Biomedical subjects

Nobuo Suzuki

Publications and source records attributed to Nobuo Suzuki.

30 records · Page 2Linked to original sources

PubMed2003

Bisphenol A suppresses osteoclastic and osteoblastic activities in the cultured scales of goldfish.

The direct effect of bisphenol A on osteoclasts and osteoblasts was examined using a culture system of goldfish scales. Tartrate-resistant acid phosphatase (TRAP) and alkaline phosphatase (ALP) were used as markers of osteoclasts and osteoblasts, respectively. Bisphenol A (10(5) M) significantly suppressed both TRAP and ALP activities. These data were reproduced in an in vivo experiment. From an analysis of a reverse transcription-polymerase chain reaction in the in vitro-cultured scales, it was demonstrated that the insulin-like growth factor (IGF)-1 mRNA expression decreased by a bisphenol A treatment. On the other hand, 17beta-estradiol (E(2)) stimulated both TRAP and ALP activities and did not change IGF-1 mRNA expression, suggesting that bisphenol A has a different effect from E(2) on bone metabolism. This study is the first to demonstrate that bisphenol A functions to suppress directly osteoblasts and osteoclasts among vertebrates, which strongly suggests that this scale in an in vitro assay system can be utilized for the evaluation of the effects of endocrine disrupters on bone cells.

Acid Phosphatase↗

PubMed2003

Increased levels of UV-induced protease activity in human UVAP-1 cells exposed to gravity-changing stress: involvement of E-64-sensitive proteases in suppression of UV mutagenicity.

Under the 1G condition, the increase in antipain-sensitive protease activity promptly after UV (mainly 254 nm wavelength) irradiation in cultured human cells is detected and found to be one of the intriguing events involved in suppression of cell mutability. It was found that two cell lines, RSa and its variant UVAP-1 cells are applicable; the former is hypermutable and not susceptible to protease activation, while the latter is hypomutable and susceptible. In the present study it was investigated whether the increase in protease activity by UV irradiation is also observed in hypomutable human UVAP-1 cells exposed to gravity-changing stress and whether the increase is involved in suppression of UV mutagenicity. Exposure of human UVAP-1 cells to gravity-changing stress such as free-fall and parabolic flight prior to UV irradiation resulted in a pronounced increase in protease activity, but not to hypergravity conditions (2 and 10G) prior to UV irradiation. To characterize the proteases, components of lysates from the cells exposed to free-fall prior to UV irradiation were fractionated by high performance liquid chromatography, indicating two separate fractions with highly increased levels of E-64-sensitive protease activity. In the cells treated with E-64 during their exposure to free-fall, K-ras codon 12 base substitution mutation was detected after UV irradiation, although the mutation was not detected after UV irradiation alone. Thus, the increase in E-64-sensitive protease activity may be involved in the suppression of UV mutagenicity in UVAP-1 cells exposed to free-fall.

Antipain↗

PubMed2003

Induction of apoptosis and ubiquitin hydrolase gene expression by human serum factors in the early phase of acute myocardial infarction.

The physiologic events leading to apoptosis in myocardial infarction and the molecules involved in the death process have not been clarified unequivocally. We developed a method to search for serum factors that induce apoptosis of human cells, using serum obtained from patients within 1 day of the onset of acute myocardial infarction (AMI). Serum factors were found to have the ability to increase the caspase-3 activity levels in human RSa cells, which are susceptible to apoptosis inducers. The factors obtained from AMI patients by elution at about 0.5 mol/L KCl from a dye-ligand column were named AMI-SFs (serum factors from AMI). Electrophoretic analysis showed DNA fragmentation in AMI-SF-treated RSa cells, but not in RSa cells treated with fractions from AMI patients 1 week after clinical onset of illness. AMI-SF-induced DNA fragmentation was also demonstrated by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling analysis, whereas a suppression of fragmentation was seen in RSa cells treated with AMI-SFs in combination with a caspase-3 inhibitor. The increase in caspase-3 activity was not inhibited by neutralizing antibodies to tumor necrosis factor-alpha, interleukin-6, human interferon-beta, or interferon-gamma. Polymerase chain reaction-based messenger RNA differential display and Northern blotting revealed an increase in the messenger RNA expression level of human ubiquitin hydrolase in AMI-SF-treated RSa cells. Antisense oligonucleotides for ubiquitin hydrolase inhibited the increase in caspase-3 activity. These findings suggested that serum from AMI patients in the acute phase contains factors that induce apoptosis, possibly by inducing the expression of the ubiquitin hydrolase gene, at least in the human cells tested.

Adult↗

PubMed2003

Increased expression of the Huntingtin interacting protein-1 gene in cells from Hutchinson Gilford Syndrome (Progeria) patients and aged donors.

Hutchinson Gilford syndrome (progeria [PG]) is a human disease associated with accelerated aging. To elucidate the acceleration mechanism, we first tried to transform a PG-derived cell line by infection of a recombinant adenovirus expressing HPV (human papilloma virus)-E6 and HPV-E7 genes. The transfected PG cells had a greater number of population doublings (PD) (>80), faster doubling time, and less staining of senescence-associated ss-galactosidase than the nontransfected PG cells. The transfected cells also showed markedly more detectable telomerase activity than the nontransformed cells. The expression levels of the genes in the E6-transduced and E7-transduced cell line were then compared with those of the nontransfected cell line using an mRNA differential display method, following reverse-transcriptase polymerase chain reaction analysis. Expression of huntingtin interacting protein-1 (HIP-1) gene was found to be increased not only in PG cells but also in fibroblast cells from aged healthy donors. Thus, HIP-1 might be a molecular assistant in the pathogenesis of the cellular senescent process in the human cells tested.

Adolescent↗

PubMed2003

Bisphenol A influences the plasma calcium level and inhibits calcitonin secretion in goldfish.

In teleosts, it is well known that plasma calcium levels increase as a result of treatment with estrogen for at least during 2 weeks and that calcitonin secretion is induced by estrogen. The present study examined the influence of bisphenol A on calcium homeostasis in goldfish and compared the above known estrogenic action. In goldfish kept in water containing bisphenol A (10(-6) M), the plasma calcium concentration increased significantly (P<0.001) at 4 days but decreased significantly (P<0.05) at 8 days. By the treatment of bisphenol A, calcitonin secretion was not induced until 4 days. At 8 days, however, plasma calcitonin, as well as calcium, decreased significantly (P<0.05), although vitellogenin was detected in the plasma. Therefore, bisphenol A influences plasma calcium levels, but its action is different from that of estrogen, which indicates that bisphenol A affects the calcium homeostasis and might bring about abnormal conditions in teleosts.

Animals↗

PubMed2003

Effect of resistance training on changes in lymphocyte subpopulation during head-down-tilt bed rest.

Head-down-tilt (HDT) bed rest imitates microgravity-like conditions for a supine human body on the ground. On the other hand, resistance training is used to eliminate unfavorable physical changes in astronauts under a microgravity environment during spaceflight. Twelve healthy males voluntarily participated in a 20-day -6 degrees HDT bed rest study. The ratio of CD3-positive peripheral blood lymphocytes were analyzed by immunofluorescence staining. Volunteers who undertook resistance-training during HDT bed rest were designated as the resistance training group (RTG), and those who did not as non-resistance-training group (N-RTG). We could not observe significant differences in the ratio of CD3-positive peripheral blood lymphocytes between RTG and N-RTG. Thus, resistance training during HDT bed rest might not have a significant effect on changes in the ratios of CD3-positive PBL.

Adult↗

PubMed2002

Involvement of human heat shock protein 90 alpha in nicotine-induced apoptosis.

There have been conflicting reports of the apoptotic effects of nicotine on human cells and those studies reporting nicotine-induced apoptosis have not unequivocally clarified the molecular mechanisms underlying the effect. However, we found here that human RSa cells, established from embryonic fibroblastic cells doubly infected with Rous sarcoma virus and Simian virus 40, underwent apoptosis when cultured with medium containing 0.06-0.6 microM nicotine. The apoptosis was assessed by cellular DNA fragmentation and caspase-3 protease activation. Viability of RSa cells was reduced by nicotine treatment, as analyzed by MTT assay and the reduction was lessened by combination treatment with a caspase-3 inhibitor, acetyl-L-aspartyl-L-glutamyl-L-valyl-L-aspart-1-al (Ac-DEVD-CHO). Levels of expression of heat shock protein 90 alpha (Hsp90 alpha) were found to be increased 20 min after the nicotine treatment, as analyzed by polymerase chain reaction-based mRNA differential display after Northern blotting analysis of mRNA amounts. Cellular contents of Hsp90 alpha were furthermore increased in the nicotine-treated RSa cells, as quantitated by Western immunoblot analysis. By contrast, in RSa cells treated with nicotine in combination with geldanamycin (GA), an inhibitor of Hsp90 alpha function, DNA fragmentation was not detected and caspase-3 protease activity levels were the same as those of mock-treated cells. Nicotine-induced caspase-3 activation and Hsp90 alpha expression, as well as suppression of the induction by GA, were also observed in a xeroderma pigmentosum patient-derived cell line, XP2OS cells. Thus, it was suggested that nicotine induces apoptosis, possibly via Hsp90 alpha expression, in human cells tested.

Apoptosis↗

PubMed2002

Effects of 3-D clino-rotation on gene expression in human fibroblast cells.

Continuous variation in the direction of the gravity vector leads to various cellular responses including modulation of gene expression. Complementary DNA (cDNA) array analyses are available to observe the variation of gene expression under different conditions. In this study, expression levels of 588 representative genes were compared using the Atlas human cDNA expression array in human fibroblast cells with and without 3-dimensional (3-D) clinostat. Five upregulated and 8 downregulated genes were detected. Among these genes, upregulation of XRCC1, and downregulation of ERB-B2 and p21(Cip1/Waf1) were confirmed by RT-PCR. These results suggested that the gene expression levels of XRCC1, ERB-B2 and p21(Cip1/Waf1) were modulated by vector-averaged microgravity induced by 3-D clinostat in human fibroblast cells. Our findings may be a basis for the biological study of 3-D culture systems.

Cell Culture Techniques↗

PubMed2002

Melatonin suppresses osteoclastic and osteoblastic activities in the scales of goldfish.

The effects of melatonin on osteoclastic and osteoblastic cells were examined using a culture system of the goldfish scale. Tartrate-resistant acid phosphatase (TRACP) and alkaline phosphatase (ALP) were used as markers of osteoclastic and osteoblastic cells, respectively. In Earle's minimum essential medium containing melatonin (10(-9) to 10(-5) m), activities of both enzymes in scales were significantly suppressed at 6 hr after incubation (TRACP: 10(-8), 10(-6), 10(-5) m; ALP: 10(-7) to 10(-5) m), but at 18 hr only ALP activity was significantly lowered (10(-8), 10(-7) m). Estradiol-17beta (E(2)) enhanced both activities, which were significantly inhibited and brought down to the level of the controls when co-incubated with E(2) and melatonin (TRACP at 6 hr: 10(-9) to 10(-5) m; ALP at 6 hr: 10(-7) m; ALP at 18 hr: 10(-8) m). Moreover, using reverse-transcription polymerase chain reaction, the mRNA expression of the estrogen receptor (ER) and insulin-like growth factor (IGF)-1, which are related to osteoblastic growth and differentiation, was decreased in the melatonin-treated scales. These results suggest that melatonin acts directly on the scale osteoclastic and osteoblastic cells where it suppresses the ALP activity via down-regulation of ER and IGF-1 mRNAs expression. This is the first report on the function of melatonin in osteoclasts and on the suppressive effect of melatonin in osteoblasts among vertebrates.

Acid Phosphatase↗

PubMed2002

Establishment and characterization of GSA-1, a human cell line highly susceptible to apoptosis after free-fall.

The induction of apoptosis by microgravity and/or gravity-changing stress is considered to be one of the important causes of cell death, although the molecular mechanisms of the apoptotic event remain unclarified. In this study, we established a cell line, GSA-1, from ethyl methanesulfonate-treated human RSa cells. GSA-1 cells were highly susceptible to apoptosis after a free-fall; 24.4% of these cells underwent apoptosis after free-fall, compared with only 6% of the RSa cells. The apoptosis of GSA-1 cells was augmented by ultraviolet (UV, principally 254-nm wavelength) irradiation before free-fall to a greater extents than those in RSa cells. The molecular mechanisms of apoptosis included p53 and Bax proteins; the expression of nuclear p53 and cytoplasmic Bax in GSA-1 cells increased at 4 h after free-fall irrespective of irradiation. In addition, the rate of removal of cyclobutane pyrimidine dimer (CPD) in UV-irradiated GSA-1 cells was higher in cells exposed to free-fall than in those under the 1-G condition. Our results suggested that in GSA-1 cells, free-fall accelerates apoptosis, and that this process is associated with the accumulation of p53 and Bax, as well as CPD removal. Thus, GSA-1 cells should be useful for investigating the mechanism of cellular response, including the induction of apoptosis under gravity-changing stress.

Apoptosis↗

PubMed2002

Enhanced expression of the LDH-A gene after gravity-changing stress in human RSa cells.

A major issue in radiation and space biology is whether gene expression levels are altered in cells exposed to gravity-changing stress. In the present study, genes up- or down-regulated in radiation-sensitive human RSa cells cultured under gravity-changing conditions, were identified using a PCR-based mRNA differential display method. Exposure of cells to gravity-changing stress was performed by free-fall with a drop-shaft facility or by an airplane-conducted parabolic flight. Among the candidates for gravity-changing stress-responsive genes obtained by the differential display analysis, the lactate dehydrogenase A gene (LDH-A) was confirmed by Northern blotting analysis to exhibit increased expression levels. The gravity-changing stress consisted of a combination of microgravity and hypergravity. However, exposure of the cells to hypergravity produced by centrifuge only slightly affected the LDH-A mRNA expression. Thus, LDH-A was found to be a candidate for the genes which play a role in the cellular response to gravity-changing stress, and mainly to microgravity.

Cell Line↗

PubMedDate not supplied

Involvement of human small fragment nuclease in the resistance of human cells to UV-C-induced cell death.

Human small fragment nuclease (Sfn) is one of the cellular proteins that were reported to degrade small, single-stranded DNA and RNA. However, the biological role of Sfn in cellular response to various stressors such as UV-C (mainly 254 nm wavelength ultraviolet ray) remains unclear. We have examined whether modulation of human SFN gene expression affects cell survival capacity against UV-C-induced cell death, analyzing colony survival ability in UV-C-sensitive human RSa cells treated with short double-stranded RNA (siRNA) specific for SFN messenger RNA (mRNA). The expression levels of SFN mRNA in the siRNA-treated RSa cells decreased to about 15% compared with those in the control siRNA-treated cells. The siRNA-treated RSa cells showed lower colony survival and higher activity of caspase-3 after UV-C irradiation than the control siRNA-treated RSa cells. Furthermore, the removal capacity of cyclobutane pyrimidine dimers (CPD) in the siRNA-treated RSa cells decreased compared with the control siRNA-treated RSa cells. There was no difference in the colony survival and CPD removal capacity after UV-C irradiation between the control siRNA-treated RSa cells and mock-treated RSa cells. These results suggest that SFN expression is involved in resistance of RSa cells to UV-C-induced cell death through the roles it plays in the DNA repair process.

14-3-3 Proteins↗