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T Shirahama

Publications and source records attributed to T Shirahama.

At least 19 recordsLinked to original sources
PubMed1997

Sphingosine induces apoptosis in androgen-independent human prostatic carcinoma DU-145 cells by suppression of bcl-X(L) gene expression.

Our recent studies have suggested that sphingosine, an endogenous protein kinase C (PKC) inhibitor, may mediate apoptosis induced by a phorbol ester (PMA) in human promyelocytic leukemia HL-60 cells [Ohta et al. Cancer Res. 1995;55:691-697], and that the apoptotic induction by both PMA and sphingosine is accompanied by down-regulation of bcl-2, a gene which acts to prevent apoptotic cell death [Sakakura et al. FEBS Lett. 1996;397:177-180]. In this study, we examined the sphingosine-induced apoptosis of the androgen-independent human prostatic carcinoma cell line DU-145, which expresses bcl-X(L) and Bax but not bcl-2, and found that treatment of DU-145 cells with sphingosine suppressed bcl-X(L) in both mRNA and protein levels but did not change bax expression at all. In contrast, in apoptotic cells treated with a PKC inhibitor, staurosporine, no effect on bcl-X(L) or bax expression was observed. The initial metabolites of sphingosine in the cells, ceramide and sphingosine 1-phosphate, failed to induce apoptosis. These results indicate that, in DU-145 cells, sphingosine, but not its metabolites, induces apoptosis through down-regulation of bcl-X(L), independently of PKC inhibition. Our present results, together with previous observations, strongly suggest that apoptosis regulatory genes differ according to cell type and apoptosis induction through sphingosine is accompanied by inhibition of either bcl-2 or bcl-X(L) activity in these cells.

Androgens↗
PubMed1997

Metanestin, a glycoprotein with metastasis-associated expression in transitional cell carcinoma of the urinary bladder.

A 60-kDa glycoprotein named metanestin was identified by molecular cloning. The glycoprotein had a twice-repeated motif of Pro-Gly-Pro-Gly and carried metastasis-associated carbohydrate epitopes. The antibody to the protein portion of metanestin strongly reacted with lymph-node metastasis of transitional-cell carcinoma of the urinary bladder, but the reactivity to the primary tumor was generally weaker and the normal urothelium was scarcely reactive. Thus both metanestin and the carbohydrate on it showed metastasis-associated expression. In addition, we identified a 100-kDa glycoprotein with a 4-times-repeated motif of Pro-Ala-Pro-Ala. The antibody to the 100-kDa glycoprotein showed reactivity similar to that to metanestin.

Amino Acid Sequence↗
PubMed1997

Overexpression of bax enhances the radiation sensitivity in human breast cancer cells.

Bax-alpha, a splice variant of bax which promotes apoptosis, is expressed in many kinds of untransformed cell lines and breast tissue, whereas only weak or no expression could be detected in breast cancer cell lines and malignant breast tissue. Human breast cancer MCF-7 cells, which have a weak bax gene expression, were stably transfected with pCX2neo bax, encoding human bax and neomycin-resistant genes, and two unique clones (MCF-7/bax-1 and MCF-7/bax-2) were thus generated which expressed different levels of bax-alpha. Sensitivity to ionizing radiation (IR) was examined and each was more sensitive to IR than the parental MCF-7 cells. The degree of enhancement in radiosensitivity was dependent on the expression level of bax, and IR was found to induce intranucleosomal DNA fragmentation in stable transfectant but not in parent cells, thus suggesting that this sensitization is due to apoptosis. We suggest that exogenous bax-alpha expression might therefore be one of the factors determining cellular radiosensitivity in MCF-7 breast cancer cells and may potentially have a therapeutic application by enhancing radiation sensitivity in breast cancer cells.

Apoptosis↗
PubMed1996

Overexpression of bax sensitizes human breast cancer MCF-7 cells to radiation-induced apoptosis.

Resistance to apoptosis plays an important role in tumors that are refractory to chemotherapy and ionizing radiation (IR). bax, which forms a heterodimer with bcl-2 and accelerates apoptosis, is not, or only weakly, expressed in most human breast cancer cells, and weak bax expression is considered to be related to the resistance of breast cancer cells to apoptosis. bax expression vector was introduced to human breast cancer MCF-7 cells, which exhibit weak expression of bax, to demonstrate its role of modulating radiation-induced apoptosis. bax overexpression in MCF-7 cells by stable transfection does not affect viability by itself, but each stable transfectant was more sensitive to IR than the parental MCF-7 cells. The degree of enhancement in radiosensitivity was dependent on the expression level of bax. IR upregulated p53 and p21WAF1 about 5- to 10-fold and downregulated bcl-2 and bcl-XL by 80-90% at 6 hr in both parent and bax stably transfected MCF-7 cells to the same degree. FACS analysis and DNA electrophoresis revealed that this sensitization was due to apoptosis. We suggest that exogenous bax expression might be one of the factors determining cellular radiosensitivity in MCF-7 breast cancer cells and may have therapeutic applications for enhancing radiation sensitivity in breast cancer cells.

Apoptosis↗
PubMed1996

Sphingosine and its methylated derivative N,N-dimethylsphingosine (DMS) induce apoptosis in a variety of human cancer cell lines.

In the study of apoptosis initiated by various signals including ligands binding to cell membrane receptors such as Fas and TNFRI, the sphingomyelin pathway and its resulting metabolites, the sphingolipids, have been suggested to be involved in the signaling pathway. In earlier studies we presented data which indicated that sphingosine (Sph) itself was increased during apoptosis induced by phorbol myristate acetate (PMA) in HL60 cells and tumor necrosis factor (TNF) in neutrophils, and when added exogenously was able to induce apoptosis. We report here that Sph and its methylated derivative N,N,-dimethylsphingosine (DMS) are able to induce apoptosis in cancer cells of both hematopoietic and carcinoma origin. In human leukemic cell lines CMK-7, HL60 and U937, treatment with 20 microM Sph for 6 hr caused apoptosis in up to 90% of cells. Human colonic carcinoma cells HT29, HRT18, MKN74 and COLO205 were shown to be more susceptible to apoptosis upon addition of DMS (>50%) than of Sph (<50%), yet were weakly or not sensitive to N,N,N-trimethylsphingosine (TMS). Under the same conditions, in the presence of serum, neither Sph-1-phosphate nor ceramide analogues C2-, C6- or C8-ceramide were able to induce apoptosis in any cell lines. However, in the absence of serum, ceramide analogues induced apoptosis in leukemia cell lines after 18 hr, yet much less so than Sph or DMS. Furthermore, apoptosis induced by Sph or DMS could not be inhibited by the ceramide synthase inhibitor fumonisin B1. Apoptosis was not induced by sphingolipids in primary culture cells, such as HUVEC or rat mesangial cells, but was apparent in transformed rat mesangial cells. Additionally, apoptosis induced by Sph, DMS or C2Cer was inhibited by protease inhibitors. Our data further support the evidence that the catabolic pathway of sphingomyelin involving Sph and other metabolites is an integral part of the apoptosis pathway.

Animals↗
PubMed1996

Suppression of bcl-2 gene expression by sphingosine in the apoptosis of human leukemic HL-60 cells during phorbol ester-induced terminal differentiation.

Our recent studies have shown that intracellular levels of sphingosine, an endogenous PKC inhibitor, increase during apoptosis resulting from phorbol ester (PMA)-induced terminal differentiation of human myeloid leukemic HL-60 cells, and have suggested that sphingosine may function as an endogenous mediator of apoptosis in these cells [Ohta, et al. (1995) Cancer Res. 55, 691-697]. We report here that apoptosis induced by PMA, sphingosine, and N,N-dimethylsphingosine (DMS) was accompanied by a concomitant decrease of bcl-2 expression in both RNA and protein levels in HL-60 cells, while expression of bcl-XL and bax mRNA did not change, and neither sphingosine nor DMS induced differentiation of HL-60 cells. In contrast, in apoptotic cells induced by pharmaceutical PKC inhibitors H7 or staurosporine, expression of bcl-2 did not change nor did the intracellular sphingosine concentration. These results suggest that sphingosine may function as an endogenous mediator of apoptotic signaling in PMA-induced terminal differentiation of HL-60 cells through bcl-2 down-regulation, probably independent from PKC inhibition.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed1996

Long-term prognosis of chronic hepatitis C after treatment with interferon alpha 2b and characterization of incomplete responders.

OBJECTIVES: During the long-term follow-up of chronic hepatitis C patients treated with interferon alpha-2b, we have identified some patients who had continuous normalization, or transient elevation of the serum alanine aminotransferase levels within the first 6 months of follow-up and subsequent normalization, but HCV RNA was still found to be positive during the follow-up period (incomplete responder; ICR). We wished to clarify the characteristics of these patients, and to investigate the factors predictive of the response to interferon. METHODS: Seventy patients were treated with 6 MU of rIFN alpha-2b, three times weekly for 24 wk. Sixty-six patients (94%) completed the entire therapy and 2-yr follow-up protocol. RESULTS: Twenty-four months after cessation of IFN treatment, 24 patients (36%) were defined as complete responders (CR; continuously normal alanine aminotransferase levels, and HCV RNA negative). Nine patients (14%) were defined as ICR. Thirty-three patients (50%) were defined as nonresponders (NR). Compared with the CR or NR groups, the patients in the ICR group tended to have the mildest histological severity and lowest histological activity index scores, but there were no significant differences in any of the other histological features. The complete response rate was significantly higher in patients with low concentration of HCV RNA, and genotypes other than type II. CONCLUSIONS: After long-term follow-up of the 66 patients, 24 patients (36%) were CR, nine (14%) were ICR, and 33 (50%) were NR. The histological stage and indices of chronic hepatitis may help to predict a patient's response to IFN therapy.

Adult↗
PubMed1995

A controlled study to determine the optimal dose regimen of interferon-alpha 2b in chronic hepatitis C.

OBJECTIVES: The efficacy of three different dose treatments of interferon (IFN) was evaluated in patients with chronic hepatitis C (HCV). METHODS: Ninety-one patients with chronic hepatitis C were allocated to receive 3, 6, or 10 U of recombinant IFN-alpha 2b three times weekly for 24 wk. The number of patients was 26, 35, and 30 in the low, middle, and high dose groups, respectively. Treatment was discontinued, however, in four patients (one in the middle dose group and three in the high dose group) because of depression occurring 1-3 months after therapy began. A total of 87 patients, therefore, was analysed and was comprised of 26 patients in the low dose group, 34 in the middle dose group, and 27 in the high dose group. RESULTS: At the end of treatment, serum levels of ALT were normal in 50% (13/26) of patients in the low dose group, 70% (24/34) in the middle dose group, and 74% (20/27) in the high dose group. Patients with persistent normalization of ALT for 12 months of follow-up were defined as sustained responders. Twelve months after treatment cessation, the percentage of sustained responders was 23% (6/23) in the low dose group, 44% (15/34) in the middle dose group, and 41% (11/27) in the high dose group. Including the four patients who required discontinuation of IFN administration, the percentage of sustained responders was 43 and 37% in the middle and high dose groups, respectively. The clearance of HCV RNA was 40% in the middle dose group and 33% in the high dose group. This decrease in the clearance of HCV RNA was prominent in the high dose group. The percentage tended to be higher in the middle dose group, but the differences were not significant. CONCLUSIONS: The results suggest that a regimen of recombinant IFN-alpha 2b in a dose of 6 U three times weekly for 24 wk may be appropriate for the treatment of chronic HCV.

Alanine Transaminase↗
PubMed1994

Intravesical instillation chemotherapy of adriamycin with or without verapamil for the treatment of superficial bladder cancer: the final results of a collaborative randomized trial.

A collaborative randomized clinical trial on the intravesical administration of Adriamycin (ADM) with or without verapamil (VR), a calcium-channel blocker, as chemotherapy of superficial bladder cancer (Ta, T1) was carried out at two universities, Okayama and Kagoshima, and their affiliated hospitals. Arm A consisted of ADM given at 50 mg/50 ml saline, and arm B consisted of ADM given at 50 mg/40 ml saline plus five ampules (25 mg/10 ml saline) of injectable VR. The drugs were instilled into the bladder for 3 consecutive days, and three such courses were given with a 4-day interval between each course for a total of nine instillations. A total of 96 patients (48 in arm A and 48 in arm B) were entered into this study. The two treatment groups showed no significant difference in background factors. Of the 40 evaluated arm-A patients, 24 (60.0%) showed a response (CR + PR), whereas 19 (48.7%) of the 39 patients in arm B responded. The difference between these response rates was not statistically significant. As for adverse reactions to the intravesical chemotherapy, local inflammatory symptoms were observed in half of the patients, although no systemic reaction was observed. No significant difference was found between arm A and arm B, except for urinary turbidity. In conclusion, at the dose employed in the present clinical trial, there was no clear enhancement of the effect of ADM combined with VR in patients with superficial bladder cancer. Further clinical studies are required to determine the optimal doses of ADM and VR for their combination in intravesical chemotherapy.

Administration, Intravesical↗
PubMed1993

Neutrophil proteases associated with amyloid fibrils.

We found significant amounts of enzymatic activity characteristic of the human neutrophil proteases, elastase and cathepsin G, associated with isolated amyloid fibrils from patients with five different types of systemic amyloidosis. Amyloid deposits in tissue sections from the patients with amyloid A, amyloid transthyretin and amyloid beta 2-microglobulin amyloidosis also stained positive with antiserum to elastase and cathepsin G. Elastase and cathepsin G, found in the azurophilic granules of the neutrophil and, to a lesser extent, the monocyte, may become associated with amyloid precursor proteins before or during the formation of amyloid fibrils. This may occur in an extracellular inflammatory microenvironment or in a phagolysosome and play a role in the formation of the fibrils.

Amyloid↗
PubMed1993

The binding site for fucose-binding proteins of Lotus tetragonolobus is a prognostic marker for transitional cell carcinoma of the human urinary bladder.

BACKGROUND: Carbohydrate signals are important in tumor metastasis. METHODS: Expression of binding sites for fucose-binding proteins (FBP) of Lotus tetragonolobus were immunohistochemically analyzed in patients with transitional cell carcinoma. RESULTS: The survival of patients with invasive bladder cancer was associated with the degree of expression of FBP binding sites. Overall actuarial 5-year survival and cancer-corrected 5-year survival rates were much worse in patients with strong expression of FBP binding sites than in those with no or weak expression (P < 0.001 and P < 0.05, respectively). Strong expression of FBP binding sites in patients with Stage T3-4 disease or disease beyond Stage T2 with lymphatic permeation also was correlated with increased disease progression. Autopsies revealed that FBP binding sites were strongly expressed in all primary tumors with metastasis and in most of the metastatic tissues. CONCLUSION: The degree of expression of FBP binding sites correlates with the increased metastatic potential of bladder cancer and with poor patient survival times.

Adult↗
PubMed1993

Expression of the multidrug transporter, P-glycoprotein, in renal and transitional cell carcinomas.

BACKGROUND: Renal cell carcinomas (RCC) respond poorly to anthracyclines, Vinca alkaloids, and other agents. P-glycoprotein is overproduced in multidrug-resistant cells and thought to function as an energy-dependent drug efflux pump. The authors thus examined the expression level of P-glycoprotein in RCC and transitional cell carcinomas (TCC). METHODS: P-glycoprotein was detected using immunoblotting with a monoclonal antibody against it, C219. RESULTS: Thirty-three of 38 patients with RCC and 3 of 17 patients with TCC had P-glycoprotein positive tumors. The expression level of P-glycoprotein in most of RCC was lower than that in the normal kidney tissues and that of P-glycoprotein in the TCC was very low. The size of P-glycoprotein in 14 RCC and 3 TCC was 5-10 kilodaltons smaller than in the normal renal tissues. The variation of P-glycoprotein size in the RCC was attributed to differential N-linked glycosylation. P-glycoprotein in a RCC was photolabeled by tritiated azidopine, and the labeling was inhibited by some organic agents. P-glycoprotein distributed on the apical or marginal cell surface of the RCC. CONCLUSIONS: These data show that P-glycoprotein was expressed in many RCC, and its expression level, glycosylation, and distribution were altered. These data also suggest that the P-glycoprotein in RCC had similar drug binding site(s) to that in multidrug-resistant cells.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1993

Recombinant human tumour necrosis factor-alpha (rhTNF-alpha) and rhTNF-alpha analogue enhance amyloid deposition in the Syrian hamster.

Tumour necrosis factor-alpha (TNF-alpha) is one of the cytokines that stimulate the production of serum amyloid A (SAA), the precursor of AA amyloid. The role of TNF-alpha in amyloidogenesis was investigated in experimental hamsters using purified recombinant human TNF-alpha (rhTNF-alpha) and rhTNF-alpha analogue different from the normal molecule by two amino acid substitutions. Daily injections of 1 microgram rhTNF-alpha resulted in elevated SAA levels but even in the presence of amyloid enhancing factor (AEF) no amyloid was deposited, indicating that apart from the AEF and one particular SAA stimulating factor an additional factor is needed to result in amyloid deposition. This factor is generated by repeated injections of E. coli lipopolysaccharide (LPS). A single intraperitoneal injection of 12.5 micrograms or more of rhTNF-alpha followed by seven daily subcutaneous injections of LPS resulted in enhanced amyloid deposition. Heat denaturation of rhTNF-alpha did abolish its AEF activity. The rhTNF-alpha analogue, having one-fifth of the cytotoxic activity of the normal rhTNF-alpha, showed a similar reduction in its SAA-inducing capacity and its amyloidogenicity. This suggests the AEF activity to be closely related to TNF-alpha activity. However, poly(I)-poly(C) (a potent inducer of IL-6) also showed AEF activity, suggesting that not a single cytokine but rather a certain combination of different cytokines could be decisive in AA amyloidogenesis.

Amyloid↗
PubMed1992

Intravesical instillation of adriamycin in the presence or absence of verapamil for the treatment of superficial bladder cancer: preliminary report of a collaborative study.

A case-controlled collaborative study on the intravesical administration of Adriamycin in the presence or absence of verapamil, a calcium-channel blocker, as chemotherapy of superficial bladder cancer was carried out at two universities, Okayama and Kagoshima, and their affiliated hospitals. Although little is known about the expression of P-glycoprotein in superficial bladder cancer, it may be a cause of multidrug resistance (MDR). Verapamil was used as an inhibitor of P-glycoprotein. Arm A consisted of Adriamycin given at 50 mg/50 ml saline, and arm B constituted Adriamycin given at 50 mg/40 ml saline plus 5 ampules (10 ml) of injectable verapamil. The drugs were instilled into the bladder for 3 consecutive days in each of 3 consecutive weeks for a total of 9 instillations. No significant difference in antitumor effects was observed between arm A and arm B. Recurrent tumors responded better than did primary tumors to both arm-A and arm-B treatments (P = 0.012). In both treatment arms, significant differences (P = 0.031) in the response rate were found between tumors with diameters of less than 1 cm and those measuring 1-3 cm in diameter. Although the number of evaluable patients was limited, recurrent subjects who had previously received Adriamycin instillations responded in both treatment arms.

Administration, Intravesical↗
PubMed1992

A metastasis-associated antigen is present on a 60 kDa glycoprotein in transitional cell carcinoma of the human urinary bladder.

We have previously shown that the degree of expression of Lex-related carbohydrate epitopes, namely, Lotus tetragonolobus agglutinin (LTA) receptors, SSEA-1 and FH6, correlates with the metastatic potential of transitional cell carcinoma of the human urinary bladder. In an effort to obtain a better reagent with which to detect a metastasis-associated epitope, monoclonal antibodies were produced against LTA receptors from BOY bladder carcinoma cells. One antigen defined by such a monoclonal antibody, MM4, indeed showed better correlation with the metastatic potential of the tumour than did other carbohydrate markers. In the LTA receptors, MM4 antigen was located only on a 60 kDa glycoprotein. In extracts from primary carcinomas and lymph node metastases, the 60 kDa glycoprotein was the principal carrier of MM4 antigen. LTA receptors from these sources were composed of arrays of glycoproteins, while the 60 kDa one was invariably present. Metastasis-associated carbohydrate epitopes on the 60 kDa glycoprotein may promote metastasis by interaction with carbohydrate-recognizing proteins such as selectins on host cells.

Antigens, Neoplasm↗
PubMed1992

Intraperitoneal amyloid formation by amyloid enhancing factor--rich macrophages in ascitic fluid.

Although resident peritoneal cells from amyloidotic mice (amyloidotic peritoneal cells) are capable of processing the precursor protein of secondary amyloidosis, serum amyloid A (SAA) to amyloid fibrils, the peritoneum is a rare site for amyloid deposition. This is considered to be due to a deficiency of SAA in the peritoneum. To increase the supply of SAA to the peritoneum, ascitic fluid containing about the same protein constituents as in the serum was induced in mice. Amyloidotic peritoneal cells were packed in a microchamber which was shielded with filter membranes, and cultured in ascitic fluid supplemented with additional inflammatory factors. On the 7th day, Congo red-positive structures which showed green birefringence under polarized light were found inside and occasionally outside the chamber. By anti-AA or -SAA immunostaining, amyloid deposits and the cell surfaces of macrophages were positive. Immunologic depletion of T- and B-lymphocytes from the amyloidotic peritoneal cells did not adversely effect the amyloid formation in microchambers. These results suggest that either ascitic fluid containing sufficient amounts of SAA, or peritoneal macrophages with a high amyloid enhancing factor (AEF) activity are indispensable for AA amyloid fibrillogenesis in the peritoneum.

Amyloid↗