Biomedical subjects

M Hirota

Publications and source records attributed to M Hirota.

At least 19 recordsLinked to original sources

PubMed2007

Development of leiomyosarcoma of the uterus in MMTV-CR-1 transgenic mice.

Overexpression of Cripto-1 (CR-1) in FVB/N mice using the MMTV-LTR promoter results in increased mammary tumourigenesis in these female transgenic mice (MMTV-CR-1). Here, we characterize uterine tumours that developed in 15/76 (19.7%) of MMTV-CR-1 female nulliparous or multiparous mice during 24 months of observation compared with 0/33 (0%) of FVB/N normal control mice observed during the same time period (p < 0.01). The uterine tumours collected from the MMTV-CR-1 mice were classified as leiomyosarcomas and found to express the CR-1 transgene by polymerase chain reaction analysis and immunohistochemistry. Detection by western blot analysis showed higher levels of phosphorylated (P) forms of c-src, Akt, GSK-3beta, and dephosphorylated (DP)-beta-catenin in lysates from MMTV-CR-1 uterine leiomyosarcomas in comparison with lysates from normal control FVB/N uteri. Immunostaining showed increased nuclear localization of beta-catenin in the MMTV-CR-1 uterine leiomyosarcomas. Increased immunostaining for CR-1 was detected in 9/13 (69.2%) cases of human leiomyosarcoma compared with staining in 3/15 (20%) human leiomyoma sections. Stronger immunostaining for P-src, P-Akt, P-GSK-3beta and increased nuclear localization of beta-catenin was also seen in human leiomyosarcomas in comparison with leiomyomas. Normal human uterine smooth muscle (UtSM) cells treated with exogenous soluble rhCR-1 showed increased levels of P-src, P-Akt, P-GSK-3beta and dephosphorylated (DP)-beta-catenin and increased proliferation (p < 0.05) and migration (p < 0.01) in comparison with untreated control UtSM cells. Inhibitors against c-src, Akt or beta-catenin, individually or in combination, significantly reduced CR-1-induced migration. These results suggest a role for CR-1 during uterine tumourigenesis either directly by activating c-src and Akt and/or via cross-talk with the canonical Wnt signalling pathway, as suggested by the increased expression of P-GSK-3beta, DP-beta-catenin, and increased nuclear localization of beta-catenin in human and MMTV-CR-1 mice leiomyosarcomas.

Animals↗

PubMed2006

Anti-inflammatory sesquiterpenes from Curcuma zedoaria.

From the methanolic extract of the rhizome of Curcuma zedoaria, we isolated anti-inflammatory sesquiterpene furanodiene (1) and furanodienone (2) along with new sesquiterpene compound 3 and known eight sesquiterpenes, zederone (4), curzerenone (5), curzeone (6), germacrone (7), 13-hydroxygermacrone (8), dehydrocurdione (9), curcumenone (10), and zedoaronediol (11). Their structures were elucidated on the basis of spectroscopic data. The anti-inflammatory effect of isolated components on 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced inflammation of mouse ears were examined. Compounds 1 and 2 suppressed the TPA-induced inflammation of mouse ears by 75% and 53%, respectively, at a dose of 1.0 micromol. Their activities are comparable to that of indomethacin, the normally used anti-inflammatory agent.

Animals↗

PubMed2005

Development of an in vitro skin sensitization test using human cell lines; human Cell Line Activation Test (h-CLAT). II. An inter-laboratory study of the h-CLAT.

Recent regulatory changes have placed a major emphasis on in vitro safety testing and alternative models. In regard to skin sensitization tests, dendritic cells (DCs) derived from human peripheral blood have been considered in the development of new in vitro alternatives. Human cell lines have been also reported recently. In our previous study, we suggested that measuring CD86 and/or CD54 expression on THP-1 cells (human monocytic leukemia cell line) could be used as an in vitro skin sensitization method. An inter-laboratory study among two laboratories was undertaken in Japan in order to further develop an in vitro skin sensitization model. In the present study, we used two human cell lines: THP-1 and U-937 (human histiocytic lymphoma cell line). First we optimized our test protocol (refer to the related paper entitled "optimization of the h-CLAT protocol" within this journal) and then we did an inter-laboratory validation with nine chemicals using the optimized protocol. We measured the expression of CD86 and CD54 on the above cells using flow cytometry after a 24h and 48h exposure to six known allergens (e.g., DNCB, pPD, NiSO(4)) and three non-allergens (e.g., SLS, tween 80). For the sample test concentration, four doses (0.1x, 0.5x, 1x, and 2x of the 50% inhibitory concentration (IC(50))) were evaluated. IC(50) was calculated using MTT assay. We found that allergens/non-allergens were better predicted using THP-1 cells compared to U-937 cells following a 24 h and a 48 h exposure. We also found that the 24h treatment time tended to have a better accuracy than the 48 h treatment time for THP-1 cells. Expression of CD86 and CD54 were good predictive markers for THP-1 cells, but for U-937 cells, expression of CD86 was a better predictor than CD54, at the 24h and the 48 h treatment time. The accuracy also improved when both markers (CD86 and CD54) were used as compared with a single marker for THP-1 cells. Both laboratories gave a good prediction of allergen/non-allergen, especially using THP-1 cells. These results suggest that our method, human Cell Line Activation Test (h-CLAT), using human cell lines THP-1 and U-937, but especially THP-1 cells at 24h treatment, may be a useful in vitro skin sensitization model to predict various contact allergens.

Allergens↗

PubMed2005

Development of an in vitro skin sensitization test using human cell lines: the human Cell Line Activation Test (h-CLAT). I. Optimization of the h-CLAT protocol.

The aim of this study is to optimize the experimental conditions for an in vitro skin sensitization test using the human cell lines THP-1 and U-937. As regards pre-culturing time, the expression of CD86 on DNCB-treated THP-1 cells tended to be higher after 48h and 72h pre-culture compared with other time points evaluated. Next, we investigated the effect of chemical treatment time, and found that induction of CD86 expression on THP-1 cells by DNCB reached a plateau after 24h. Augmentation of CD86 expression is often observed when cells are treated with a subtoxic dose of allergens. To determine the appropriate dose of test samples, the cytotoxicity of test samples to THP-1 and U-937 cells was assessed with MTT assay, and the 50% inhibitory concentration (IC50) of each test sample was calculated. Based on the cytotoxicity assay data, four concentrations in the range between toxic and non-toxic were selected (0.1x, 0.5x, 1x and 2x IC50). Several kinds of antibodies were tested for staining THP-1 and U-937 cells treated with allergens/non-allergens (e.g., DNCB, Ni/SLS), and suitable antibodies for staining CD86 and CD54 were selected. We confirmed that the working dilutions of the selected CD86 and CD54 antibodies were appropriate for use in our method. The effect of an FcR blocking procedure was also evaluated. The mean fluorescence intensity (MFI value) was decreased by the FcR blocking procedure, which indicated that non-specific staining was blocked. Therefore, this procedure should be included in the method. Based on our findings, the protocol for this assay was optimized and the experimental conditions to be used in a future validation study were identified. We propose to call this kind of in vitro skin sensitization test h-CLAT, which is short for human Cell Line Activation Test.

Allergens↗

PubMed2005

Involvement of MMP-7 in invasion of pancreatic cancer cells through activation of the EGFR mediated MEK-ERK signal transduction pathway.

AIMS: To clarify the involvement of matrix metalloproteinase-7 (MMP-7) in cell dissociation and the subsequent invasion of pancreatic cancer cells. METHODS: Western blotting, in vitro invasion assay, immunocytochemistry, and immunohistochemistry were performed in pancreatic cancer cell lines or pancreatic cancer tissue. RESULTS: The active form of the MMP-7 protein was expressed exclusively in the conditioned medium of dissociated (PC-1.0 and AsPC-1) pancreatic cancer cells, whereas proMMP-7 protein was only detected in the conditioned medium of non-dissociated (PC-1 and Capan-2) cells. Both intracellular and conditioned medium localised MMP-7 was greatly reduced by treatment with the epidermal growth factor receptor (EGFR) inhibitor AG1478 and the mitogen activated protein kinase kinase (MEK) inhibitor U0126 in pancreatic cancer cells. MMP-7 treatment significantly induced the disruption of tight junction (TJ) structures and subsequent cell dissociation, and activation of the EGFR mediated MEK- ERK (extracellular signal regulated protein kinase) signalling pathway in the non-dissociated pancreatic cancer cells. Moreover, the strong in vitro invasiveness of dissociated cells was inhibited by AG1478 and U0126 treatment, whereas the weak invasiveness of non-dissociated cells was apparently induced by MMP-7 treatment. In addition, MMP-7 expression was stronger at the invasive front than at the centre of human pancreatic tumours. CONCLUSION: MMP-7 is involved in cell dissociation and the subsequent invasion of pancreatic cancer cells. It induces the disruption of TJ structures and forms a positive feedback loop with activation of the EGFR mediated MEK-ERK signalling pathway.

Adenocarcinoma↗

PubMed2005

Hepatic portal venous gas associated with nonocclusive mesenteric ischemia in a hemodialysis patient.

Hepatic portal venous gas (HPVG) has been rarely described in chronic hemodialysis patients. We report a case of HPVG in a 59-year-old female patient with hemodialysis-dependent chronic renal failure due to diabetes who presented with acute onset of abdominal pain. Abdominal CT demonstrated the presence of gas in the portal veins. However, on laparotomy, no evidence of bowel necrosis or perforation could be found. HPVG seemed to be caused by nonocclusive mesenteric ischemia (NOMI), an increasingly recognized complication in hemodialysis patients. The patient responded favorably to intravenous hyperalimentation and antibiotics.

Diagnosis, Differential↗

PubMed2004

Critical progressive small-graft injury caused by intrasinusoidal pressure elevation following living donor liver transplantation.

In adult-to-adult living liver transplantation, small-for-size graft syndrome sometimes occurs. The relationship between the hemodynamic changes and histologic findings has not been studied in patients with failure of small-for-size grafts. We analyzed the relationship between the postoperative hemodynamic changes and pathologic findings in patients with small-for-size grafts that ended in graft failure. From March 1999 to December 2002, adult-to-adult living-donor liver transplantation with small-size grafts (graft volume/standard liver volume less than 40%) was performed in eight patients. Three patients died from graft failure caused by overperfusion, which was diagnosed from pathologic findings. We analyzed the relation between hepatic hemodynamic parameters, such as portal venous blood velocity or splenic arterial pulsatility index, and histologic changes in patients with graft failure. Severe portal hyperperfusion (90 cm/sec at the umbilical portion) was observed on postoperative day 1. Among patients with graft failure, critical hemodynamic changes, such as sudden onset of extremely deteriorated portal venous blood flow, occurred during the early postoperative period (postoperative day 5, 3, 6, respectively). Histologic examination revealed vacuolar changes in the cytoplasm of hepatocytes, and submassive necrosis indicated intrasinusoidal pressure elevation. These changes were not observed in the biopsy obtained soon after reperfusion. In conclusion, critically decreased vascular beds may cause intrasinusoidal pressure elevation and sinusoidal circulatory disturbances.

Adult↗

PubMed2003

Hemichrome formation observed in human haemoglobin A under various buffer conditions.

AIM: To observe hemichrome formation in human haemoglobin A under various buffer conditions. METHOD: Hemichrome formation of human oxyhaemoglobin A (HbO2) was studied spectrophotometrically in 0.1 m buffer at various temperatures and pH values. RESULTS: Following autoxidation in ferrous HbO2, it was evident that formation of hemichrome, which tends to precipitate, occurred at various stages during the course of the autoxidation reaction namely at initial, intermediate or final stages, depending on temperature and pH of the solution. By varying temperature of the solution from 35 to 55 degrees C and pH from 4.5 to 10.5, it is shown here that HbO2 exhibits high susceptibility for hemichrome formation and its occurrence is a function of pH, temperature and progress of autoxidation of HbO2. Unlike HbO2 and its separated haemoglobin chains, monomeric bovine heart myoglobin (MbO2) did not easily form hemichrome. CONCLUSION: These findings provide a clue on the crucial role of haemoglobin molecule for senescent cell recognition or homeostasis in the blood circulation.

Animals↗

PubMed2003

Novel OK-432-conjugated tumor vaccines induce tumor-specific immunity against murine tongue cancer.

Priming with tumor antigens is one of the most important strategies in cancer immunotherapy. To enhance tumor antigenicity, OK-432, a streptococcal preparation, was coupled to squamous cell carcinoma (KLN-205) by means of a 0.2% glutaraldehyde method. The purpose of this study was to investigate whether OK-432-conjugated tumor vaccines could induce tumor-specific immunity. Our originally developed mouse tongue cancer model was used throughout this work for the analysis of antitumor effects. Prepared OK-432-conjugated KLN-205 vaccines were immunized 3 times to DBA/2 mice. The results showed that the KLN-205 vaccines induced cytolytic activity and strongly suppressed both KLN-205 tumor incidence and growth, and survival of the mice was improved. Moreover, the histological results showed that a greater number of lymphocytes had infiltrated around tumor cells by 24 hours after tumor inoculation in the vaccine group. These results suggest that immunizations with KLN-205 vaccines increase the antitumor effects against tongue cancer.

Animals↗

PubMed2002

In vivo thyroid 131I monitoring system using an imaging plate.

The counting efficiency and detection limit of an in vivo thyroid 131I monitoring system using an imaging plate (IP) were estimated using an anthropomorphic thyroid neck phantom. The counting efficiency of the IP system was approximately constant for thyroid volumes between 11.7 and 20 ml and neck diameters between 10 and 14 cm. The detection limits were distributed from 288 to 451 Bq depending on the combination of neck diameter, thyroid volume and tissue thickness. The IP monitoring system gives a reliable counting efficiency notwithstanding the variation of thyroid volume and neck diameter. The IP system is a new option for thyroid 131I monitoring.

Humans↗

PubMed2001

Differential uptake of grepafloxacin by human circulating blood neutrophils and those exudated into tissues.

The uptake of the antimicrobial quinolone agent, grepafloxacin, both by human circulating blood neutrophils and by those exudated into tissues, was evaluated in vitro by comparing the intracellular drug concentrations. In circulating blood neutrophils, the uptake of grepafloxacin was rapid and saturable at 37 degrees C. The uptake of grepafloxacin into circulating blood neutrophils was reduced by lowering the environmental temperature or by the presence of metabolic inhibitors, suggesting the involvement of an active transport mechanism. Furthermore, the uptake of grepafloxacin by tissue (salivary) neutrophils was also partially temperature-dependent and was significantly greater than that by circulating blood neutrophils, i.e. exudation of neutrophils into tissue results in a markedly enhanced transport mechanism for grepafloxacin. This phenomenon may be related to the higher defense activity against infection seen in exudated tissue neutrophils.

Anti-Infective Agents↗

PubMed2001

Expression of survivin is correlated with cancer cell apoptosis and is involved in the development of human pancreatic duct cell tumors.

BACKGROUND: Survivin is a new member of the inhibitor of apoptosis family of antiapoptotic proteins. This protein was expressed selectively in all the most common human carcinomas but not in normal adult tissues. To our knowledge, the relation between survivin expression and apoptosis or tumorigenesis has not yet been studied in pancreatic neoplasms. METHODS: The authors investigated the expression of survivin in 4 pancreatic carcinoma cell lines and 56 human pancreatic tissues (5 cases of normal, 12 cases of chronic pancreatitis [CP], 26 cases of pancreatic duct cell adenocarcinoma [PDC], 16 lesions of 13 intraductal papillary-mucinous tumor [IPMT] by immunohistochemistry, immunoblotting, and reverse transcription-polymerase chain reaction (RT-PCR) to examine the association of its expression with tumor apoptosis and/or tumorigenesis. RESULTS: Survivin expression was found in the tumor cells but not in the nonneoplastic pancreatic tissues (normal and CP tissues). Survivin expression was observed in 20 of 26 cases of PDC (76.9%) and in 9 of 16 IPMT lesions that ranged from adenoma to invasive (56.3%) by immunohistochemistry. Survivin was more frequently expressed in malignant tumors than in benign tumors (P = 0.0089). In PDC, high levels of survivin expression were associated significantly with a reduction in the apoptotic index of the tumor cells (0.445% +/- 0.150% vs. 0.961% +/- 0.378%; P < 0.0001). CONCLUSIONS: These data suggest that the expression of survivin may be upregulated during an early step of tumorigenesis and during the development of cancer by reducing the cancer cell apoptosis.

Adenocarcinoma↗

PubMed2001

Genetic mutations in exons 3 and 4 of the pancreatic secretory trypsin inhibitor in patients with pancreatitis.

PURPOSE: We hypothesized that mutations in the pancreatic secretory trypsin inhibitor (PSTI) gene could promote autodigestion, leading to acute or chronic pancreatitis. Our investigation involved mutation analysis of the PSTI gene in patients with acute or chronic pancreatitis. METHODS: Mutation analysis for the PSTI gene was performed in patients with acute or chronic pancreatitis. Unrelated healthy volunteers and family members of a chronic pancreatitis patient with point mutations in the PSTI gene were also analyzed. RESULTS: Two types of single-point mutation in the PSTI gene were observed in one patient with chronic pancreatitis: 34Asn (AAT)-to-Ser (AGT) (101 A > G N34S: N34S) in exon 3, and 67Arg (CGC)-to-Cys (TGC) (199 C > T R67C: R67C) in exon 4. No mutations with amino-acid substitution were found in other patients or in the volunteer group. In the patient with the PSTI gene mutations, no additional mutations were observed in the cationic trypsinogen gene. The family study revealed that the mother and a maternal uncle were homozygotes for the N34S mutation, while the father and brother were compound heterozygotes for the N34S and R67C mutations. The uncle (N34S/N34S) showed clinical manifestations of pancreatitis, but the other family members did not. CONCLUSIONS: The N34S mutation may cause a predisposition to pancreatitis, with incomplete penetrance. However, with the limited information available, it is not known whether the R67C mutation promotes pancreatitis.

Adolescent↗

PubMed2001

A new procedure of percutaneous microwave coagulation therapy under artificial hydrothorax for patients with liver tumors in the hepatic dome.

Percutaneous microwave coagulation therapy (PMCT) has been widely used as an effective minimal invasive therapy for small liver tumors. The occurrence of a sonographic masked space due to the presence of the lung, however, has become a major obstacle to visualizing the whole tumor in the hepatic dome. To facilitate the use of PMCT for liver tumors in the hepatic dome, we developed PMCT in combination with the artificial hydrothorax method (percutaneous transdiaphragmatic MCT: PTD-MCT). Our new approach for PMCT to the hepatic tumors located in Couinaud's segments VIII or VII just under the diaphragm resulted in a successful treatment. The separation of the lung from the diaphragm by the infusion of saline into the pleural cavity enabled us not only to visualize the whole tumor in the hepatic dome to accurately target the tumor, but also helped us to avoid injuring the lung. PTD-MCT is therefore strongly recommended for the treatment of liver tumors in the hepatic dome.

Aged↗

PubMed2001

Comparison of antimycobacterial activity of grepafloxacin against Mycobacterium avium with that of levofloxacin: accumulation of grepafloxacin in human macrophages.

The bactericidal activity of two new quinolones, grepafloxacin and levofloxacin, against five strains of Mycobacterium avium was investigated in vitro. The minimum inhibitory concentrations (MICs) of these two quinolones, determined by the broth microdilution method, were comparable for all strains tested. In contrast, grepafloxacin suppressed the intracellular growth of all the strains in monocyte-derived macrophages more strongly than levofloxacin, when the cells infected with these strains were incubated for 7 days in the presence of various concentrations of the two new quinolones. To find the reason for the strengthened intracellular killing activity of grepafloxacin, we determined the ratio of the concentration of the new quinolones in the cells to that in the medium (C/M concentration ratio). The C/M concentration ratio of grepafloxacin was increased to 34.7 by 7 days, whereas that of levofloxacin at 7 days was only 12.3. These data suggested that a higher level of intraphagocytic accumulation of grepafloxacin endows it with greater mycobactericidal activity.

Anti-Infective Agents↗

PubMed2001

Feasibility of in vivo thyroid 131I monitoring with an imaging plate.

A new in vivo thyroid 131I monitoring method was devised by using an imaging plate (IP). A thyroid image obtained with a realistic neck-thyroid phantom showed a unique shape characteristic of the thyroid gland. A 131I thyroid imaging allows visual confirmation of thyroid accumulation of 131I. The detection limit of the IP system of 290 Bq was about 1/100 of the screening level of 30 kBq in cases of public emergencies. The IP system is applicable for thyroid 131I monitoring.

Humans↗

PubMed2001

Expression of cyclin-dependent kinase inhibitors in taste buds of mouse and hamster.

Taste buds are specialized epithelial cell clusters in the oral squamous cell epithelium. Although taste buds have been reported to renew rapidly, the mechanism of cell cycle control in these specialized structures remains unresolved. To clarify the cell cycle status and role of cyclin-dependent kinase inhibitors (CDKI) for cell cycle control in the taste buds, we analyzed cell proliferation activity using bromodeoxyuridine (BrdU) and Ki-67 immunostainings and the expression of the Cip/Kip family of CDKI (p21Cip1, p27Kip1, and p57Kip2) in the circumvallate papillae of mouse and hamster. BrdU-positive cells were detected in the basal layer of the oral epithelium. In the taste buds, Ki-67-positive cells were seen in the basal area, with only a very few positive cells in the taste buds. Both p21Cip1 and p27Kip1 positive cells were seen in the suprabasal layer of the non-gustatory oral epithelium. In the taste buds, stronger p27Kip1 staining was detected than in the non-gustatory epithelium. Western blotting analysis revealed that p27Kip1 was abundant in the mucosal tissues from circumvallate papillae. Thus, our study suggests that the taste bud cells except for basal cells are post-mitotic cells and that the cell cycle arrest associated with taste bud cell differentiation could be regulated predominantly by p27Kip1.

Animals↗

PubMed2001

Altered cytokine response in rat acute pancreatitis complicated with endotoxemia.

We demonstrated that the dynamic aspects of cytokine production in rat acute pancreatitis, which was induced by cerulein and aggravated by subsequent lipopolysaccharide (LPS) injection. A priming effect by induction of mild pancreatitis with cerulein enhanced the subsequent cytokine production by LPS injection. Alternatively, after induction of severe pancreatitis with cerulein and LPS, cytokine production was markedly suppressed for > or = 90 hours. Production of interleukin-2 (IL-2) by splenocytes decreased, and mortality rate after cecal ligation and puncture (CLP) increased significantly after induction of severe acute pancreatitis. These results suggest that the suppression of a cytokine response in severe acute pancreatitis may alter the defense system and, as a result, increase mortality after CLP.

Acute Disease↗