Biomedical subjects

T Sugimura

Publications and source records attributed to T Sugimura.

At least 181 records · Page 10Linked to original sources

PubMed1997

Suppression of nitric oxide production in lipopolysaccharide-stimulated macrophage cells by omega 3 polyunsaturated fatty acids.

Although nitric oxide (NO) is an important biological mediator, its excessive production in inflammation is thought to be a causative factor for cellular injury and, over the long term, cancer. In the present study, the effects of several fatty acids on NO production in murine macrophage cell line RAW264 cells stimulated with lipopolysaccharide were examined. Suppression of NO production was observed with the omega 3 polyunsaturated fatty acids (PUFAs), docosahexaenoic acid, eicosapentaenoic acid and alpha-linolenic acid, in a dose-dependent fashion. In contrast, no inhibition was observed with omega 6 PUFA (linoleic acid), omega 9 PUFA (oleic acid) or a saturated fatty acid (stearic acid). Western and northern blot analyses suggested that suppression of the induction of inducible NO synthase gene expression is responsible for the inhibition of NO production by omega 3 PUFAs. The inhibitory effect of omega 3 PUFA on NO production in activated macrophages could contribute to their cancer chemopreventive influence.

Animals↗

PubMed1997

Identification of canine calicivirus capsid protein and its immunoreactivity in western blotting.

A canine calicivirus (CaCV) isolated in Japan, designated as CaCV No. 48 strain, was propagated in MDCK cells and purified by CsCl equilibrium gradient centrifugation. Sodium dodecyl sulfate-polyacrylamide gel electrophoretic analysis of the purified samples revealed the presence of only one major species of viral protein of about 60 kilodaltons after Coomassie staining. The same band, presumably that of the capsid protein, was detected by western blotting using a mouse hyperimmune serum. This capsid protein was synthesized in MDCK cells as early as 2 hr post-inoculation. Experimental infection of dogs resulted in the production of anti-CaCV antibodies which were detected by microneutralization test and western blotting. Likewise, serosurvey revealed not only the presence of neutralizing antibodies but also reactivity of the field sera against the capsid protein of the purified virus. These results indicate that the capsid protein of CaCV No. 48 strain is immunogenic and could be detected by antibodies in western blotting.

Animals↗

PubMed1997

Effective prevention of thrombocytopenia using adenovirus-mediated transfer of HST-11FGF-4 gene: in vivo and in vitro studies.

A novel biological function of HST-1 protein (FGF-4) was investigated by constructing an adenovirus vector containing the HST-1 cDNA and applied for thrombocytopenia as a gene therapy. A single intraperitoneal injection of the replication-deficient adenovirus containing the HST-1 gene (Adex1HST-1) into mice caused a two-fold increase in peripheral platelet count for 30 days, and effectively prevented experimentally induced thrombocytopenia. Studies of Adex1HST-1-infected or HST-1 protein-treated megakaryocytic Dami cells suggested that HST-1 protein promotes megakaryocyte maturation, and increases cytokine secretion from megakaryocyte and adhesive interactions between megakaryocyte and endothelial cells. Colony assay revealed that HST-1 protein stimulated CFU-MK (colony-forming unit of megakaryocyte) not alone but synergistically with early-acting cytokines such as IL-3 or Tpo (c-mpl ligand) as a megakaryocyte potentiating factor. These results have important implications for clinical application of the Adex1HST-1 for thrombocytopenia.

Adenoviridae↗

PubMed1997

Suppression of Ki-ras p21 levels leading to growth inhibition of pancreatic cancer cell lines with Ki-ras mutation but not those without Ki-ras mutation.

Ki-ras point mutation characteristically occurs frequently in human pancreatic cancer. To clarify the effect of antisense Ki-ras RNA on various pancreatic cancer cell lines, a plasmid expressing an antisense Ki-ras gene fragment (AS-K-ras-LNSX) was transduced into seven human pancreatic cancer cell lines (AsPC-1, MIA PaCa-2, PANC-1, PSN-1, BxPC-3, Hs 700T, and Hs 766T) by liposome-mediated transfection. Western blot analysis showed that transfection of AS-K-ras-LNSX led to a significant reduction in the amounts of Ki-ras p21 protein in all the pancreatic cancer cell lines except BxPC-3. The growth of pancreatic cancer cell lines having Ki-ras point mutations (AsPC-1, MIA PaCa-2, PANC-1, and PSN-1) was suppressed after transduction of AS-K-ras-LNSX, whereas the effect of the antisense construct on the growth was not significant in cell lines with a wild-type Ki-ras gene (BxPC-3, Hs 700T, and Hs 766T). These results suggest that the pancreatic cancer cells with activated Ki-ras depend heavily on a Ki-ras p21-mediated growth signal pathway for their growth because they were far more susceptible to the suppression of the Ki-ras p21 protein than the cells with wild-type Ki-ras. The remarkably increased dependence of the cancer-cell growth circuitry on one or a few crucial regulatory molecules may thus be a common feature of the cancer cells and implies a novel rationale for the targeting of cancer therapy.

Cell Division↗

PubMed1997

[A case of incomplete Kearns-Sayre syndrome with a stroke like episode].

A 32-year-old woman developed chronic progressive hearing impairment, trunkal ataxia, bilateral ptosis and external ophthalmoplegia. She also showed slowly progressive mild to moderate proximal dominant muscle weakness and atrophy. ECG showed incomplete right bundle branch block. An aerobic exercise test showed abnormal blood lactate elevation and muscle biopsy revealed ragged-red fibers in addition to the myopathic change. Analysis of mitochondrial DNA extracted from biopsied muscle and fibroblast samples revealed a 1,758bp deletion from the cytochrome b to ND6 coding regions. Common mutations in tRNALeu(UUR) coding region to the mitochondrial encephalomyopathy, lactic acidosis and stroke-like episodes (MELAS) were not present. She was diagnosed as having incomplete Kearns-Sayre syndrome (KSS). Since the age of 35, she developed complex partial seizure attacks with secondary generalization frequently and at the age of 42, she had a severe generalized seizure with delayed consciousness loss followed by left hemiplegia. MRI showed wide T2-high signal lesions in the right temporo-parieto-occipital area. The proton MR-spectroscopy showed prominent increase of lactate beyond the lesions detected by MRI, indicating diffuse aerobic metabolic dysfunction in the central nervous system. We reviewed two other KSS cases with a stroke like episode, who also had epilepsy and large deletion but no tRNALeu(UUR) mutation, in mitochondrial DNA. Patients with KSS who have seizure may develop the stroke-like episode as seen in MELAS patients.

Adult↗

PubMed1997

No involvement of APC gene mutations in ulcerative colitis-associated rat colon carcinogenesis induced by 1-hydroxyanthraquinone and methylazoxymethanol acetate.

A rat model for human ulcerative colitis (UC) has been developed by using 1-hydroxyanthraquinone (1-HA) to cause severe inflammation of colonic mucosa. 1-HA also has synergistic effects on the carcinogenicity of methylazoxymethanol (MAM) acetate in the rat colon. In this study, four adenomas and 16 adenocarcinomas induced in male F344 rats by 1-HA and MAM acetate were examined for mutations in the entire coding regions and introns flanking coding exons of the APC gene by polymerase chain reaction (PCR)-single strand conformation polymorphism (SSCP) and PCR-restriction-SSCP analyses. No mutations were found. These results, together with our previous observations of a relative lack of Ki-ras gene mutations in the same tumors, are similar to those found in human UC-associated colon cancer, suggest a common pathway in these two systems, although they are different in their implication of p53 mutations. Therefore, this model may have some relevance and application to the study of colon cancer in human inflammatory bowel disease, which is not associated with APC mutations or with Ki-ras or p53 mutations.

Animals↗

PubMed1997

Combined analysis of microsatellite instability and K-ras mutation increases detection incidence of normal samples from colorectal cancer patients.

Microsatellite instability (MI) and K-ras oncogene mutation have been widely used as biomarkers of genetic changes in colorectal cancer (CRC). Each of these biomarkers was independently found in normal-appearing colonic mucosa at stages preceding the development of CRC, albeit at a relatively low incidence. To assess the potential value of combined MI and K-ras mutation analysis in the detection of normal-appearing colonic mucosa samples taken from patients with CRC, we have chosen to analyze multiple (3-7) normal colonic mucosa samples and the respective colorectal tumor tissues from 20 patients with CRC. As a control, we have used 54 normal mucosa samples obtained from 9 autopsies of patients without CRC. In at least 1 of 5 loci analyzed, MI was found in 8 of 20 patients via analysis of multiple normal-appearing colonic mucosa samples from each patient. Combined analysis of MI and mutant ras alleles in normal-appearing colonic mucosa samples enabled the identification of 11 of 20 patients with CRC. None of the 54 normal colonic mucosa samples obtained from 9 patients without CRC were found to carry mutant ras or MI. The ability to detect 55% of patients with CRC via the analysis of normal mucosa samples provides an important advance in our approach toward early detection of individuals who may be at risk to develop this tumor type.

Aged↗

PubMed1996

Identification and characterization of three isotypes of protein phosphatase inhibitor-2 and their expression profiles during testis maturation in rats.

cDNAs for three isotypes of inhibitor-2(I-2), I-2 alpha 1, I-2 alpha 2 and I-2 beta were isolated from a rat testis library. I-2 alpha 2 and I-2 beta are new forms. The former is an alternatively spliced form of I-2 alpha 1, encoding a protein with substitution of three amino acids for 14 amino acids of the I-2 alpha 1 protein at the C-terminus. The latter is derived from a different gene and encodes a 126-amino-acid protein having highly conserved regions with the I-2 alpha 1 protein from amino acid positions 22-47 and 111-126. I-2 alpha 2 and I-2 beta are expressed exclusively in the testis, and the expressions of all three forms of I-2 coincide with sperm cell maturation. The half-maximal inhibitory concentration of the GST-I-2 alpha 2 fusion protein on the PP1 gamma 2 catalytic subunit is the same as that of GST-I-2 alpha 1, being 10 nM. However, the half-maximal inhibitory concentration of GST-1-2 beta is 100-fold higher, being 1 microM. GST-I-2 beta showed no competition with GST-I-2 alpha 1. and its biological significance is unknown.

Amino Acid Sequence↗

PubMed1996

Long-term consequences of Kawasaki disease. A 10- to 21-year follow-up study of 594 patients.

BACKGROUND: The long-term consequences of the cardiovascular sequelae in Kawasaki disease remain uncertain. METHODS AND RESULTS: We identified 594 consecutive children with acute Kawasaki disease between 1973 and 1983, and this cohort was followed up for 10 to 21 years (mean, 13.6 years). In all patients, we evaluated coronary lesions by coronary angiography just after the acute stage. One hundred and forty-six patients (24.6%) were diagnosed as having coronary aneurysms. A second angiogram was performed 1 to 2 years later in all 146 patients who previously had coronary aneurysms, which demonstrated that 72 (49.3%) of these 146 had regression in the coronary aneurysm. A third angiogram was performed for 62 patients, a fourth for 29, and a fifth for 17. By 10 to 21 years after the onset of the illness, stenosis in the coronary aneurysm had developed in 28 patients. Myocardial infarction occurred in 11 patients, 5 of whom died. In the 26 patients with giant coronary aneurysms, stenotic lesions developed in 12, and no regression occurred. The 448 patients with normal findings at the first angiogram subsequently never developed any abnormal cardiac findings. Systemic artery aneurysms developed in 13 patients (2.2%), and valvular heart disease appeared in 7 (1.2%). CONCLUSIONS: The incidence of coronary aneurysm in acute Kawasaki disease was 25%, 55% of which showed regression. During follow-up, ischemic heart disease developed in 4.7% and myocardial infarction in 1.9%. Death occurred in 0.8%.

Aneurysm↗

PubMed1996

Use of von Willebrand factor promoter to transduce suicidal gene to human endothelial cells, HUVEC.

Angiogenesis is an essential component of multifactorial carcinogenesis and thus a potential target of therapeutic intervention. To develop a novel cancer gene therapy strategy based on suppression of tumor angiogenesis, we examined the feasibility of targeting and preferential killing of proliferating endothelial cells by use of the von Willebrand factor (vWf) promoter and herpes simplex virus thymidine kinase gene (HSV-TK). Based on previous reports on the vWf promoter, we tested two putative vWf promoter regions. The luciferase assay showed that the shorter region, which encompasses most of the first noncoding exon, had stronger activity in endothelial cells. Although the promoter activity was low when employed as an internal promoter for retroviral and adenoviral vectors, endothelial cell specificity was suggested; the promoter, when used to drive the HSV-TK gene, could preferentially suppress endothelial cell growth in the presence of prodrug ganciclovir, suggesting the feasibility of designing an anti-angiogenesis gene therapy using the vWf promoter and the suicide gene/prodrug strategy.

Cell Division↗

PubMed1996

Cloning, expression and chromosomal localization of Wnt-13, a novel member of the Wnt gene family.

The Wnt genes, encoding structurally-related secreted glycoproteins, are implicated in mammary carcinogenesis induced by mouse mammary tumor virus. In search of the Wnt gene(s) expressed in human gastric cancer, a WTGC1 cDNA fragment sharing 66.9% amino-acid homology with human and mouse Wnt-2 was isolated by degenerate polymerase chain reaction. The human gene corresponding to WTGC1 was designated as Wnt-13 and overlapping Wnt-13 cDNAs were cloned. Nucleotide sequence analysis indicated that the Wnt-13 gene encodes the protein of 372 amino acids, including a signal peptide, two potential N-glycosylation sites and 24 cystein residues highly conserved among members of the Wnt gene family. The Wnt-13 mRNA of 2.5 kb in size was detected in heart, brain, placenta, lung, prostate, testis, ovary, small intestine and colon of adult human and also in brain, lung and kidney of fetal human. Among various cancer cell lines, the Wnt-13 mRNA was detected in HeLa (cervical cancer), MKN28 and MKN74 (gastric cancer). The Wnt-13 gene has been mapped to human chromosome 1p13. These results suggest that the Wnt-13 gene may be involved in normal human development or differentiation as well as in human carcinogenesis.

Adult↗

PubMed1996

Organ variation in the mutagenicity of MeIQ in Big Blue lacI transgenic mice.

2-Amino-3,4-dimethylimidazo[4,5-f]quinoline (MeIQ), which is a heterocyclic amine found in food and the potent mutagen in S. typhimurium TA98, was examined for its genotoxic potential using lacI transgenic mice (Big Blue, C57BL/6N lineage). Female mice, at 7 weeks of age, were given a diet containing 0.03% MeIQ for 1, 4 and 12 weeks, and mutant frequencies (MF) were analyzed in the bone marrow, liver, forestomach, colon and heart. The MF increased in a feeding period-dependent manner. Relative to untreated mice, the MF after a 12-week-feeding of MeIQ was 38 times higher in the colon, 5.8 times higher in the bone marrow, 4.6 times higher in the liver, and 2.6 times higher in the forestomach. No increase in MF was detected in the heart, where no tumors develop.

Animals↗

PubMed1996

HST-1/FGF-4 stimulates proliferation of megakaryocyte progenitors synergistically and promotes megakaryocyte maturation.

Megakaryocyte (MK) development is dependent on the complex interaction of MK progenitors, various cytokines and stromal elements. We previously reported that an injection of replication-deficient adenovirus containing HST-1/FGF-4 cDNA (Adex1HST-1) into mice caused a twofold increase in peripheral platelet count for 30 days without any other hematological or histological abnormality. In the present study using Adex1HST-1-infected human megakaryocytic Dami cells, we demonstrated for the first time that HST-1/FGF-4 promoted MK maturation, inducing increases in DNA ploidy, cytoplasmic and membrane maturation, and platelet-like particle release. Moreover, HST-1/FGF-4 acted on megakaryocytic cells to induce secretion of IL-6 and TNF-alpha, and increased adhesion of megakaryocytic cells to human endothelial cells primarily via VLA-4 and LFA-1 molecules; both mechanisms have been shown to lead to MK maturation. We also showed that HST-1/FGF-4 stimulates the proliferation of MK progenitors not alone but synergistically with IL-3 via IL-6 and with c-mpl ligand (thrombopoietin) not via IL-6. This result supports the hypothesis of the presence of two distinct populations of MK progenitors: IL-3-dependent and Tpo-dependent. All these results suggest that HST-1/FGF-4 can regulate MK development not only as an MK potentiating factor, but also as an inducer of cytokine secretion from MK, and as a modulator of adhesive interactions with endothelial cells.

Adenoviruses, Human↗

PubMed1996

A rat genetic map constructed by representational difference analysis markers with suitability for large-scale typing.

Representational difference analysis (RDA) was applied to isolate chromosomal markers in the rat. Four series of RDA [restriction enzymes, BamHI and HindIII; subtraction of ACI/N (ACI) amplicon from BUF/Nac (BUF) amplicon and vice versa] yielded 131 polymorphic markers; 125 of these markers were mapped to all chromosomes except for chromosome X. This was done by using a mapping panel of 105 ACI x BUF F2 rats. To complement the relative paucity of chromosomal markers in the rat, genetically directed RDA, which allows isolation of polymorphic markers in the specific chromosomal region, was performed. By changing the F2 driver-DNA allele frequency around the region, four markers were isolated from the D1Ncc1 locus. Twenty-five of 27 RDA markers were informative regarding the dot blot analysis of amplicons, hybridizing only with tester amplicons. Dot blot analysis at a high density per unit of area made it possible to process a large number of samples. Quantitative trait loci can now be mapped in the rat genome by processing a large number of samples with RDA markers and then by isolating markers close to the loci of interest by genetically directed RDA.

Alleles↗

PubMed1996

Genetic alterations in rat colon tumors induced by heterocyclic amines.

BACKGROUND: In rat colon tumors induced by the cooked food mutagens 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) and 2-amino-3-methylimidazo[4,5-f]quinoline (IQ), ras and p53 alterations are rarely detected. To investigate the roles of the APC gene and microsatellite instability (MI) in PhIP-induced colon carcinogenesis, mutations of the APC gene and alterations of microsatellites were examined. METHODS: Complimentary DNA sequence of the rat APC gene were determined by polymerase chain reaction (PCR) using primers based on the human APC sequence. PCR-single strand conformation polymorphism (SSCP) analysis was performed using primers based on sequences of flanking introns and exon 15. Microsatellite alterations were also analyzed using 85 microsatellite sequences dispersed through most of the rat chromosomes. RESULTS: Five mutations in the APC gene were detected in four of eight PhIP-induced rat colon tumors. All five mutations involved deletion of a guanine base in a 5'-GGGA-3' sequence. Only 2 of 13 IQ-induced colon tumors had mutations of the APC gene and these were base substitution mutations. Seven of eight PhIP-induced colon tumors had microsatellite alterations in at least one locus, whereas no alterations were observed in the IQ-induced colon tumors. CONCLUSIONS: The specific 5'-GGGA-3' to 5'-GGA-3' mutation and MI demonstrated in this study are strong evidence of a mutational fingerprint of PhIP.

Animals↗

PubMed1996

32P-Postlabeling analysis of a DNA adduct, an N2-acetyl derivative of guanine, formed in vitro by methylglyoxal and hydrogen peroxide in combination.

Methylglyoxal is a direct-acting mutagen in Salmonella typhimurium TA100 and its mutagenicity is markedly enhanced in the presence of hydrogen peroxide. In addition, a mixture of methylglyoxal and hydrogen peroxide reacts with 2'-deoxyguanosine to form N2-acetyl-2'-deoxyguanosine. We examined whether the guanine residues in DNA were acetylated by methylglyoxal in the presence of hydrogen peroxide using the 32P-postlabeling method. First, N2-acetyl-2'-deoxyguanosine 3'-monophosphate and N2-acetyl-2'-deoxyguanosine 3,5'-diphosphate were chemically synthesized as standard compounds for the analysis. Then calf thymus DNA (3.24 micromol) was treated with methylglyoxal (64.8 micromol) at pH 7.4 for 3 h at 37 degrees C, and subsequently with hydrogen peroxide (64.8 micromol) at 37 degrees C for 2 h. The adduct formation was analyzed using HPLC in combination with the 32P-postlabeling method under the standard conditions. N2-Acetyl-2'-deoxyguanosine was detected at levels of 2/10(6) nucleotides in double-stranded DNA and 1/10(5) nucleotides in single-stranded DNA. The estimated limit of detection by our method was 3 per 10(8) nucleotides.

Acetylation↗

PubMed1996

Inhibitory potential of pregnancy and lactation on mammary carcinogenesis induced by a food-derived carcinogen, 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine, in Sprague-Dawley rats.

Effects of pregnancy and lactation on a food-derived carcinogen 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP)-induced mammary carcinogenesis were investigated in female Sprague-Dawley rats. When rats were administrated PhIP in the diet (100 ppm) for 15 weeks, palpable subcutaneous tumors were first detected at week 24 in three nulliparous rats, while tumor latency was apparently increased in animals undergoing delivery and weaning during the PhIP administration period. However, since the incidence of palpable tumors was not consistently increased in the nulliparous animals, possibly due to the short PhIP-treatment, and the fact that spontaneous tumors developed in parous animals after week 44, the final incidences and multiplicities of histopathologically confirmed adenocarcinomas at week 48 were not significantly different between the two groups. These findings art partly supported by the epidemiological evidence that breast cancer is more prevalent in nulliparous than in parous women.

Adenocarcinoma↗

PubMed1996

Inhibition of protein serine/threonine phosphatases by fumonisin B1, a mycotoxin.

Fumonisin B1 (FB1), a mycotoxin produced by the fungus Fusarium moniliforme, which is a common contaminant of corn, is suspected to be a cause of human esophageal cancer. FB1 is hepatotoxic and hepatocarcinogenic in rats, and although the mechanisms involved have not been clarified, the latter is associated with a weak initiating activity. The effects of FB1 on the activity of protein serine/threonine phosphatases (PPs) (PP1, PP2A, PP2B, PP2C and PP5/T/K/H) were investigated in the present study. Inhibition of dephosphorylation was noted for all five PPs with IC50 values of 80 microM-3000 microM. Among the five PPs examined, PP5 was most sensitive with an IC50 of 80 microM. This concentration is comparable to that estimated to be reached in the rat body by feeding FB1 to obtain hepatic tumors. Inhibition of PP5 could thus play important roles in the toxicity and carcinogenic action of FB1.

Animals↗