Biomedical subjects

H Nagano

Publications and source records attributed to H Nagano.

At least 55 records · Page 3Linked to original sources

PubMed2000

Functional analysis of the type III secretion system in enteropathogenic Escherichia coli O157:H45.

A mass outbreak of Escherichia coli O157:H45 was first reported in Japan in 1998. This pathogen was classified as an enteropathogenic E. coli (EPEC) O157 because it was characterized by the Shiga toxin gene (stx)-negative and bundle-forming pilus (bfp) gene-positive genotypes. In this study, we investigated the type III secretion system in EPEC O157. Although no type III secreted proteins, Esps (E. colisecreted proteins), in EPEC O157:H45 were detectable in culture supernatant, secreted proteins were induced by the introduction of an EPEC plasmid-encoded regulator, per. In further contrast to EHEC O157:H7, EPEC O157:H45 triggered the accumulation of tyrosine phosphorylated proteins beneath the adherent bacteria. These results suggest that regulation of the type III secretion apparatus and host signal transduction events between E. coli O157:H45 and O157:H7 are completely different.

Adhesins, Bacterial↗

PubMed2000

[Effect of thiabendazole (TBZ) on glutathione (GSH) and GSH related enzymes in mice liver].

The present study examines the effects of thiabendazole (TBZ), its metabolites, 5-hydroxythiabendazole (5-OH TBZ) and 2-acetylbenzimidazole (ABI), and structural related compounds, thiazoles and thioamides on glutathione (GSH) concentration and GSH-related enzymes in the livers of ICR 11 week-old female mice. GSH concentration in liver and kidney of mice given orally TBZ 0.65 mol/kg (TBZ group) increased significantly compared with control mice from 24 h to 48 h after administration of TBZ. Even in mice to which TBZ at 0.175 mol/kg was administered in combination with L-buthionine sulfoximine (BSO) 4 mmol/kg (i.p.) (BSO-TBZ group), kidney GSH showed significant increase compared with BSO-control mice 48 h after the administration of TBZ. gamma-Glutamylcysteine synthetase (gamma-GCS) activity in the livers of the TBZ group markedly increased at 48 h and that of BSO-TBZ group increased from 24 h to 48 h. gamma-GCS in mice liver is thus enhanced by TBZ regardless of BSO administration. Hepatic glutathione peroxidase activity of the TBZ group did not change in response to cumene hydroperoxide assubstrate. That of BSO-treated mice decreased by TBZ-coadministration and significant differences was noted between BSO-control and BSO-TBZ group from 1 h to 48 h later. Hepatic glutathione S-transferase (GST) activity toward 1,2-dichloro-4-nitrobenzene (DCNB) was significantly elevated 24 h after administrations of TBZ in TBZ and BSO-TBZ groups. GST activity toward 1,2-epoxy-3-(p-nitrophenoxy) propane of TBZ group increased from 0.5 h to 24 h. Hepatic GST activity toward DCNB and 1-chloro-2,4-dinitrobenzene did not change by administration of 0.65 mol/kg 5-OH TBZ or ABI but increased by administrations of 0.33 mol/kg of thiazole, 4-methylthiazole, 4,5-dimethylthiazole or 2,4-dimethylthiazole. Increase in GSH concentration and GST activity in mice liver by TBZ administration may be considered to provide protection from TBZ or its active metabolites.

Animals↗

PubMed2000

Purification of collagenase and specificity of its related enzyme from Bacillus subtilis FS-2.

A collagenase in the culture supernatant of B. subtilis FS-2, isolated from traditional fish sauce, was purified. The enzyme had a molecular mass of about 125 kDa. It degraded gelatin with maximum activity at pH 9 and a temperature of 50 degrees C. The purified enzyme was stable over a wide range of pH (5-10) and lost only 15% and 35% activity after incubation at 60 degrees C and 65 degrees C for 30 min, respectively. Slightly inhibited by EDTA, soybean tripsin inhibitor, iodoacetamide, and iodoacetic acid, the enzyme was severely inhibited by 2-beta-mercaptoethanol and DFP. The protease from B. subtilis FS-2 culture digested acid casein into fragments with hydrophilic and hydrophobic amino acids as C-terminals, in particular Asn, Gly, Val, and Ile.

Bacillus subtilis↗

PubMed2000

Assessment of Stanniocalcin-1 mRNA as a molecular marker for micrometastases of various human cancers.

We assessed Stanniocalcin-1 (STC-1) mRNA expression in human normal tissues, various types of human cancer cell lines, and cancer tissues obtained during surgery. Using a reverse transcriptase-polymerase chain reaction (RT-PCR) assay developed to detect STC-1 mRNA, the transcripts were detected in 20 out of 21 cancer cell lines and in all tumor tissues from various types of cancer. Semi-quantitative analyses with multiplex RT-PCR showed that STC-1 mRNA tended to be enhanced in cancer tissues of hepatocellular carcinoma and colorectal cancer compared to background cancer-free tissues. Analysis of blood samples obtained from 11 patients with hepatocellular carcinoma before, after and during hepatectomy showed STC-1 mRNA expression in 8 out of 11 patients at least one time. Normal donor blood samples (n=31) were all-negative for STC-1 mRNA expression. Our results indicate that STC-1 mRNA might be a useful molecular marker for detection of tumor cells in blood from patients with various types of malignancies.

Aged↗

PubMed2000

Outcome of pancreatic cancer patients based on genetic lymph node staging.

We prospectively assessed the clinical value of genetic staging of lymph node metastasis in patients with pancreatic adenocarcinoma who underwent curative surgery. K-ras gene mutations were detected in the primary tumors in 18 of 25 patients with pancreatic adenocarcinoma. Among these 18 patients, mutated K-ras gene was also found in at least one lymph node in 13 patients. Of these 13 patients, seven had no evidence of histological nodal involvement and six had histological lymph node metastasis. Although there was no significant difference in overall survival rates between the pathological node-negative and -positive patients, overall survival of the five patients with nodes-negative for the mutated K-ras gene were significantly better than that of the 13 patients with genetically metastasis-positive nodes (p<0.001). Furthermore, overall survival of the six patients with genetically metastasis-positive nodes limited to peripancreatic area was significantly better than that of seven patients with genetical metastasis in lymph nodes beyond the peripancreatic areas (p=0.018). These findings suggest that detection of K-ras gene mutations in lymph nodes may be clinically useful to assess the accurate tumor staging and to stratify the patient with pancreatic adenocarcinoma who are at high or low risk for recurrence after curative surgery.

Adenocarcinoma↗

PubMed2000

Expression of interferon alpha/beta receptor in human hepatocellular carcinoma.

Interferon-alpha (IFNalpha) plays a crucial role in the antiproliferation and immunoregulatory activity through the specific cell surface receptor, interferon-alpha/beta receptor (IFNalpha/betaR). We examined the immunohistochemical expression of IFNalpha/betaR in 91 hepatocellular carcinoma (HCC), HCV-related chronic hepatitis (n=38) and cirrhosis (n=53), dysplastic nodules (n=5), and normal liver (n=9). The level of IFNalpha/betaR increased in chronic hepatitis and cirrhosis compared with normal liver. All the dysplastic nodules showed moderate or high expression. In HCCs, 26% (24/91) of patients showed high IFNalpha/betaR expression while the remaining 38% (35/91) showed moderate, and 35% (32/91) no or faint expression. Clinicopathological survey demonstrated a significant correlation between IFNalpha/betaR expression and differentiation of carcinoma (P=0.0008) although there was no correlation between IFNalpha/betaR expression in HCC and survival or disease-free survival. Thus, IFNalpha/betaR was expressed not only in chronic hepatitis or liver cirrhosis but in HCC and its expression was significantly correlated with tissue differentiation of carcinoma.

Blotting, Western↗

PubMed2000

Genetic detection for micrometastasis in lymph node of biliary tract carcinoma.

The presence of regional lymph node metastasis is one of the most significant poor-prognosis factors in patients with biliary tract carcinoma. To establish a sensitive reverse transcription (RT)-PCR assay to detect micrometastases in lymph nodes of biliary tract carcinoma, we first investigated the optimal markers in biliary tract carcinoma. The expressions of the six candidates for a suitable RT-PCR marker [mammaglobin B, carcinoembryonic antigen (CEA), cytokeratin (CK) 20, prostate-specific antigen, and melanoma antigens (MAGE-1 and MAGE-3)] were evaluated in two bile duct cancer cell lines and human biliary tract carcinoma tissues. Of 32 carcinoma tissues, mammaglobin B, CEA, prostate-specific antigen, MAGE-1, MAGE-3, and CK 20 were expressed in 28 (88%), 26 (81%), 4 (13%), 5 (16%), 7 (22%), and 9 (28%), respectively. Mammaglobin B and CEA were considered to be good markers of the six candidates. We then examined 209 lymph nodes obtained from 15 patients with biliary tract carcinoma by RT-PCR assay using both mammaglobin B and CEA and compared the results with those of histological examination. All of 20 histologically positive lymph nodes for metastasis displayed the PCR product(s) of marker genes. Of 189 histologically negative nodes, 24 (13%) nodes expressed mammaglobin B and/or CEA mRNA, suggesting the presence of micrometastasis. Our findings suggest that mammaglobin B and CEA could be useful RT-PCR markers for the detection of lymph node micrometastases in biliary tract carcinomas. Our RT-PCR assay allows accurate clinical staging necessary for patient stratification with respect to adjuvant therapy after surgery.

Aged↗

PubMed2000

Augmentation of antitumor activity of 5-fluorouracil by interferon alpha is associated with up-regulation of p27Kip1 in human hepatocellular carcinoma cells.

Several clinical trials have demonstrated the effectiveness of combination therapy with 5-fluorouracil (5-FU) and IFN-alpha in colon cancer, hepatocellular carcinoma (HCC), and other malignancies. In our preliminary clinical studies, we have observed outstanding effects with this combination therapy in patients with advanced HCC. However, the underlying mechanism by which IFN-alpha modulates the effects of 5-FU is unknown. We, therefore, conducted a mechanistic study using two HCC cell lines, PLC/PRF/5 and HuH7. IFN-alpha significantly enhanced the growth inhibitory effect of 5-FU in PLC/PRF/5 cells but not in HuH7 cells, and the isobolographic analysis indicated that this effect was synergistic. Flow cytometric analysis showed a delay in the progression of G0-G1 to S phase in PLC/PRF/5, and a sustained, induction of the cyclin-dependent kinase inhibitor p27-Kip1 and down-regulation of cyclin D1 was observed. Moreover, increased expression of p27Kip1 was associated with reduced CDK-2-associated kinase activity. Another difference in the two cell types was that PLC/PRF/5 expressed abundant IFN receptors, but HuH7 did not. Apoptosis assays were not helpful in explaining the mechanism. Our results suggest that the synergistic effects of 5-FU and IFN-alpha may in part be attributable to alterations in cell cycle progression via up-regulation of p27Kip1.

Apoptosis↗

PubMed2000

[Asthma death among adults in Japan 1995-1997. Analysis of 295 cases reported questionnaires sent to hospitals with more than 100 beds. Asthma Death Investigation Committee].

To clarify recent trends in adult asthma mortality, the Asthma Death Investigation Committee of Japan studied the clinical characteristics of 295 patients who died of asthma between 1995 and 1997. Males were slightly more than females among the death cases. Approximately half of the patients ranged in age from 60 to 79 years. Tendency to increase of death among young male adults continued. One third of the patient deaths involved the asphyxic type, while status asthmaticus was the cause death in 21.9%. Half of the asthmatics died in hospitals or emergency rooms, and places where the fatal attacks occurred were mainly patients' houses. The main cause fatal asthma attacks was respiratory infection, followed by fatigue, stress, and discontinuation of medication. Most of the patients were classified moderate or severe type of asthma 1 month before death. Histories of life-threatening attacks and hospitalization due to severe attacks, irregular visits to the hospital, low compliance, and insufficiency of corticosteroid treatment were suggested as the main risk factors in adult asthma deaths.

Adolescent↗

PubMed1999

The cognate coat protein is required for cell-to-cell movement of a chimeric brome mosaic virus mediated by the cucumber mosaic virus movement protein.

Cucumber mosaic cucumovirus (CMV) and brome mosaic bromovirus (BMV) have many similarities, including the three-dimensional structure of virions, genome organizations, and requirement of the coat protein (CP) for cell-to-cell movement. We have shown that a chimeric BMV with the CMV 3a movement protein (MP) gene instead of its own cannot move from cell to cell in Chenopodium quinoa, a common permissive host for both BMV and CMV. Another chimeric BMV was constructed by replacing both MP and CP genes of BMV with those of CMV (MP/CP-chimera) and tested for its infectivity in C. quinoa, to determine whether the CMV CP has some functions required for the CMV MP-mediated cell-to-cell movement and to exhibit functional difference between CPs of BMV and CMV. Cell-to-cell movement of the MP/CP-chimera occurred, and small local lesions were induced on the inoculated leaves. A frameshift mutation introduced in the CMV CP gene of the MP/CP-chimera resulted in a lack of cell-to-cell movement of the chimeric virus. These results indicate that the viral movement mediated by the CMV MP requires its cognate CP. Deletion of the amino-terminal region in CMV CP, which is not obligatory for CMV movement, also abolished cell-to-cell movement of the MP/CP-chimera. This may suggest some differences in cell-to-cell movement of the MP/CP-chimera and CMV. On the other hand, the sole replacement of BMV CP gene with that of CMV abolished viral cell-to-cell movement, suggesting a possibility that the viral movement mediated by the BMV MP may also require its cognate CP. Functional compatibility between MP and CP in viral cell-to-cell movement is discussed.

Amino Acid Sequence↗

PubMed1999

Participation of the liver in generation of a vigorous anti-donor response after inoculation of donor spleen cells.

BACKGROUND: There is a general agreement that a preferential accumulation of alloantigens within the liver could induce hyporesponsiveness to the inoculated antigens. Entrapment of antigens in the liver may evoke an unique immune response in the organ and play a key role in determination of the fate of the transplanted grafts. To understand the immune response in the liver after inoculation of allogeneic donor antigens, we examined the immune response to systemically inoculated alloantigen in rats whose sensitized liver was replaced with that of naive rats or in naive rats whose liver was replaced with that of sensitized rats. METHODS: Using implantation of syngeneic liver (alloantigen-accumulated/naive) in rats (naive/alloantigen-sensitized), we compared the immune responses to alloantigen between rats with hepatic/extrahepatic alloantigen at 24 hr after alloantigen inoculation. This was called sensitized-liver-grafted (SLG)/sensitized-liver-removed (SLR) treatment. The immune response to donor alloantigen in this model was evaluated by survival of skin or heart grafts, complement-dependent cytotoxicity (CDC) titer and delayed-type hypersensitivity (DTH) response. RESULTS: Compared with the mean survival time (MST) in donor spleen cell inoculated (DSI) rats (skin and heart, MST: 8.2+/-1.1 and 10.7+/-2.3 days), SLG rats rejected allografts in an accelerated fashion (skin and heart, MST: 5.5+/-0.5 and 4.2+/-0.8 days), associated with higher CDC titer and DTH response. In contrast, allograft survival was moderately prolonged in SLR (skin and heart, MST: 16.5+/-2.6 and 29.5+/-3.7 days) associated with suppressed CDC titer and DTH response. The survival of third-party allograft after SLG or SLR treatment (skin, MST: 9.3+/-1.5 or 9.7+/-0.6 days) indicated that immunological hyper/hyporesponsiveness was donor-specific. CONCLUSIONS: A strong anti-donor immune response was induced by the transfer of donor antigen-baring liver to naive rats 24 hr after alloantigen inoculation, whereas removal of the liver suppressed alloimmune response. Our results indicate that vigorous anti-alloimmune response occurred in the liver after systemic inoculation of donor spleen cells.

Adoptive Transfer↗

PubMed1999

Pretreatment of crude pancreatic islets with mitomycin C prolongs graft survival time in xenogeneic rat-to-mouse model.

BACKGROUND: Rejection of pancreatic islet grafts is still a serious problem. We evaluated the effect of mitomycin C (MMC) on the survival of crude islets grafts after xenogeneic islet transplantation. METHODS: WS (RT1k) rat islets pretreated with various concentrations of MMC (0, 1, 3.2, 10, 32, 50, 100, 320, and 1,000 microg/ml) were transplanted into C57BL/6 mice with streptozotocin-induced diabetes. In vivo graft function was assessed by a daily measurement of nonfasting blood glucose concentration in each animal. We also examined the separate effect of MMC on purified islets and contaminants present in the crude islet preparation. RESULTS: MMC at doses of 10, 32, 50, and 100 microg/ml resulted in a significant prolongation of the mean graft survival time from a control of 12.4+/-2.5 days to 23+/-7.4, 17.5+/-5.4, 25.5+/-14.7, and 26.7+/-8.9 days, respectively. Deterioration of glucose metabolism was noted when the dose exceeded 32 microg/ml, whereas at 320 microg/ ml, MMC failed to restore normoglycemia. Prolongation of survival time of crude islets was the result of its effect on islets and contaminant components of the crude islet preparation. In vitro study showed that MMC treatment at a higher concentration than 10 microg/ml reduces the stimulatory as well as proliferative capacity of lymph node cells. CONCLUSIONS: Pretreatment of pancreatic islets with MMC at 10 microg/ml prolongs xenograft survival without deterioration of in vivo graft function. This novel treatment modality represents a new strategy for the modulation of immunity of islets and contaminants in crude islet preparations.

Animals↗

PubMed1999

Exploitation of the continuum between early ischemia/reperfusion injury and host alloresponsiveness: indefinite kidney allograft survival by treatment with a soluble P-selectin ligand and low-dose cyclosporine in combination.

BACKGROUND: We have shown previously that sPSGL, a soluble glycoprotein ligand for P and E selectins, reduces the events associated with ischemia/reperfusion injury of the kidney. In the present study, we have attempted to modulate differentially early inflammatory influences and later host alloresponsiveness in an LBNF1-Lewis renal graft model by treatment with sPSGL in combination with a marginally effective dose of cyclosporine (CsA). METHODS: Four experimental groups were studied: group 1=control animals receiving vehicle only; group 2=sPSGL monotherapy alone; group 3=low-dose CsA; group 4=sPSGL plus low-dose CsA. Grafts were removed at 1, 3, 5, and 7 days (n=3/time point) and assessed by histology, immunohistology, and reverse transcriptase-polymerase chain reaction. Long-surviving grafts in recipients of groups 3 and 4 were followed functionally for more than 28 weeks. RESULTS: Graft function was prolonged indefinitely in recipients in group 4, all of which survived for more than 200 days. In contrast, survival of animals in groups 1 and 2 was not increased substantially, whereas only 4 of 17 animals in group 3 (23.5%) survived more than 24 days (P<0.01). Five days after engraftment, necrosis was relatively minimal in group 4 organs but pronounced in those of the other groups. By immunohistology, numbers of infiltrating CD4+ and CD8+ T cells and ED1+ macrophages were significantly diminished in group 4 allografts compared with those of the other groups. Serial assessment of chemokine and cytokine mRNA expression confirmed these findings. The long-term effects of CsA treatment alone were compared with those of sPSGL in combination with CsA. Proteinuria remained virtually absent in group 4 recipients. Morphologically, the few long-surviving grafts in group 3 showed signs of chronic rejection; those in group 4 remained relatively normal. CONCLUSIONS: Although treatment with sPSGL alone showed no apparent influence on the acutely rejecting transplants, at least by the parameters examined in this study, it produced indefinite survival of kidney grafts when used in combination with low-dose CsA. The data support the influence of early nonspecific injury on later immunological rejection.

Animals↗

PubMed1999

Injection of mitomycin-C-treated spleen cells induces donor-specific unresponsiveness to cardiac allografts in rats.

BACKGROUND: In this study, preoperative mitomycin-C- (MMC) treated donor-specific transfusion (DST) was examined for its ability to induce unresponsiveness to cardiac allografts in rats. METHODS: DA (RT1a) rats were used as donors, BUF (RT1b) or WS (RT1k) rats as recipients, and Lew (RT1l) rats as third party donors. BUF or WS rats were given i.v. injection of DA spleen cells (SPCs) suspension (5x10(7)/l ml) with or without MMC treatment 10 days before cardiac transplantation. Delayed-type hypersensitivity and complement-dependent cytotoxicity assays were carried out in these animals separately to examine in vivo immunosuppressive effect. Suppressor assay was also examined to determine in vitro immunosuppressive effects in allogeneic mixed leukocyte culture. RESULTS: In the full allogeneic DA-to-BUF rat strain combination, preoperative i.v. administration of MMC-treated donor SPCs led to a significant prolongation of graft survival over the control (110+/-66 versus 7.2+/-0.8 days: P<0.01), although administration of nontreated donor SPCs did not (9.3+/-1.0 days). This beneficial effect of MMC treatment was also seen in the DA-to-WS rat combination (31+/-16 days versus donor-specific transfusion alone; 11+/-1.5 days or untreated control; 12+/-1.5 days; P<0.05). However, injection of third party DA SPCs in the Lew-to-BUF combination induced no significant prolongation of cardiac allograft survival compared with the untreated control (11+/-0.6 versus 11+/-2.0 days; NS), indicating that this prolongation effect was induced in an antigen-specific manner. The immunosuppressive effect was also secured for both delayed-type hypersensitivity response and anti-donor cytotoxic antibody production. Moreover, addition of MMC-treated SPCs to mixed lymphocyte culture led to antigen-specific suppression. CONCLUSIONS: Preoperative i.v. injection of MMC-treated donor SPCs is promising for inducing unresponsiveness in rat cardiac allograft model.

Adoptive Transfer↗

PubMed1999

Microchimerism in thymus is associated with up-regulated T helper type 1 cytokine transcription during cardiac allograft rejection in rats.

BACKGROUND: Intrathymic microchimerism (MC) is thought to be responsible for inducing allograft tolerance. However, the role of MC in the thymus gland after transplantation, particularly in the rejection response, is unknown. We investigated serial changes in intrathymic cytokine production associated with MC and allograft rejection. METHODS: Donor-specific cell injection (DSI) and heterotopic heart transplantation (HTx) were performed in the fully allogeneic combination using DA rats (RT1a) as donors and WS rats (RT1k)as recipients. MC was checked by polymerase chain reaction (PCR) using a donor RT1.Bbeta domain 1 region sequence-specific primers. Reverse transcription (RT)-PCR analysis of cytokine (interleukin [IL]-2, interferon-gamma, IL-4, and IL-10) profiles of the thymus was performed in animals given DSI, HTx, or DSI/HTx. RESULTS: DSI alone resulted in an immediate development of MC, detected by PCR, in various organs including the thymus, spleen, liver, and blood, of most rats, lasting for over 2 months. However, DSI-induced MC selectively disappeared in the thymus on day 7 after grafting, several days before the rejection of cardiac allograft. RT-PCR analysis of cytokine profiles showed that the levels of Th1 (IL-2 and interferon-gamma) cytokines transcribed in the thymus were higher than in the spleen. MC reappeared in the thymus on day 21 after grafting, but was not associated with elevation of Th1 cytokine transcription when allograft was replaced by fibrosis. CONCLUSIONS: Intrathymic MC does not always confer unresponsiveness to alloantigen, but can be eliminated after anti-donor response.

Animals↗

PubMed1999

Expression and prognostic roles of the G1-S modulators in hepatocellular carcinoma: p27 independently predicts the recurrence.

Expression of cell-cycle modulators at the G1-S boundary, retinoblastoma gene product (pRb), p21, p16, p27, p53, cyclin D1, and cyclin E was investigated with 104 hepatocellular carcinomas (HCC), as well as 90 of their adjacent noncancerous lesions and 9 normal liver control specimens. The labeling indices (LI) of pRb, p21, p16, and p27 were higher in HCC lesions than in the adjacent noncancerous lesions and normal controls. Especially, p27 LI in noncancerous lesions was significantly higher than that in normal livers (P =.011). Aberrant p53 expression and cyclin D1 and E overexpression were observed exclusively in HCC lesions. pRb was positive in 85.6% of the HCC cases and was not related to any clinicopathological parameters. The p21 LI was generally low (average, 5.5 +/- 9.8). Although a negative regulator, p21 LI was higher in cases with intrahepatic metastasis (P =.0359). The p16 LI was significantly decreased (P =.0121) in cases with advanced stage. p27 LI was significantly decreased in cases with portal invasion (P =.0409), poor differentiation (P <.0001), larger size (P =.0421), and intrahepatic metastasis (P =.0878, borderline significance). On the other hand, aberrant p53 expression showed positive relationships with poor differentiation (P =.0004) and Ki-67 LI (P =. 0047). Cyclin D1 overexpression was found in 32.6% of the cases and occurred more frequently in those with high Ki-67 LI (P =.0032), pRb expression (P =.0202), poor differentiation (P =.0612, borderline significance), and intrahepatic metastasis (P =.0675, borderline significance). Cyclin E was overexpressed in 35.5% and had positive relationships with Ki-67 LI (P =.0269) and stage (P =.0125). In univariate analysis, cases with p27 LI < 50 (P =.0004), cyclin D1 overexpression (P =.0041), and cyclin E overexpression (P =.0572, borderline significance) showed poorer outcomes for disease-free survival (DFS). In multivariate analysis, p27 expression could be recognized as an independent prognostic marker for DFS. These findings suggest that in HCC: 1) p27 is active against HCC progression in early phases and, possibly, hepatocarcinogenesis as a negative regulator and can be a novel prognostic marker for DFS; and 2) cyclin D1 predominantly works for cell-cycle progression at the G1-S boundary.

Adult↗

PubMed1999

Detection and differentiation of Aino and Akabane Simbu serogroup bunyaviruses by nested polymerase chain reaction.

Reverse transcription-polymerase chain reaction (RT-PCR) and nested PCR were developed to detect and differentiate Aino (AINO) and Akabane (AKA) virus S RNA. Two pairs of AINO- and AKA-specific primers for nested PCR were synthesized and examined for their capacity to amplify PCR products using 7 Simbu serogroup viruses isolated in Japan and Australia. RT- and nested PCR using AKA-specific primers amplified cDNA from Tinaroo virus RNA as well as homologous RNA. Nested PCR products were differentiated by Hph I and Bst EII digestion. Peaton (PEA) virus S cDNA was also amplified using AINO- specific primers. The nested PCR products of PEA virus were not digested by 3 restriction enzymes (Ava II, Eco RI and Hae II), whereras those of AINO virus were digested as expected. Using this technique, AINO and AKA viruses were detected at concentrations as low as 10(-3) plaque-forming units (PFU) and 10(-5) PFU, respectively, in a supernatant of virus-infected cells. It was possible to detect AINO and AKA genome from various tissues of experimentally infected mice, and also the AKA nested PCR products from serum samples from sentinel cattle naturally infected with AKA virus. The present nested PCR appears a simple, rapid and valuable method for diagnosing AINO and AKA infection.

Animals↗

PubMed1999

Improvement of molar ratio of branched-chain amino acids to tyrosine (BTR) associated with liver fibrosis in chronic hepatitis C patients treated with interferon-alpha.

The molar ratio of branched-chain amino acids (BCAA) to tyrosine (BTR), which has recently been possible to easily determine by an enzymatic method, can be clinically used as a substitute for that of BCAA to aromatic amino acids (AAA). We used this method to investigate the diagnostic implications of serum BTR in patients with chronic hepatitis C treated with interferon-alpha. There was a good correlation between BTR and staging (fibrosis) scores both before and after treatment. BTR increased significantly at 24 months after treatment in the responder group (P < 0.01). Moreover, a significant increase in BTR was observed in patients with improved staging (P = 0.015), but not grading (necroinflammatory), scores. BTR in chronic hepatitis C patients seems to be reflected by the extent of liver fibrosis, and, in the responder group, it can be improved, together with liver fibrosis, by interferon-alpha treatment.

Adult↗