induced diabetic Wistar rats and OLETF rats reduced the GSH content in aortic tissue. This suggested reduced glutathione redox cycle function of aorta.
Neutrophils are often the first cells of the immune system to encounter an invader, such as bacteria and fungi. Lidocaine-epinephrine induced transient potentiation of the production of superoxide anion, while prilocaine-felypressine induced persistent inhibition of the production in neutrophils. Moreover, lidocaine-epinephrine inhibited the production of hydrogen peroxide in spite that it potentiated the production of superoxide anion, while prilocaine-felypressine inhibited the production of hydrogen peroxide as well as superoxide anion. By contrast, lidocaine-epinephrine and prilocaine-felypressine are both effective in significantly inhibiting adhesion and phagocytosis. Using flow cytometric analysis, both local anesthetics were found to be effective in inhibiting the expression of Mac-1 (CD11b/CD18) in neutrophils. These results suggest that lidocaine-epinephrine and prilocaine-felypressine differentially modulate the production of superoxide anion, and could similarly inhibit adhesion, phagocytosis, and the production of hydrogen peroxide by neutrophils.
15-Deoxy-Delta(12,14)-prostaglandin J(2) (dPGJ(2)) is a metabolite of prostaglandin D(2), that binds to peroxisome proliferator-activated receptor gamma (PPARgamma). PPARgamma and prostaglandin D(2) synthase, which is required for dPGJ(2) synthesis, are predominantly expressed in macrophages. In contrast, IL-10 and IL-12 produced by macrophages stimulate Th1 and Th2 immune response, respectively. This study investigated the effect of dPGJ(2) on IL-10 and IL-12 production by macrophages in response to lipopolysaccharide (LPS). Our data clearly demonstrated that dPGJ(2) inhibits LPS-induced IL-10 and IL-12 production by macrophages. A different agonist of PPARgamma, 13-hydroxyoctadecadienoic acid, similarly inhibited the production of IL-10 and IL-12 in response to LPS. Further, dPGJ(2) did not appear to act through the PGD(2) receptor. These results suggest that dPGJ(2) may inhibit LPS-induced IL-10 and IL-12 production by macrophages through PPARgamma.
Gab-1 is a multiple docking protein that is tyrosine phosphorylated by receptor tyrosine kinases such as c-Met, hepatocyte growth factor/scatter factor receptor, and epidermal growth factor receptor. We have now demonstrated that cell-cell adhesion also induces marked tyrosine phosphorylation of Gab-1 and that disruption of cell-cell adhesion results in its dephosphorylation. An anti-E-cadherin antibody decreased cell-cell adhesion-dependent tyrosine phosphorylation of Gab-1, whereas the expression of E-cadherin specifically induced tyrosine phosphorylation of Gab-1. A relatively selective inhibitor of Src family kinases reduced cell-cell adhesion-dependent tyrosine phosphorylation of Gab-1, whereas expression of a dominant-negative mutant of Csk increased it. Disruption of cell-cell adhesion, which reduced tyrosine phosphorylation of Gab-1, also reduced the activation of mitogen-activated protein kinase and Akt in response to cell-cell adhesion. These results indicate that E-cadherin-mediated cell-cell adhesion induces tyrosine phosphorylation by a Src family kinase of Gab-1, thereby regulating the activation of Ras/MAP kinase and phosphatidylinositol 3-kinase/Akt cascades.
cDNA clones of MT5-matrix metalloproteinase (MT5-MMP) and a related protein (designated MT5-MMP-del) were isolated by screening the cDNA library and by 3'-rapid amplification of cDNA ends using an ovary RNA of the medaka fish Oryzias latipes. The MT5-MMP clone encodes a protein of 546 amino acids while the MT5-MMP-del clone encodes a protein of 431 amino acids. Compared with mammalian counterparts, the fish MT5-MMP and MT5-MMP-del both lack the signal peptide and a part of the prodomain. The fish MT5-MMP and MT5-MMP-del were different in that the latter did not have the stem/transmembrane/cytoplasmic domain. The two fish MMPs were expressed in the ovary, testis, brain, and intestine. In the ovary, MT5-MMP mRNA was expressed in the oocytes of small growing follicles. In contrast, MT5-MMP-del mRNA was found in the stromal interstitial cells. These results strongly suggest that a MT5-MMP/gelatinase A cascade may possibly operate in the process of spawning and/or other events associated with ovulated oocytes or fertilized eggs of the medaka fish.
The effects of the local blood circulation and absolute torque on muscle endurance at different knee-joint angles were determined. The rate of muscle deoxygenation (using near-infrared spectroscopy), and the rate of muscle fatigue (using the slope of integrated electromyography, iEMG) were evaluated concurrently. Nine healthy subjects performed submaximal (50% maximal voluntary contraction. MVC) static knee extension at 50 degrees (extended position, EXT) and 90 degrees (flexed position, FLEX) joint angles until the target torque could no longer be maintained: that time was measured as the endurance time. They exercised with the circulation occluded (OCCL), and without (FREE) to study the possible effects of the local circulation. Although MVC torque was independent of joint angle [mean (SD) FLEX 250.6 (51.7) Nm and EXT 246.5 (46.6) N x m], significantly shorter (P<0.01) endurance time in FLEX [FREE 71.1 (10) s and OCCL 63.1 (8.8) s] than at EXT [FREE 115.3 (30) s and OCCL 106.7 (29.1) s] were obtained in both circulatory conditions. The iEMG-time slope was significantly greater in FLEX at the proximal and distal portion (P<0.05) in both circulatory conditions. Muscle deoxygenation rate in OCCL was significantly greater (P<0.05) at FLEX [20.8 (8.0)%] than EXT [10.9 (4.0)%]. The results would suggest that different knee-joint angle affects muscle endurance even if the local circulation is controlled. Circulatory disturbance would further reduce muscle endurance in EXT, but not in FLEX. Because of the greater muscle internal force in FLEX, local blood flow might be already limited even with a free circulation. The greater muscle deoxygenation and muscle fatigability would be related to the shorter muscle endurance in FLEX.
We describe a rare case of a benign pseudotumorous lesion (fibroangiomyomatous hyperplasia with elastosis) in the gallbladder in a 44-year-old Japanese woman, and discuss the rarity of elastosis in the gallbladder. To our knowledge, this case may be the first report of a pseudotumorous lesion of the gallbladder with elastosis in Japan.
A case of a penetration of the duodenum by a needle with migration to the pancreas in a 50-year-old man is reported herein. The patient was referred to us with a chief complaint of diarrhea. An abdominal plain roentgenogram showed a needle in the upper abdominal area. An abdominal computed tomography scan and contrast X-ray revealed the foreign body to be located outside of the duodenum and in the head of the pancreas. An emergency operation was therefore performed on the first day and the needle in the head of the pancreas was thus extirpated safely. A perforation of the gastrointestinal tract by an ingested foreign body is difficult to accurately and quickly diagnose when no peritonitis or abscess formation is observed. Therefore, the use of contrast X-ray is considered to be useful in the diagnosis of such a perforation.
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The innate host defense system is regulated in part by the number and activation of the function of neutrophils and macrophages. The immunological effects of a variety of quinolones on host defense functions of macrophage are reported. Ofloxacin (OFLX), lomefloxacin (LFLX), tosufloxacin (TFLX), fleroxacin (FLRX), sparfloxacin (SPFX) and levofloxacin (LVFX) significantly inhibited phagocytosis of Escherichica coli by macrophages. Moreover, TFLX markedly potentiated the expression of the adhesion molecule Mac-1 by macrophages. No significant alteration was detected in the adherence and the expression of adhesion molecule Mac-1 in macrophages treated with the other quinolones. In contrast, OFLX, LFLX, TFLX, and LVFX were effective in significantly increasing the production of hydrogen peroxide, while the other agents did not. These results suggest that the quinolones at a therapeutic concentration differentially affect phagocytosis, adhesion, and the production of hydrogen peroxide by macrophages.