Biomedical subjects

Yutaka Shimada

Publications and source records attributed to Yutaka Shimada.

At least 19 recordsLinked to original sources

PubMed2006

STAT1 activation-induced apoptosis of esophageal squamous cell carcinoma cells in vivo.

BACKGROUND: The induction of apoptosis might be a promising treatment for cancers refractory to conventional therapies, such as esophageal cancer. In this study, we examined whether epidermal growth factor-induced growth inhibition results from apoptosis of esophageal squamous cell carcinoma (SCC) cells as a result of STAT1 activation and evaluated whether interferon gamma (IFN-gamma) can induce apoptosis of cancer cells in vivo. METHODS: To assess the function of STAT1, we established stable transfectants expressing dominant-negative STAT1. Apoptosis was assessed by several experimental techniques, including flow cytometry. Differentiation was evaluated by Western blot test with involucrin used as a marker. In vivo, cancer cells were injected into male BALB/c nu/nu mice. Two weeks later, the mice started to receive injections of IFN-gamma or saline into a tail vein four times per week. Concentrations of IFN-gamma in the tumors were analyzed by enzyme-linked immunosorbent assay. Apoptosis was evaluated by TUNEL (terminal deoxynucleotidyl transferase dUTP nick-end labeling) staining. RESULTS: Epidermal growth factor inhibited the growth of esophageal SCC cells by causing apoptosis through several pathways involving STAT1 activation. IFN-gamma induced the apoptosis of cancer cells, but it also promoted the differentiation (not apoptosis) of primary cultured cells derived from normal esophageal epithelium. IFN-gamma also inhibited the growth of xenograft tumors of esophageal SCC cells in vivo. CONCLUSIONS: Our results suggest that IFN-gamma is one candidate for cytokine-based therapy of cancer. IFN-gamma-induced STAT1 activation might be involved in the apoptosis of esophageal SCC cells and in the terminal differentiation of normal squamous cells. Further studies of STAT1 signaling pathways may provide the basis for new targeted therapeutic strategies for esophageal SCC.

Animals↗

PubMed2006

Severe pulmonary metastasis in obese and diabetic mice.

Although obesity is known as a risk factor for several human cancers, the association of obesity with cancer recurrence and metastasis remains to be characterized. Here, B16-BL6 melanoma and Lewis lung carcinoma cells were intravenously injected into diabetic (db/db) and obese (ob/ob) mice. The number of experimental lung colonies was markedly promoted in these mice when compared with C57BL/6 mice. In contrast, tumor growth at the implanted site was comparable when cells were inoculated orthotopically. The use of B16-BL6 cells stably transfected with the luciferase gene revealed that the increased metastasis reflected a difference mainly within 6 hr after the intravenous inoculation of tumor cells. Administration of recombinant leptin in ob/ob mice abolished the increase in metastasis early on as well as the decrease in the splenic NK cell number. In addition, depletion of NK cells by an anti-asialo-GM1 antibody abrogated the enhanced metastasis in db/db mice. These results demonstrate that metastasis is markedly promoted in diabetic and obese mice mainly because of decreased NK cell function during the early phase of metastasis.

Animals↗

PubMed2006

Gli-1 expression is associated with lymph node metastasis and tumor progression in esophageal squamous cell carcinoma.

OBJECTIVE: We investigated the expression and function of the hedgehog (Hh) pathway in human esophageal squamous cell carcinoma (ESCC). METHODS: The expression of Hh pathway molecules were detected in 34 human ESCC cell lines by RT-PCR. Subsequently, we investigated the effects of cyclopamine, a specific inhibitor of the Hh pathway, on cell proliferation, migration and invasion. Next, the effects of siRNA targeting Gli-1 were examined. Immunohistochemically, the expression of Gli-1 was studied in 104 ESCC specimens and compared with the clinicopathological characteristics of the patients. RESULTS: Gli-1 were expressed in 31 of 34 cell lines (91%), while Sonic hedgehog (SHh), Patched (Ptch), and Smoothened (Smo) expression was noted in all 34 cell lines. Cyclopamine significantly inhibited cell proliferation and migration in ESCC cells that expressed Gli-1. siRNA targeting Gli-1 inhibited cell growth in ESCC cells. Gli-1 was expressed in 52 of 104 cancer specimens (50%). Gli-1 expression was associated with tumor depth (p < 0.001), positive lymph node metastasis (p = 0.004) and a poor prognosis (p = 0.0047). CONCLUSION: Our results raise the possibility that the inhibition of the Hh pathway could be a novel target for esophageal cancer therapy.

Antineoplastic Agents↗

PubMed2006

Cholesterol-fed rabbit as a unique model of nonalcoholic, nonobese, non-insulin-resistant fatty liver disease with characteristic fibrosis.

BACKGROUND: The number of patients suffering from metabolic syndrome is increasing rapidly. Metabolic syndrome causes severe pathological changes in various organs, including the liver, and its main phenotype is nonalcoholic fatty liver disease (NAFLD). NAFLD has a broad spectrum ranging from simple fatty change to severe steatohepatitis with marked fibrosis. Recently, several experimental animal models for NAFLD have been proposed. However, most were established by rather artificial conditions such as genetic alteration. In the present study, we tried to establish a unique animal model mimicking some of the physiopathological features of NAFLD using high-cholesterol-fed rabbits. METHODS: Male rabbits fed with standard rabbit food containing 1% cholesterol for 8 weeks and 12 weeks were compared to controls (six rabbits/group). The weight of food was strictly restricted to 100 g/rabbit per day. RESULTS: Body weights and fasting plasma insulin levels showed no significant differences among the groups. In contrast, characteristic fine fibrosis was extended from perivenular to pericellular areas, and microvesicular fatty change with ballooning degeneration was observed in perivenular areas in livers of the cholesterol-fed rabbits. Increase of serum cholesterol level, activation of hepatic stellate cells, and exposure to oxidative stress were also recognized. CONCLUSIONS: Cholesterol-fed rabbits share several physiopathological features of NAFLD. Because this model did not show insulin resistance or obesity, it may be useful for elucidating the mechanism of NAFLD related mainly to hyperlipidemia.

Animals↗

PubMed2006

Protective effects of keishibukuryogan on the kidney of spontaneously diabetic WBN/Kob rats.

Keishibukuryogan, one of the traditional herbal formulations, is used clinically to improve blood circulation. It consists of the following five crude drugs: Cinnamomi Cortex, Poria, Moutan Cortex, Persicae Semen and Paeoniae Radix. In this study, the effects of keishibukuryogan against renal damage in spontaneously diabetic WBN/Kob rats were examined. Oral administration of keishibukuryogan significantly attenuated urinary protein excretion and serum creatinine levels. It did not affect body weight loss and blood glucose levels, but it suppressed renal and hepatic weights of WBN/Kob rats. Keishibukuryogan also reduced fibronectin and transforming growth factor beta(1) (TGF-beta(1)) protein expression in the renal cortex. Furthermore, lipid peroxidation levels in both kidney and liver were significantly lower than those of untreated control WBN/Kob rats. Urinary excretion of 8-hydroxy-deoxyguanosine was suppressed by keishibukuryogan treatment. These results suggest that keishibukuryogan reduces oxidative stress by hyperglycemia, and that it protects renal function and suppresses fibronectin deposition induced by TGF-beta(1) production in WBN/Kob rats.

8-Hydroxy-2'-Deoxyguanosine↗

PubMed2006

Astaxanthin ameliorates features of metabolic syndrome in SHR/NDmcr-cp.

Glucose and lipid metabolic parameters play crucial roles in metabolic syndrome and its major feature of insulin resistance. This study was designed to investigate whether dietary astaxanthin oil (ASX-O) has potential effects on metabolic syndrome features in an SHR/NDmcr-cp (cp/cp) rat model. Oral administration of ASX (50 mg/kg/day) for 22 weeks induced a significant reduction in arterial blood pressure in SHRcp. It also significantly reduced the fasting blood glucose level, homeostasis index of insulin resistance (HOMA-IR), and improved insulin sensitivity. The results also showed an improved adiponectin level, a significant increase in high-density lipoprotein cholesterol, a significant decrease in plasma levels of triglycerides, and non-esterified fatty acids. Additionally, ASX showed significant effects on the white adipose tissue by decreasing the size of the fat cells. These results suggest that ASX ameliorates insulin resistance by mechanisms involving the increase of glucose uptake, and by modulating the level of circulating lipid metabolites and adiponectin.

Adiponectin↗

PubMed2006

Effect of d-pseudoephedrine on cough reflex and its mode of action in guinea pigs.

d-Pseudoephedrine (PSE) is one of the main ingredients of Ephedrae herba. Although PSE is widely applied for patients with a common cold and upper respiratory inflammation as a decongestant, the effects of PSE on cough have never been reported. In this study, we investigated the antitussive effects of intraperitoneal injection of PSE on the cough reflex induced by microinjection of citric acid into the larynx of guinea pigs. PSE decreased the number of cough reflexes dose-dependently (-18.3 +/- 5.0% at 20 mg/kg, P<0.05; -41.1 +/- 7.2% at 60 mg/kg, P<0.01). Furthermore, PSE (60 mg/kg) increased the threshold intensity for inducing fictive cough by electrical micro-stimulation of the nucleus tractus solitarius (+72.7 +/- 8.4%, P<0.01). On the afferent discharge of the superior laryngeal nerve, PSE suppressed the increases of amplitude and frequency when stimulated by citric acid at laryngeal mucosa. These results demonstrate that PSE possesses an antitussive effect that might be derived from both central and peripheral actions.

Animals↗

PubMed2006

Nicotine induces the fragile histidine triad methylation in human esophageal squamous epithelial cells.

The fragile histidine triad (FHIT) gene has been proposed to have an important role in very early carcinogenesis. Methylation of the FHIT gene is associated with transcriptional inactivation in esophageal squamous cell carcinoma, and FHIT inactivation has been linked to smoking-related carcinogenesis. In this study, we confirmed methylation of the FHIT gene in human esophageal squamous epithelial cells (HEECs) and examined whether nicotine induced alteration of FHIT. Methylation status in the promoter region of the FHIT gene and p16(INK4A) gene was determined by methylation-specific PCR in HEECs exposed to nicotine under various conditions. Methylation status of the FHIT gene was confirmed by DNA-sequencing analysis. Protein expression of Fhit and the DNA methyltransferases (DNMTs) DNMT1 and DNMT3a were assessed by immunoblot analysis. In the absence of nicotine, methylation of the FHIT gene and attenuation of Fhit protein were not detected in HEECs. Nicotine induced the methylation of FHIT gene and attenuated Fhit protein in association with increased expression of DNMT3a. Reexpression of Fhit protein in HEECs was found after cessation of moderate- to long-term exposure to nicotine. Our results show that nicotine induces methylation of the FHIT gene followed by loss of Fhit protein expression in HEECs. Continuous smoking may thus increase the risk of esophageal cancer.

Acid Anhydride Hydrolases↗

PubMed2006

The biological role of the low-affinity p75 neurotrophin receptor in esophageal squamous cell carcinoma.

UNLABELLED: In this study, we investigated the clinicopathologic significance of the low-affinity p75 neurotrophin receptor (p75NTR; which is expressed in the stem/progenitor cell fraction of normal esophageal epithelial cells) in 187 resected esophageal squamous cell carcinoma (ESCC) specimens and found that approximately 50% of ESCC expressed p75NTR. Our investigation using ESCC cell lines showed that p75NTR was intensely expressed in the cells with high colony-forming capacity but they were sensitive to cell death on inhibition of p75NTR expression with transient transfection of small interfering RNA (siRNA). These findings suggest that p75NTR is necessary for survival and maintenance of ESCC tumors, providing us with a potential target for novel therapies. PURPOSE: p75NTR is expressed in a stem/progenitor cell fraction of human normal esophageal epithelial cells. In this study, we investigated the expression and biological role of p75NTR in ESCC. EXPERIMENTAL DESIGN: The expression of p75NTR in 187 resected ESCC specimens was immunohistochemically investigated. The expression of p75NTR in 30 ESCC cell lines (KYSEs) was assessed by reverse transcription-PCR, immunocytochemistry, and flow cytometry. The p75NTR-bright and p75NTR-dim/negative cells were isolated from KYSE150 by magnetic beads and colony formation was investigated. The role of p75NTR in KYSEs was assessed by transient transfection of siRNA. RESULTS: p75NTR was expressed in 92 of 187 (49.2%) tumors. In well-differentiated tumors, positive staining was apparent in the first one to two layers from infiltrative margin of the tumors where most of the cells were actively proliferating. In moderately differentiated tumors, p75NTR was expressed in wider range from the margin of the tumors whereas p75NTR was diffusely distributed in poorly differentiated tumors. p75NTR was expressed in all examined KYSEs and the mean proportion of the p75NTR-bright fraction was 30.1%. The size of p75NTR-positive colonies was larger than that of p75NTR-negative colonies derived from KYSE150 (P<0.0001). The purified p75NTR-bright cells formed p75NTR-positive large colonies more frequently than the p75NTR-dim/negative cells (P<0.0001). Down-regulation of p75NTR expression by siRNA resulted in marked growth inhibition with induction of apoptosis. CONCLUSIONS: Our findings suggest that p75NTR is necessary for survival and maintenance of ESCC tumors, providing us with a potential target for novel therapies.

Adult↗

PubMed2006

Chenodeoxycholic acid stimulates the progression of human esophageal cancer cells: A possible mechanism of angiogenesis in patients with esophageal cancer.

Bile acids are known to promote the growth of gastrointestinal cancer. However, the underlying mechanism remains unclear. We examined whether bile acids induce tumor growth via the cyclooxygenase (COX)-2 angiogenic pathway. In vitro, esophageal squamous cell carcinoma (ESCC) cells and esophageal adenocarcinoma cells were studied. Production of prostaglandin E2 (PGE2) and vascular endothelial growth factor (VEGF) in response to treatment with chenodeoxycholic acid (CDCA) was assessed by enzyme-linked immunosorbent assay (ELISA). COX-2 protein and VEGF protein were measured by immunoblot analysis, and COX-2 activity was measured by ELISA. In vivo, CDCA was administered to ESCC cell-bearing mice. Tumor tissues were analyzed immunohistochemically, and microvessel density was evaluated. Clinically, 134 patients with ESCC who underwent esophagectomy were studied. In vitro, CDCA induced the production of PGE2 and VEGF in dose- and time-dependent manners, and these effects were attenuated by a selective COX-2 inhibitor, mitogen-activated protein kinases inhibitor, or epidermal growth factor receptor inhibitor. CDCA-induced COX-2 in the cell lysate increased the secretion of VEGF into the culture medium. In vivo, CDCA markedly enhanced tumor growth and increased vascularization. Clinically, patients whose tumors expressed both COX-2 and VEGF had poor outcomes. Our results suggest that bile acids, important constituents of duodenal fluid, stimulate the development of human esophageal cancer by promoting angiogenesis via the COX-2 pathway.

Adult↗

PubMed2006

Effects of eppikahangeto, a Kampo formula, and Ephedrae herba against citric acid-induced laryngeal cough in guinea pigs.

To evaluate the efficacy of three common antitussive Kampo formulas, eppikahangeto (EPP), bakumondoto (BAK), and shoseiryutogomakyokansekito (SGM), a new cough model of guinea pig was used, which could specifically induce a laryngeal cough by microinjection of citric acid solution into the larynx. Kampo extract was dissolved in water and the animals were given access ad libitum for 3 days, and then the number of coughs during 10 min was counted. EPP extract decreased the number of coughs dose-dependently (0.3% extract, -22.9 +/- 6.6%, P<0.01; 1.0% extract, -32.4 +/- 5.5%, P<0.01). BAK extract and SGM extract had no significant effect. Intraperitoneal injection of codeine (60 mg/kg) also decreased the number of coughs (-36.1 +/- 9.1%, P<0.05). Furthermore, Ephedrae herba (EH) extract reduced the number of coughs (-18.3 +/- 6.0%, P<0.05), but the extract of EPP without EH did not. These results suggest that EPP has an antitussive effect against laryngeally-induced cough in guinea pigs, and the crucial herbal medicine is EH.

Animals↗

PubMed2006

Esophageal cancer: evaluation with triple-phase dynamic CT--initial experience.

PURPOSE: To prospectively assess which phase of a triple-phase dynamic contrast material-enhanced multi-detector row computed tomography (CT) protocol is optimal for visualization of esophageal cancer. MATERIALS AND METHODS: The study was supported by the local ethical committee; all patients gave written informed consent. Thirty-one lesions in 28 consecutive patients (26 men, two women; mean age, 65 years; range, 53-87 years) with histopathologically confirmed esophageal cancer were evaluated with triple-phase dynamic CT performed at 5, 35, and 65 seconds (first arterial, second arterial, and venous phases) after attenuation of 200 HU was obtained at the descending aorta. Qualitative image analysis was performed to assess appearance and conspicuity of the tumor. Appearances of all 31 lesions were classified into three categories-not identifiable, focal enhancement with or without minimal (<1 cm) wall thickening, and focal mass lesion or obvious (>1 cm) wall thickening. Results were compared with surgical or endoscopic ultrasonographic findings. Quantitative assessment included regions-of-interest measurement of the tumor and normal esophageal wall and the difference between those measurements. A paired t test was used to determine which phase showed the highest tumor attenuation and tumor-to-normal esophageal wall attenuation differences. RESULTS: At visual assessment, 30 lesions were identified in the second arterial phase. Of these 30 lesions, eight were focal enhancements; the best conspicuity was during the second arterial phase. Furthermore, seven of these eight lesions were T1 cancers. The remaining 22 lesions were enhanced masses or wall thickening. Twenty-one of these 22 tumors also showed best conspicuity in the second arterial phase. The greatest attenuation of tumors in the second arterial phase was 130.0 HU, and the difference in attenuation between tumor and normal esophageal wall was 50.6 HU in the second arterial phase, which were significantly higher than those in the other two phases (P<.01, each). CONCLUSION: The second arterial phase of dynamic CT is the optimal phase for visualization of esophageal cancer.

Adenocarcinoma↗

PubMed2006

The transcription factor Cdx2 regulates the intestine-specific expression of human peptide transporter 1 through functional interaction with Sp1.

H(+)/peptide cotransporter 1 (PEPT1, SLC15A1) localized at the brush-border membranes of intestinal epithelial cells plays important roles in the intestinal absorption of small peptides and a variety of peptidemimetic drugs. We previously demonstrated that transcription factor Sp1 functions as a basal transcriptional regulator of human PEPT1. However, the factor responsible for the intestine-specific expression of PEPT1 remains unknown. In the present study, we investigated the effect of the intestinal transcription factors on the transcription of the PEPT1 gene and found that only Cdx2 markedly trans-activated the PEPT1 promoter. However, the promoter region responsible for this effect lacked a typical Cdx2-binding sequence, but instead, possessed some Sp1-binding sites. In vitro experiments using Caco-2 cells showed that (1) mutation of the Sp1-binding site diminished the effect of Cdx2, (2) co-expression of Cdx2 and Sp1 synergistically trans-activated the PEPT1 promoter and (3) Sp1 protein was immunoprecipitated with Cdx2 protein. These results raise the possibility that Cdx2 modulates the PEPT1 promoter by interaction with Sp1. The significance of Cdx2 in vivo for PEPT1 regulation was shown by the determination of mRNA levels of Cdx2 and PEPT1 in human tissue. In gastric samples, some with intestinal metaplasia, the levels of PEPT1 and Cdx2 mRNA were highly correlated. Taken together, the present study suggests that Cdx2 plays a key role in the transcriptional regulation of the intestine-specific expression of PEPT1, possibly through interaction with Sp1.

Binding Sites↗

PubMed2006

An inducible short-hairpin RNA vector against osteopontin reduces metastatic potential of human esophageal squamous cell carcinoma in vitro and in vivo.

PURPOSE: To elucidate the clinical significance of osteopontin and the effect of conditional down-regulation of osteopontin in esophageal squamous cell carcinoma (ESCC), we investigated osteopontin expression in tumors and tested an inducible osteopontin-short-hairpin RNA (shRNA) expression vector in an ESCC cell line. EXPERIMENTAL DESIGN: Osteopontin mRNA expression was extracted from gene expression profiles of 23 tumors determined by cDNA microarray and was analyzed. Paraffin sections of 144 tumors were immunohistochemically investigated. Osteopontin protein expression in 34 cell lines was examined by Western blot. A doxycycline-inducible osteopontin-shRNA vector was stably transfected into HSA/c cells to assess the role of osteopontin in cell motility, invasion in vitro, tumor formation, and lymph node metastasis in nude mice. RESULTS: cDNA microarray revealed that high osteopontin mRNA expression was associated with poor survival of ESCC patients (P = 0.029). In immunohistochemistry, osteopontin protein expression was associated with poor prognosis (P < 0.001), distant lymph node metastasis (P = 0.0004), tumor staging (P = 0.027), and histologic grade (P = 0.024). Multivariate analysis showed that osteopontin overexpression was the strongest independent prognostic factor among nine clinicopathologic variables (P < 0.001). Among cell lines tested, 30 had overexpressed osteopontin protein compared with a normal esophageal epithelial cell line. An inducible shRNA vector against osteopontin successfully down-regulated osteopontin expression by 71% to 88% and repressed cell motility by 69% to 97%, cell invasion by 59% to 71%, tumor formation by 56% to 92%, and lymph node metastasis by 50% to 67% in HSA/c cells. CONCLUSIONS: Our findings suggest that osteopontin overexpression may play an important role in progression of ESCC and osteopontin could be a potential target of ESCC therapy.

Administration, Oral↗

PubMed2006

Polo-like kinase and survivin are esophageal tumor-specific promoters.

For developing successful cancer gene therapy strategies, tumor-specific gene delivery is essential. In this study, we used esophageal cancer (EC) cells to identify and evaluate esophageal tumor-specific gene promoters. Four genes (polo-like kinase-1/PLK, survivin/BIRC5, karyopherin alpha 2/KPNA2, and pituitary tumor transforming gene protein 1/PTTG1) were identified by a microarray analysis as highly expressed in EC cell lines vs. five normal organ tissues (liver, lung, kidney, brain, and heart). By quantitative RT-PCR, the average mRNA expression levels of these four genes in 20 primary ECs were 2.7-fold (PLK), 6.1-fold (survivin), 2.6-fold (KPNA2), and 2.4-fold (PTTG1) higher than that of each gene in 24 different normal organs. By dual luciferase assay, the promoter activity of PLK and survivin in EC cell lines was 18.9-fold and 28.5-fold higher, respectively, than in normal lung and renal cells. The promoters of PLK and survivin could be useful tools for developing EC-specific gene therapy vectors.

Biomarkers, Tumor↗

PubMed2006

A new specific gene expression in squamous cell carcinoma of the esophagus detected using representational difference analysis and cDNA microarray.

OBJECTIVES: To detect new specific gene expressions in squamous cell carcinoma of the esophagus. METHODS: Representational difference analysis of cDNA (cDNA RDA) was applied to a human esophageal cancer cell line (KYSE170) and a human esophageal epithelial cell line (HEEC-1). RESULTS: LAGE-1 was expressed specifically in KYSE170, but not in HEEC-1. It is also expressed in 27% of esophageal cancer cell lines (3/11) and 33% of esophageal cancer tissues (10/30), but not in other HEECs, normal esophageal epithelium, or other normal tissues except testis, ovary and kidney. The expression of LAGE-1 is strongly correlated with that of MAGE-A1 (p = 0.013, Fisher's exact probability test). Fibronectin, cytokeratin 6B, cytokeratin 19, cyclin D2 and Ten-m2 were detected as candidates for downregulated genes. Reduced expression profiles of them were also identified using cDNA microarrays. The expression of LAGE-1 was induced by 5'-aza-2'-deoxycytidine (5Aza-dC) and trichostatin A (TSA) in esophageal cancer cell lines, which did not express LAGE-1. In HEECs, 5Aza-dC induced LAGE-1 expression, but TSA did not. CONCLUSIONS: LAGE-1 expression was detected in esophageal cancer by cDNA RDA. LAGE-1 might have the potential to be a target antigen for anti-tumoral immunotherapy in esophageal cancers because of its tumor-specific expression similar to that of MAGE-A1.

Antigens, Neoplasm↗

PubMed2006

Cisplatin-dependent upregulation of death receptors 4 and 5 augments induction of apoptosis by TNF-related apoptosis-inducing ligand against esophageal squamous cell carcinoma.

TNF-related apoptosis-inducing ligand (TRAIL) is a member of the TNF superfamily known to induce apoptosis in a variety of cancers. The purpose of our study was to examine the effects of TRAIL in combination with cisplatin against esophageal squamous cell carcinoma (ESCC) cell lines in vitro and in vivo, and to elucidate underlying molecular mechanisms. Expression profiles of TRAIL receptors were investigated in 19 ESCC (KYSE) cell lines using RT-PCR. Crystal violet staining assays were performed to reveal the sensitivity against TRAIL. Flow cytometric analyses of apoptosis induction and TRAIL receptor expression were performed. Furthermore, Western blot was used to clarify the apoptosis pathway involved, and a nude-mouse xenograft model was used to show effects in vivo. Results show that death receptors (DR) 4 and 5 were expressed in 100% of the cell lines, and 79% (15/19) expressed 4 TRAIL receptors. There was only 1 cell line without decoy receptor expression. Eighteen cell lines were resistant to TRAIL, but in some, the combination treatment with cisplatin could overcome this resistance. They underwent apoptosis via activation of caspase-8 and -3, and cisplatin-dependent upregulation of DR4 and 5 was detected. Furthermore, pretreatment with cisplatin followed by TRAIL resulted in significant tumoricidal effects. Finally, systemic administration of TRAIL with cisplatin synergistically suppressed tumor growth of ESCC xenografts in nude mice. These results provide a significance of cisplatin-induced upregulation of death receptors as apoptosis-inducing machinery, and it was suggested that sequential administration of cisplatin and TRAIL might be a feasible chemotherapeutic regimen against ESCC.

Animals↗

PubMed2006

Prospective antitumor effects of the combination of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and cisplatin against esophageal squamous cell carcinoma.

PURPOSE: Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is a member of the TNF family, which binds to death receptor (DR) 4 and DR5 to mediate apoptosis. Previously, we showed that the combination of TRAIL and cisplatin was effective against esophageal squamous cell carcinoma (ESCC) cell lines in vitro and in vivo, using one of the ESCC cell lines (KYSE 170). KYSE 110 is another ESCC cell line, but it lacks expression of decoy receptors. Thus, by using KYSE 110, we can eliminate any effects from two decoy receptors. METHODS: We used reverse transcription-polymerase chain reaction (RT-PCR) to reveal the expression of TRAIL receptors. Crystal violet staining and flow cytometry were done to confirm cytotoxicity and induction of apoptosis. KYSE 110 xenografted in nude mice was treated with TRAIL and cisplatin. The tumors were subsequently removed for microscopic studies. RESULTS: ESCC sensitive to the combination treatment in vitro was also sensitive to the treatment in vivo. Furthermore, induction of apoptosis resulted via the caspase cascade and the mitochondrial pathway. Low doses of both cisplatin and TRAIL sufficed to obtain these effects. CONCLUSION: These findings imply that in clinical application, low doses of both agents could be administered to minimize side effects, while augmenting tumoricidal activities.

Animals↗