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Biomedical subjects

K Kaji

Publications and source records attributed to K Kaji.

At least 73 records · Page 4Linked to original sources
PubMed1994

Increase in production of hepatocyte growth factor by human embryonic lung fibroblasts in the process of aging in culture.

It was determined whether human hepatocyte growth factor (hHGF)-producing ability would change in the human embryonic lung fibroblast cell strains (MRC-5 and IMR-90) until the cells senesced in culture. The effects of phorbol 12-myristate 13-acetate (PMA), dexamethasone, and transforming growth factor-beta 1 (TGF-beta 1) on hHGF production were also studied in these cell strains. For stimulation of DNA synthesis of adult rat hepatocytes in primary culture, hHGF secreted by MRC-5 cells at 39.9 and 69.8 population doubling levels (PDLs) showed almost the same activity as recombinant hHGF. Secretion of hHGF by MRC-5 cells increased about threefold between 37.3 and 67.8 PDLs. IMR-90 cells also showed about a threefold increase in hHGF secretion with increased passage from 37.8 to 66.0 PDL. Both cell strains showed almost the same ratio of hHGF amount in the cell extracts to that secreted into the medium around 40 and 70 PDLs. Northern blot analysis showed that the transcriptional level of the hHGF gene in MRC-5 cells increased about three-fold from 42.0 to 73.6 PDL in culture. These findings indicated that hHGF production increased in both cell strains with aging in culture. Production of hHGF in both cell strains was remarkably stimulated by treatment with 10 nM PMA. On the other hand, hHGF production in both cell strains was slightly suppressed by treatment with 1 microM dexamethasone. TGF-beta at a concentration of 5 ng/ml prominently inhibited hHGF production in both cell strains. The response of both cell strains to these regulators for hHGF production was almost the same around 40 and 70 PDLs in culture.

Animals↗
PubMed1994

Protection of linoleic acid hydroperoxide-induced cytotoxicity by phenolic antioxidants.

The protective effects of phenolic antioxidants on linoleic acid hydroperoxide (LOOH)-induced toxicity to cultured human umbilical vein endothelial cells were examined. Our previous results were confirmed that for tocopherol homologs, lipophilicity and the presence of a phenolic hydroxyl group and two alkyl groups at its ortho positions are critical for protection against LOOH-induced cytotoxicity. Probucol and butylated hydroxytoluene (BHT) were more effective than other simple alkylated phenols. It was found that the protective effects of alkylated phenols were depended on by the presence of two alkyl groups; in particular, two tert-butyl groups, at positions ortho to a hydroxyl group and an alkyl group at the para position. Among alpha-tocopherol, 2,2,5,7,8-pentamethylchroman-6-ol, and BHT, the relative effectiveness of protection against the cytotoxicity (1.0:0.86:0.58, respectively) was inconsistent with the previously reported, relative antioxidant activity in homogeneous solution (1.0:1.2:0.004, respectively). Probably, the effectiveness of protection by phenolic antioxidants against the cytotoxicity depend primarily on their incorporation rate into cells due to their lipophilicity, secondly on their antioxidant activity, and thirdly on their orientation in biomembranes.

Antioxidants↗
PubMed1994

Programmed cell death in response to alkyllysophospholipids in endothelial cells.

Addition of the alkyllysophospholipid ET16-OMe, a putative antitumor drug, to the culture medium of human vascular endothelial cells resulted in apoptotic cell death. The death was characterized as programmed cell death since the process was inhibited by the addition of an inhibitor of protein synthesis. The mechanism responsible for apoptosis induced by alkyllysophospholipid has unique characteristics, as compared to those of apoptosis induced by other antitumor drugs, since the drug caused fragmentation of dying cells and its effect could be overcome by the presence of a survival factor, namely, fibroblast growth factor.

Antineoplastic Agents↗
PubMed1994

Activation of the alternative pathway of human complement by apoptotic human umbilical vein endothelial cells.

Complement activation generally does not occur on homologous cells. We observed C3 deposition on cultured human umbilical vein endothelial cells (HUVEC) when those which had died of apoptosis were treated with human serum. The C3 deposition on apoptotic HUVEC required Mg2+ but not Ca2+. In addition, the incubation of apoptotic HUVEC with purified C3, B, and D in the presence of Mg2+ resulted in C3 deposition. These results indicated that the C3 deposition on apoptotic HUVEC is mediated by the activation of the alternative complement pathway. C3 contains an intrachain thioester bond in the alpha chain (110 kDa) and upon activation to C3b, binds with membrane molecules by forming an ester or amide bond. Western blotting of reduced C3b-membrane molecule complexes, isolated from serum-treated apoptotic HUVEC by immunoprecipitation with anti-C3, revealed the covalent binding of C3b to several membrane molecules. Most of the C3b-membrane molecule complexes were cleaved by hydroxylamine, suggesting covalent binding via an ester bond. The molecular mass of the major alpha chain fragment released by hydroxylamine treatment was not 105 kDa but 68 kDa, which corresponds to the alpha 1 fragment of iC3b. These results indicate that most of the C3b on HUVEC was cleaved at its alpha' chain to yield iC3b, which consists of three disulfide-linked polypeptide chains and is a ligand of the complement receptor type 3 (CR3) of phagocytes. These results suggest that apoptotic HUVEC can activate the alternative pathway of the homologous complement and that the complement is related to the clearance of apoptotic cells by phagocytes.

Apoptosis↗
PubMed1994

Classification of signals for blocking apoptosis in vascular endothelial cells.

The survival and death of human umbilical vascular endothelial cells in culture are affected by several factors, such as fibroblast growth factor (FGF), serum, phorbol ester (TPA), and vanadate. In order to identify common aspects of the various signal-transduction processes during the course of apoptotic or programmed cell death, we designed experiments to distinguish between these factors in terms of the pathway that is responsible for the processing of each stimulus. We found, for example, that the effect of removal of FGF was specifically overcome by the addition of the phorbol ester. Our results indicated that two distinct pathways were operative, one specific for signal transduction initiated by FGF and phorbol ester and another specific for signal transduction initiated by serum and vanadate. These two pathways merged down-stream of the individual signal-processing pathways.

Apoptosis↗
PubMed1994

[Autoimmune hepatitis with hepatitis C virus markers (anti HCV antibody or HCV-RNA) and interferon therapy].

Autoimmune hepatitis (AIH) with hepatitis C virus (HCV) markers are seen frequently after the establishment of the method to detect HCV markers. According to the mechanism of the liver injury, we divide these cases of AIH with HCV markers to three groups; 1) the cases of AIH with HCV infection incidentally, 2) the cases of AIH triggered by HCV infection, and 3) the cases of chronic hepatitis C with immunological abnormalities. As far as we don't have any markers to differentiate these cases, we now think we should treat such cases with steroids at first before administering interferon, when we are not sure about the mechanism of the liver injury, because interferon is thought to be more risky than steroid for the cases who may have immunological abnormalities.

Autoimmune Diseases↗
PubMed1994

Hepatitic bile duct injuries in chronic hepatitis C: histopathologic and immunohistochemical studies.

Interlobular bile ducts are reported to be damaged in viral hepatitis. Such damages are called hepatitic duct injuries and mimic chronic nonsuppurative destructive cholangitis of primary biliary cirrhosis. In this study, hepatitic bile duct injuries were evaluated histopathologically and immunohistochemically in 149 needle liver biopsy specimens from patients with chronic hepatitis C and compared immunohistochemically with primary biliary cirrhosis. Fifty-one of the needle biopsies from patients with chronic hepatitis C (34.2%) showed hepatitic bile duct injuries which were distributed focally in the liver and showed variable epithelial damages such as cytoplasmic swelling, vacuolation and acidophilia, nuclear pleomorphism, and loss of nuclear polarity. Some of the injured bile ducts were embedded within lymphoid aggregates, whereas others were surrounded by lymphocytes as well as other inflammatory cells to varied degrees. The majority of lymphoid cells around hepatitic bile duct injury were B- and T-cells mixed in various proportions, and activated T-cells were occasionally found within the biliary epithelial layer. Histopathologic and serial section studies disclosed that bile duct loss was rare in chronic hepatitis C. Statistical analysis revealed that advancement of chronic hepatitis and the degree of necroinflammatory processes of the liver, particularly in the portal tracts, were positively correlated with the occurrence of hepatitic bile duct injuries. Immunohistochemical studies demonstrated that, whereas strong ectopic expression of HLA-DR and enhanced expression of HLA-A,B,C, and pyruvate dehydrogenase E2-complex in biliary epithelial cells were frequently observed in chronic nonsuppurative destructive cholangitis of primary biliary cirrhosis, such unusual expressions were generally absent or mild, even if present, in bile duct injuries in chronic hepatitis C.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1994

Frequent occurrence of hepatocellular carcinoma in cirrhotic livers after surgical resection of atypical adenomatous hyperplasia (borderline hepatocellular lesion): a follow-up study.

OBJECTIVES: Adenomatous hyperplasia (AH) of the liver occurs in chronic advanced liver diseases, and has been suspected to be a precancerous lesions in human hepatocarcinogenesis. METHODS: In this study, the incidence of hepatocellular carcinoma (HCC) after surgical resection of AH was evaluated in 24 patients with cirrhosis and AH in the absence of HCC (mean follow-up period, 31.4 months; range, 12-77 months). AH were classified as ordinary AH lacking hepatocellular atypia (OAH), atypical AH with structural and cellular atypia insufficient for carcinoma (AAH), and atypical AH with focal malignancy containing areas of HCC (FM). RESULTS: HCC was noted in a few years (follow-up period range, 12-77 months; mean, 31.4 months) in all 3 patients whose resected nodules were classified as FM, in 4 (36%) of 11 with AAH resected nodules, and none of 10 with OAH resected nodules. The incidence of HCC in the patients with FM or AAH nodules was significantly higher than that in those with OAH nodules. CONCLUSIONS: These results suggest that our classification of AH is useful in the assessment of the risk of HCC in patients with cirrhosis and AH, and that those with AAH or FM resected nodules must be followed up more frequently than those with OAH nodules, because of their higher risk of HCC. The frequent occurrence of HCC after resection of FM or AAH nodules may imply that HCC is of multicentric origin.

Adult↗
PubMed1993

Augmentation of endothelin-1, prostacyclin and thromboxane A2 secretion associated with in vitro ageing in cultured human umbilical vein endothelial cells.

Differences in the secretion of the vascular regulators endothelin-1 (ET-1, prostacyclin (PGI2) and thromboxane A2 (TXA2) associated with ageing were investigated in cultured endothelial cells isolated from normal human umbilical veins (HUVECS). HUVECS at different population doubling levels (PDLs) were cultured in medium MCDB-104 supplemented with FBS and ECGF. Cell saturation density was determined by a Coulter counter, and concentrations of ET-1, PGI2 and TXA2 in the media were determined by radioimmunoassay. The cellular and nuclear size of HUVECS increased with advancing age, and the dividing ability decreased. Cell saturation density of HUVECS decreased 5-fold between PDLs 7 and 67 (P < 0.01). The secretion of ET-1 by HUVECS at a young stage of growth (PDL 7.6) increased linearly between 0 and 36 h of incubation (P < 0.01). ET-1 secretion increased approximately 3-fold between PDLs and 67 (P < 0.01). PGI2 secretion increased 6-fold between PDLs 7 and 67 (P < 0.01), and TXA2 secretion increased 18-fold between PDLs 7 and 67 (P < 0.01). The ratio of PGI2 to TXA2 secretion decreased 3-fold between PDLs 7 and 40 (P < 0.01), and remained at the lower ratio between PDLs 40 and 67. This data indicates that the anti-thrombotic or anti-vasoconstrictive role of endothelial cells may decrease during in vitro ageing.

Aging↗
PubMed1993

Reduction of nitric oxide producing activity associated with in vitro aging in cultured human umbilical vein endothelial cell.

To investigate the involvement of nitric oxide (NO) in the pathogenesis of the cardiovascular system with aging, we studied the basal release of NO in cultured endothelial cells isolated from normal human umbilical veins (HUVECs) at different population doubling levels (PDLs). The release of NO was assayed by a newly established, highly sensitive assay system monitoring guanosine 3':5'-cyclic monophosphate (cGMP) formation in a pig kidney epithelial cell line cocultured with HUVECs. The cGMP formation by NO released from HUVECs decreased 3.3-fold between PDLs 8 (14.1 pmol/ml) and 50 (4.3 pmol/ml) (P < 0.01). The result suggests that the vaso-constrictive activity of endothelial cells increases during in vitro aging which may reflect the increased vascular disorders in aged people.

1-Methyl-3-isobutylxanthine↗
PubMed1993

Induction of apoptosis by hemorrhagic snake venom in vascular endothelial cells.

Vascular degeneration appears to play crucial roles in producing many vascular malfunctions (1-3). In order to identify specific inducers of programmed death in vascular endothelial cells (VEC), examinations were made of the effects of substances that are known to affect the vascular system by using VEC in culture (4,5). We found that hemorrhagic snake venoms induced apoptotic cell death or programmed cell death of VEC. By contrast, neurotoxic snake venoms did not induce programmed cell death but caused necrosis at much higher doses of the venoms. No effect of hemorrhagic venom was observed with many types of cultured cells other than VEC. Thus, hemorrhagic snake venom appears to be a useful tool for studies of the molecular mechanisms of vascular apoptosis. The results also suggest a possible mechanism of action of hemorrhagic snake venom on the vascular system.

Animals↗
PubMed1993

Protective effect of lipophilic derivatives of ascorbic acid on lipid peroxide-induced endothelial injury.

The protective effects of ascorbic acid and its lipophilic derivatives on the cytotoxicity of linoleic acid hydroperoxide (LOOH), a primary product of lipid peroxidation, was investigated in cultured human umbilical vein endothelial cells. When cells were incubated with ascorbic acid for 24 h before treatment with LOOH, ascorbic acid did not protect cells from oxidative damage by LOOH. Its lipophilic derivatives such as 6-O-palmitoyl,6-O-stearoyl, and 2,6-O-dipalmitoyl esters and 2-O-octadecyl ether (CV3611) had an apparent protective effect on the toxicity of LOOH. When cells were incubated in medium containing both antioxidant and LOOH, the protective effect of lipophilic derivatives became weak. An interaction between alpha-tocopherol and ascorbic acid was not observed in this system using endothelial cells. However, in the case where the cells were preincubated with alpha-tocopherol and 6-O-palmitoylascorbic acid, the ascorbic acid derivative had a synergistic effect to alpha-tocopherol in protecting against LOOH-induced damage. These results suggest that lipophilic antioxidants incorporated into cellular membranes are effective protectors against LOOH, whereas lipophobic antioxidants located outside the membrane do not have any protective effect.

Ascorbic Acid↗
PubMed1993

Inhibition of cell proliferation by a unique lysophosphatidic acid, PHYLPA, isolated from Physarum polycephalum: signaling events of antiproliferative action by PHYLPA.

The unique Physarum lysophosphatidic acid, PHYLPA, having a cyclopropane in the fatty acid moiety and a cyclic phosphate at C-2 and C-3 positions of the glycerol, inhibited proliferation of human fibroblast cells, TIG-3 and TIG-7, which were cultured in a chemically defined (serum-free) medium. The cells at S- and M-phases proceeded to G2- and G1-phases, respectively, and most of cells were arrested at G1- or G2-phase during PHYLPA treatment. The growth was recovered when PHYLPA was removed from the medium. In the presence of serum, PHYLPA did not show obvious inhibitory effects, indicating the existence of a factor(s) which neutralizes the antiproliferative activity of PHYLPA. PHYLPA elicited an increase in 3',5'-cyclic adenosine monophosphate (cAMP) in a biphasic fashion in fibroblast cells. It also elicited inositol phosphate accumulation, as well as a transient rise in cytoplasmic free Ca2+ ion.

Animals↗
PubMed1992

Inhibition of eukaryotic DNA polymerase alpha with a novel lysophosphatidic acid (PHYLPA) isolated from myxoamoebae of Physarum polycephalum.

A specific inhibitor of DNA polymerase alpha was isolated from the lipid fraction prepared from myxoamoebae of a true slime mold, Physarum polycephalum. The purified substance was subjected to structural studies by fast atom bombardment mass spectroscopy, infrared spectroscopy, and two-dimensional nuclear magnetic resonance spectroscopy. The structure of this substance was thereby suggested to be a novel lysophosphatidic acid (LPA) composed of cyclic phosphate and cyclopropane-containing hexadecanoic acid. Then we named this substance PHYLPA (Physarum LPA). PHYLPA inhibited more than 80% of the affinity-purified calf thymus DNA polymerase alpha activity at a concentration of 10 micrograms/ml (approximately 20 microM). Inhibition was observed for DNA polymerase alpha but not for DNA polymerase beta or gamma from various eukaryotic species, nor did it inhibit DNA polymerase I from E. coli. From kinetic analyses, the inhibition was considered to be caused by the interaction of PHYLPA with the template DNA.

Animals↗
PubMed1992

Salvage of infarcted myocardium by angiogenic action of basic fibroblast growth factor.

Coronary collateral vessels reduce damage to ischemic myocardium after coronary obstruction. Factors that stimulate collateral formation are expected to have ameliorating effects on myocardial infarction. In a canine experimental myocardial infarct model, intracoronary injection of basic fibroblast growth factor (bFGF) improved cardiac systolic function and reduced infarct size. Treatment with bFGF increased the number of arterioles and capillaries in the infarct. Thus, the angiogenic action of bFGF might lead to a reduction in infarct size. The application of bFGF might bring about a therapeutic modality for the salvage of infarcted myocardium.

Animals↗
PubMed1992

Species-specific differences in the mitogenic activity of heparin-binding growth factors in the sera of various mammals.

Sera from different mammalian species displayed great differences in mitogenic activity, as measured by stimulation of DNA synthesis in BALB/c 3T3 cells (3T3 cells). Among the sera examined, fetal bovine serum was least active, and increasing activity was detected in calf serum, human serum, rat serum and mouse serum, in that order. Rat and mouse sera exhibited extremely high mitogenic activity with 3T3 cells, but when TIG-1 human fetal lung fibroblasts were used for the DNA assay instead, the activity levels of all of the sera were lower, and the differences between them were smaller. To determine the reasons for these differences, the heparin-binding growth factors in each serum were separated on a heparin affinity column. Five peaks of DNA-stimulating activity were obtained. Three of these were found in all sera examined, with both 3T3 cells and TIG-1 cells. Two other peaks were found only with 3T3 cells; one was peculiar to rat and mouse sera, with extremely high activity in the rat, and the other was specific to fetal serum. The dependence of the activity of these peaks on the cells used for the test was confirmed using normal rat lung fibroblasts and immortalized rat kidney cells. These findings adequately explain the species-specific differences in mitogenic activity of whole sera, and the variation in activity depending on the cells used for assay of DNA synthesis.

3T3 Cells↗
PubMed1992

Transforming growth factor beta 1-specific binding proteins on human vascular endothelial cells.

Transforming growth factor beta (TGF beta) regulates the growth of human umbilical vein endothelial cells (HUVEC) differently depending on the isoform of TGF beta and the culture conditions. The cells are resistant to growth inhibition by TGF beta when the cells are cultured on substratum coated with gelatin. However, the proliferation of HUVEC cultured on substratum without a gelatin coating is inhibited by TGF beta, depending on the isoform and concentration of TGF beta. Binding assays with 125I-TGF beta 1 reveal that HUVEC contain a single class of high-affinity (Kd = 4.4 pM) TGF beta 1 binding sites with 8500 sites per cell. Affinity cross-linking studies demonstrate that HUVEC express 180 and 80 kDa TGF beta 1 binding sites that do not bind TGF beta 2. The reduction and the removal of glycosaminoglycans does not affect the electrophoretic mobility of the 180-kDa binding protein cross-linked with 125I-TGF beta 1. Therefore, the 180-kDa TGF beta 1 binding protein is not related to the type III TGF beta receptor, but might be a novel TGF beta 1-specific receptor/binding protein expressed on vascular endothelial cells. The expression of TGF beta 1 binding sites is not affected by the presence or absence of the gelatin coating on the culture substratum. The data suggest that a gelatin coating does not regulate the susceptibility of HUVEC to TGF beta 1 at the level of the receptor/binding proteins, and that growth inhibition of HUVEC by TGF beta 1 is linked to the regulation of extracellular matrices required for the interaction between the cells and the substratum.

Binding Sites↗