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Biomedical subjects

M Hirata

Publications and source records attributed to M Hirata.

At least 127 records · Page 7Linked to original sources
PubMed1999

Antibodies against the PH domain of phospholipase C-delta1 inhibit Ins(1,4,5)P3-mediated Ca2+ release from the endoplasmic reticulum.

The pleckstrin homology domain (PH domain) is now well known as a structural module for the binding of inositol compounds. In the present study, polyclonal antibodies against the peptide KVKSSSWRRERFYK, derived from the N-terminal of the PH domain of phospholipase C-delta1 (PLC-delta1), were raised in rabbits. These were then tested for their ability to inhibit the binding of inositol 1,4,5-trisphosphate [Ins(1,4,5)P3] to the binding proteins including the receptor molecule. The Fab fragment of the antibodies but not the whole molecule inhibited the binding of Ins(1,4,5)P3 not only to PLC-delta1 but also to the Ins(1,4,5)P3 receptor, indicating that the antibodies raised recognized the binding site for Ins(1,4, 5)P3 in the receptor. Rat basophilic leukemic cells were permeabilized with saponin and assayed for Ins(1,4,5)P3-mediated Ca2+ release. Pretreatment of permeabilized RBL cells with the Fab fragment of the antibodies diminished the release of Ca2+ caused by Ins(1,4,5)P3, and further absorption experiments using a variety of synthetic peptides suggested that the tripeptide KVK is the epitope of the antibodies. Structural information about KVK will help in screening for Ins(1,4,5)P3 antagonists.

Animals↗
PubMed1999

Response of DNA ploidy to chemotherapy in primary and metastatic lesions in human osteosarcomas.

Primary and pulmonary metastatic and pulmonary metastatic tumors (two synchronous and seven metachronous metastases) in nine patients with osteosarcomas were studied by DNA cytofluorometry. All patients were treated with both pre and postoperative chemotherapy. The results showed that all five diploid osteosarcomas and three of the four aneuploid tumors did not markedly change their ploidy pattern after preoperative chemotherapy, and had almost the same ploidy patterns as the pulmonary metastatic lesions. Those eight tumors showed poor histologic response and chemoresistance by the doxorubicin binding assay. Only one aneuploid osteosarcoma showing good histologic response and chemosensitivity changed its ploidy pattern to diploid, with the disappearance of aneuploid tumor cells and its synchronous pulmonary metastatic tumor also showed conversion to a diploid pattern with massive tumor necrosis. It is evident that those tumors showing no change in their ploidy pattern after chemotherapy were resistant to the chemotherapy. Therefore, we conclude that regardless of whether the pulmonary metastatic tumors were synchronous or metachronous, they showed the same change in their ploidy pattern as well as their chemosensitivity as the primary human osteosarcoma from which they were derived.

Adolescent↗
PubMed1999

Relationship between P-glycoprotein positivity, doxorubicin binding ability and histologic response to chemotherapy in osteosarcomas.

We previously reported that the doxorubicin binding ability detected by the doxorubicin (adriamycin) binding assay was closely correlated with the chemosensitivity of human osteosarcomas. In this study, we undertook to clarify the relationship between P-glycoprotein positivity (%PPG) and doxorubicin binding ability (%DB) in human osteosarcomas in order to determine which is a more sensitive index of histologic response to chemotherapy. Ten primary osteosarcomas were analyzed by the doxorubicin binding assay and by immunofluorescence to detect cellular P-glycoprotein positivity. Three good responders to chemotherapy containing doxorubicin showed a %DB greater than 90% (average: 96.43%), whereas the seven poor responders had values less than 80% (average: 35.31%). The difference between the two groups was statistically significant (P = 0.0167). However, the average %PPG of the three good responders was 6.73%, whereas the %PPG of the seven poor responders was 14.27%. There was no significant difference in %PPG between the two groups (P = 0.3051). No negative correlation between the %DB and the %PPG of all osteosarcomas (r = 0.536, P = 0.1104) was found, although there was a trend that those tumors with a high %PPG showed a low %DB. These results suggest that osteosarcomas showing a low %DB and %PPG with poor response to chemotherapy, may have multidrug resistance mechanisms other than P-glycoprotein. Therefore, we conclude that doxorubicin binding ability, which reflects all of the doxorubicin-resistant mechanisms, was more sensitive than P-glycoprotein positivity in predicting the chemosensitivity of human osteosarcoma.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1999

Prognostic significance of DNA ploidy pattern in osteosarcomas in association with chemotherapy.

In this study, we analysed the DNA ploidy of osteosarcomas at biopsy and attempted to clarify the relationship between DNA ploidy pattern and prognosis. Thirty patients with non-metastatic osteosarcoma of an extremity were studied. All underwent intensive chemotherapy with doxorubicin, cisplatin and methotrexate, in addition to wide tumor resection. DNA ploidy was detected by DNA cytofluorometry, using isolated and smeared cells of biopsied tumor tissue. Twelve tumors showed a diploid ploidy pattern and 18 showed a non-diploid pattern such as aneuploidy (15 tumors) and euploid-polyploidy (3 tumors). The event-free survival rate at 9 years was 63.5% in non-diploid osteosarcoma patients and 13.3% in diploid osteosarcoma patients. There was a statistically significant difference between the two groups (P = 0.0278). These results lead us to conclude that a non-diploid osteosarcoma may be more sensitive to chemotherapy than a diploid tumor.

Adolescent↗
PubMed1999

Cytofluorometric DNA ploidy analysis in giant cell tumor of bone: histologic and prognostic value.

DNA ploidy analysis by DNA cytofluorometry was performed on 41 tumors obtained from 37 patients with primary giant cell tumor of bone (GCT). Histologically, 26 of the tumors from primary or recurrent lesions were evaluated as grade I, and 13 tumors as grade II. Among the 33 primary GCT patients, 4 patients had local recurrence or pulmonary metastasis. The DNA ploidy pattern and the percentage of hyperdiploid cells showing a greater DNA content than diploid cells, were obtained from DNA cytofluorometry. All of the 33 primary tumors were diploid. Of 6 recurrent tumors, 4 were diploid and 2 were euploid-polyploid. One of the two pulmonary metastatic tumors was diploid, but another that demonstrated a malignant transformation to malignant fibrous histiocytoma was aneuploid. The percentage of hyperdiploid cells was significantly different between primary and recurrent tumors (P = 0.0188) and between grade I and grade II tumors (P = 0.0052), while there was no difference between primary tumors in the cases that recurred or metastasized and those that did not. Thus, these data indicate that cell proliferative activity is closely correlated with biological aggressiveness and histological grading, although DNA ploidy is not useful for predicting prognosis.

Adolescent↗
PubMed1999

Identification of myosin II as a binding protein to the PH domain of protein kinase B.

Myosin II was identified as a binding protein to the pleckstrin homology (PH) domain of protein kinase B (PKB) in CHO cell extract by using the glutathione S-transferase-fusion protein as a probe. When myosin II purified from rabbit skeletal muscle was employed, myosin II was shown to bind almost exclusively to the PH domain of PKB among the PH domain fusion proteins examined. The purified myosin II bound to the PH domain of PKB with a Kd value of 1.1 x 10(-7) M. Studies with a series of truncated molecules indicated that the whole structure of the PH domain is required for the binding of myosin II, and the binding to the PH domain was inhibited by phosphatidylinositol 4,5-bisphosphate. These results suggest that myosin II is a specific binding protein to the PH domain of particular proteins including PKB.

Animals↗
PubMed1999

The human and rat forms of multiple inositol polyphosphate phosphatase: functional homology with a histidine acid phosphatase up-regulated during endochondral ossification.

We have derived the full-length sequences of the human and rat forms of the multiple inositol polyphosphate phosphatase (MIPP); their structural and functional comparison with a chick histidine acid phosphatase (HiPER1) has revealed new information: (1) MIPP is approximately 50% identical to HiPER1, but the ER-targeting domains are divergent; (2) MIPP appears to share the catalytic requirement of histidine acid phosphatases, namely, a C-terminal His residue remote from the RHGxRxP catalytic motif; (3) rat MIPP mRNA is up-regulated during chondrocyte hypertrophy. The latter observation provides a context for proposing that MIPP may aid bone mineralization and salvage the inositol moiety prior to apoptosis.

Acid Phosphatase↗
PubMed1999

Clinical outcome of total scapulectomy in 10 patients with primary malignant bone and soft-tissue tumors.

BACKGROUND AND OBJECTIVES: Limb reconstruction after total scapulectomy for malignant bone and soft-tissue tumors around the scapula is difficult. This study was undertaken to clarify the clinical results of total scapulectomy in patients with malignant bone and soft-tissue tumors around the shoulder girdle in our institute between 1984 and 1998. METHODS: Ten patients undergoing total scapulectomy had an age range of 12-82 years (average = 56 years). There were 5 cases of bone tumor and 5 cases of soft-tissue tumor. The follow-up period ranged from 8 months to 13 years 5 months. RESULTS: Seven patients are currently alive; the remaining 3 patients died of other diseases. One case of local recurrence was detected. The 2-year survival rate of all cases was 78.8%, and the 5-year survival rate was 52.5%. The average function evaluated by Enneking's criteria was 64.6%. Although the range of motion in the shoulder joint was seriously limited in all patients, the elbow and hand functions were almost normal. Recently, we have used a bone-anchoring system to suture between the clavicle and muscles, including the biceps, triceps, and deltoid muscles. CONCLUSIONS: Patients who undergo total scapulectomy may achieve much better upper limb function than those who undergo forequarter amputation (interscapulothoracic amputation).

Adolescent↗
PubMed1999

Extended blood half-life of monomethoxypolyethylene glycol-conjugated hen lysozyme is a key parameter controlling immunological tolerogenicity.

The blood half-life of a protein is prolonged by conjugating a protein with a linear amphiphilic polymer, monomethoxypolyethylene glycol (mPEG). The conjugation gives a protein immunotolerogenicity; hence, it is likely that the long half-life is crucial for the tolerogenicity. We prepared a tolerogenic mPEG conjugate of hen egg lysozyme (mPEG1.5-HEL), which is conjugated 1.5-fold the molecular weight of mPEG against that of HEL, and evaluated the relationship between in vivo stability and the tolerogenicity. mPEG1.5-HEL retained immunogenicity to prime HEL-specific T cell and antibody responses and had a long blood half-life, more than 27 times that of native HEL. The tolerant state was maintained as long as mPEG1.5-HEL was detected in sera. With a decrease in the blood mPEG1.5-HEL level, the tolerant state returned gradually to the responsive state; however, reinjection of mPEG1.5-HEL again restored the tolerance. Thus, the extended blood half-life of HEL by mPEG conjugation is probably vital for establishing and maintaining the tolerant states.

Animals↗
PubMed1999

Medial plantar nerve conduction velocities among patients with vibration syndrome due to chain-saw work.

OBJECTIVE: The present study examined the effect of the vibration syndrome (VS) on the peripheral nervous system in the lower extremities. METHODS: Thirty-eight patients with VS due to previous exposure to vibration from chain-saw work and 55 age-matched controls were examined for sensory nerve conduction velocities in the medial plantar nerve (SCV-P). The patient group was divided into two subgroups, one with (n=19) and the other without vibration-induced white finger (VWF; n=19). RESULTS: Analysis of variance of SCV-P for the three groups showed significant difference (F(2,89)=10.65, P < 0.0001). A significant difference was found between the controls and the VWF(+) group (P < 0.0001) but not between the controls and the VWF(-) group (P=0.0508) by multiple comparison using Scheffe's method. CONCLUSION: These findings suggest that VS affects the peripheral nervous system function in the lower extremities via mediation of circulatory disturbance manifested as VWF.

Action Potentials↗
PubMed1999

Membrane association of a new inositol 1,4,5-trisphosphate binding protein, p130 is not dependent on the pleckstrin homology domain.

The 130-kDa protein was isolated as a novel inositol 1,4,5-trisphosphate (Ins(1,4,5)P3) binding protein from rat brain and was molecularly cloned to be found similar to phospholipase C-delta 1 (Kanematsu, T., Takeya, H., Watanabe, Y., Ozaki, S., Yoshida, M., Koga, T., Iwanaga, S. and Hirata, M., 1992. Putative inositol 1,4,5-trisphosphate binding proteins in rat brain cytosol, J. Biol. Chem. 267, 6518-6525; Kanematsu, T., Misumi, Y., Watanabe, Y., Ozaki, S., Koga, T., Iwanaga, S., Ikehara, Y. and Hirata, M., 1996. A new inositol 1,4,5-trisphosphate binding protein similar to phospholipase C-delta 1, Biochem. J. 313, 319-325). The 130-kDa protein and its deleted protein expressed in COS-1 cells were seen in both the membrane and the cytosol fractions. Truncation of 232 residues from the N-terminus, the protein molecule lacking the pleckstrin homology (PH) domain was also localized in the membrane fraction as much as seen with a full-length protein and other deleted proteins, thereby indicating that the PH domain is not primarily involved in the membrane localization. The addition of Mg2+ to homogenates of COS-1 cells caused the translocation of expressed proteins from the cytosol to the membrane fraction, yet further addition of AlF4- which induced the activation of GTP binding proteins did not cause a further translocation. The protein translocated to the membrane by the addition of Mg2+ was hardly extracted with Triton X-100. The inclusion of Ins(1,4,5)P3 or phosphatidylinositol 4,5-bisphosphate in cell homogenates caused the very small reduction in the amounts of membrane-associated proteins expressed by some constructs. These results indicate that (i) the PH domain is not primarily involved in the membrane localization of the 130-kDa protein, (ii) the activation of GTP binding protein does not appear to cause the translocation of the 130-kDa protein, and (iii) intrinsic phosphatidylinositol 4,5-bisphosphate present in the membrane appears to be involved in the membrane association of the 130-kDa protein to a very small extent, probably through the binding site in the PH domain.

Adenosine Diphosphate Ribose↗
PubMed1999

Use of phosphorofluoridate analogues of D-myo-inositol 1,4,5-trisphosphate to assess the involvement of ionic interactions in its recognition by the receptor and metabolising enzymes.

D-myo-inositol 1,4,5-trisphosphate [Ins(1,4,5)P3] analogues fluoridated at 4- or 5-phosphate or both were analysed to assess the involvement of ionic interactions between the phosphates of Ins(1,4,5)P3 and the proteins that recognize it, such as metabolic enzymes and the InsP3 receptor. These analogues were effective in inhibiting type I Ins(1,4,5)P3 5-phosphatase activity with much the same potency as Ins(1,4,5)P3, although the enzyme showed a lower Km value as pH values increased. In contrast, the analogues were less potent ligands than Ins(1,4,5)P3 in both the assay of [3H]Ins(1,4,5)P3 binding to the receptors and the phosphorylation of [3H]Ins(1,4,5)P3 catalysed by Ins(1,4,5)P3 3-kinase. These results suggest that ionic interactions with the dianionic 4- and 5-phosphates of Ins(1,4,5)P3 are involved in recognition by the receptor and the kinase, but not by the phosphatase.

Binding, Competitive↗