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Biomedical subjects

M Fujino

Publications and source records attributed to M Fujino.

At least 55 records · Page 3Linked to original sources
PubMed2000

Inhibition of Fas-mediated fulminant hepatitis in CrmA gene-transfected mice.

Hyperimmune response via Fas/Fas-ligand and perforin/granzyme pathways may be essential in pathogenesis of virus-induced fulminant hepatitis. CrmA inhibits activation of caspases and granzyme B, suggesting it may block these pathways. We investigated whether CrmA expression would inhibit Fas-associated lethal hepatitis in mice. We successfully generated AxCALNLCrmA, a recombinant adenovirus expressing CrmA gene with a Cre-mediated switching cassette. We increased CrmA expression level in the liver transfected with AxCALNLCrmA (10(9) pfu) by increasing administration dose (10(7)-10(9) pfu) of AxCANCre, a recombinant, adenovirus-expressing Cre gene. Injection of anti-Fas antibody into the control mice rapidly led to animal death due to massive liver apoptosis, while the apoptosis was dramatically reduced in the CrmA-expressed mice. The animal survival increased with an increase of CrmA expression. The formation of active caspase-3 was markedly inhibited in the crmA-transfected hepatocytes in vitro. These results suggest that crmA is an effective gene that can inhibit immune-related liver apoptosis.

Adenoviridae↗
PubMed2000

Stimulation of corticotropin-releasing hormone-mediated adrenocorticotropin secretion by central administration of prolactin-releasing peptide in rats.

Prolactin-releasing peptide (PrRP) is a recently isolated hypothalamic peptide which is an endogenous ligand to an orphan receptor. We previously demonstrated that PrRP neurons are widely distributed throughout the rat brain and suggested that PrRP may have important functions in the central nervous system. To analyze the function of PrRP, we studied the effect of intracerebroventricular (i.c.v.) PrRP administration on c-Fos protein accumulation in the rat brain. The results clearly indicated that c-Fos protein accumulation was dramatically increased in the nuclei of corticotropin-releasing hormone (CRH)-positive parvocellular neurosecretory cells in the paraventricular nucleus (PVN). We also demonstrated synapse-like contact between PrRP neurons and CRH cell bodies in the PVN, which suggests that PrRP31 has some effect on CRH secretion. We therefore investigated the effect of i.c.v. administration of PrRP31 on the CRH-mediated increase in adrenocorticotropin (ACTH) levels, and found that plasma ACTH levels were indeed increased by i.c.v. PrRP31. In addition, animals pre-treated with intravenous alpha-helical CRH, a potent CRH antagonist, showed attenuated plasma ACTH responses after i.c.v. PrRP31 administration. These results strongly suggest that PrRP affects the hypothalamic-pituitary-adrenal axis.

Adrenocorticotropic Hormone↗
PubMed2000

Discovery of novel, potent, and selective small-molecule CCR5 antagonists as anti-HIV-1 agents: synthesis and biological evaluation of anilide derivatives with a quaternary ammonium moiety.

The search for new small-molecule CCR5 antagonists by high-throughput screening (HTS) of the Takeda chemical library using [(125)I]RANTES and CHO/CCR5 cells led to the discovery of lead compounds (A, B) with a quaternary ammonium or phosphonium moiety, which were synthesized to investigate new MCP-1 receptor antagonists. A series of novel anilide derivatives 1 with a quaternary ammonium moiety were designed, synthesized, and tested for their CCR5 antagonistic activity. Through the optimization of lead compounds, we have found N,N-dimethyl-N-[4-[[[2-(4-methylphenyl)-6, 7-dihydro-5H-benzocyclohepten-8-yl]carbonyl]amino]benzyl]tetrahydr o-2 H-pyran-4-aminium chloride (1r, TAK-779) as a highly potent and selective nonpeptide CCR5 antagonist with a IC(50) value of 1.4 nM in the binding assay. Compound 1r also inhibited the replication of macrophage (M)-tropic HIV-1 (Ba-L strain) in both MAGI-CCR5 cells and PBMCs with EC(50) values of 1.2 and 3.7 nM, respectively. The synthesis and structure-activity relationships of 1r and its related compounds are detailed.

Amides↗
PubMed2000

Protease inhibitor reduces loss of tensile strength in rat anastomosis with peritonitis.

BACKGROUND: The tensile strength in intestinal anastomoses decreases postoperatively in association with degradation of the extracellular matrix, and these changes would be expected to be more intense in the presence of peritonitis. MATERIALS AND METHODS: In this study, we investigated extracellular matrix degradation and tensile strength in a rat model of intestinal anastomosis with peritonitis. In the chemical peritonitis model, peritonitis was induced 24 h earlier with intraperitoneal HCl. A serine protease inhibitor, nafamostat mesilate (NM), was given intraperitoneally to some animals every 12 h from immediately after the operation for 3 days. Immunostaining was performed by the standard streptavidin-biotin-peroxidase method after fibronectin (Fn) and factor XIII antigen retrieval on paraformaldehyde-fixed, paraffin-embedded tissue sections. RESULTS: In comparison with controls, administration of NM reduced the loss of tensile strength on Day 3 in a dose-dependent manner, and high-dose NM (20/mg/kg) significantly prevented the loss of tensile strength on Day 3 (P < 0. 05). In the control group, degradation of the collagen layer in the anastomosis was associated with disappearance of Fn and factor XIII staining on Day 3. The administration of NM attenuated these changes with intense immunostaining for Fn and factor XIII seen particularly between collagen fibers on both sides of the anastomosis on Day 3. In the chemical peritonitis model, administration of NM also significantly prevented the loss of tensile strength on Day 3 without disappearance of collagen fibers. CONCLUSION: These findings suggest that NM may be clinically useful for preventing intestinal leakage, particularly when anastomoses are performed under protease-activating conditions, such as intestinal edema and inflammation.

Anastomosis, Surgical↗
PubMed2000

Adenovirus-mediated gene therapy for mucopolysaccharidosis VII: involvement of cross-correction in wide-spread distribution of the gene products and long-term effects of CTLA-4Ig coexpression.

Recombinant adenoviruses expressing human beta-glucuronidase (AxCAhGUS) and CTLA-4Ig (AxCACTLA-4Ig) were generated and therapeutic efficacy was investigated using a murine model of mucopolysaccharidosis type VII (MPSVII). Seven days after the intravenous administration of AxCAhGUS, high levels of beta-glucuronidase (GUSB) activity were observed in the liver, spleen, heart, lung, kidney, and serum, while viral DNA was predominantly detected in the liver. To investigate the contribution of in vivo cross-correction of GUSB between the liver and other organs, we injected the serum obtained from the transduced mice into untreated MPSVII mice. Similar distributions of GUSB activity were observed in the serum-injected mice, suggesting that GUSB activities detected in the extrahepatic organs were due to the cross-correction rather than the direct gene transduction. This result also suggested that maintaining high levels of GUSB in the systemic circulation was essential for the effective treatment of MPSVII. To achieve this, we injected AxCAhGUS and AxCACTLA-4Ig into MPSVII mice. Serum GUSB activity was sustained at high levels for more than 200 days and morphological normalization of the liver and spleen was observed for a year. This suggests that long-term therapeutic efficacy in visceral organs of MPSVII is achievable by coexpression of CTLA-4Ig through an in vivo cross-correction pathway.

Abatacept↗
PubMed2000

New neuropeptides containing carboxy-terminal RFamide and their receptor in mammals.

Only a few RFamide peptides have been identified in mammals, although they have been abundantly found in invertebrates. Here we report the identification of a human gene that encodes at least three RFamide-related peptides, hRFRP-1-3. Cells transfected with a seven-transmembrane-domain receptor, OT7T022, specifically respond to synthetic hRFRP-1 and hRFRP-3 but not to hRFRP-2. RFRP and OT7T022 mRNAs are expressed in particular regions of the rat hypothalamus, and intracerebroventricular administration of hRFRP-1 increases prolactin secretion in rats. Our results indicate that a variety of RFamide-related peptides may exist and function in mammals.

Amino Acid Sequence↗
PubMed2000

Bacterial translocation and tumor necrosis factor-alpha gene expression in experimental hemorrhagic shock.

OBJECTIVE: To investigate whether bacterial translocation is the causative mechanism underlying cytokine production during hemorrhagic shock. DESIGN: Prospective, randomized, unblinded animal study. SETTING: Surgical research laboratories of Shiga University of Medical Science. SUBJECTS: Male Sprague-Dawley rats. INTERVENTIONS: The rats were randomly divided into three groups. Each animal was anesthetized with pentobarbital, given a continuous infusion of 0.9% saline, and monitored for blood pressure. The normoxic and sham shock groups breathed room air, whereas the hyperoxic shock group was administered 100% oxygen. Except in the sham shock group, blood was withdrawn to induce a hemorrhagic shock state, then the shed blood was reinfused. Sixty minutes after the induction of hemorrhagic shock, arterial blood cultures were performed in all three groups. The animals were then killed, and their mesenteric lymph nodes (MLNs) were harvested for bacterial culture. The terminal ileum, liver, spleen, kidney, lung, and MLNs were also collected for histologic study by in situ hybridization. MEASUREMENTS AND MAIN RESULTS: In the bacteriologic study, the prevalence of bacterial translocation was 0% (0/11) in the hyperoxic shock group, 55% (6/11) in the normoxic shock group, and 0% (0/9) in the sham shock group. In the in situ hybridization study, tumor necrosis factor-alpha gene expression was detected only in the ileal tissue, MLNs, and spleens of the normoxic shock group. Blood cultures were sterile in all three groups. CONCLUSIONS: Bacterial translocation occurred in MLNs within 1 hr of hemorrhage. Hemorrhagic shock causes tumor necrosis factor-alpha gene expression as well as bacterial translocation in MLNs, but not in the liver, in this model. Bacterial translocation was prevented by hyperoxia early in the course of hemorrhagic shock. Hyperoxia also prevented tumor necrosis factor-alpha gene expression along the bacterial invasion route.

Animals↗
PubMed2000

Apelin, the natural ligand of the orphan seven-transmembrane receptor APJ, inhibits human immunodeficiency virus type 1 entry.

In addition to the CCR5 and CXCR4 chemokine receptors, a subset of primary human immunodeficiency virus type 1 (HIV-1) isolates can also use the seven-transmembrane-domain receptor APJ as a coreceptor. A previously identified ligand of APJ, apelin, specifically inhibited the entry of primary T-tropic and dualtropic HIV-1 isolates from different clades into cells expressing CD4 and APJ. Analysis of apelin analogues demonstrated that potent and specific antiviral activity was retained by a 13-residue, arginine-rich peptide. Antiviral potency was influenced by the integrity of methionine 75, which contributes to APJ-binding affinity, and by the retention of apelin residues 63 to 65. These studies demonstrate the ability of a small peptide ligand to block the function of APJ as an HIV-1 coreceptor, identify apelin sequences important for the inhibition, and provide new reagents for the investigation of the significance of APJ to HIV-1 infection and pathogenesis.

Acquired Immunodeficiency Syndrome↗
PubMed2000

Analyses for susceptibility of rat anterior pituitary cells to prolactin-releasing peptide.

We validated the effect of prolactin-releasing peptide (PrRP) on prolactin (PRL) secretion from rat anterior pituitary cells in in vitro culture. We found that culture conditions considerably influenced the response of the anterior pituitary cells to PrRP. Longer culture term (4 d) was required to obtain better responses of the anterior pituitary cells to PrRP in comparison to thyrotropin-releasing hormone (TRH). Under the culture conditions employed here, PrRP was comparable to TRH in the potency promoting PRL secretion, and the action of PrRP was very specific for PRL secretion. The susceptibility of the anterior pituitary cells to PrRP varied in female rats depending on the process of reproduction: the cells prepared from lactating rats were the most sensitive to PrRP compared with those from random-cycle and pregnant rats. Because the expression levels of PrRP receptor mRNA in the pituitary varied during the reproductive process, we speculated that the susceptibility of the anterior pituitary cells would reflect cellular changes including the expression level of PrRP receptors. In addition, treatment with estrogen in vivo enhanced the susceptibility of the cultured anterior pituitary cells in male rats. Our results indicate that the susceptibility of the rat anterior pituitary cells to PrRP is regulated by physiological mechanisms.

Animals↗
PubMed1999

Isolation and cDNA cloning of a novel galanin-like peptide (GALP) from porcine hypothalamus.

Galanin is a widely distributed neuropeptide with a variety of physiological functions. Three galanin receptor subtypes, GALR1, GALR2, and GALR3, have been reported. We isolated a novel galanin-like peptide (GALP) from porcine hypothalamus by observing its activity for increasing [(35)S]GTPgammaS binding to a membrane preparation of GALR2-transfected cells. The peptide had 60 amino acid residues and a non-amidated C terminus. The amino acid sequence of GALP-(9-21) was completely identical to that of galanin-(1-13). A cloned porcine GALP cDNA indicated that GALP was processed from a 120-amino acid GALP precursor protein. The structures of rat and human GALP-(1-60) were deduced from cloned cDNA, which indicated that the amino acid sequences 1-24 and 41-53 were highly conserved between humans, rats, and pigs. Receptor binding studies revealed that porcine GALP-(1-60) had a high affinity for the GALR2 receptor (IC(50) = 0.24 nM) and a lower affinity for the GALR1 receptor (IC(50) = 4.3 nM). In contrast, galanin showed high affinity for the GALR1 (IC(50) = 0.097 nM) and GALR2 receptors (IC(50) = 0.48 nM). GALP is therefore an endogenous ligand that preferentially binds the GALR2 receptor, whereas galanin is relatively non-selective.

Amino Acid Sequence↗
PubMed1999

Central administration of prolactin-releasing peptide stimulates oxytocin release in rats.

The prolactin-releasing peptide (PrRP) is a novel hypothalamic peptide that has been purified as a ligand of an orphan receptor which is expressed in pituitary cells, and is known to stimulate prolactin release both in vitro and in vivo. We previously determined the immunocytochemical localization of PrRP neurons in the rat brain and our results suggest that PrRP takes part in a variety of brain functions. Additionally, in rats we have demonstrated the synaptic contact of PrRP neurons with oxytocin cell bodies in the paraventricular hypothalamic nucleus (PVH) and supraoptic nucleus (SON). This observation indicates that PrRP may regulate oxytocin secretion. In the present study, we performed intra-cerebroventricular administration of PrRP to conscious rats, and examined the effect of PrRP on the plasma levels of oxytocin and vasopressin. Our results show that central administration of PrRP increased the plasma oxytocin and vasopressin levels in female rats, but in male rats only oxytocin was increased. These results suggest that the PrRP acts as a neuro-modulator of the function of magnocellular neurons, especially oxytocin neurons, in the brain.

Animals↗
PubMed1999

Apelin, the natural ligand of the orphan receptor APJ, is abundantly secreted in the colostrum.

By using a strategy that we have developed to search for the ligands of orphan seven-transmembrane-domain receptors [S. Hinuma et al., Nature 393 (1998) 272-276], we have recently identified a natural ligand, apelin, for the orphan 7TMR, APJ [K. Tatemoto et al., Biochem. Biophys. Res. Commun. 251 (1998) 471-476]. In this paper, we isolated rat and mouse apelin cDNAs, and analyzed the tissue distribution of apelin mRNA in rats. Although apelin mRNA was widely detected in a variety of tissues, the highest expression of apelin mRNA was detected in the mammary gland of pregnant rats. In the mammary gland, biologically active apelin and its mRNA considerably increased during pregnancy and lactation, and reached a maximal level around parturition. Moreover, a large amount of apelin (14-93 pmol/ml) was found to be secreted in the bovine colostrum, and it was still detectable even in commercial bovine milk. Since apelin partially suppressed cytokine production by mouse spleen cells in response to T cell receptor/CD3 cross-linking, the oral intake of apelin in the colostrum and milk might modulate immune responses in neonates.

Adipokines↗
PubMed1999

Tissue distribution of prolactin-releasing peptide (PrRP) and its receptor.

Prolactin-releasing peptide (PrRP) is a novel bioactive peptide, originally isolated from bovine hypothalamus by utilizing an orphan seven-transmembrane-domain receptor expressed in the human pituitary gland. In this paper, we analyzed the tissue distribution of rat and human PrRP and their receptor mRNAs by quantitative reverse transcription-polymerase chain reaction (RT-PCR) and Northern blotting. In RT-PCR analysis, rat PrRP receptor mRNA was detected in the central nervous system, and the highest expression was detected in the pituitary gland. In addition, in situ hybridization revealed that rat PrRP receptor mRNA was highly expressed in the anterior lobe of the pituitary. On the other hand, rat PrRP mRNA was most abundantly expressed in the medulla oblongata, while significant levels of expression were widely detected in other tissues. In Northern blot analyses, human PrRP receptor mRNA was detected only in the pituitary gland among tissues examined. Human PrRP mRNA was detected in the medulla oblongata and in the pancreas. In contrast to the pattern of mRNA expression, the highest content of bioactive PrRP was found in the hypothalamus rather than the medulla oblongata in the rat brain, indicating that PrRP mRNA does not always parallel with mature PrRP in tissue distribution. The wide distribution of PrRP and its receptor suggests that they have various functions not only in the pituitary gland but also in the other tissues.

Amino Acid Sequence↗