Biomedical subjects

Hiroshi Nomoto

Publications and source records attributed to Hiroshi Nomoto.

At least 19 recordsLinked to original sources

PubMed2006

Processing of nerve growth factor: the role of basic amino acid clusters in the pro-region.

Neurotrophins are synthesized first as precursors called pro-neurotrophins, and their propeptides are then proteolytically removed to form mature neurotrophins. However, a significant proportion of total neurotrophins has been shown to be secreted as pro-neurotrophins. Furthermore, pro- and mature neurotrophins have been shown to elicit opposite effects on cell survival. Thus, the processing step of neurotrophins is very important. In order to understand the mechanism of neurotrophin processing, we focused on the two basic amino acid clusters in the pro-region of nerve growth factor (NGF). Various NGFs mutated at basic amino acids in the pro-region were introduced in COS7 and PC12 cells. The results indicated that these basic amino acid clusters were actually cleaved in the cells by furin, but that their cleavage contributed little to the production of mature NGF. However, one of the two sites was considered to contribute to mature NGF production depending on conditions used.

Amino Acids↗

PubMed2006

Autocrine activation of cultured macrophages by brain-derived neurotrophic factor.

To elucidate a significance of the expression of brain-derived neurotrophic factor (BDNF) in the activated microglia/macrophages of the injured central nervous system, we examined BDNF actions on or BDNF synthesis by macrophages cultured from the mouse peritoneal cavity. They synthesized BDNF and neurotrophin-3 (NT-3) in addition to expressing high-affinity neurotrophin receptors, full-length TrkB (FL), truncated TrkB (TK(-)), and TrkC, thus suggesting an autocrine influence of BDNF and NT-3. BDNF, but not NT-3, enhanced phagocytic activity and stimulated synthesis/secretion of interleukin-1beta in the same manner as lipopolysaccharide (LPS). Furthermore, there was a significant correlation of the phagocytic activity with the expression of BDNF or TrkB (FL). These results imply that the phagocytic activity of macrophages depends on BDNF synthesis and/or TrkB (FL) expression, suggesting that BDNF participates in the activation processes of macrophages by acting in an autocrine manner.

Animals↗

PubMed2006

Identification of AMP N1-oxide in royal jelly as a component neurotrophic toward cultured rat pheochromocytoma PC12 cells.

An extract of royal jelly (RJ) induced processes from cultured rat pheochromocytoma PC12 cells. Active components were isolated, and identified as adenosine monophosphate (AMP) and AMP N1-oxide. AMP N1-oxide was more than 20 times as active as AMP, judging from the minimal concentration to elicit activity. AMP N1-oxide was thought to be responsible for about half of the process-forming activity of whole RJ. Chemically-synthesized AMP N1-oxide was active similarly to the molecule purified from RJ, confirming AMP N1-oxide as the active entity. AMP N1-oxide also suppressed proliferation of PC12 cells and stimulated expression of neurofilament M, a specific protein of mature neurons, demonstrating the stimulatory activity of AMP N1-oxide to induce neuronal differentiation of PC12 cells. Pharmacological experiments suggested that AMP N1-oxide actions are mediated by adenyl cyclase-coupled adenosine receptors, including A2A. Thus AMP N1-oxide is a key molecule that characterizes RJ, and is not found in natural products other than RJ.

Adenosine Monophosphate↗

PubMed2005

Involvement of glial cell line-derived neurotrophic factor in activation processes of rodent macrophages.

The physiological roles of glial cell line-derived neurotrophic factor (GDNF) expressed in the microglia/macrophages of the injured spinal cord have not yet been clarified. mRNA expression of chemokines, including monocyte chemoattractant protein (MCP)-1, was evoked within 1 hr after transection of the spinal cord, and GDNF mRNA expression was similarly up-regulated. Immunohistochemical analysis showed that GDNF was coexpressed with MCP-1 in the CD11b-positive cells. Therefore, we examined further the effects of GDNF on cultured rat peritoneal macrophages. GDNF enhanced the phagocytic activity of the macrophages via GFRalpha-1, glycosylphosphatidylinositol-anchored specific binding site of GDNF, in a c-Ret-independent manner. The influence of autocrine and/or paracrine GDNF synthesis was evaluated by performing activation experiments using macrophages cultured from heterozygous (+/-) GDNF gene-deficient mice or wild-type (+/+) mice. There were no morphological differences dependent on genetic types or stimulators. However, the GDNF mRNA level, but not the MCP-1 or GFRalpha-1 mRNA level, was substantially lower in the mutant macrophages than in the +/+ cells irrespective of stimulation with MCP-1 or lipopolysaccharide (LPS). The phagocytic activity enhanced by MCP-1 or LPS was significantly lower in the mutant cells (+/-) than in the +/+ ones, demonstrating the involvement of endogenous GDNF in the activation processes of macrophages in vitro and suggesting that not only neuroprotective function but also activation of macrophages is effected by the GDNF produced after a spinal cord injury.

Animals↗

PubMed2005

Inflammation-induced GDNF improves locomotor function after spinal cord injury.

Activation of microglia/macrophages after injury occurs limitedly in the CNS, which finding may explain unsuccessful axonal regeneration. Therefore, the relationship between lipopolysaccharide (LPS)-induced inflammation and recovery of locomotor function of rats after spinal cord injury was examined. High-dose LPS improved locomotor function greater than low-dose LPS, being consistent with the expression of neurotrophic factor (GDNF) in microglia/macrophages. Experiments using GDNF gene mutant mice confirmed that the increase in the GDNF mRNA level, rather than the reduction in the mRNA level of inducible NO synthase, could be correlated with the restoration activity of locomotor function. These results suggest that a higher degree of inflammation leads to a higher degree of repair of CNS injuries through GDNF produced by activated microglia/macrophages.

Animals↗

PubMed2005

Oral administration of royal jelly facilitates mRNA expression of glial cell line-derived neurotrophic factor and neurofilament H in the hippocampus of the adult mouse brain.

Royal jelly (RJ) is known to have a variety of biological activities toward various types of cells and tissues of animal models, but nothing is known about its effect on brain functions. Hence, we examined the effect of oral administration of RJ on the mRNA expression of various neurotrophic factors, their receptors, and neural cell markers in the mouse brain. Our results revealed that RJ selectively facilitates the mRNA expression of glial cell line-derived neurotrophic factor (GDNF), a potent neurotrophic factor acting in the brain, and neurofilament H, a specific marker predominantly found in neuronal axons, in the adult mouse hippocampus. These observations suggest that RJ shows neurotrophic effects on the mature brain via stimulation of GDNF production, and that enhanced expression of neurofilament H mRNA is involved in events subsequently caused by GDNF. RJ may play neurotrophic and/or neuroprotective roles in the adult brain through GDNF.

Administration, Oral↗

PubMed2005

Stimulation of production of glial cell line-derived neurotrophic factor and nitric oxide by lipopolysaccharide with different dose-responsiveness in cultured rat macrophages.

To understand the molecular basis of inflammation-induced neurotrophic influences, we investigated the effects of lipopolysaccharide (LPS) on production of glial cell line-derived neurotrophic factor (GDNF) in the injured rat spinal cord or in cultured rat macrophages in comparison with the effects on synthesis/secretion of inducible nitric oxide synthase (iNOS) and nitric oxide (NO). We found that GDNF mRNA expression lasted longer than that of iNOS mRNA in the injured spinal cord after injection of the high-dose LPS that had improved locomotor function, suggesting that the GDNF expression and its balance with NO generation were critical for injury regeneration. Therefore, we next investigated the effects of LPS on cultured macrophages. Levels of iNOS mRNA and secreted NO were enhanced by LPS at lower concentrations (10 ng/mL and above), whereas mRNA expression and secretion of GDNF were elevated only at higher concentrations (100 ng/mL and above). The culture medium of macrophages treated with 10 ng/mL of LPS was actually neurotoxic against cultured cortical neurons, whereas that conditioned at 1000 ng/mL was not. These observations suggest that neurotoxicity partly based on NO is induced by a lower degree of inflammation, whereas neurotrophic effects based on GDNF are manifested at a higher degree of inflammatory activity.

Animals↗

PubMed2004

Cyclic AMP/protein kinase a signal attenuates Ca(2+)-induced fibroblast growth factor-1 synthesis in rat cortical neurons.

Fibroblast growth factor (FGF)-1 is increased in particular brain regions after birth, suggesting an involvement of some regulatory neuronal circuits. To address the neuronal activity responsible for FGF-1 synthesis, effects of various neurotransmitter receptor activation on cellular FGF-1 content were examined using cultured rat cortical neurons. Histamine, glutamate, carbachol, serotonin or gamma-aminobutyric acid (GABA) caused an increase of FGF-1 content. Because this effect was mimicked by (1) N-methyl-D-aspartate, a glutamatergic agonist; (2) Ca(2+) ionophore; (3) depolarization with high concentration of KCl, but was abolished in Ca(2+)-free medium, Ca(2+) influx was thought to trigger FGF-1 synthesis. Such Ca(2+)-mediated enhancement of FGF-1 synthesis, however, did not occur in the presence of norepinephrine (NE), but was restored by KT-5720, an inhibitor of protein kinase A (PKA), suggesting an interplay between Ca(2+)-activated and cAMP/PKA signals for neuronal FGF-1 synthesis. This mechanism was proved to function in vivo by stimulation of FGF-1 expression in neurons of the cerebral cortex after intracerebral administration of propranolol, an antagonist of adrenergic beta receptors. This demonstrates that FGF-1 synthesis is essentially upregulated by Ca(2+) influx through excitatory neuronal activities, but such an effect is abolished by neurotransmission that evokes cAMP/PKA signals. FGF-1 produced is thought to act on establishment and maintenance of particular neuronal circuits in the brain, which may be one of the ways neurotransmitters regulate brain function.

Adrenergic beta-Antagonists↗

PubMed2004

Orally administered bovine lactoferrin induces caspase-1 and interleukin-18 in the mouse intestinal mucosa: a possible explanation for inhibition of carcinogenesis and metastasis.

We have previously demonstrated that oral administration of bovine lactoferrin (bLF) markedly inhibits lung metastatic colony formation, and that this inhibition was possibly due to the activation of T and NK cells. Furthermore, we found that interleukin-18 (IL-18) is induced in epithelial cells of the small intestine by bLF. The present study was undertaken to confirm cytokine production in response to bLF and to assess the underlying mechanisms. Markedly elevated IL-18 levels were found in the small intestine 1-3 h after a single administration of bLF, its pepsin hydrolysate (bLFH), or bTF. Importantly, while IL-18 was significantly increased after a regimen of seven daily administrations of bLF or bLFH, administration of bTF over the course of seven days had little or no effect. In addition to IL-18, a significant increase in caspase-1 activity and interferon-gamma (IFN-gamma) was found in the small intestine after administration of bLF. Similarly, in peritoneal macrophages, bLF markedly enhanced caspase-1 activity and IL-18 levels. Finally, a caspase-1 inhibitor significantly decreased bLF mediated induction of IL-18 in vitro. (bTF had no effect on either caspase-1 or IFN-gamma or on IL-18 in vitro.) These results demonstrate the possibility that elevation of caspase-1 activity by bLF and its hydrolysate may be important for production of mature IL-18 in vivo, and thus in potentiating the killing activity of T and NK cells against tumor cells.

Administration, Oral↗

PubMed2004

Preferential mammary carcinogenic effects of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) in human c-Ha-ras proto-oncogene transgenic rats.

In order to clarify the susceptibility of the Hras128 rat harboring copies of the human c-Ha-ras proto-oncogene to 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), Hras128 rats were intragastically treated with 100 mg/kg PhIP 8 times (females) or 80 mg/kg PhIP 10 times (males) over a 9-week period, then sacrificed at weeks 12 and 30. Multiple mammary tumors of adenocarcinoma type were induced in all females, while 83% of treated males developed adenocarcinomas, sarcomas and transitional carcinosarcomas, as evidenced by casein and vimentin immunoreactivity. All tumors examined had mutations in the c-Ha-ras transgene, while the endogenous rat c-Ha-ras gene was intact. Our results indicate that 1) Hras128 rats of both sexes are preferentially susceptible to mammary carcinogenesis with PhIP; 2) activation of the transgene, but not the endogenous c-Ha-ras gene, may be important in this regard; 3) the variety of tumor types evident in male rats indicates that immature mammary gland cells of the terminal end buds may be a target of PhIP; 4) although the transgene is expressed in all organs, susceptibility to PhIP is limited to mammary glands.

Animals↗

PubMed2004

Chemoprevention of colorectal cancer by grape seed proanthocyanidin is accompanied by a decrease in proliferation and increase in apoptosis.

Effects of proanthocyanidin (PA), procyanidin B-2 (B-2), and epigallocatechin gallate (EGCG) on azoxymethane (AOM)-induced colonic preneoplastic aberrant crypt foci (ACF) formation were investigated using F344 rats. The numbers of total ACF in rats treated with 0.002% PA and 0.05% B-2 were significantly decreased compared with the AOM alone group (control). Cell proliferation in the colon, as shown by proliferating cells nuclear antigen (PCNA), was also reduced in those treatments. The single-stranded DNA (ssDNA) labeling index, a marker for apoptosis, was significantly increased in 0.002% PA and 0.05% B-2 groups compared with control. Moreover, the numbers of CD11b/c+ cells (macrophages) and NKR-P1A+ cells (NK cells) in the all groups were significantly increased compared with control. In an in vitro study using rat colon cancer cell line RCN-9, PA, especially 5-10mer of PA (PA5/10), strong growth inhibition was shown. PA5/10 caused the most remarkable apoptosis as cleared by FACS analysis. These cells showed significantly increased caspase-3 activity. The results would suggest that the PA, especially PA5/10, might strongly enhance caspase-3 activity and cause apoptosis in cancer cells. PA at fairly low doses in the long term might serve as an effective means for preventing colon carcinogenesis.

Animals↗

PubMed2004

Cancer prevention by natural compounds.

Increasing attention is being paid to the possibility of applying cancer chemopreventive agents for individuals at high risk of neoplastic development. For this purpose by natural compounds have practical advantages with regard to availability, suitability for oral application, regulatory approval and mechanisms of action. Candidate substances such as phytochemicals present in foods and their derivatives have been identified by a combination of epidemiological and experimental studies. Plant constituents include vitamin derivatives, phenolic and flavonoid agents, organic sulfur compounds, isothiocyanates, curcumins, fatty acids and d-limonene. Examples of compounds from animals are unsaturated fatty acids and lactoferrin. Recent studies have indicated that mechanisms underlying chemopreventive potential may be combinations of anti-oxidant, anti-inflammatory, immune-enhancing, and anti-hormone effects, with modification of drug-metabolizing enzymes, influence on the cell cycle and cell differentiation, induction of apoptosis and suppression of proliferation and angiogenesis playing roles in the initiation and secondary modification stages of neoplastic development. Accordingly, natural agents are advantageous for application to humans because of their combined mild mechanism. Here we review naturally occurring compounds useful for cancer chemprevention based on in vivo studies with reference to their structures, sources and mechanisms of action.

Animals↗

PubMed2003

Growth arrest of PC12 cells by nerve growth factor is dependent on the phosphatidylinositol 3-kinase/Akt pathway via p75 neurotrophin receptor.

We recently isolated mutant PC12 cell clones (PC84 cells) by transfection of PC12 cells with nerve growth factor (NGF) cDNA. These cells secreted active NGF and extended short processes, but proliferated faster than the parental PC12 cells. Because the expression level of p75, a low-affinity receptor for NGF, was significantly low, we suspected that NGF signaling via p75 was necessary for the growth arrest of the PC12 cells, and this was shown to be the case by repressing p75 function in PC12 cells. In this study, we examined the downstream signaling of p75, which would ultimately evoke the growth arrest. NGF is known to induce rapid phosphorylation of MAP kinase and Akt in PC12 cells, whereas in PC84 cells, MAP kinase was phosphorylated but the phosphorylation level of Akt was very low under the serum-free condition. This finding suggested that the low expression level of p75 in PC84 cells was the reason for the low Akt activation. Because Akt is known to be activated via phosphatidylinositol (PI) 3-kinase, we treated PC12 cells with a PI3-kinase inhibitor, Wortmannin, and found these cells did not cease proliferation in the presence of NGF. Furthermore, anti-p75 neutralizing antibody reduced NGF-induced phosphorylation of Akt in PC12 cells under the serum-free condition. Because we had already shown that PC12 cells treated with anti-p75 neutralizing antibody did not cease proliferation in the presence of NGF, these results suggest that NGF activates Akt via p75, which is necessary for the NGF-induced growth arrest of PC12 cells.

Androstadienes↗

PubMed2003

Neurotrophins facilitate synthesis of choline acetyltransferase and tyrosine hydroxylase in cultured mouse neural stem cells independently of their neuronal differentiation.

Effects of three neurotrophins, i.e., nerve growth factor, brain-derived neurotrophic factor, and neurotrophin-3, on the expression of four neurotransmitter-synthesizing enzymes, i.e. choline acetyltransferase (ChAT), tyrosine hydroxylase (TH), dopamine beta hydroxylase (DBH), and glutamate decarboxylase 65 were investigated in cultured mouse neural stem cells. All three neurotrophins enhanced the mRNA expression of ChAT, TH, or DBH of the cells caused to differentiate by the removal of fibroblast growth factor (FGF)-2 from the culture medium, and increased the protein and mRNA levels of ChAT and TH of even the undifferentiated proliferating neural stem cells due to the presence of FGF-2. These results demonstrate that neurotrophins stimulate the synthesis of ChAT and TH of the neural stem cells prior to neuronal differentiation, and suggest that neurotrophins may play roles in the commitment to neuronal cells and choice of specific neurotransmitter phenotypes in early stages of neurogenesis.

Animals↗

PubMed2003

Neurotrophins facilitate neuronal differentiation of cultured neural stem cells via induction of mRNA expression of basic helix-loop-helix transcription factors Mash1 and Math1.

Neurogenesis is promoted by basic helix-loop-helix (bHLH) transcription factors Mash1, Math1, or NeuroD but suppressed by another set, Hes1 and Hes5. It remains unknown what kinds of extracellular signals are involved in their regulation; therefore, the effects of neurotrophins on the expression of bHLH factors and neuronal differentiation were investigated by the use of cultured mouse neural stem cells. Each neurotrophin increased Mash1 and Math1 mRNAs of the stem cells growing in the presence of fibroblast growth factor-2 (FGF-2), but did not alter Hes1, Hes5, or NeuroD mRNA levels. Simultaneously, most of the cells expressed nestin but not microtubule-associated protein 2 (MAP2), and remained undifferentiated. FGF-2 removal from the medium reduced the levels of Hes1 and Hes5 mRNAs and increased those of Mash1, Math1, and NeuroD mRNAs, resulting in substantial neuronal differentiation. When the cells were pretreated with brain-derived neurotrophic factor, a neurotrophin, FGF-2 removal enhanced earlier NeuroD expression and generated many more MAP2-positive cells. The high level of Mash1 and Math1 that had been elevated at FGF-2 withdrawal accelerated NeuroD expression in cooperation with the reduced Hes1 and Hes5 expression. Our present results suggest that neurotrophins stimulate neuronal differentiation by altering the balance of expression of various bHLH transcription factors.

Animals↗

PubMed2003

Carcinogenesis and its modification by environmental endocrine disruptors: in vivo experimental and epidemiological findings.

Although a great deal of concern has been raised about the hazard potential of endocrine disruptors present in the environment, the in vivo data available from both experimental and epidemiological studies suggest that the majority of those agents do not pose a risk with regard to cancer development. Indeed, naturally occurring examples such as isoflavonoids even appear to exert protective effects. Only for xenobiotics such as 1,1,1-trichloro-2,2-bis(p-chlorophenyl)ethane (DDT), polychlorinated biphenyls (PCBs) and tetrachloro-p-dioxin (TCDD) and special cases of phenols and phthalates is there unequivocal evidence of carcinogenicity and this appears to be directly linked to their toxicity. Thus, careful in vivo assessment is required before drawing any conclusions regarding agents capable of affecting the mammalian endocrine system.

Animals↗

PubMed2002

4-Methylcatechol stimulates phosphorylation of Trk family neurotrophin receptors and MAP kinases in cultured rat cortical neurons.

Effects of 4-methycatechol (4MC), a potent stimulator of nerve growth factor and brain-derived neurotrophic factor (BDNF) synthesis, on phosphorylation of cellular molecules in cultured rat cortical neurons were examined. 4MC stimulated tyrosine phosphorylation of various proteins of molecular weight from 10-300 kDa including Trks, which are high-affinity neurotrophin receptors. Moreover, 4MC enhanced the phosphorylation of serine 133 of mitogen-activated protein kinase (MAPK/ERK) in a dose-dependent manner. Pretreatment of cultures with PD98059, a selective inhibitor of MAPK kinase (MEK-1), inhibited 4MC-induced phosphorylation of ERKs, demonstrating MEK-1-mediated activation. Therefore, it seems that 4MC triggered the phosphorylation of Trks, resulting in the activation of the subsequent MAPK/ERK signal cascade, or perhaps the involvement of BDNF action as 4MC can stimulate neuronal BDNF synthesis. The phosphorylation of MAPK/ERK was unaffected, however, in the presence of cycloheximide, a protein synthesis inhibitor, and K252a, a selective inhibitor of Trks, suggesting that the effect of newly synthesized BDNF was negligible on this event, and that primary sites of 4MC actions are not limited only to Trks. These results suggest that 4MC primarily activates multiple signal transduction molecules such as tyrosine kinases, including Trks. A significant increase in the survival rate of cortical neurons in the presence of 10 or 100 nM 4MC supported this idea, because the concentrations were much lower than those for stimulation of BDNF synthesis. Our results strongly suggest that the neurotrophic actions of 4MC found so far are mediated predominantly by direct activation of some intracellular signals including MAPK/ERK rather than by neurotrophin synthesis.

Aging↗

PubMed2002

Role of low-affinity p75 receptor in nerve growth factor-inducible growth arrest of PC12 cells.

Mutant PC12 cell clones (PC84 cells) were obtained by transfection with nerve growth factor (NGF) cDNA. These cells secreted active NGF, extended short processes, and proliferated faster than the parental PC12 cells. These features are of great interest because the parental PC12 cells cease proliferation and extend long processes when transfected with NGF cDNA. PC84 cells expressed a high level of acetylcholinesterase activity and neurofilament M, which indicates that PC84 cells were differentiated. The inhibition of TrkA by K252a diminished the short processes of PC84 cells but had no effect on their fast proliferation. The expression level of TrkA in PC84 cells was comparable to that in PC12 cells; whereas that of another NGF receptor, p75, was significantly lower. These data suggest that the decrease of p75 contributed to the continuous growth of PC84 cells, which was confirmed by suppressing p75 activity of PC12 cells with the antisense oligonucleotide of p75 or with anti-p75 neutralizing antibody. The treated cells did not cease proliferation in the presence of NGF and extended short processes. Our results suggest that NGF signaling via TrkA affects the differentiation characteristics of PC12 cells but that an additional signaling via p75 is necessary for the growth arrest of the cells.

Acetylcholinesterase↗