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Biomedical subjects

Shinji Takai

Publications and source records attributed to Shinji Takai.

At least 73 records · Page 4Linked to original sources
PubMed2005

The absence of Rhodococcus equi in Mongolian horses.

In native Mongolian horses, the incidence and distribution of Rhodococcus equi are poorly understood. One hundred and fourteen equine fecal samples and 71 soil samples were collected from the camp sites of 26 nomadic families located in three areas less than 100 km from Ulaanbaatar, Mongolia. Five fecal samples were also collected from foals of Przewalski's Horses introduced into the Hustai National Park, Mongolia. No R. equi was isolated from the Mongolian horses or the soil samples. However, three colonies of R. equi were isolated from two fecal samples collected from foals of Przewalski's Horses. These isolates were avirulent, with neither 15- to 17-kDa antigens (VapA) nor a 20-kDa antigen (VapB) genes being detected. We concluded that native Mongolian horses and their environment appear free from contamination with R. equi.

Animals↗
PubMed2005

Variation in the N-terminal region of an M-like protein of Streptococcus equi and evaluation of its potential as a tool in epidemiologic studies.

OBJECTIVE: To develop a method for typing Streptococcus equi on the basis of the DNA sequence of the genes that produce an M-like protein and to compare isolates among the United States, Japan, and other countries. SAMPLE POPULATION: S equi strains CF32, Hidaka/95/2, and NCTC9682 as well as 82 other isolates from the United States, Japan, and other countries obtained during 1975 to 2001. PROCEDURE: DNA sequences of the structural genes ( SeM and SzPSe) that produce M-like proteins were determined for 3 representative strains to find a variable region. Variability in this region of SeM was then determined for the other isolates. Amino acid sequences were deduced and analyzed phylogenetically by use of the neighbor-joining method. RESULTS: Sequence diversity was detected in the N-terminal region of SeM but not in SzPSe of the 3 representative strains. Base substitutions in the variable region of SeM varied in a nonsynonymous manner, resulting in variation in the amino acid sequence. Eighty-five isolates were categorized as 32 types of SeM on the basis of differences in the deduced amino acid sequences. CONCLUSIONS AND CLINICAL RELEVANCE: This study documented a region in the N-terminal portion of SeM that varies in a nonsynonymous manner. This information should be useful in molecular epidemiologic studies of S equi.

Amino Acid Sequence↗
PubMed2004

Therapeutic applications of chymase inhibitors in cardiovascular diseases and fibrosis.

Chymase activates not only angiotensin I to angiotensin II but also latent transforming growth factor-beta-binding protein to transforming growth factor-beta. In dog grafted veins, chymase activity and angiotensin II concentration along with vascular proliferation were significantly increased, while they were significantly suppressed by a chymase inhibitor. After balloon injury in dog arteries, chymase activity was significantly increased in the injured artery, and a chymase inhibitor and an angiotensin AT(1) receptor antagonist were effective in preventing the vascular proliferation, but an angiotensin-converting enzyme inhibitor was ineffective. In fibrotic models, the tissue fibrosis was reduced by chymase inhibitors. In adhesion models, the transforming growth factor-beta concentration and adhesion formation were suppressed by chymase inhibitors. Therefore, chymase inhibitors may be useful for preventing cardiovascular diseases and fibrosis via inhibition of angiotensin II formation and transforming growth factor-beta activation.

Animals↗
PubMed2004

A specific chymase inhibitor, NK3201, suppresses bleomycin-induced pulmonary fibrosis in hamsters.

We evaluated whether a chymase inhibitor, 2-(5-formylamino-6-oxo-2-phenyl-1,6-dihydropyrimidine-1-yl)-N-[(3,4-dioxo-1-phenyl-7-(2-pyridyloxy))-2-heptyl]acetamide (NK3201), suppressed bleomycin-induced pulmonary fibrosis. Hamsters were orally administered NK3201 (30 mg/kg per day) or placebo, beginning 5 days before intratracheal instillation of bleomycin (10 mg/kg). Four weeks after the instillation of bleomycin, pulmonary chymase activity in placebo-treated hamsters was significantly higher than in control hamsters, whereas the activity in NK3201-treated hamsters was significantly lower than in placebo-treated hamsters. The ratio of fibrotic area to total area in NK3201-treated hamsters was significantly decreased to 54.0% of the ratio in placebo-treated hamsters. Therefore, NK3201 may be useful in the prevention of pulmonary fibrosis.

Acetamides↗
PubMed2004

A specific chymase inhibitor, 2-(5-formylamino-6-oxo-2-phenyl-1,6-dihydropyrimidine-1-yl)-N-[[3,4-dioxo-1-phenyl-7-(2-pyridyloxy)]-2-heptyl]acetamide (NK3201), suppresses development of abdominal aortic aneurysm in hamsters.

In this study, we investigated the effect of a specific chymase inhibitor, 2-(5-formylamino-6-oxo-2-phenyl-1,6-dihydropyrimidine-1-yl)-N-[[3,4-dioxo-1-phenyl-7-(2-pyridyloxy)]-2-heptyl]acetamide (NK3201), in the development of abdominal aortic aneurysm in a hamster experimental model. The abdominal aortic aneurysm was induced by application of elastase onto the abdominal aorta in hamster. Each hamster was administered NK3201 (30 mg/kg/day p.o.) or placebo beginning 4 days before application of elastase and continuing through the experiments. Sham-operated hamsters received saline application onto the abdominal aorta. Two weeks after application of elastase, the aortic diameter in the placebo-treated group was significantly increased to 1.6-fold compared with the value for the sham-operated group, whereas that in the NK3201-treated group was significantly reduced. The chymase activities in the sham-operated and the placebo-treated groups were 0.35 +/- 0.01 and 3.44 +/- 0.62 mU/mg protein, respectively, and this difference was significant. NK3201 significantly reduced the chymase activity in the placebo-treated group. Here, we demonstrated for the first time that a chymase inhibitor prevented the development of abdominal aortic aneurysm in a hamster experimental model.

Acetamides↗
PubMed2004

Significance of chymase inhibition for prevention of adhesion formation.

To clarify the role of chymase in adhesion formation, we investigated whether a chymase inhibitor could prevent adhesion formation after surgery in hamsters. Hamsters received a lesion produced by uterus scraping. A specific chymase inhibitor, 2-[4-(5-fluoro-3-methylbenzo[b]thiophen-2-yl)sulfonamido-3-methanesulfonylphenyl]oxazole-4-carboxylicacid (TY-51184), or placebo was injected into the abdomen before closing and scores for adhesion formation were assessed at 1, 4, and 12 weeks. A single peritoneal administration of TY-51184 significantly decreased the adhesion scores even at 12 weeks (placebo, 2.80+/-0.20; chymase inhibitor, 1.60+/-0.31). Thus, chymase inhibitors may be a novel strategy to prevent adhesion formation.

Animals↗
PubMed2004

An antiarrhythmic effect of a chymase inhibitor after myocardial infarction.

Chymase plays an important role in the regulation of local angiotensin (Ang) II formation in the cardiac tissue. We recently found that cardiac chymase was activated significantly and survival rate markedly improved by treatment with chymase inhibitors after myocardial infarction (MI) in hamsters. However, the mechanisms for this effect have not been established. Because lethal arrhythmias are generally believed to contribute to sudden cardiac death, we assessed whether inhibition of cardiac chymase would provide an antiarrhythmic effect during the 8-h ischemic period after 2-[4-(5-fluoro-3-methylbenzo-[b]thiophen-2-yl)sulfonamide-3-methanesulfonylphenyl]oxazole-4-carboxylicacid (TY51184) (a specific chymase inhibitor, 1 mg/kg i.v.) treatment by ligation of left anterior descending coronary artery (LAD) in dogs. Effects of candesartan (an Ang II type 1 receptor antagonist, 1 mg/kg i.v.) in this model were also assessed. Total Ang II-forming activity and chymase activity in the infarcted heart were increased significantly 8 h after LAD ligation. A time-dependent elevation of Ang II in plasma was also observed. A decrease in plasma Ang II levels after TY51184 treatment occurred concomitantly with suppression of cardiac chymase activity. LAD ligation resulted in a large number of ventricular arrhythmias (VAs). TY51184 and candesartan treatments largely suppressed the appearance of VAs, and the efficacy of the two agents was similar. These findings demonstrate that chymase inhibition can provide an antiarrhythmic effect after MI, and the reduction of Ang II by TY51184 may be mainly responsible for this beneficial effect. An antiarrhythmic effect of chymase inhibitors may contribute to reductions in the mortality rate during the acute phase after MI.

Animals↗
PubMed2004

Pivotal role of nuclear factor kappaB signaling in anti-CD40-induced liver injury in mice.

Nuclear factor kappaB (NF-kappaB) has a central role in coordinating the expression of a wide variety of genes that control immune responses and is also recognized as an antiapoptotic transcription factor. Here, we focused on the role of the NF-kappaB signaling pathway in the interaction between inflammatory cells and hepatocytes in liver inflammation. We found that pretreatment of mice with adenoviruses expressing a mutant form of the inhibitor kappaB superrepressor (Ad5IkappaB), a NF-kappaB inhibitor, reduced the migration of inflammatory cells and cytokine and chemokine expression in the liver 12 hours after a single intravenous injection of an anti-CD40 antibody (alphaCD40) compared with mice infected with control adenoviruses (Ad5LacZ). We also confirmed reductions in cytokine production by macrophages, T cells, and natural killer (NK) cells in the liver of Ad5IkappaB-treated mice by FACS analysis. However, alphaCD40 treatment in Ad5IkappaB-infected mice induced elevation of serum alanine aminotransferase at 24 hours, and the liver injury was associated with massive hepatocyte apoptosis. Furthermore, interferon gamma (IFN-gamma) production by NK cells and T cells was increased and stimulated tumor necrosis factor alpha (TNF-alpha) production by macrophages in the Ad5IkappaB-infected liver. Moreover, the liver injury was completely suppressed by the administration of anti-IFN-gamma and anti-TNF-alpha. These results suggest that inhibition of NF-kappaB activity suppressed alphaCD40-induced liver inflammation at an early phase, resulting in a reduction in cytokine and chemokine production, whereas it sensitized hepatocytes to TNF-alpha-induced apoptosis and exacerbated liver injury at the late phase. In conclusion, NF-kappaB exerts pivotal activities at inflammatory sites, and caution should be exercised in NF-kappaB-targeted therapy of liver disease.

Animals↗
PubMed2004

Leflunomide protects from T-cell-mediated liver injury in mice through inhibition of nuclear factor kappaB.

Leflunomide is a novel immunosuppressive and anti-inflammatory agent for the treatment of autoimmune disease. The aim of this study was to investigate whether leflunomide protects from liver injury induced by concanavalin A (Con A), a T-cell-dependent model of liver damage. BALB/c mice were injected with 25 mg/kg Con A in the presence or absence of 30 mg/kg leflunomide. Liver injury was assessed biochemically and histologically. Levels of circulating cytokines and expressions of cytokine messenger RNA (mRNA) in the liver and the spleen were determined. Treatment with leflunomide markedly reduced serum transaminase activities and the numbers of dead liver cells. Leflunomide significantly inhibited increases in plasma tumor necrosis factor alpha (TNF-alpha) and interleukin 2 concentrations, and also reduced TNF-alpha mRNA expression in the liver after administration of Con A. These findings were supported by the results in which leflunomide administration decreased the number of T lymphocytes infiltrating the liver as well as inhibiting their production of TNF-alpha. Activation of nuclear factor kappaB (NF-kappaB), which regulates TNF-alpha production, was inhibited in the liver of mice treated with leflunomide, resulting in a reduction of TNF-alpha production from lymphocytes infiltrating the liver. In conclusion, leflunomide is capable of regulating T-cell-mediated liver injury in vivo and that this event may depend on the decrease of TNF-alpha production in the liver through inhibition of NF-kappaB activation caused by leflunomide.

Animals↗
PubMed2004

Effect of chymase-dependent transforming growth factor beta on peritoneal adhesion formation in a rat model.

PURPOSE: To clarify the role of chymase produced from mast cells, which are closely related to adhesion formation, we investigated the preventive effect of a chymase inhibitor on adhesion formation in a rat model. METHODS: A lesion was created in rats by uterus scraping, and a chymase inhibitor, Suc-Val-Pro-Phep(OPh)2 (10 microM), or a placebo was injected into the abdomen. The level of transforming growth factor beta (TGF-beta) in the peritoneal fluid was also measured. RESULTS: By 2 weeks after the operation, the scores for adhesion formation in the chymase inhibitor-treated group were significantly lower than those in the placebo-treated group, at 1.64 +/- 0.34 and 3.27 +/- 0.19, respectively (P < 0.01). After scraping the uterus, the level of TGF-beta in the peritoneal fluid was significantly higher in the placebo-treated group, whereas it was significantly suppressed by the chymase inhibitor. CONCLUSIONS: Chymase may play an important role in adhesion formation aided by TGF-beta.

Animals↗
PubMed2004

Role of chymase on growth of cultured canine Tenon's capsule fibroblasts and scarring in a canine conjunctival flap model.

Chymase is a chymotrypsin-like serine protease contained in the secretory granules of mast cells. Recently, we reported that chymase activity and the number of chymase-positive mast cells in conjunctival tissues were significantly increased during the wound healing process in a hamster model of glaucoma surgery. However, it has been unclear the role of chymase on conjunctival scarring. In the present study, we evaluated the effect of dog chymase on cell proliferation of fibroblasts established from canine Tenon's capsule and the effect of a chymase inhibitor on scarring in a canine conjunctival flap model. After a fibroblast cell culture was established from canine Tenon's capsules, the fibroblasts were incubated in the presence of dog chymase (5-20 ng ml(-1)). Cell proliferation was evaluated by bromodeoxyuridine incorporation. In a canine conjunctival flap model, a sponge treated with a chymase inhibitor, Suc-Val-Pro-Phe(P)(OPh)(2), or placebo was placed in between the conjunctiva and sclera and the conjunctival incision was closed. One week after the surgery, adhesion degree was assessed, and chymase activities in the conjunctival lesion and in the areas of the conjunctiva and sclera were measured. In cultured canine Tenon's capsule fibroblasts, dog chymase significantly increased cell proliferation, and this chymase-dependent proliferation was completely suppressed by the chymase inhibitor. In the canine surgical model, chymase activity in placebo-treated eyes was significantly increased compared to control eyes, while it was significantly decreased by treatment with the chymase inhibitor. Scores for adhesion degree in the chymase inhibitor-treated eyes were significantly decreased in comparison with those in placebo-treated eyes. The conjunctival area in the chymase inhibitor-treated eyes was also suppressed to 52.6% compared with that in placebo-treated treated eyes. In conclusion, chymase stimulates proliferation of fibroblasts derived from canine Tenon's capsule and chymase may play an important role in scarring after glaucoma surgery.

Animals↗
PubMed2004

Chymase as a novel target for the prevention of vascular diseases.

In vascular tissues, chymase catalyzes the production of angiotensin II, which plays a crucial role in vascular diseases. Recent clinical studies and animal models of vascular proliferation and atherosclerosis have provided evidence that angiotensin II formed by chymase is involved in these processes. These observations suggest that chymase might promote the development of vascular proliferation and atherosclerosis. Chymase also activates matrix metalloproteinase 9, which promotes aortic aneurysm and angiogenesis, and thus chymase inhibitors might also prevent the progression of abdominal aortic aneurysm and angiogenesis. We propose that chymase is a novel target for preventing vascular diseases.

Animals↗
PubMed2004

Attenuation of adhesion formation after cardiac surgery with a chymase inhibitor in a hamster model.

OBJECTIVE: Chymase is one of the inflammatory mediators and is released from mast cells, which are closely associated with adhesion formation. Chymase also activates transforming growth factor beta1, which promotes tissue fibrosis. However, the role of chymase in cardiac adhesion formation has not yet been elucidated. We have assessed whether a specific chymase inhibitor, Suc-Val-Pro-Phe(p) (OPh)(2), prevents postoperative cardiac adhesions in hamsters. METHODS: In 66 hamsters the epicardium was abraded, and then either chymase inhibitor or placebo was injected into the left thoracic cavity, leaving the pericardium open. Cardiac chymase activity, the level of transforming growth factor beta1 in the pleural fluid, and the density of epicardial mast cells were measured 3 days postoperatively. The degree of adhesion formation was evaluated macroscopically and histologically 2 weeks postoperatively by using a grading score ranging from 0 (no adhesions) to 4 (severe adhesions). RESULTS: The cardiac chymase activity and level of transforming growth factor beta1 were lower in the chymase inhibitor-treated group compared with in the placebo-treated group (45.8 +/- 18.7 vs 79.7 +/- 13.7 microU/mg protein [P <.025] and 15.6 +/- 6.5 vs 33.2 +/- 9.8 microg/mL [P <.01], respectively). The density of mast cells was higher in the placebo-treated group, and there was suppression to 60% of this value in the chymase inhibitor-treated group. The adhesion scores were lower in the chymase inhibitor-treated group compared with in the placebo-treated group (1.3 +/- 1.3 vs 3.0 +/- 1.1, P <.01). CONCLUSION: Use of a chymase inhibitor suppresses not only cardiac chymase activity but also the level of transforming growth factor beta1, and this results in a reduction in postoperative cardiac adhesion.

Animals↗
PubMed2004

Tear chymase in vernal keratoconjunctivitis.

PURPOSE: To determine the levels of mast cell chymase and tryptase activity in the tears of patients with vernal keratoconjunctivitis (VKC). METHODS: Subjects were 38 VKC patients and 18 healthy controls whose chymase and tryptase activity in tears was measured by enzyme assay. VKC severity was quantified based on the following clinical signs: papillary hypertrophy, conjunctival hyperemia, edema, punctate keratitis, Trantas dots, and mucus production. Of the 38 VKC patients, the degree of disease severity was mild in 13, moderate in 18, and severe in 7. RESULTS: Mean chymase activity and standard deviation in tears was 0.23+/-0.07mU in mild VKC, 0.68+/-0.22mU in moderate VKC, 1.91+/-0.71 mU in severe VKC, and 0.11+/-0.05 mU in healthy controls. The increase in all VKC stages was statistically significant compared to that in healthy control. The degree of chymase activity in tears correlated significantly with VKC severity (r = 0.9245, p < 0.001). High tryptase activity was also detected in the tears of VKC patients, although increased tryptase activity in tears did not correlate with disease severity (r = 0.1999). CONCLUSIONS: Chymase activity in tears may thus be a sensitive marker for determining the severity of VKC.

Adolescent↗
PubMed2004

Necrotic death of Rhodococcus equi-infected macrophages is regulated by virulence-associated plasmids.

Rhodococcus equi is a gram-positive intracellular pathogen that can cause severe bronchopneumonia in foals and AIDS patients. It has been reported that advanced infection of foals is characterized by tissue necrosis, coinciding with the presence of degenerate bacteria-laden macrophages. Here, we report that the possession of the VapA-expressing plasmid, which has been previously correlated with a high level of virulence for foals and mice, strongly increases cytotoxicity of R. equi for murine macrophage-like (J774E) cells. Isolates containing different, VapB-expressing plasmids are less virulent and also have a lower cytotoxic potential. Isogenic strains lacking either plasmid are avirulent and have a very low cytotoxic potential. We show, using fluorescence-activated cell sorter analysis (annexin V/7-amino-actinomycin D and sub-G1-analysis), Western blotting [poly(ADP-ribose) polymerase processing analysis], and electron microscopy (macrophage and nucleus morphologies) that the deaths of murine macrophages are the result of necrotic rather than apoptotic events. We demonstrate that the bacteria must be alive in order to act cytotoxic. Therefore, one effect of the virulence-associated plasmids during infection with R. equi is the promotion of necrotic damage to the host.

Animals↗
PubMed2004

Chymase-derived angiotensin II and arrhythmias after myocardial infarction.

Mast cell-derived chymase seems to be important in the regulation of local angiotensin (A) II formation in cardiovascular tissues. In human heart, chymase accounts for 80% of A II formation. Therefore, the chymase-dependent A II pathway may play an important role in the pathogenesis of A II-related cardiovascular diseases. For example, cardiac chymase was activated earlier than ACE and this activation lasted longer than that of ACE after myocardial infarction (MI) in hamsters. Treatment with a specific chymase inhibitor treatment, but not an ACE inhibitor, improved post-MI survival as well as cardiac function and the extent of the beneficial effects was similar to that observed for an AT1-receptor antagonist treatment in this model. The survival benefit after MI seems to be related to an antiarrhythmic effect of the chymase inhibitor because chymase inhibition reduces the incidence of ventricular arrhythmias during periods of heart ischemia in a dog MI model. Thus, an antiarrhythmic effect of the chymase inhibitor may contribute to a reduction in mortality rate during the acute phase after MI.

Angiotensin II↗
PubMed2004

A single treatment with a specific chymase inhibitor, TY-51184, prevents vascular proliferation in canine grafted veins.

In this study, we evaluated whether a specific chymase inhibitor, TY-51184 (2-[4-(5-fluoro-3-methylbenzo[b]thiophen-2-yl)sulfonamido-3-methanesulfonylphenyl]oxazole-4-carboxylicacid), prevents the vascular proliferation in canine grafted veins. In the placebo-and chymase inhibitor-treated groups, the external jugular vein was infiltrated with saline and 10 microM TY-51184, respectively, and then it was grafted to the ipsilateral carotid artery. The non-surgical dogs were used as the control group. By 28 days after grafting, the chymase and ACE activities were significantly increased in the injured arteries. TY-51184 significantly reduced the chymase activity in the grafted veins, while it did not affect the ACE activity. The intimal areas in the placebo- and TY-51184-treated groups were 3.32 +/- 0.16 and 1.96 +/- 0.52 mm(2), respectively, and this difference was significant. The ratios of intimal area to medial area in the placebo- and TY-51184-treated groups were 66.8 +/- 3.5% and 34.9 +/- 9.2%, respectively, and this difference was also significant. There was a significant relationship between vascular proliferation and chymase activity, but not ACE activity. In this study, we demonstrated that a single treatment with a specific chymase inhibitor, TY-51184, could prevent the vascular proliferation in canine grafted veins.

Animals↗