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Biomedical subjects

T Wakabayashi

Publications and source records attributed to T Wakabayashi.

At least 73 records · Page 4Linked to original sources
PubMed2000

Production of a single-chain variable fragment antibody recognizing type III mutant epidermal growth factor receptor.

The type III deletion mutant of the epidermal growth factor receptor (EGFR) is a potential target in diagnostic and therapeutic approaches for those glioblastomas characterized by its expression. We previously raised a mouse monoclonal antibody, 3C10 (IgG2b) specifically recognizing this mutant EGFR. In this study, a single-chain variable fragment (scFv) antibody was produced. Partial determination of its N-terminal amino acid sequence and preparation of adequate primers for variable heavy chain (V(H)) and variable light chain (V(L)) genes were performed to allow cloning by means of reverse transcriptase-polymerase chain reaction. The genes cloned were assembled with a linker, (Gly4Ser)3, and ligated into a bacterial expression vector to express the scFv as cytoplasmic inclusion bodies. After appropriate refolding, the antibody activity of the V(H)-V(L) scFv was examined in an enzyme-linked immunosorbent assay. 3C10 scFv showed a selective reactivity with the mutant peptide, similarly to the parental 3C10 antibody. A mouse transfectant expressing the type III mutant EGFR and a glioblastoma with type III deletion-mutant EGFR were positively stained by immunofluorescence. By Biacore analysis, the affinity (K(A)) of the parental 3C10 for the mutant peptide was 9.7 x 10(7) M(-1), while that of 3C10 scFv was 2.45 - 2.48 x 10(7) M(-1), being approximately 4-fold weaker. The results together suggested that the scFv antibody retained the appropriate structure to recognize a conformational epitope of the mutant receptor, similarly to the parental antibody.

3T3 Cells↗
PubMed2000

Improving public awareness of organ transplantation in Japan.

The goals of the Japanese chapter of the Transplant Recipients International Organization are to act as a mediator in the provision of mutual aid to transplant recipients, candidates, and their families and to provide education in both the public and professional arenas to promote organ transplantation in Japan. Since 1991, members of the Japanese chapter of the Transplant Recipients International Organization have acted as volunteer facilitators for more than 150 transplant candidates and recipients. The organization's activities have included fund-raising to cover the costs of transplantation, preparing an original donor card for distribution, and petitioning members of the Diet (the Japanese legislative body) 14 times, including speaking at public hearings twice and submitting a petition requesting early passage of the organ transplant bill. The Japanese Diet finally passed an organ transplant bill in June 1997. The activities of the Japanese chapter of the Transplant Recipients International Organization have contributed to the promotion of organ transplantation in Japan and to understanding and support for transplantation within the Japanese community as a whole.

Attitude to Health↗
PubMed2000

Japanese organ transplant law: a historical perspective.

Although organ transplantation following brain death has progressed in the West, it has lagged far behind in Japan, following the first such case in 1968. As effective immunosuppressants made transplantation a better option, Japanese patients increasingly sought treatment overseas. Japanese physician groups studied issues related to transplantation but did not succeed in making brain-dead donor transplants available to patients, and the matter was referred to the government. However, transplant medicine was still marked by controversy, and as political pressure was applied the controversy deepened, splintering public opinion. At the same time, transplant groups continued working to establish structures to allow transplantation to proceed. Public awareness and knowledge of brain death grew, and acceptance widened. Eventually, legislation was passed in June 1997 that allowed organ donations from some brain-dead donors. The law is restrictive, and such organ transplants in Japan are still limited.

Attitude to Health↗
PubMed2000

Sarcomatous transformation of a prolactinoma associated with development of a fatal internal carotid artery pseudoaneurysm--case report.

A 21-year-old female with a 5-year history of prolactinoma was referred to our hospital because of cerebrospinal fluid leakage and meningitis immediately following a third transsphenoidal operation for a rapidly growing tumor. Histological examination of the tumor removed at the second transsphenoidal operation found atypical epithelial and sarcomatous components. Administration of bromocriptine, radiotherapy, and chemotherapy were initiated. Emergent craniotomy was required for rapid regrowth of the tumor. Histological examination found predominant sarcomatous components. Tumor growth was difficult to control. The patient died of subarachnoid hemorrhage due to rupture of a pseudoaneurysm involving the C1 portion of the right internal carotid artery. Pituitary adenoma rarely shows malignant transformation. In this case, prolactinoma underwent malignant change to sarcoma. Development of the pseudoaneurysm may have resulted from surgical manipulation, radiotherapy, or tumor invasion.

Adult↗
PubMed2000

Luminacins: a family of capillary tube formation inhibitors from Streptomyces sp. I. Taxomony, fermentation, isolation, physico-chemical properties and structure elucidation.

A new family of capillary tube formation inhibitors, designated luminacins, has been discovered in the fermentation broth of a soil bacterium. The strain was identified as Streptomyces sp. Mer-VD1207 from taxonomic studies. By means of a series of chromatographic procedures, fourteen structurally related components, luminacins A1, A2, B1, B2, C1, C2, D, E1, E2, E3, F, G1, G2, and H, were isolated, and their structures were elucidated on the basis of spectroscopic analyses.

Angiogenesis Inhibitors↗
PubMed2000

Luminacins: a family of capillary tube formation inhibitors from Streptomyces sp. II. Biological activities.

Twelve of the fourteen isolated components of the luminacin family were assayed for activity to inhibit capillary tube formation in vitro. Seven of them showed potent activity with IC50 values of less than 0.1 microg/ml in a rat aorta matrix culture model. Luminacin D, the strongest inhibitor, inhibited both endothelial cell proliferation and capillary tube formation. Morphological observation suggested that luminacin D inhibited the rearrangement of endothelial cells in the initial stage of tube formation. Luminacins and their derivatives are good candidates for application as angiogenesis inhibitors with a novel mechanism of action.

Angiogenesis Inhibitors↗
PubMed2000

[Studies of gastric acid secretion and gastric mucosal change after Helicobacter pylori eradication--the short and long-term observation].

We studied the gastric acid secretion and the histological findings of gastric mucosa in 48 patients with peptic ulcer before and after successful Helicobacter pylori (H. pylori) eradication. The pH of fasting gastric juice significantly decreased from 2.46 to 1.65 four weeks after eradication (p < 0.001). The factors such as disease, atrophic border, inflammatory cell involvement and histological atrophy were analyzed by logistic regression. Only the atrophic border was a significant and independent factor to convert the acidity after eradication, and the pH of gastric juice prominently decreased in the patients with widespread atrophic mucosal area. In the 24-hour intragastric pH monitoring, the pH3 holding time overnight remarkably decreased from 54.1% to 22.3% after eradication (p < 0.005). In the long follow-up of 17 patients (the mean follow-up period was 15.2 months), the pH of gastric juice kept a lower titer of 1.66. The mean atrophic scores in the upper body reduced from 1.5 to 0.9 (p < 0.05). In addition to the functional recovery of parietal cells after eradication, it was considered that the histological improvement of atrophy in oxyntic mucosa was associated with the recovery of pH.

Adult↗
PubMed2000

[Evaluation of chemotherapy for gastric cancer by endoscopic ultrasonography].

We judged the efficacy of chemotherapy using endoscopic ultrasonography (EUS) in 26 cases of gastric cancer. Treatment efficacy was evaluated according to the General Rules for the Gastric Cancer Study, based on the reduction of the largest cross-sectional area of the tumor. A reduction of over 50% was rated as U-PR and a reduction of -25-50% was rated as U-NC. Our findings showed U-PR in 11 cases and U-NC in 15 cases. Three cases initially considered to be NC according to the rules were judged to be U-PR based on EUS findings. Marked therapeutic efficacy in these 3 cases was demonstrated clinically and this was confirmed by EUS findings. Generalized Wilcoxon test showed a significant difference in the cumulative survival rate between U-PR and U-NC cases (p < 0.05). EUS provides an objective means of evaluating the efficacy of chemotherapy in gastric cancer patients, including those with lesions that cannot be evaluated by the General Rules for the Gastric Cancer Study.

Adult↗
PubMed2000

Structural changes of mitochondria during free radical-induced apoptosis.

The initial proposal for apoptosis stressed nuclear change (condensation of chromatin) and the intactness of intracellular organelles, including mitochondria, based on light and electron microscopic observations. However, data have accumulated to demonstrate that the opening of megachannels of mitochondrial membranes, resulting in the swelling of the organelles, notably by Ca2+ and free radicals, is the crucial step in the apoptotic processes of the cell. Application of fluorescent dyes to mitochondria, combined with flow cytometry, has made it possible to detect subtle changes in the structure and function of the organelles related to apoptosis. The present article overviews structural aspects of mitochondria related to apoptosis, including the free radical-induced formation of megamitochondria.

Animals↗
PubMed2000

[Correlation between DNA topoisomerase II alpha expression and sensitivity to etoposide in human glioma cell lines].

The expression of topoisomerase II alpha (Topo II alpha) was investigated in six human glioma cell lines as a function of doubling time, cell cycle distribution, and sensitivity to an antineoplastic agent, etoposide (ETP). The Topo II alpha level was determined by immunohistochemical and flow cytometric studies using monoclonal antibody, 8D2, specific for Topo II alpha. In all cell lines, the Topo II alpha level correlated directly with sensitivity to ETP, but not with doubling time or percentage of cells in any phase of the cell cycle. These findings suggest that the cytocidal activity of ETP may be mediated by qualitative and quantitative changes in Topo II alpha in human glioma cells. We speculate that better knowledge of Topo II alpha expression in surgical specimens may lead to more individualized use of ETP in patients with malignant glioma.

Antigens, Neoplasm↗
PubMed2000

[Effectiveness of DeVIC chemotherapy for recurrent primary central nervous system lymphoma].

Six cases with recurrent or refractory primary central nervous system lymphoma were treated with a new chemotherapeutic regimen "DeVIC (dexamethasone, VP16, ifosfamide, carboplatin)". Five recurrent cases had a remission period for an average of 18 months after initial treatment, but relapse occurred. One refractory case had no response after initial treatment. Then these 6 cases were treated with 1-3 courses of DeVIC chemotherapy at intervals of 4 weeks. Two cases achieved complete remission, and 3 cases attained partial remission (response rate was 83%). One case showed no response after 1 course of DeVIC chemotherapy. However, in all cases re-relapse occurred 1-5 months after remission, and only 1 case is still alive. DeVIC chemotherapy produced a high response rate for recurrent central nervous system lymphoma, but re-relapse occurred after only a few months. The establishment of maintenance chemotherapy is required.

Adolescent↗
PubMed2000

Clinical factors related to the aqueous levels of vascular endothelial growth factor and hepatocyte growth factor in proliferative diabetic retinopathy.

PURPOSE: To investigate clinical factors related to the aqueous humor levels of vascular endothelial growth factor (VEGF) and hepatocyte growth factor (HGF) in patients with proliferative diabetic retinopathy (PDR). METHODS: Undiluted aqueous humor was obtained during ocular surgery from 46 eyes of 46 patients and the levels of growth factors were measured using enzyme-linked immunosorbent assays. VEGF and HGF levels were compared with the number of photocoagulation. VEGF and HGF levels in patient groups stratified according to the existence of vitreous hemorrhage, retinal detachment, and fibrovascular membrane were analyzed. And clinical parameters associated with the growth factors were determined by multiple regression analysis. RESULTS: The levels of VEGF decreased significantly as the extent of photocoagulation increased and showed significant positive correlation with the existence of vitreous hemorrhage. There was no significant correlation between VEGF levels and the existence of fibrovascular membrane and traction retinal detachment. The levels of HGF also decreased significantly as the extent of photocoagulation increased, but increased significantly when fibrovascular membrane existed. There were no significant correlations between HGF levels and the existence of vitreous hemorrhage and traction retinal detachment. Concerning systemic conditions, each growth factor has no significant correlation with duration and type of diabetes mellitus, the treatment regimen, the control of hemoglobin A1C, and the existence of hypertension or renal dysfunction. CONCLUSION: The therapeutic effect of panretinal photocoagulation on PDR might be partly exerted by reduction of the levels of VEGF and HGF in ocular fluid. Since the clinical parameters associated with VEGF were different from those associated with HGF, these growth factors might influence the progression of retinopathy in different ways.

Adult↗
PubMed1999

Effects of coenzyme Q10 on changes in the membrane potential and rate of generation of reactive oxygen species in hydrazine- and chloramphenicol-treated rat liver mitochondria.

Effects of CoQ10 and cycloheximide (CHX) on hydrazine- and chloramphenicol (CP)-induced morphological and some functional changes of mitochondria using cultured rat hepatocytes and effects on the process of recovery from CP intoxication using mouse liver were examined. Results obtained are summarized as follows: (1) The formation of megamitochondria induced in the hepatocytes cultured for 22 h in the presence of 2 mM hydrazine or CP (300 microgram/ml) was suppressed by pretreatment of hepatocytes with CoQ10 (1 microM) or CHX (0.5 microgram/ml). This was proved by electron microscopic analysis of mitochondria. (2) Treatment of hepatocytes with hydrazine for 48 h or longer caused decreases in the membrane potential of mitochondria, which were suppressed by CoQ10. (3) Treatment of hepatocytes with hydrazine for 22 h or longer caused remarkable increases in intracellular levels of reactive oxygen species in hepatocytes, which were suppressed by CoQ10. (4) The process of recovery from the CP-induced changes of mitochondria in mouse liver was accelerated by CoQ10 and CHX.

Animals↗
PubMed1999

Synergism between mild hyperthermia and interferon-beta gene expression.

In the present study, the synergistic effect of mild hyperthermia in combination with gene expression of interferon-beta (IFN-beta) was examined in vitro in the human glioma cell line U87MG. The cells transiently expressed the IFN-beta gene under the control of the mouse mammary tumor virus promoter and were then subjected to temperature elevation (41 degrees C for 1 h). In terms of the cell killing effect, the optimum scheme was obtained by transfection for 4 days before hyperthermia, i.e. rate in the time course of IFN-beta gene expression. The relative specific growth rate decreased to 32% compared with the control while it was only 40% under the hyperthermic conditions. These observations suggest that IFNbeta expression was able to enhance the sensitivity of transfected glioma cells to mild hyperthermia.

Animals↗
PubMed1999

Cycloheximide and 4-OH-TEMPO suppress chloramphenicol-induced apoptosis in RL-34 cells via the suppression of the formation of megamitochondria.

Toxic effects of chloramphenicol, an antibiotic inhibitor of mitochondrial protein synthesis, on rat liver derived RL-34 cell line were completely blocked by a combined treatment with substances endowed with direct or indirect antioxidant properties. A stable, nitroxide free radical scavenger, 4-hydroxy-2,2,6, 6-tetramethylpiperidine-1-oxyl, and a protein synthesis inhibitor, cycloheximide, suppressed in a similar manner the following manifestations of the chloramphenicol cytotoxicity: (1) Oxidative stress state as evidenced by FACS analysis of cells loaded with carboxy-dichlorodihydrofluorescein diacetate and Mito Tracker CMTH2MRos; (2) megamitochondria formation detected by staining of mitochondria with MitoTracker CMXRos under a laser confocal microscopy and electron microscopy; (3) apoptotic changes of the cell detected by the phase contrast microscopy, DNA laddering analysis and cell cycle analysis. Since increases of ROS generation in chloramphenicol-treated cells were the first sign of the chloramphenicol toxicity, we assume that oxidative stress state is a mediator of above described alternations of RL-34 cells including MG formation. Pretreatment of cells with cycloheximide or 4-hydroxy-2,2, 6,6-tetramethylpiperidine-1-oxyl, which is known to be localized into mitochondria, inhibited the megamitochondria formation and succeeding apoptotic changes of the cell. Protective effects of cycloheximide, which enhances the expression of Bcl-2 protein, may further confirm our hypothesis that the megamitochondria formation is a cellular response to an increased ROS generation and raise a possibility that antiapoptotic action of the drug is exerted via the protection of the mitochondria functions.

Animals↗
PubMed1999

Detailed localization of photoreceptor guanylate cyclase activating protein-1 and -2 in mammalian retinas using light and electron microscopy.

Guanylate cyclase activating proteins, GCAP-1 and GCAP-2, have a pivotal role in the activation of guanylate cyclase in phototransduction. Previous studies on the localization of GCAP-1 and GCAP-2 are contradictory. In this study, we tried to avoid possible artifacts accompanied by immunocytochemistry. Immunolabeling of a GCAP was carried out using antibodies pre-adsorbed with a different type of GCAP. In addition, immunolabeling was performed using three different animal species under different fixation and embedding. Electron microscopic immunocytochemistry was also performed to reveal subcellular localization of GCAPs as well as confirming data obtained by light microscopy. All data indicate that anti-GCAP-1 antibody binding sites were found predominantly in cone outer segments, in particular, in disk membrane regions. Sparse labeling was observed in rod outer segments, but the labeling was much lower than that seen in cone outer segments. Less labeling is also found in synaptic regions and inner segments of cones. No labeling was detected in connecting cilia and its cytoplasmic extensions. Such labeling patterns were similar among human, monkey and bovine retinas. The localization of GCAP-1 is consistent with the pattern of a recently reported human cone-specific degeneration. Anti-GCAP-2 antibody binding sites were detected in both inner and outer segments of rods and cones of all three animals although the labeling density was slightly different among species. Cryo-immuno-labeling of GCAP-2 in bovine retinas revealed that labeling sites were more concentrated in rods than those of cones, and that synaptic regions were also labeled. The different localization of GCAPs suggest that roles of GCAP-1 and GCAP-2 may be different.

Animals↗