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Biomedical subjects

F Shibata

Publications and source records attributed to F Shibata.

At least 37 records · Page 2Linked to original sources
PubMed1996

Differential changes in the concentrations of cytokine-induced neutrophil chemoattractant (CINC)-1 and CINC-2 in exudate during rat lipopolysaccharide-induced inflammation.

Recently we have purified four neutrophil chemotactic factors from conditioned medium of rat granulation tissue. Two chemokines besides cytokine-induced neutrophil chemoattractant-1 (CINC-1, formerly called CINC) and CNC-3/macrophage inflammatory protein-2 (MIP-2) were novel chemoattractants, designated as CINC-2 alpha and CINC-2 beta. In the present report, we developed an enzyme-linked immunosorbent assay (ELISA) specific for CINC-2. The biotin-streptavidin sandwich ELISA for CINC-2 beta detected CINC-2 alpha and CINC- 2 beta equally well between 1 ug/mI and 300 ng/ml, but did not show cross-reactivity with CINC-1 and CINC-3. The concentrations of CINC-1 and CINC-2 in the exudate during rat lipopolysaccharide (LPS)-induced inflammation were determined. The CINC-1 concentration in the exudate reached a maximum at 4 h after LPS injection, whereas the CINC-2 level steadily increased up to 8 h. The number of infiltrated cells in the exudate increased linearly until 6 h and gradually up to 8 h. Increase in the cell number was correlated with total concentrations of CINC-1 and CINC-2. The results suggest that CINC-2 as well as CINC-1 plays an important role in accumulation of neutrophils into the inflammatory lesion of LPS- induced inflammation in rats.

Animals↗
PubMed1996

Evidence for two putative receptors mediating chemotactic activity of cytokine-induced neutrophil chemoattractants (CINCs) for rat neutrophils.

Recently we have found that only cytokine-induced neutrophil chemoattractant (CINC)-3 induces an increase in intracellular [Ca2+] in rat neutrophils stimulated first with CINC-1, CINC-2 alpha or CINC-2 beta, and hypothesized that rat neutrophils have another receptor specific for CINC-3 in addition to a common receptor for all CINCs [Shibata F., et al., Eur. J. Biochem., 231, 306 (1995)]. To evaluate this hypothesis, chemotactic activity of these CINCs in vitro was determined by using rat neutrophils together with each CINC. In the presence of CINC-1, migration of neutrophils toward CINC-1, CINC-2 alpha and CINC-2 beta was inhibited, but that toward CINC-3 was not. Similar results were obtained when CINC-2 alpha or CINC-2 beta was added to neutrophils. However, addition of CINC-3 to neutrophil suspension in the upper chamber resulted in the complete inhibition of the neutrophil migration toward each CINC. The results support our hypothesis and indicate both the putative receptors can mediate chemotaxis of rat neutrophils.

Animals↗
PubMed1995

Recombinant production and biological properties of rat cytokine-induced neutrophil chemoattractants, GRO/CINC-2 alpha, CINC-2 beta and CINC-3.

Recently we found four cytokine-induced neutrophil chemoattractants, CINC-1, CINC-2 alpha, CINC-2 beta and CINC-3/macrophage inflammatory protein 2 (MIP-2), in conditioned medium of granulation tissue obtained from carrageenin-induced inflammation in rats [Nakagawa, H., Komorita, N., Shibata, F., Ikesue, A., Konishi, K., Fujioka, M. & Kato, H. (1994) Biochem. J. 301, 545-550]. In the present report, we describe recombinant production of CINC-2 alpha, CINC-2 beta and CINC-3 in Escherichia coli, and biological properties of these chemokines. Neutrophil chemotactic activities of CINC-2 alpha and 2 beta in vitro were the same as the activity of CINC-1. CINC-3 had an activity comparable to other CINCs, but showed a decrease at high concentrations. Stimulation of neutrophils with CINCs induced an increase in intracellular [Ca2+] dose-dependently. CINC-3 was more potent than the other CINCs and still induced an increase in intracellular [Ca2+] in rat neutrophils stimulated first with other CINCs. CINC-2 alpha, CINC-2 beta and CINC-3 induced a comparable response to CINC-1 in the release of cathepsin G from rat neutrophils. Injection of CINC-2 alpha, 2 beta and 3 into preformed air-pouch on the back of rat induced infiltration of neutrophils to an extent similar to that caused by the injection of CINC-1. These data indicate CINC-2 alpha, 2 beta and 3 as well as CINC-1 are chemoattractants specific for neutrophil in vivo.

Animals↗
PubMed1994

Transcriptional regulation of basic fibroblast growth factor gene by p53 in human glioblastoma and hepatocellular carcinoma cells.

Mutations of the p53 gene are found in various human cancers. The frequency of its mutation is reported to increase during tumor progression in most tumors. In human gliomas, mutations of the p53 gene are found in about one-third of the malignant forms and in few of the benign ones, indicating their possible involvement in tumor progression. On the other hand, we have recently shown that basic fibroblast growth factor (basic FGF) plays a crucial role in tumor progression as an autocrine growth factor in tissues of human gliomas. Therefore, we hypothesized that p53 might regulate the promoter activity of the basic FGF gene, which has several GC boxes and no typical TATA box. In this study, cotransfection assays using human glioblastoma and hepatocellular carcinoma cells and establishment of stable cell lines expressing mutant-type p53 were performed. The basic FGF gene promoter was demonstrated to be regulated by p53 at the transcriptional level and its basal core promoter was found to be responsive to p53. Expression of endogenous basic FGF was also demonstrated to be activated by mutant type p53. Wild-type p53 repressed gene expression of the basic FGF and its mutant activated it in vitro, implying one of the possible pathways in tumor progression.

Carcinoma, Hepatocellular↗
PubMed1994

Identification of cytokine-induced neutrophil chemoattractants (CINC), rat GRO/CINC-2 alpha and CINC-2 beta, produced by granulation tissue in culture: purification, complete amino acid sequences and characterization.

Four basic neutrophil chemotactic factors (chemokines) have been purified from conditioned medium of granulation tissue obtained from carrageenin-induced inflammation in the rat. On the basis of their N-terminal amino acid sequences, one of the chemokines was identical with rat GRO/cytokine-induced neutrophil chemoattractant (CINC) which we reported previously, and another was identical with rat macrophage inflammatory protein-2 (MIP-2). Two other chemokines were novel chemoattractants related to MIP-2. The novel chemokines are referred to as rat GRO/CINC-2 alpha and CINC-2 beta, and consequently CINC and rat MIP-2 are renamed rat GRO/CINC-1 and CINC-3 respectively. The complete amino acid sequences of purified CINC-2 alpha and CINC-3 were determined by analysis of the fragments isolated from proteinase V8-treated CINCs. The cDNA for CINC-2 beta was cloned by reverse transcription/PCR amplification using specific primers starting with total RNA extracted from lipopolysaccharide-stimulated rat macrophages. A comparison of the amino acid sequence encoded by the cDNA with the N-terminal amino acid sequence of purified CINC-2 beta revealed that mature CINC-2 beta is a 68-residue chemoattractant produced by cleavage of a 32-residue signal peptide. The difference in amino acid sequences between CINC-2 alpha and CINC-2 beta consisted of only three C-terminal residues. Rat GRO/CINC-2 alpha is a major chemokine, and the four purified chemokines have similar chemotactic activity, suggesting that they contribute to neutrophil infiltration into inflammatory sites in rats.

Amino Acid Sequence↗
PubMed1993

Molecular cloning and characterization of a human carboxylesterase gene.

A cDNA encoding human liver carboxylesterase and its gene were isolated. Nucleotide sequence analyses of the cDNA revealed that the predicted enzyme protein consists of 567 amino acids, including 18 amino acids of a putative signal peptide. Comparison of the deduced amino acid sequences of this enzyme with those of seven other carboxylesterases in various mammalian species, together with experimental data from several other laboratories, showed that these enzymes can be classified into three groups depending on the sequences at their carboxyl terminals and the presence or absence of one exon. A human carboxylesterase gene was found to span approximately 30 kb and to have 14 small exons. Alignments of this gene with those of human cholinesterase and rat cholesterol esterase indicated insertional sites at some introns and homologous amino acid sequences around them, although these genes have different numbers of exons. Thus the results supported the conclusion that these esterases evolved from a common ancestral gene.

Amino Acid Sequence↗
PubMed1991

Functional characterization of the human basic fibroblast growth factor gene promoter.

In order to understand the regulation of basic fibroblast growth factor (bFGF) gene expression, we have cloned and characterized the human bFGF gene and its regulatory elements. Using restriction endonuclease digestion, we have mapped the entire gene and sequenced all intron/exon boundaries to confirm authenticity and to determine organization. The data show that intron 1 is at least 16 kb long while intron 2 is 16 kb long. The human bFGF gene, including its three exons, is therefore at least 36 kb long. There are five GC boxes which may represent SP-1 binding sites and one potential AP-1 binding site within the core promoter region. Primer extension analysis indicates the presence of one bFGF-RNA transcription start site. We used a standard bacterial CAT gene expression system to identify the DNA sequence containing the functional bFGF gene promoter. Deletion analysis suggests the presence of two negative regulatory elements; one in the non-transcribed 5'-promoter region and the other within transcribed (but non-translated) sequences 3' of the promoter core.

Amino Acid Sequence↗
PubMed1990

Structural characterization of a follicle-stimulating hormone action inhibitor in porcine ovarian follicular fluid. Its identification as the insulin-like growth factor-binding protein.

Recently, an inhibitory polypeptide that could block the follicle-stimulating hormone-induced estradiol and progesterone production in rat ovary granulosa cells has been isolated from porcine ovarian follicular fluid. Amino-terminal sequence analysis of the purified inhibitor suggests that it could be the porcine congener of the 53-kDa subunit of the growth hormone-dependent insulin-like growth factor binding protein (IGF-BP3). Using amino acid sequence information derived from the purified inhibitor to construct oligonucleotide probes, we have now identified the complementary deoxyribonucleic acids (cDNAs) encoding the inhibitory polypeptide from a porcine liver and a porcine ovary library. The nucleotide and predicted amino acid sequences revealed that the cDNAs indeed encode the porcine homolog of the recently characterized human IGF-BP3. The mature polypeptide consists of 266 amino acids, which is 2 amino acids longer than the human sequence. Between the two species, there are 42 amino acid substitutions, but the 18 cysteines and the three Asn-linked glycosylation sites are totally conserved. A single mRNA species of 2.6 kilobases encoding the IGF-BP3 was detected in porcine gonadal, brain, and liver tissues by Northern analysis.

Amino Acid Sequence↗
PubMed1990

[Plasma and red blood cell cyanide concentrations during hypotension induced by sodium nitroprusside or by a nitroprusside-trimetaphan mixture in rabbits].

Plasma and red blood cells cyanide concentrations during hypotension induced by sodium nitroprusside and a nitroprusside-trimetaphan mixture were studied in 29 male rabbits under halothane anesthesia. They were randomly divided into three groups; Nitroprusside (group N; n = 10), A nitroprusside-trimetaphan mixture (group M; n = 10), Controls (group C; n = 9). No changes were noted in plasma and red blood cells cyanide concentrations in group C throughout the experiment. During and after induced hypotension, in group N, plasma cyanide concentration was significantly higher than the control value. The maximum increase occurred 60min after induction of hypotension and the highest concentration of plasma cyanide was six times the control value. In contrast, in group M, plasma cyanide concentration was unchanged from the control value. However, during and after induced hypotension, red blood cells cyanide concentrations of group N and group M were significantly higher compared with the control values. Red blood cells concentrations of cyanide increased for 30 and 60 min during induced hypotension in group N (27.11 +/- 3.9 micrograms.ml-1, P less than 0.001, 46, 73 +/- 4.7 micrograms.ml-1, P less than 0.001, respectively) and in group M (0.31 +/- 0.05 micrograms.ml-1, P less than 0.05, 0.29 +/- 0.03 micrograms.ml-1, P less than 0.05, respectively). In conclusion, the data suggest that a nitroprusside-trimetaphan mixture is a safe method for hypotensive anesthesia.

Animals↗
PubMed1988

Complementary DNA cloning and sequencing of rat ovarian basic fibroblast growth factor and tissue distribution study of its mRNA.

Three cDNA clones encoding rat basic fibroblast growth factor (FGF) were isolated from 10(6) independent clones prepared from a pregnant mare serum gonadotropin (PMSG)-stimulated rat ovarian cDNA library. One of the cDNA clones contained the entire coding sequence for basic FGF. The other two possessed the sequence coding the carboxy terminal 61 amino acids of rat basic FGF, the putative upstream intron sequence, and a 3'-noncoding region. The cDNAs encoding rat basic FGF predict a molecule consisting of 154 amino acid residues, which is one amino acid shorter than the human and bovine basic FGF. Otherwise, there are only 5 conservative amino acid substitutions between the rat and the human/bovine sequences. Poly A+ RNA from brain cortex and hypothalamus show a single 6.0 kb band that hybridizes to the cloned cDNA probe by Northern analyses. The observation that basic FGF mRNA is below the limits of detection in adrenal, spleen, heart, lung, kidney, liver, stomach, small intestine, large intestine, testis, and ovary support the notion that the that the high levels of the protein found in these tissues is due to storage of the mitogen in the extracellular matrix and not continuous gene expression. The significance of the abundance of mRNA in tissues which are not undergoing either active angiogenesis or cell proliferation (hypothalamus and brain cortex) is unclear but emphasizes the potential neuronotrophic function of basic FGF.

Animals↗