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Biomedical subjects

S Okada

Publications and source records attributed to S Okada.

At least 163 records · Page 9Linked to original sources

Analysis of all exons of TSC1 and TSC2 genes for germline mutations in Japanese patients with tuberous sclerosis: report of 10 mutations.

Twenty-seven Japanese patients with the tuberous sclerosis complex (TSC), consisting of 23 sporadic and 4 familial cases, were tested for mutations in the TSC1 and TSC2 genes, using single-strand conformational polymorphism analysis and direct sequencing. Four possible pathogenic mutations were found in the TSC1 gene, including three frame shifts and a nonsense mutation in a familial case. All mutations were expected to result in a truncated hamartin gene product. The TSC2 gene analysis identified six possible pathogenic mutations only in the sporadic cases, including two frame shifts, one in-frame deletion, and three missense mutations. Two of the TSC2 mutations were expected to result in a truncated tuberin gene product. These results of the Japanese TSC patients were compatible with the reports from Europe and the United States, i.e., (1) TSC1 mutations are rarer in sporadic cases than in familial cases, (2) substantial numbers of sporadic cases arise from mutations in the TSC2 gene, and (3) mutations of the TSC1 gene may cause premature truncation of hamartin.

Adolescent↗

Molecular characterization of squalene synthase from the green microalga Botryococcus braunii, race B.

The green microalga Botryococcus braunii produces large amounts of liquid hydrocarbons and is classified into three races, depending on the type of the hydrocarbon produced. The B race produces two types of triterpenoid hydrocarbons, squalene and botryococcene, both of which are putative condensation products of farnesyl diphosphate. In an attempt to better understand the regulation involved in the production of squalene and botryococcene, we have isolated and characterized a squalene synthase (SS) gene from the B race of B. braunii. A 366-bp cDNA fragment was initially obtained from the B race utilizing a reverse transcription/polymerase chain reaction and degenerate primers based on conserved amino acid sequences found in all SS enzymes. Using this putative SS fragment as a probe, a 2632-bp cDNA clone was isolated from a cDNA library. This cDNA contained an open reading frame coding for a protein with 461 amino acids and a predicted molecular mass of 52.5 kDa. Comparison of the Botryococcus SS (BSS) with SS from different organisms showed 52% identity with Nicotiana tabacum, 51% with Arabidopsis thaliana, 48% with Zea mays, 40% with rat, 39% with yeast, and 26% with Zymomonas mobilis. Expression of full-length and carboxy-terminus truncated BSS cDNA in Escherichia coli resulted in significant levels of bacterial SS enzyme activity but no botryococcene synthase activity. RNA blot hybridization analysis of algal cultures during a culture cycle indicated that BSS gene expression is preferential during rapid growth. Given that the DNA blot analysis indicated only a single copy of the SS gene in the algal genome, these results suggest either that there exists coordinate expression of separate synthase genes for squalene and botryococcene biosynthesis or that there might be unique physiological conditions controlling the SS vs botryococcene synthase activity of a single peptide species.

Algal Proteins↗

Reactivation of human herpesvirus 6 by infection of human herpesvirus 7.

We have attempted to reactivate human herpesvirus 6 (HHV-6) by infection with HHV-7 using childhood exanthem subitum patients in vitro. Peripheral blood mononuclear cells (PBMCs) were collected from children who had a history of exanthem subitum(ES) by HHV-6 and were infected by human herpesvirus 7 (HHV-7) in vitro. The antigen positive rate to HHV-6 started to increase 7 days after the infection and reached a maximum by Day 15 using an immunofluorescence antibody test. The copy number of HHV-6 DNA also increased in the samples in 10 days after infection in vitro. No antigen or increase in DNA was detected in PBMCs, that were mock-infected or infected with supernatant of stock virus after ultracentrifugation, suggesting that an infection by HHV-7 is necessary to reactivate HHV-6. In the paired sera samples during the acute and the convalescent phases of ES, seven to ten bands, that were specific for HHV-6, were recognized in samples from the acute phase, and at least 5 dominant polypeptides were found more intensively after HHV-7 infection.

Blotting, Southern↗

Monochloramine enhances Fas (APO-1/CD95)-induced apoptosis in Jurkat T cells.

Monochloramine derivatives are physiological oxidants produced by activated neutrophils. We report the effects of chemically prepared monochloramine (NH2Cl) on Fas-induced apoptosis in Jurkat T cells. When the cells were pretreated with NH2Cl (20-70 microM), subsequent addition of apoptosis-inducing anti-Fas antibody resulted in a synergistic enhancement of apoptosis. Treatment of NH2Cl (50-70 microM) alone resulted in a slight but definite apoptosis. Caspase activities, as measured by DEVD and IETD cleavage activities, were also elevated synergistically by NH2Cl + anti-Fas antibody stimulation. Moreover, a broad caspase inhibitor, Z-VAD-fmk, almost completely inhibited the apoptosis induced by NH2Cl and/or anti-Fas antibody. Fas expression on the Jurkat cell surface was not affected by the NH2Cl treatment. After 3 h of NH2Cl treatment, when the apoptosis was beginning to increase, the cells showed cytochrome c release from mitochondria, proteolytic activation of caspase 9, and poly (ADP-ribose) polymerase cleavage, regardless of Fas stimulation. Z-VAD-fmk almost completely inhibited this poly (ADP-ribose) polymerase cleavage, but not cytochrome c release. By contrast, Fas stimulation alone resulted in neither cytochrome c release nor caspase 9 activation at 3 h, and the increase in the DEVD cleavage activity and apoptosis became evident at later time points. These results suggested that NH2Cl enhanced Fas-induced apoptosis through the cytochrome c release and caspase 9 activation at the early stage of apoptosis. Chloramines derived from acute inflammation may modify immune reactions, such as cell-mediated cytotoxicity and some autoimmune diseases, by the enhancement of Fas-induced apoptosis.

Apoptosis↗

Radiotherapy for hepatocellular carcinoma.

PURPOSE: To retrospectively evaluate the effectiveness of radiotherapy with or without transarterial embolization (TAE) and/or percutaneous ethanol injection (PEI) in patients with hepatocellular carcinoma who were ineligible for surgery. PATIENTS AND METHODS: From October 1984 to November 1997, 62 patients underwent radiotherapy receiving 50 to 70 Gy in 25 to 35 treatments with or without transarterial embolization and/or percutaneous ethanol injection and were followed for a median period of 8.6 months (1.5 to 92 months). RESULTS: Overall median survival rates were 9.5 months. Significant prognostic factors were the extent of pretreatment liver function impairment, radiation field size and the existence of tumor thrombosis. Six-month and 1-year local control rates were 67 and 54%, respectively. Seven of the 8 patients who suffered from hepatic failure had poor pretreatment liver functions. CONCLUSION: Radiotherapy with or without transarterial embolization and/or percutaneous ethanol injection appears effective in controlling hepatocellular carcinoma and prolonged survival. Individualized treatment strategies are presented depending on the tumor presentation and the degree of liver function impairment.

Aged↗

Combined modality therapy including surgery for stage III small-cell lung cancer on the basis of the sensitivity assay in vitro.

Fifty-nine consecutive patients with clinical stage (cStage) I-III resectable small-cell lung cancer (SCLC) underwent surgery with adjuvant chemotherapy based on an in vitro sensitivity assay between April 1982 and March 1992. In 42 (71%) of these patients, a stable passage of cancer cells from resected specimens was possible and the sensitivity of these cultured SCLC cells to anticancer drugs was examined by the MTT method. In the sensitivity assay, vincristine (VCR) showed the most intense specific efficacy for SCLC, followed by cyclophosphamide (CPM) and cisplatin (CDDP). The 5-year survival rates for pathological stage (pStage) I, pStage II, and pStage III were 55%, 33%, and 23%, respectively. The 5-year survival of the patients with pStage III operated on in the first 5 years was 7% (1/14). On the other hand, 6 of the 16 pStage III patients (38%) operated on during the second 5-year period, who were generally treated with pre- and postoperative adjuvant chemotherapy combined with three drugs, survived over 5 years. In conclusion, these results suggest that combined modality therapy including surgery is necessary, and adjuvant chemotherapy combined with VCR, CPM, and CDDP may be useful in the treatment of Stage III SCLC disease, for the purpose of achieving a long-term survival with both a good performance status and quality of life for the patients.

Adult↗

Cloning of the cyclodextrin glucanotransferase gene from alkalophilic Bacillus sp. A2-5a and analysis of the raw starch-binding domain.

The cyclodextrin glucanotransferase (CGTase) gene of alkalophilic Bacillus sp. A2-5a was cloned and expressed in Bacillus subtilis ANA-1 as a host. The DNA region included an open reading frame encoding a 704-amino-acid polypeptide with a typical raw starch-binding motif in its C-terminal region. The CGTase purified from Bacillus sp. A2-5a bound to raw starch as strongly as porcine pancreas alpha-amylase, as expected from the sequence motif. A chromosomal region (a DNA fragment of about 14.1 kbp) including the CGTase gene was also cloned and the nucleotide sequence was determined. Possible cyclodextrinase and putative cyclodextrin-binding protein genes were found in the flanking region of the CGTase gene, which implied that the novel starch-degradation pathway postulated for a gram-negative bacterium [Klebsiella oxytoca; Fiedler et al. (1996) J Mol Biol 256: 279-291] also exists in a gram-positive bacterium i.e. Bacillus.

Amino Acid Sequence↗

Measurement of serum hyaluronate as a predictor of human liver failure after major hepatectomy.

Serum hyaluronate can be used as an index of hepatic sinusoidal endothelial cell function. This study was designed to evaluate its application as a predictor of liver failure after major hepatectomy. Thirty-six patients who underwent right liver lobectomy after percutaneous transhepatic right branch portal vein embolization were divided into two groups based on their postoperative clinical course (groups 1 and 2, with and without postoperative liver failure, n = 6 and n = 30, respectively). We serially measured serum hyaluronate levels using a sandwich binding protein assay system before and after hepatectomy and determined relations with progression of the underlying chronic liver disorder, portal venous pressure, and liver growth of the left lobe after portal embolization. Serum hyaluronate levels were significantly elevated, in line with the degree of severity of the underlying chronic liver disorder, and correlated well with the portal venous pressure and the hypertrophic ratio of the left lobe subsequent to portal embolization. Serum hyaluronate levels in group 1 were significantly higher than those in group 2 before surgery and increased steeply during the early period after hepatectomy. These results suggest that the serum hyaluronate reflects the hepatic functional reserve, and serial measurement of this parameter after hepatectomy can serve as a simple indicator for early detection of posthepatectomy liver failure.

Biomarkers↗

Interferon treatment on glomerulonephritis associated with hepatitis C virus.

We report on a 10-year-old girl with glomerulonephritis associated with hepatitis C virus infection, who was treated with interferon-alpha. On the first renal biopsy at 8 years of age, mild mesangial hypercellularity in a segmental to semiglobal pattern was present in all glomeruli. After 6 months interferon-alpha therapy, proteinuria diminished completely. However, mesangial proliferation was advanced on the second biopsy at 10 years of age. We concluded that the interferon-alpha was effective in the treatment of proteinuria despite the lack of pathological improvement.

Antiviral Agents↗

Botryoid Wilms tumor: case report and review of literature.

A rare case of botryoid Wilms tumor is presented. The main clinical manifestations were persistent low-grade fever, malaise, and proteinuria associated with microhematuria. Ultrasonography revealed an echogenic mass in the right kidney, and a contrast-enhanced mass was found in the dilated collecting system by contrast-enhanced computed tomography. The surgically resected tumor was a polypoid, light-yellow, glistening mass that occupied a large part of the renal pelvis and originated from the pelvicaliceal wall. Part of the tumor extended to the proximal ureter, resulting in hydronephrosis in the involved kidney. No parenchymal lesion was observed. Microscopic examination revealed epithelial, stromal, and blastemal components, which indicated Wilms tumor. Infection had occurred in the hydronephrotic kidney, which presumably had caused the major presenting symptoms. The prognosis of our patient and previously reported cases of botryoid Wilms tumor was good compared with that of typical Wilms tumor, since the botryoid type can be detected at an early stage.

Hematuria↗

Long-term survival of a poor-risk octogenarian following wedge resection under VATS for small-cell lung cancer: report of a case.

We describe herein the case of an 81-year-old man who has remained disease-free for more than 3 years after undergoing a wedge resection of cStage I small-cell lung cancer (SCLC) under video-assisted thoracoscopic surgery (VATS), with no adjunct chemotherapy or radiotherapy. The patient had compromised pulmonary function and was a poor surgical risk. As he could not have endured a conventional lobectomy or intensive chemotherapy, a nonanatomical wedge resection of the area of lung involved by the primary tumor was carried out under VATS. Cancer cells from the resected tumor were cultured and the growth characteristics and sensitivity to 12 anticancer drugs were examined. The majority of primary cultured cells proliferated in a monolayer, like paving stones, resembling the growth pattern of non-small-cell carcinoma cells in vitro. The subcultured cells were resistant to most of the drugs, but showed weak sensitivity to cisplatin (CDDP), adriamycin (ADR), and vincristine (VCR). Therefore, the patient was discharged with no adjunct postoperative therapy and was followed up at an outpatient clinic. He has remained alive and disease-free for more than 3 years. Thus, we considered that performing wedge resection under VATS for a primary tumor could be appropriate treatment for selected patients with cStage I SCLC in a peripheral region, especially if they are elderly and a poor surgical risk.

Age Factors↗

The significance of surgery for bulky N2 small-cell lung cancer: a clinical and in vitro analysis of long-term survivors.

Until recently, stage III small-cell lung cancer (SCLC) has not been considered an indication for surgical treatment, however, stable subculture has become possible in about 80% of the patients with SCLC in our hospital. Therefore, we have been performing surgery for all patients with cStage I-III resectable tumors and giving combined-modality therapy based on the sensitivity assay in vitro since April 1982. In the present study, we reviewed 30 consecutive patients with cStage III and pStage III SCLC, including 27 with N2 disease who underwent surgery between 1982 and 1992, 7 of whom (23%) survived disease-free for over 5 years. We examined the cell characteristics in vitro and the actual treatment of five patients with bulky N2 lesions, four of whom were long-term survivors and one of whom died 11 months after surgery, as controls for comparison with the long-term survivors. Based on the results of the cell characteristics in vitro, SCLC was determined to be a heterogeneous tumor. Thus, surgical excision of the primary tumor with adjuvant therapy might be necessary to achieve long-term survival and maintain good performance status, and to improve quality of life in patients with bulky N2 SCLC by eliminating drug-resistant tumor cells within the primary tumor.

Aged↗

Long-term survival of a patient with stage IV pulmonary large cell carcinoma achieved by combined-modality therapy: report of a case.

We describe herein the case of a 59-year-old-man with stage IV pulmonary large cell carcinoma and a giant brain metastasis, in whom two sublines with different growth characteristics and drug sensitivities in vitro were established from the primary tumor. Disease-free survival for more than 5 years after surgery was achieved by combined-modality therapy together with surgery to remove the primary tumor, radiation to the brain metastasis, and chemotherapy to presumed hematogenous dissemination. Subline 1 proliferated in a monolayer of epithelial-like cells, while subline 2 showed a floating colony pattern of proliferation, resembling the typical growth characteristics of small cell lung cancer (SCLC) cells in vitro. Subline 2 was sensitive to a number of drugs, namely, vincristine (VCR), cyclophosphamide (CPM), adriamycin (ADR), and cisplatin (CDDP), whereas subline 1 was resistant to many drugs. The patient was treated with a combination of 44 Gy of whole-brain irradiation and a number of cycles of chemotherapy comprised of ADR, VCR, and CPM, followed by CDDP, VCR, and CPM, based on the results of sensitivity testing of the subline 2 cells. As a result, the patient has been disease-free for more than 5 years postoperatively. In conclusion, this case report serves to demonstrate that meticulous combined-modality treatment taking tumor heterogeneity in human cancers into account may be necessary to achieve breakthroughs in current cancer therapy for advanced lung cancer.

Antineoplastic Combined Chemotherapy Protocols↗

Mutation analysis of two Japanese patients with Fanconi-Bickel syndrome.

Fanconi-Bickel syndrome (FBS), or glycogen storage disease type XI, is a rare autosomal recessive disorder characterized by hepatorenal glycogen accumulation, Fanconi nephropathy, and impaired utilization of glucose and galactose. Recently, this disease was elucidated to link mutations in the glucose transporter 2 (GLUT2) gene. Only three mutations in three FBS families have been reported. Therefore, it is important to elucidate mutations in the GLUT2 gene in FBS by answering the question of whether the syndrome is a single gene disease. In this report, we describe two patients in two unrelated families clinically diagnosed with FBS. No mutation in the entire protein coding region of the GLUT2 gene was detected in patient 1, which suggested that no mutation existed in the GLUT2 gene, or that some mutations had affected the expression of the GLUT2 gene. In patient 2, a novel homozygous nonsense mutation (W420X, Trp at codon 420 to stop codon) was detected. These results support the correlation between GLUT2 gene mutation and FBS syndrome. However, many patients must be analyzed to determine whether other genes are involved in FBS.

Adult↗