Biomedical subjects

Shin-Ichi Miyatake

Publications and source records attributed to Shin-Ichi Miyatake.

25 records · Page 2Linked to original sources

PubMed2003

Over expression of bone morphogenetic protein-3b (BMP-3b) using an adenoviral vector promote the osteoblastic differentiation in C2C12 cells and augment the bone formation induced by bone morphogenetic protein-2 (BMP-2) in rats.

BMP-3b is a novel BMP-3-related protein and its biological functions are unknown. In order to investigate the biological actions of BMP-3b, we constructed a BMP-3b-expressing recombinant adenoviral vector (AxCAKBMP-3b). We show that over expression of BMP-3b stimulated the induction of differentiation and the osteoinduction activity of a human BMP-2-expressing recombinant adenoviral vector (AxCAOBMP-2). C2C12 cells were infected in vitro with AxCAKBMP-3b, AxCAOBMP-2 and a control vector containing no foreign genes (AxCAwt). Cells infected with AxCAOBMP-2 and AxCAKBMP-3b produced more alkaline phosphatase and secreted more osteocalcin into the culture medium than cells infected with AxCAOBMP-2 and AxCAwt. When AxCAOBMP-2, AxCAKBMP-3b, and AxCAwt were injected into the calf muscles of nude rats (F 344/N Jcl-rnu), the osteoinduction seen with AxCAOBMP-2 and AxCAKBMP-3b was greater than that seen with AxCAOBMP-2 and AxCAwt.

Adenoviridae↗

PubMed2003

The early successful treatment of glioblastoma patients with modified boron neutron capture therapy. Report of two cases.

We very effectively treated two patients with recurrent glioblastoma with modified boron neutron capture therapy (BNCT). In this paper, we describe the effectiveness of this treatment, and discuss the ways in which we modified the treatment. A 61-year-old man had a first operation for a right temporal glioblastoma, followed by full-dose chemo-radiotherapy. One year after the operation a partial removal was performed for the recurrent tumor at the same site. Fifty days after the second surgery, the patient received BNCT. We used an epithermal neutron beam as the neutron source, and used both sodium borocapate and boronophenylalanine as boron compounds with the craniotomy. Forty-eight hours after the BNCT, the follow-up MRI was applied to estimate the early effect of this treatment, which showed a 70% reduction in the contrast enhanced lesion, compared with the pretreatment MRI. In addition, the lesion/normal brain ratio of thallium-SPECT had improved markedly. No serial sequelae appeared after this treatment, and the patient remains healthy 6 months after the treatment. A 29-year-old young lady had a right temporal brain tumor, which was partially resected and followed by stereotactic radiosurgery for the residual mass. Seven months after the radiosurgery, a second operation was performed, which revealed the glioblastoma as diagnosis. We applied BNCT uneventfully for this patient with epithermal beam and two kinds of boron compounds as described above. For the treatment of the patient irradiation was applied without craniotomy with marked reduction of tumor volume immediately after the treatment.

Adult↗

PubMed2003

Efficient proliferation and adipose differentiation of human adipose tissue-derived vascular stromal cells transfected with basic fibroblast growth factor gene.

Human vascular stromal (VS) cells obtained from mature adipose tissue were transfected with an adenovirus vector carrying the basic fibroblast growth factor (bFGF) gene. bFGF protein was observed in VS cell nuclei 24 h after transfection and in the cytoplasm and extracellular space 72 h after transfection. Naive VS cells were almost static in vitro and proliferated in a dose-dependent manner on stimulation with recombinant bFGF (rbFGF). However, bFGF-transfected VS cells proliferated spontaneously to the same extent as naive VS cells when stimulated with rbFGF at 100 ng/ml. The former cells started to proliferate on day 3 after transfection and the proliferation pattern was similar to that of the latter cells, although only a slight amount of bFGF protein was detected in the culture medium when the bFGF-transfected cells started to proliferate. The proliferation of bFGF-transfected VS cells was completely inhibited by bFGF neutralizing antibody, which also completely inhibited the proliferation of naive VS cells stimulated with rbFGF. Under conditions favoring differentiation to adipocytes, bFGF-transfected VS cells stopped proliferating and started to accumulate lipid in the cytoplasm. bFGF-transfected VS cells, which spontaneously and efficiently proliferate while preserving their ability to differentiate into adipocytes, may be an adequate cell source for human adipose tissue regeneration.

Adipose Tissue↗

PubMed2003

Transcriptional expression of survivin and its splice variants in brain tumors in humans.

OBJECT: Survivin, one of the apoptosis inhibitor proteins, has been detected in most cancers in humans. In addition, two splice variants (survivin-2B and survivin-deltaEx3) have been identified. The authors investigated the transcription levels of survivin messenger (m)RNA and its splice variants in nine tumor cell lines, including gliomas, and in 25 brain tumor samples, by performing quantitative reverse transcription-polymerase chain reaction. The correlation between transcript expression levels and pathological findings were also analyzed. METHODS: Transcription levels were measured using primer pairs specific for survivin and either of its splice variants and were normalized to the glyceraldehyde 6-phosphate dehydrogenase. Among the tumor cell lines tested, glioblastoma cell lines showed the highest levels of survivin expression. Among brain tumor samples studied, survivin was preferentially expressed in malignant brain tumors and gliomas. The relative expression level of survivin-deltaEx3/survivin was significantly higher in malignant than in benign brain tumor samples. Expression patterns were dominant for survivin-deltaEx3 in malignant brain tumors and dominant for survivin-2B in benign ones. A significant linear correlation between survivin mRNA expression and MIB-1 labeling index was demonstrated in all brain tumor samples. CONCLUSIONS: The authors' results indicate that quantifying the levels of survivin and its splice variants is useful for the prediction of the cell biological malignancy of gliomas, independent of their pathological features.

Antibodies, Antinuclear↗

PubMed2002

Overexpression of basic fibroblast growth factor and Bcl-xL with adenoviral vectors protects primarily cultured neurons against glutamate insult.

OBJECTIVE: Excitatory amino acid (EAA) toxicity seems to be an important mechanism of neuronal cell death after cerebral infarction. We examined the inhibitory effects of neuronal cell death caused by EAA in vitro by means of adenoviral gene transfer of neurotrophic basic fibroblast growth factor (bFGF) and antiapoptotic Bcl-xL. METHODS: Recombinant adenoviral vectors expressing human bFGF gene with secretory signals of interleukin-2 and human Bcl-xL gene were constructed. Primarily cultured rat neuronal cells were treated with glutamate to cause EAA, and the neuroprotective effects of gene transfer by these adenoviral vectors were investigated at several time points of infection. RESULTS: Each adenoviral infection to primarily cultured neuronal cells exhibited neuroprotective effects against EAA caused by glutamate. Both gene transfer of bFGF with secretory signal and Bcl-xL transfer to neuronal cells exhibited the synergistic neuroprotective effects against EAA. These effects were most prominent with gene transfer 4 hours before glutamate insult; gene transfer performed simultaneously with and up to 4 hours after the insult exhibited definite neuroprotective effects. CONCLUSION: These experiments revealed marked neuroprotective effects of adenoviral gene transfer of bFGF and Bcl-xL into neuronal cells in vitro. The findings may lead to new approaches for treating occlusive cerebrovascular disease.

Adenoviridae↗

PubMed2002

Gene therapy using tissue-specific replication competent HSV.

Tissue- or cell-specific targeting of vectors is critical to the success of gene therapy. I describe a novel approach to viral-mediated gene therapy, where viral replication and associated cytotoxicity are limited to a specific cell-type by the regulated expression of an essential immediate-early viral gene product. This is illustrated with two herpes simplex virus type 1 vectors (G92A and d12.CALP) whose growth are restricted to albumin- or calponin-expressing cells, respectively. G92A was constructed by inserting an albumin enhancer/promoter--ICP4 transgene into the thymidine kinase gene of mutant herpes simplex virus type 1 d120, deleted for both copies of the ICP4 gene. This vector also contains the Escherichia coli lacZ gene under control of the thymidine kinase promoter, a viral early promoter, to permit easy detection of infected cells containing replicating vector. In the adult, albumin is expressed uniquely in the liver and in hepatocellular carcinoma and is transcriptionally regulated. G92A efficiently replicated in vitro in two human hepatoma cell lines expressing albumin, but not in three human non-hepatoma, albumin-non-expressing tumor cell lines, while all cell lines were equally susceptible to a tissue non-specific HSV recombinant, hrR3. In vivo, G92A replicated well in subcutaneous xenografts of human hepatoma cells (Hep3B) in athymic mice, but not in non-hepatoma subcutaneous tumors (PC3 and HeLa), whereas, hrR3 replicated well in both tumor types. Intratumoral inoculation of G92A inhibited the growth of established subcutaneous hepatoma tumors in nude mice, but not prostate tumors. D12CALP also revealed the cell-specific replication to leiomyosarcoma in which calponin expression was augmented. Using hrR3, we demonstrated inhibition of re-stenosis of rat carotid arteries caused by balloon injury. The antiproliferative effects of this virus was marked in the proliferating smooth muscle cells, however, there still remained the fear for the injury of the endothelial cells. Confining a productive, cytotoxic viral infection to a specific cell-type should be useful for tumor therapy and the ablation of specific cell-types for the generation of animal models of disease. Further experiments using d12CALP will be focused on the arteriosclerosis due to balloon angioplasty or organ transplantation.

Angioplasty, Balloon↗

PubMedDate not supplied

Enhanced detection of malignant glioma xenograft by fluorescein-human serum albumin conjugate.

OBJECT: During the surgical resection of malignant gliomas, it is important to make clear the border zone between the tumor tissue and normal brain tissue. For this purpose, we conjugated fluorescein and human serum albumin (FLS-HSA) and compared its effectiveness against that of fluorescein-sodium (FLS-Na) alone in detecting human glioma xenografts in SCID mice through a fluorescence microscope. METHODS: We made FLS-HSA conjugate using carbodiimide as a linking reagent. SCID mice, with U251MG cells transplanted subcutaneously, were prepared as tumor models. The animals were sacrificed 15, 30, 60, 180, 360, or 720 min after the intravenous administration of either the FLS-HSA conjugate or FLS-Na alone (n = 3). Fluorescence images were taken with a digital camera, and the brightness of the tumor and that of the peripheral tissue in each image were quantified. In the group of tumor-bearing mice that received FLS-Na, the fluorescence of tumor tissue disappeared 60 min after the reagent was administered, and there was no significant difference in brightness between the tumor and peripheral tissue at any time point. On the other hand, injection of FLS-HSA revealed relative tumor-selective brightness and sufficient contrast between the tumor and surrounding tissue 60 and 360 min after administration. CONCLUSION: FLS-HSA has the advantages of specificity and persistence of fluorescence over FLS-Na for the purpose of identifying glioma nodules in xenogenic subcutaneous tumor transplantation models.

Animals↗