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J Hamada

Publications and source records attributed to J Hamada.

At least 19 recordsLinked to original sources
PubMed2003

Activation of caspase-12 by endoplasmic reticulum stress induced by transient middle cerebral artery occlusion in mice.

We sought to clarify the involvement of caspase-12, a representative molecule related to endoplasmic reticulum (ER) stress-induced cell-death signaling pathways, in neuronal death resulting from ischemia/reperfusion in mice. Transient focal cerebral ischemia (1 h) was produced by intraluminal occlusion of the middle cerebral artery (MCA). We assessed the expression patterns of caspase-12, Bip/GRP78, an ER-resident molecular chaperone whose expression serves as a good marker of ER stress, and caspase-7 by Western blotting and/or immunohistochemistry. Double-fluorescent staining of caspase-12 immunohistochemistry and the terminal deoxynucleotidyl transferase-mediated DNA nick-end labeling (TUNEL) method was performed to clarify the involvement of caspase-12 in cell death. We confirmed that ER stress was induced during reperfusion in our model, as witnessed by up-regulated Bip/GRP78 expression in the MCA territory. Western blot analysis revealed that caspase-12 activation occurred at 5-23 h of reperfusion, and immunoreactivity for caspase-12 was enhanced mainly in striatal neurons on the ischemic side at the same time points. We found the co-localization of caspase-12 immunoreactivity and DNA fragmentation detectable by the TUNEL method. We did not detect the presence of caspase-7 in the ER fraction at the period of caspase-12 cleavage. Our results imply that cerebral ischemia/reperfusion induces ER stress and that caspase-12 activation concurred with ER stress. Caspase-12 seems to be involved in neuronal death induced by ischemia/reperfusion. Caspase-7 is not likely to contribute to the cleavage of caspase-12 in our experimental model.

Animals↗
PubMed2003

The neuroprotective effect of a novel calmodulin antagonist, 3-[2-[4-(3-chloro-2-methylphenyl)-1-piperazinyl]ethyl]-5,6-dimethoxy-1-(4-imidazolylmethyl)-1H-indazole dihydrochloride 3.5 hydrate, in transient forebrain ischemia.

A novel calmodulin (CaM) antagonist DY-9760e, (3-[2-[4-(3-chloro-2-methylphenyl)-1-piperazinyl]ethyl]-5,6-dimethoxy-1-(4-imidazolylmethyl)-1H-indazole dihydrochloride 3.5 hydrate), with an apparent neuroprotective effect in vivo, potently inhibits CaM-dependent nitric oxide synthase in situ. In the present study, we determined whether DY-9760e inhibits nitric oxide (NO) production and protein nitration by peroxynitrite (ONOO(-)) formation in the hippocampal CA1 region of gerbils after transient forebrain ischemia. In freely moving gerbils, NO production after 10-minute forebrain ischemia was monitored consecutively with in vivo brain microdialysis. Pretreatment with DY-9760e (50 mg/kg i.p.) significantly decreased the increased levels of NO(x)(-) (NO metabolites, NO(2)(-) plus NO(3)(-)) immediately after, 24 h after cerebral ischemia-reperfusion to the control levels of sham-operated animals. Western blot and immunohistochemical analyses using an anti-nitrotyrosine antibody as a marker of ONOO(-) formation indicated a marked increase in nitrotyrosine immunoreactivity in the pyramidal neurons of the CA1 region 2 h after reperfusion, and DY-9760e significantly inhibited increased nitrotyrosine immunoreactivity. Coincident with the inhibition of the NO production and protein tyrosine nitration, pretreatment with DY-9760e rescued the delayed neuronal death in the hippocampal CA1 region. These results suggest that the inhibitory effects of DY-9760e on the NO-ONOO(-) pathway partly account for its neuroprotective effects in cerebral ischemia.

Animals↗
PubMed2001

Expression and characterization of hypoxia-inducible factor (HIF)-3alpha in human kidney: suppression of HIF-mediated gene expression by HIF-3alpha.

Hypoxia inducible factors (HIFs) are heterodimeric transcription factors that regulate a number of adaptive responses to low oxygen tension. They are composed of alpha- and beta-subunits that belong to the basic helix-loop-helix-PAS superfamily. Here we examined the expression of HIF-alpha subunit proteins in the human kidney and found that, in addition to HIF-1alpha and -2alpha, HIF-3alpha was also expressed. The sequence analysis revealed that, like mouse HIF-3alpha, human HIF-3alpha has high similarity with HIF-1alpha and -2alpha in the bHLH and PAS domains, but lacks structures for transactivation found in the C-terminus of HIF-1alpha and -2alpha. Furthermore, we performed reporter gene analysis and showed that HIF-3alpha suppresses hypoxia-inducible HIF-mediated gene expression. HIF-3alpha might be a negative regulator of hypoxia-inducible gene expression in the human kidney.

Amino Acid Sequence↗
PubMed2001

Intravenously injected FK506 failed to inhibit hippocampal calcineurin.

FK506 (tacrolimus) is known as an inhibitor for calcineurin and is used in numerous research fields. It is not clear whether intravenously injected FK506 inhibits neuronal calcineurin. We measured the calcineurin activities of normal and postischemic rat hippocampi after intravenous injection of FK506 (3 mg/kg). Intravenously injected FK506 had no inhibitory effect on calcineurin activity in the hippocampi of normal and postischemic rats, whereas FK506 inhibited the calcineurin in vitro (purified enzyme, hippocampal homogenate, and hippocampal slice culture homogenate). Thus, it is considered that intravenously injected FK506 does not act on intraneuronal calcineurin and that several effects of FK506 are not due to the inhibition of neuronal calcineurin.

Animals↗
PubMed2001

Expression of angiopoietin-2 in human glioma cells and its role for angiogenesis.

In highly vascular malignant glioma, glioma cells themselves may express angiogenic factors and induce angiogenesis. Recent studies have shown that novel angiogenic factors, angiopoietin-1 (Ang1) and -2 (Ang2), play important roles in the modulation of vasculogenesis and angiogenesis. In this study, we determined Ang2 mRNA expression in cultured human malignant glioma cells (U105, U251, and U373 MG) by reverse transcriptase-PCR. Western blot analysis and immunocytochemical analysis with antihuman Ang2 antibody revealed that Ang2 protein was expressed and secreted by these cells. Furthermore, hypoxia increased the Ang2 protein level in cultured glioma cells. Serial sections of 32 human glioma tissues (14 glioblastomas, eight anaplastic astrocytomas, seven astrocytomas, and three pilocytic astrocytomas) were immunostained against Ang2, vascular endothelial growth factor, Tie2, von Willebrand factor, and alpha smooth muscle actin. The immunoreactivity of each angiogenic factor was higher in malignant gliomas than in low-grade gliomas. Ang2 protein was detected not only in endothelial cells but also in glioma cells, and its expression was prominent in both the area surrounding the necrosis and the periphery of glioblastomas. In the area surrounding necrosis, Ang2 was highly expressed and tumor vessels showed regression. In the tumor periphery, Ang2 was highly expressed and many small vessels stained positively for von Willebrand factor but not for alpha smooth muscle actin, suggesting angiogenesis. Statistical analysis revealed that the Ang2 expression was negatively correlated with vessel maturation in malignant gliomas and that vascular endothelial growth factor expression was positively correlated with vessel maturation in low-grade gliomas (P < 0.05). These results suggest that glioma cells themselves express Ang2 and that expression may be induced by hypoxic stimulation and may play a crucial role in the vessel maturation, angiogenesis, and vessel regression in malignant glioma.

Adolescent↗
PubMed2001

Interaction of MCC2, a novel homologue of MCC tumor suppressor, with PDZ-domain Protein AIE-75.

AIE-75 is a protein identified as an autoantigen in patients with autoimmune enteropathy and as a colon cancer-related antigen. It has recently been assigned to be a causative gene for Usher type 1C congenital syndromic hearing loss. The novel protein has three PSD-95/Dlg/ZO-1 (PDZ) protein-protein interaction domains and is therefore implicated to function as a molecular anchor or sorter. We have identified a novel protein that binds to AIE-75 by yeast two-hybrid screening. The protein has a high homology to the tumor suppressor MCC (mutated in colon cancer; or MCC1 hereafter) and was named MCC2. MCC2 protein binds the first PDZ domain of AIE-75 with its C-terminal amino acids -DTFL. Since the MCC1 does not bind to AIE-75 and the MCC2 displays different expression patterns in various organs compared to MCC1, they appear to play distinct roles in cells. The MCC2 gene is located on chromosome 19p13 in the vicinity of APCL gene, while MCC1 maps near to APC tumor suppressor gene. Because of negative expression of MCC2 in a panel of cancer cell-lines compared to the corresponding normal tissues, we suggest that further study is necessary to investigate a possible role of MCC2 as a tumor suppressor.

Adaptor Proteins, Signal Transducing↗
PubMed2001

Increased oxidative DNA damage in mammary tumor cells by continuous epidermal growth factor stimulation.

BACKGROUND: Growth factors can enhance the malignant potential of tumor cells. To examine the relationship between growth factors and tumor progression, we previously established a weakly malignant cell line, ER-1. We found that a 24-hour exposure of ER-1 cells to epidermal growth factor (EGF) induced malignant properties (tumor progression) that were reversible but that, after a 1-month exposure, these changes were irreversible. In this study, we investigated the irreversible changes induced in ER-1 cells by a 1-month exposure to EGF and the possible involvement of oxidative stress. METHODS: ER-1 cells were treated with EGF (100 ng/mL) for 1 month in the presence or absence of an antioxidant, N-acetylcysteine or selenium, and compared with untreated control ER-1 cells. We assessed tumor progression by measuring intracellular peroxide levels, 8-hydroxydeoxyguanosine (a marker for oxidative DNA damage) levels, in vitro invasiveness, and in vivo tumorigenicity and metastatic ability. All statistical tests are two-sided. RESULTS: After ER-1 cells were treated for 1 month with EGF, levels of intracellular peroxide and 8-hydroxyguanosine in the DNA of treated cells were higher than those in the DNA of control cells, and treated ER-1 cells were more tumorigenic and metastatic in vivo and more invasive in vitro than untreated control cells (all P<.001). Levels of 8-hydroxyguanosine in DNA increased as the length of the EGF treatment increased (P<.001). However, when N-acetylcysteine or selenium was added with EGF for 1 month, levels of intracellular peroxide and 8-hydroxyguanosine in DNA were comparable to those in control cells (r =.795). Both tumorigenicity (P =.008) and metastatic ability (P<.001) decreased after addition of N-acetylcysteine or selenium. CONCLUSION: The irreversible changes caused by continuous EGF stimulation of ER-1 cells result from increased oxidative damage in the DNA, which generates tumor cells with more malignant characteristics.

8-Hydroxy-2'-Deoxyguanosine↗
PubMed2001

Gelsolin functions as a metastasis suppressor in B16-BL6 mouse melanoma cells and requirement of the carboxyl-terminus for its effect.

Gelsolin, an actin-binding protein, is implicated as a critical regulator in cell motility. In addition, we have reported that cellular levels of gelsolin are decreased in various tumor cells, and overexpression of gelsolin by gene transfer suppresses tumorigenicity. We sought to assess the effects of gelsolin overexpression on metastasis and to determine the importance of a carboxyl-terminus that confers Ca(2+) dependency on gelsolin for effects of its overexpression. Expression vectors with cDNA encoding either full-length wild-type or His321 mutant form, isolated from a flat revertant of Ras-transformed cells and a carboxyl-terminal truncate, C-del of gelsolin, were transfected into a highly metastatic murine melanoma cell line, B16-BL6. Expression of introduced cDNA in transfectants was confirmed using Western blotting, 2-dimensional gel electrophoresis and reverse transcription-polymerase chain reaction (RT-PCR). We characterized phenotypes of transfectants, such as growth rate, colony formation in soft agar, cell motility and metastasis formation in vivo. Transfectants expressing the wild-type, His321 mutant and C-del gelsolin exhibited reduced growth ability in soft agar. Although expression of integrin beta1 or alpha4 on the cell surface of transfectants was not changed, wild-type and His321 mutant gelsolin, except for C-del gelsolin, exhibited retardation of cell spreading, reduced chemotatic migration to fibronectin and suppressed lung colonization in spontaneous metastasis assay. Gelsolin may function as a metastasis suppressor as well as a tumor suppressor gene. The carboxyl-terminus of gelsolin is important for retardation of cell spreading, reduced chemotasis and metastasis suppression.

Animals↗
PubMed2001

Development of a yeast stop codon assay readily and generally applicable to human genes.

We established a yeast-based method to screen chain-terminating mutations that is readily applicable to any gene of interest. Based on the finding that 18- to 24-base-long homologous sequences are sufficient for gap repair in vivo in yeast, we used a strategy to amplify a test-gene fragment with addition of 24-bp sequences homologous to both cut-ends of a yeast expression vector, pMT18. After co-transformation with the amplified fragment and the linearized pMT18, each yeast (Saccharomyces cerevisiae) cell automatically forms a single-copy circular plasmid (because of CEN/ARS), which expresses a test-gene::ADE2 chimera protein. When the reading frame of the test-gene contains a nonsense or frameshift mutation, truncation of the chimera protein results in lack of ADE2 activity, leading to formation of a red colony. By using a nested polymerase chain reaction using proofreading Pfu polymerase to ensure specificity of the product, the assay achieved a low background (false positivity). We applied the assay to BRCA1, APC, hMSH6, and E-cadherin genes, and successfully detected mutations in mRNA and genomic DNA. Because this method--universal stop codon assay--requires only 4 to 5 days to screen a number of samples for any target gene, it may serve as a high-throughput screening system of general utility for chain-terminating mutations that are most prevalent in human genetic diseases.

Adenomatous Polyposis Coli Protein↗
PubMed2001

Management outcome for ruptured anterior circulation aneurysms with a Hunt and Hess clinical grade of III in patients in the 9th decade of life.

BACKGROUND: This study was conducted to define general guidelines for rational clinical decision-making with respect to patients older than 80 years who present with ruptured anterior circulation aneurysms and a Hunt and Hess clinical grade of III. METHODS: The 29 consecutive patients enrolled in this study were treated at 4 participating centers between 1995 and 1998. All were 80 years or older on admission and met 7 eligibility criteria. The decision to treat surgically or non-surgically was made individually at each center. The outcome at discharge was assessed using the Glasgow Outcome Scale (GOS); the Barthel Score (BS) was used to determine the patients' quality of life after discharge. RESULTS: Of 15 surgically treated patients, 4 died before discharge and 8 were discharged in poor condition. Of these, 7 died of unrelated causes within 2 years of the SAH and one is living in a nursing home. Three patients with middle cerebral artery (MCA) aneurysms made a good recovery; they are currently living at home. Of the 14 conservatively treated patients, 10 died during their hospital stay mainly because of rebleeding that occurred within 3 weeks of the subarachnoid hemorrhage (SAH). Three others who were discharged in poor condition died of systemic illness within 2 years. The remaining patient is living in a nursing home with a BS value of 0. CONCLUSIONS: The outcomes for patients treated conservatively were catastrophic. Even in surgically treated patients, favorable outcomes were rare; the best results were achieved in surgically treated patients with MCA aneurysms.

Aged↗
PubMed2001

Activation of Akt/protein kinase B contributes to induction of ischemic tolerance in the CA1 subfield of gerbil hippocampus.

Apoptosis plays an important role in delayed neuronal cell death after cerebral ischemia. Activation of Akt/protein kinase B has been recently reported to prevent apoptosis in several cell types. In this article the authors examine whether induction of ischemic tolerance resulting from a sublethal ischemic insult requires Akt activation. Sublethal ischemia gradually and persistently stimulated phosphorylation of Akt-Ser-473 in the hippocampal CA1 region after reperfusion. After lethal ischemia, phosphorylation of Akt-Ser-473 showed no obvious decrease in preconditioned gerbils but a marked decrease in nonconditioned gerbils. Changes in Akt-Ser-473 phosphorylation were correlated with changes in Akt activities, as measured by an in vitro kinase assay. Intracerebral ventricular infusion of wortmannin before preconditioning blocked both the increase in Akt-Ser-473 phosphorylation in a dose-dependent manner and the neuroprotective action of preconditioning. These results suggest that Akt activation is induced by a sublethal ischemic insult in gerbil hippocampus and contributes to neuroprotective ischemic tolerance in CA1 pyramidal neurons.

Androstadienes↗
PubMed2001

Neuroprotective effect of sodium orthovanadate on delayed neuronal death after transient forebrain ischemia in gerbil hippocampus.

In transient forebrain ischemia, sodium orthovanadate as well as insulinlike growth factor-1 (IGF-1) rescued cells from delayed neuronal death in the hippocampal CA1 region. Adult Mongolian gerbils were subjected to 5-minute forebrain ischemia. Immunoblotting analysis with anti-phospho-Akt/PKB (Akt) antibody showed that phosphorylation of Akt at serine-473 (Akt-Ser-473) in the CA1 region decreased immediately after reperfusion, and in turn transiently increased 6 hours after reperfusion. The decreased phosphorylation of Akt-Ser-473 was not observed in the CA3 region. The authors then tested effects of intraventricular injection of orthovanadate and IGF-1, which are known to activate Akt. Treatment with orthovanadate or IGF-1 30 minutes before ischemia blocked delayed neuronal death in the CA1 region. The neuroprotective effects of orthovanadate and IGF-1 were associated with preventing decreased Akt-Ser-473 phosphorylation in the CA1 region observed immediately after reperfusion. Immunohistochemical studies with the anti-phospho-Akt-Ser-473 antibody also demonstrated that Akt was predominantly in the nucleus and was moderately activated in the cell bodies and dendrites of pyramidal neurons after orthovanadate treatment. The orthovanadate treatment also prevented the decrease in phosphorylation of mitogen-activated protein kinase (MAPK). Pretreatment with combined blockade of phosphatidylinositol 3-kinase and MAPK pathways totally abolished the orthovanadate-induced neuroprotective effect. These results suggest that the activation of both Akt and MAPK activities underlie the neuroprotective effects of orthovanadate on the delayed neuronal death in the CA1 region after transient forebrain ischemia.

Androstadienes↗
PubMed2001

Possible involvement of endothelin-1 and nitric oxide in the pathogenesis of proliferative diabetic retinopathy.

BACKGROUND: Overproduction of endothelin-1 (ET-1) and nitric oxide (NO) in the retina is demonstrated in experimental diabetic animals. To clarify the possible involvement of ET-1 and NO in the pathogenesis of diabetic retinopathy, the authors examined the vitreous levels of these principal endothelium-derived vasoactive substances in patients with proliferative diabetic retinopathy (PDR). METHODS: Vitreous fluid was taken from patients with PDR (ET-1, n = 12; NO, n = 12) and from patients with macular holes as controls (ET-1, n = 10; NO, n = 10) at vitreous surgery. Endothelin-1 and NO metabolites were measured by radioimmunoassay and high-performance liquid chromatography based on the Griess method, respectively. RESULTS: Endothelin-1 levels (mean +/- SE) were 21.5 +/- 1.7 pg/mL in the vitreous of patients with PDR and 16.7 +/- 0.7 pg/mL in the vitreous of patients with macular hole. There was a significant difference between patients with PDR and controls (P = 0.009, Mann-Whitney). Nitrate (NO3) was 49.8 +/- 5.0 micromol/L in patients with PDR and 24.2 +/- 2.8 micromol/L in patients with macula hole; it was also significantly elevated in patients with PDR (P = 0.004, Mann-Whitney), whereas nitrite (NO2) was not detected in this study. CONCLUSION: These results indicate that ET-1 and NO may be related in the pathogenesis of PDR.

Adult↗
PubMed2001

Inhibition of nitric oxide production during global ischemia ameliorates ischemic damage of pyramidal neurons in the hippocampus.

We examined the relationship between nitric oxide (NO) production and delayed neuronal death (DND), in the rat hippocampus induced by 21 minutes of transient global ischemia produced by the occlusion of both of the common carotid arteries combined with systemic hypotension. NO production during ischemia and reperfusion was investigated by quantifying the nitrite (NO2-) levels of the in vivo microdialysis samples collected ever 3 minutes from the hippocampus. To determine the origin of NO production, we studied the effects of the focal administration of NG-nitro-L-arginine methyl ester (L-NAME), an inhibitor of the constitutive NO synthase (NOS). We also carried out systemic administration of a selective neuronal NOS inhibitor, 7-nitroindazole (7-NI). Rats were grouped as follows: group 1 (n = 22), vehicle; group 2 (n = 19), L-NAME; group 3 (n = 12), 7-NI; and group 4 (n = 12), a sham operation. The role of NO in the hippocampal DND was investigated histologically one week after ischemia. The level of NO production was significantly decreased in groups 2 and 3 as compared to group 1 in which NO production was significantly increased (p < 0.05). The density of remaining neurons in the CA1 area was significantly reduced only in group 1 (p < 0.01). Taken together, it can be concluded that NO production by neuronal NOS during ischemia and reperfusion resulted in DND in the CA1 region of the rat hippocampus.

Animals↗
PubMed2001

[Endovascular treatment for facial arteriovenous malformations using new particles: report of two cases].

Facial arteriovenous malformations (FAVM) are difficult to treat because of their highly vascular networks. Intravascular treatment using liquid material to occlude the FAVM occasionally results in skin necrosis after embolization. The use of particulate materials to obliterate the nidus often fails to obtain a permanent cure due to arterial recanalization. We report two patients with FAVM who were successfully treated with endovascular embolization using a new type of particulate material. One patient was treated with embolization only, the other was treated with embolization followed by surgical resection. Both patients showed clinical and angiographic improvement. Intravascular treatment using particles with a smooth surface and optimal size is safe and effective in the treatment of patients with FAVM.

Adolescent↗
PubMed2001

[Successful treatment with bypass and interventional surgery for a ruptured pseudo carotid artery aneurysm after transsphenoidal surgery: a case report].

We report case of a patient who suffered a pseudo internal carotid artery (IC) aneurysm following transsphenoidal surgery. He was successfully treated with bypass surgery and IC occlusion involving the pseudoaneurysm using Guglielmi detachable coils (GDCs). This 50-year-old man with recurrent FSH-releasing pituitary adenoma suffered profuse arterial bleeding during transsphenoidal surgery. The hemorrhage was managed, using oxidized cellulose with bio-bond. His postoperative course was uneventful, but, he developed massive epistaxis 20 days after surgery. Cerebral angiograms showed a pseudoaneurysm arising from the C4 portion of the left IC. He could not tolerate the balloon occlusion test. Using GDCs, we immediately performed left IC occlusion involving the pseudoaneurysm followed by bypass surgery between the left EC and the left middle cerebral artery. Postoperative angiograms showed that the pseudoaneurysm was completely occluded and the bypass was fully patent. When massive arterial bleeding is encountered during transsphenoidal surgery, the patient should be carefully monitored to detect early the development of a pseudoaneurysm. When such an aneurysm is found or has ruptured, interventional surgery has proved effective in the management of this complication.

Adenoma↗
PubMed2001

Analysis of calcium deposits in calcific periarthritis.

OBJECTIVE: To determine if hydroxyapatite (HAP), octacalcuim phosphate (OCP), or tricalcium phosphate (TCP) can be found in the calcium deposits in calcific periarthritis. METHODS: Thirty-six specimens from 34 patients who had acute inflammation and roentgenographically recognized calcification in soft tissue were analyzed. Twenty-three patients with calcific tendinitis in the shoulder and 11 with calcific periarthritis at other sites were included. We prepared 2 kinds of samples from each specimen; a dried sample (washed and dried calcific deposit), and a sample heated to 1,000 degrees C. All were analyzed by X-ray diffraction, Raman spectroscopy, infrared absorption spectroscopy, and X-ray fluorescence spectrometry for calcium and phosphorus molar ratio. Synthetic HAP was used as the control in each analysis. RESULTS: The X-ray diffraction patterns of all dried samples were similar to those of HAP and carbonate apatite. We found no diffraction patterns of OCP or TCP. However, an OH- group at 3570cm(-1) was observed with Raman spectroscopy for samples heated to 1,000 degrees C and synthetic HAP, but not for the dried samples. Infrared absorption spectroscopy also confirmed an OH- group for samples heated to 1,000 degrees C and synthetic HAP, and confirmed that dried samples contained carbonate. CONCLUSION: Calcium deposits are composed of carbonate apatite. HAP, OCP, and TCP were not identified in any deposits.

Adult↗