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Biomedical subjects

M Namba

Publications and source records attributed to M Namba.

At least 181 records · Page 10Linked to original sources
PubMed1992

Effects of barbiturates with or without liver-tumor-promoting activity on survival and DNA synthesis of suckling and adult rat hepatocytes in serum-free primary culture.

The effects of four barbiturates with or without liver-tumor-promoting activity were examined on survival and deoxyribonucleic acid (DNA) synthesis of suckling and adult rat hepatocytes in serum-free primary culture: Of the four barbiturates, two promoters, phenobarbital and barbital, enhanced DNA synthesis of suckling rat hepatocytes at low concentrations of 0.5-2 mM or 0.5 mM, but suppressed it at high concentrations of 3 mM or 1.5-4 mM. DNA synthesis of adult rat hepatocytes was, however, only suppressed by phenobarbital within the dose range tested of 1-3 mM. On the other hand, two remaining non-promoters, barbituric acid and amobarbital, did not increase but only suppressed DNA synthesis of suckling rat hepatocytes within the dose ranges of 0.5-4 mM and 0.05-0.5 mM respectively. Phenobarbital and amobarbital were effective for supporting survival and maintaining morphological features of suckling and adult rat hepatocytes at the relatively high concentrations of 3-4 mM and 0.5-0.75 mM respectively. However, barbital and barbituric acid were ineffective for maintenance of hepatocytes. The results show that the ability to support survival of primary cultured hepatocytes is not a common property of liver-tumor-promoter barbiturates but is a common property of some barbiturates with high lipophilicity, and that the maintenance of hepatocytes by phenobarbital or amobarbital is not due to a counterbalance of stimulated proliferation and death of the cells.

Aging↗
PubMed1992

Increase of S-(1,2-dicarboxyethyl)glutathione in regenerating rat liver.

Changes in the level of S-(1,2-dicarboxyethyl)glutathione (DCE-GS) with time were determined during regeneration of rat liver after partial hepatectomy. The DCE-GS level increased in regenerating rat liver, reaching a maximum level (4.7-fold) on day 2 and reverted to the normal level in 1 week. During the liver regeneration process, the activity of DCE-GS-synthesizing enzyme in the liver retained its control level, while the level of glutathione--a substrate of the enzyme in the liver--fluctuated in parallel with that of DCE-GS.

Animals↗
PubMed1992

Effects of intracellular cyclic AMP and cyclic GMP levels on DNA synthesis of young-adult rat hepatocytes in primary culture.

Possible roles of dibutyryladenosine 3',5'-cyclic monophosphate (cAMP) and dibutyryl-guanosine 3',5'-cyclic monophosphate (cGMP) in regulation of hepatocyte DNA synthesis were examined using primary cultures of young-adult rat hepatocytes maintained in arginine-free medium. Throughout the experimental period, nonparenchymal cells were hardly observed in the selective medium. When epidermal growth factor (EGF) was added to the cultures, a transient increase in the intracellular cAMP level preceded the elevation of hepatocyte DNA synthesis. EGF-stimulated hepatocyte DNA synthesis was remarkably enhanced by the elevation of the intracellular cAMP level induced by treatment with cAMP alone or a combination of cAMP and theophylline, an inhibitor of cyclic nucleotide phosphodiesterase. Furthermore, the early elevation of intracellular cAMP alone, which was induced by treatment with the combination of cAMP and theophylline, caused a remarkable increase in hepatocyte DNA synthesis. On the other hand, addition of EGF to the cultures caused a rapid decrease in the intracellular cGMP level followed by an increase in hepatocyte DNA synthesis. EGF-stimulated hepatocyte DNA synthesis was severely suppressed or completely inhibited by the elevation of the intracellular cGMP level induced by treatment with cGMP alone or a combination of cGMP and dipyridamole, a specific inhibitor of cGMP phosphodiesterase. These findings indicate that cAMP and cGMP act oppositely on the regulation of DNA synthesis of young-adult rat hepatocytes in primary culture: cAMP plays a positive role, whereas cGMP plays a negative role. Also it is strongly suggested that an early elevation of the intracellular cAMP level is essential for the onset of DNA synthesis in hepatocyte primary cultures.

Age Factors↗
PubMed1992

Human hepatocyte growth factor stimulates the growth of HUH-6 clone 5 human hepatoblastoma cells.

The effects of human hepatocyte growth factor (hHGF), a potent mitogen for rat and human hepatocytes in primary culture, on proliferation of human hepatoma and hepatoblastoma cells were examined. Out of five cell lines; HLE, HuH-6 clone 5, HuH-7, PLC/PRF/5, and Hep G2, only HuH-6 Clone 5 cells were stimulated by recombinant hHGF. Both native and recombinant hHGFs caused dose-dependent increases in cell number and DNA synthesis of cells. This stimulation was strongly inhibited by anti-hHGF monoclonal antibody.

Animals↗
PubMed1992

Primary culture of liver cancer tissues with or without transcatheter arterial embolization and establishment of a cell strain.

More epithelial-like cells are found in primary cultures of transcathetel arterial embolization (TAE)-untreated human liver cancer tissues than in those of TAE-treated tissues. A new cell strain, HUH-33, established from the former, grows slowly and is untransplantable into nude mice and secretes alpha-fetoprotein, albumin and some other tumor markers. Chromosome numbers are widely distributed. HUH-33 expresses hepatocyte type keratin and contains desmosome- or intermediate filament bundle-like structures.

Aged↗
PubMed1992

Antiproliferative effects of suramin on human cancer cells in vitro and in vivo.

The present experiment was undertaken to study what types of human cancers are responsive to the antiproliferative effects of suramin. The human malignant cells used were as follows: cervical cancer (HeLa), mammary cancer (MCF-7), bladder cancer (EJ), hepatoma (HuH-7, PLC/PRF/5), embryonal carcinoma (PA-1), in vitro transformed fibroblasts (KMST-6, SUSM-1, VA-13), five myeloma cell lines (KMM-1, KMS-5, KMS-11, KMS-12, RPMI 8226), Burkitt's lymphoma (Raji), acute promyelocytic leukemia (HL-60), chronic myelocytic leukemia (K562), Epstein-Barr virus nuclear antigen positive lymphoblastoid cells (KMS-9). The cells were treated with 25 to 100 micrograms/ml suramin for 72h. Proliferation of HuH-7 and two human myeloma cells (KMS-11 and KMS-12) was remarkably inhibited, and that of PA-1, PLC/PRF/5, KMST-6, two other myeloma cell lines (KMM-1 and KMS-5), Raji and HL-60, was moderately inhibited. In order to confirm part of the results obtained from in vitro experiments, in vivo experiments were also undertaken. The growth of HuH-7 cells transplanted subcutaneously into nude mice was significantly suppressed by intravenous injection of suramin. We discussed the possibility that certain types of human cancers, the growth of which seemed to be more or less dependent on polypeptide growth factors, might be sensitive to the antiproliferative effects of suramin.

Animals↗
PubMed1992

[Clinical assessment of urinary free L-fucose levels].

We measured urinary levels of free L-fucose in healthy subjects, patients with benign diseases, and patients with cancer using an automated analyzer and a newly isolated L-fucose dehydrogenase, and evaluated the clinical usefulness of the results. The values obtained were corrected for urinary creatinine as micromoles per gram of creatinine. The cutoff value, set at the mean + 2SD for the healthy subjects, was 250 mumol/g.Cr. Patients with gallbladder cancer, bile-duct cancer, liver cancer, pancreatic cancer, or cirrhosis of the liver had significantly higher levels of L-fucose than the healthy subjects. The diagnostic sensitivity for these five diseases, taken together, was 68% (144/213). Specificity for the detection of cancer was calculated by use of false positives for patients with cholelithiasis, hepatitis, and pancreatitis: it was 73% (76/104). Diagnostic accuracy for these seven diseases taken together was therefore 69% (220/317). We compared the positive ratio of the L-fucose level with that of the tumor markers AFD and CA19-9. The positive ratio of an L-fucose value above the cutoff was higher than the positive ratio of either marker in bile-duct cancer, gallbladder cancer, liver cancer, and pancreatic cancer. The results suggested that the urinary levels of free L-fucose reflected the metabolism of sugar chains of glycoconjugates, and may be usefully clinically as a tumor marker.

Biomarkers, Tumor↗
PubMed1992

[Cell culture and its application primary culture of human hepatocytes and its application].

Difficulty in obtaining human liver specimens has made a delay in the development of methodology for isolation and primary culture of human hepatocytes compared to animal hepatocyte cultures. Recently the collagenase-liver-perfusion technique, which was developed for rat hepatocyte isolation, has been applied to prepare isolated human hepatocytes. This method greatly improved the yield and viability of hepatocytes of human liver tissues. Since then, the isolated human hepatocytes and their primary culture have been widely used for studies of hepatic metabolism, regeneration, transplantation, hepatitis B virus infection, hepatotoxins, detection of human carcinogens, hepatocarcinogenesis and so forth. These current studies were reviewed.

Cell Division↗
PubMed1992

[Neo-adjuvant chemotherapy with tegafur suppository for rectal cancer--evaluation of the antitumor effects, tissue levels of 5-FU and inhibition of thymidylate synthase. Tochigi Colorectal Cancer Study Group].

We evaluated antitumor effect histologically and assayed the tissue levels of 5-fluorouracil (5-FU) and thymidylate synthase (TS) activity using surgical specimens obtained from the patients with rectal cancer, who were given tegafur suppositories prior to surgery. The antitumor effect was evaluated histologically according to classification of the general rules for the gastric cancer study (Japanese Research Society for Gastric Cancer). In 39 patients, 16 tumor specimens revealed no effect (grade-0), 22 tumors grade-1 effect, and one was not evaluable because of the severe inflammatory changes. In 23 of these patients, resected specimens were available for the assay. 5-FU levels in cancer tissues were significantly higher than those in normal tissues, and TS inhibition rates (TSIR) were almost identical, averaging around 20%, in both cancer and normal tissues. Comparing the 5-FU levels and TS activity according to the histological effects (i.e.: 'grade-0' vs 'grade-1'), the 5-FU levels in the tumors achieved grade-1 were significantly higher than in the tumors showed 'grade 0' (p less than 0.01), and TSIR in the former were relatively greater than in the latter (p = 0.053). It is suggested that both tissue levels of 5-FU and TSIR may be useful parameters to predict the anti-tumor effect against rectal cancer after administration of 5-FU and its derivatives.

Chemotherapy, Adjuvant↗
PubMed1992

Gene rearrangement and overexpression of PRAD1 in lymphoid malignancy with t(11;14)(q13;q32) translocation.

The proto-oncogene PRAD1 (parathyroid adenoma 1) on chromosome 11q13 was found to be overexpressed in all five B-cell lines with t(11;14)(q13;q32) translocation tested. One B-cell lymphoma and four myeloma cell lines with this translocation demonstrated more than 10-fold overexpression as determined by Northern blot analysis, when compared with normal lymphoid tissues such as thymus, spleen and lymph node. Hematopoietic cell lines without the translocation were also examined, but none of these demonstrated the overexpression, confirming that overexpression of the PRAD1 gene is associated with t(11;14) translocation. A truncated form of mRNA was seen in one of five cell lines with the translocation, SP-49. Hybridization with different regions of the PRAD1 cDNA revealed that the truncated form of mRNA retained the coding region but had lost the 3' untranslated region. Southern blot analysis demonstrated a gene rearrangement in this SP-49 cell line. To study the genetic alteration responsible for the truncated form of mRNA in this cell line, the rearranged allele as well as the germline allele were cloned. The restriction map revealed that the rearranged portion was at the 3' end of the PRAD1 gene, eliminating the mRNA-destabilizing signal AUUUA. Human-rodent hybrid cell analysis demonstrated that the region introduced 3' of PRAD1 was derived from chromosome 11, suggesting that the PRAD1 gene region is deleted at the 3' end. Over-expression of the PRAD1 gene in association with t(11;14)(q13;q32) translocation suggested that in these cases the regulation of PRAD1 was altered by the juxtaposed gene, most likely the immunoglobulin heavy-chain gene from chromosome 14.

Base Sequence↗
PubMed1991

Stable expression of human tissue-type plasminogen activator regulated by beta-actin promoter in three human cell lines: HeLa, WI-38 VA13 and KMS-5.

A high-level and stable expression system of human tissue-type plasminogen activator (t-PA) was accomplished in human cells by selecting a promoter and a host cell line. First, we have constructed two types of t-PA expression plasmids containing 3 kb of the human beta-actin promoter region or 0.3 kb of SV40 early promoter region and these plasmids were transfected into HeLa cells, respectively, and the resulting transfectants were found to secrete various amounts of t-PA derived from the plasmids to the culture media. Southern blot analysis revealed that the beta-actin promoter was more efficient than the SV40 early promoter with regard to the expression level per single copy of the t-PA gene in the transfected HeLa cells. Next, the t-PA expression plasmid containing the beta-actin promoter was also transfected into WI-38 VA13 cells, a human fibroblastic cell line, and KMS-5 cells, a human lymphoid cell line, in order to compare the expression ability of the promoter among these three cell lines. Some of the transfectants from both cell lines were also found to produce t-PA. It was also found that the expression levels in HeLa and WI-38 VA13 seemed to be more efficient than that in KMS-5.

Actins↗
PubMed1991

Extending effects of phospholipids, cholesterol, and ethanolamines on survival of adult rat hepatocytes in serum-free primary culture.

In a serum-free primary culture, membrane lipids, such as phosphatidylethanolamine, phosphatidylcholine, or cholesterol, effectively prolonged the survival of adult rat hepatocytes. These lipids effectively prevented hepatocytes from morphologic degeneration observed in control cultures, such as enlargement of cell surface, degranulation of cytoplasm, and multinucleation. The maintenance effect of phospholipid precursors, ethanolamine, or phosphoethanolamine on the primary-cultured hepatocytes was similar to that of phospholipids. These effects appear to be due to stabilization of the plasma membrane.

Animals↗
PubMed1991

Expression of liver-specific functions and secretion of a hepatocyte growth factor by a newly established rat hepatoma cell line growing in a chemically-defined serum-free medium.

A rat hepatoma cell line 3'-mRLh-2 was established from 3'-methyl-4-dimethylaminoazobenzene-induced hepatoma. Cells proliferated well in 5Fs-DM-160, a chemically-defined serum-free medium; population doubling time was 68.5 h, and modal chromosome number was 81 (21%). The cells were transplantable, and the transplanted tumors were histologically diagnosed as hepatocellular carcinoma. They were cytochemically positive for gamma-glutamyl transpeptidase. The cells possessed about 30% of tyrosine aminotransferase activity level in the rat liver, and showed 5.5 to 7.4-fold induction of this enzyme activity in response to dexamethasone. Also, the cells secreted alpha-fetoprotein, albumin, transferrin, alpha 1-acid glycoprotein, C-reactive protein, fibrinogen, complement component C3, and other five-serum proteins. Furthermore, the conditioned medium stimulated DNA synthesis in primary cultures of adult rat hepatocytes in a dose-dependent, saturable manner and in the absence of epidermal growth factor. These properties of the cell line 3'-mRLh-2 were compared with those of the popular rat hepatoma cell lines, such as H4-II-E-C3 from Reuber hepatoma H35 and HTC from Morris hepatoma 7288C.

Animals↗
PubMed1991

Regulation of collagen gene expression by transformed human fibroblasts: decreased type I and type III collagen RNA transcription.

The regulation of collagen gene expression in normal diploid human fetal fibroblasts (KMS-6 cells), and fibroblasts immortally transformed by treatment of KMS-6 with Co-60 gamma rays (KMST-6 cells) was compared to that of ones tumorigenically transformed by treatment of KMST-6 cells with Harvey murine sarcoma virus (KMST-6-Ras cells). Synthesized collagenous protein decreased to approximately 30% of that of normal fetal fibroblasts in both transformed cell lines, and the relative rate of collagen synthesis to total protein synthesis decreased about sixfold in KMST-6 cells and twelvefold in KMST-6-Ras cells. The m-RNA levels of type I collagen in both of these cell lines decreased to approximately 20% of that of the control fibroblasts, whereas type III collagen m-RNA levels decreased to only 9% of that of the control. The copy number of the collagen gene in both transformed cell lines was unaltered. The transcriptional rates of collagen alpha 1(I) and collagen alpha 1(III) in both cell lines decreased to 20% and 7% respectively of that of control. These data indicate that collagen synthesis was reduced at the transcriptional level in these transformed human fibroblasts.

Cell Line, Transformed↗
PubMed1991

Analysis of methylation in the c-MYC gene in five human myeloma cell lines.

Genomic alterations of the human c-MYC gene were analysed in five human myeloma cell lines established in Kawasaki Medical School and compared with those of normal lymphocytes, Raji cells from Burkitt's lymphoma, and an Epstein-Barr virus positive lymphoblastoid cell line (LCL). Although no structural chromosome aberrations at 8q24, the c-MYC locus, were distinct, the mRNA level of c-MYC in these myeloma cell lines was 30-50-fold that in normal peripheral blood lymphocytes. Regarding the methylation of c-MYC, DNAs of the myeloma cell lines were digested with MspI plus EcoRI or HpaII plus EcoRI, and hybridized with three genomic 32P-labelled probes; the first, second and third exons of the human c-MYC gene, respectively. The extent of methylation in cytosine at a single CCGG site in the third exon substantially decreased in these myeloma cell lines as compared with that in normal tonsillar B, LCL and Raji cells. No significant differences in hypomethylation between these myeloma, normal B, LCL and Raji cells was detected in the first and second exon of c-MYC. These results suggest that the hypomethylation in the third exon of c-MYC might be related to the enhanced expression of c-MYC in these human myeloma cell lines.

Aged↗
PubMed1991

Effects of antioxidants on survival of adult rat hepatocytes under various oxygen tensions in serum-free primary culture.

Effects of antioxidants, such as superoxide dismutase, vitamin C, vitamin E, 4-(0-benzylphenoxy)-N-methylbutylamine hydrochloride (bifemelane), and selenite on survival of adult rat hepatocytes were examined under normoxic and hyperoxic conditions in serum-free primary culture. The tested antioxidants, except for vitamin C, significantly increased the survival rate of hepatocytes under the normoxic condition (under air). Thus, even the normoxic culture condition is hyperoxic for hepatocytes. Elevation of oxygen tension (40% O2) caused severe morphologic degeneration of hepatocytes and remarkable decrease in the survival rate of the cells. Addition of the antioxidants effectively protected hepatocytes from the morphologic degeneration, and significantly improved the survival of the cells under the hyperoxic condition. These findings indicate that the antioxidants can maintain the long-term survival of hepatocytes in serum-free primary culture.

Animals↗
PubMed1991

[Effects of suramin on cell proliferation of various types of human malignant cells].

Suramin, a polyanionic compound used clinically for the treatment of African trypanosomiosis and onchocerciasis, has been shown to inhibit the action of various growth factors such as platelet-derived growth factor, epidermal growth factor, fibroblast growth factor and transforming growth factor-beta to stimulate DNA synthesis of cells. Therefore, we investigated effects of suramin on cell proliferation of various types of human malignant cells in culture. Cell lines used were as follows: cervical cancer (HeLa), mammary cancer (MCF-7), bladder cancer (EJ), hepatoma (HuH-7, PLC/PRF/5), embryonal carcinoma (PA-1), and three in vitro transformed human fibroblast lines (KMST-6, SUSM-1, and VA-13). A serum-free defined medium, ASF103, was used when the effect of suramin on proliferation of cells was investigated. This culture medium contains only bovine serum albumin (0.1%), transferrin (5 micrograms/ml) and insulin (5 micrograms/ml) as peptide factors. On day 1, the drug was added to culture medium at the concentration of 25-100 micrograms/ml and 72-96 hr later, the number of cells was counted. The growth inhibition was expressed as the percentage of cells surviving after treatment of cells with suramin, with survival in the control condition representing 100 percent. Proliferation of HuH-7 cells was prominently inhibited and those of PA-1, PLC/PRF/5 and KMST-6 were moderately inhibited under the same conditions of treatment. On the other hand, other five cell lines were not responsive to up to 100 micrograms/ml suramin.

Breast Neoplasms↗