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Objective:To explore the changes of serum B7 homolog 2 (B7-H2), tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), interleukin(IL)-37 and IL-17A levels in patients with primary immune thrombocytopenia (ITP), and to analyze the clinical guiding significance of each index level in the diagnosis and treatment of ITP.Methods:A total of 90 patients with ITP treated in Jining Hospital of Traditional Chinese Medicine from September 2018 to September 2020 were retrospectively selected as the research group, and 90 healthy patients during the same period were selected as the normal control group. The levels of serum B7-H2, TRAIL, IL-37, IL-17A and platelet count (PLT) in the two groups were compared, and the clinical guidance significance of each index level in the diagnosis and treatment of ITP were analyzed by receiver operating characteristic(ROC) curve.Results:The serum levels of B7-H2, TRAIL, IL-37 and IL-17A in the research group were higher than those in the normal control group, and PLT was lower than that in the normal control group: (25.12 ± 4.29) μg/L vs. (12.26 ± 3.10) μg/L, (0.92 ± 0.20) μg/L vs.(0.46 ± 0.13) μg/L, (145.02 ± 43.18) ng/L vs. (50.23 ± 14.07) ng/L, (21.63 ± 5.06) ng/L vs. (7.71 ± 2.04) ng/L, (16.12 ± 4.61) × 10 9/L vs. (200.86 ± 61.4) × 10 9/L, there were statistical differences ( P<0.05). After treatment for 3 months, 73 patients obtained remission, and 13 patients were non-remission. The levels of B7-H2, TRAIL, IL-37, IL-17A in the non-remission group before and after treatment were higher than those in the remission group, and PLT was lower than that in the remission group, there were statistical differences ( P<0.05). Pearson correlation analysis showed that the levels of serum B7-H2, TRAIL, IL-37 and IL-17A were negatively correlated with PLT ( P<0.05). The ROC curve analysis showed that the area under the curve of serum B7-H2, TRAIL, IL-37 and IL-17A in prognostic prediction was 0.916, the sensitivity and the specificity were 94.12% and 86.30%. Conclusions:The serum levels of B7-H2, TRAIL, IL-37 and IL-17A in patients with ITP are significantly elevated, and are closely related to the level of PLT. Combined detection can help clinically predict the direction of disease outcome.
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The rapidly developing resistance of cancers to chemotherapy agents and the severe cytotoxicity of such agents to normal cells are major stumbling blocks in current cancer treatments. Most current chemotherapy agents have significant cytotoxicity, which leads to devastating adverse effects and results in a substandard quality of life, including increased daily morbidity and premature mortality. The death receptor of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) can sidestep p53-dependent pathways to induce tumor cell apoptosis without damaging most normal cells. However, various cancer cells can develop resistance to TRAIL-induced apoptosis via different pathways. Therefore, it is critical to find an efficient TRAIL sensitizer to reverse the resistance of tumor cells to TRAIL, and to reinforce TRAIL's ability to induce tumor cell apoptosis. In recent years, traditional Chinese medicines and their active ingredients have shown great potential to trigger apoptotic cell death in TRAIL-resistant cancer cell lines. This review aims to collate information about Chinese medicines that can effectively reverse the resistance of tumor cells to TRAIL and enhance TRAIL's ability to induce apoptosis. We explore the therapeutic potential of TRAIL and provide new ideas for the development of TRAIL therapy and the generation of new anti-cancer drugs for human cancer treatment. This study involved an extensive review of studies obtained from literature searches of electronic databases such as Google Scholar and PubMed. "TRAIL sensitize" and "Chinese medicine" were the search keywords. We then isolated newly published studies on the mechanisms of TRAIL-induced apoptosis. The name of each plant was validated using certified databases such as The Plant List. This study indicates that TRAIL can be combined with different Chinese medicine components through intrinsic or extrinsic pathways to promote cancer cell apoptosis. It also demonstrates that the active ingredients of traditional Chinese medicines enhance the sensitivity of cancer cells to TRAIL-mediated apoptosis. This provides useful information regarding traditional Chinese medicine treatment, the development of TRAIL-based therapies, and the treatment of cancer.
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Objective:To explore the mechanism of circular permuted tumor necrosis factor-related apoptosis-inducing ligand (CPT) reversing the resistance to imatinib in chronic myeloid leukemia (CML) cells.Methods:Five patients with CML in the Affiliated Hospital of Inner Mongolia Medical University from 2016 to 2020 were selected, and heparinized bone marrow blood samples were collected at the first diagnosis and imatinib resistance phase, and mononuclear cells were isolated. The mononuclear cells collected at the first diagnosis were named A1-E1, and the mononuclear cells collected after imatinib resistance were named A2-E2. Human CML wild-type K562 cell line (K562-W) was given gradually increasing small doses of low-concentration imatinib to obtain imatinib-resistant K562 cells (K562-R). K562-R cells were cultured with 20 μg/L CPT and these cells were set as CPT-K562-R group. The CCK-8 method was used to detect the half inhibitory concentration ( IC50) of cells for imatinib. K562-W and K562-R cells were used to establish CML xenografts nude mice models, then the nude mice were divided into K562-W, K562-R and CPT-K562-R xenograft groups. Imatinib was perfused orally in all three groups, and CPT was injected subcutaneously in the CPT-K562-R group at the same time. The tumor volume of the three groups of nude mice before and 4 weeks after treatment with imatinib, and the survival time of the three groups of nude mice were compared. Western blot was used to detect the changes of tyrosine protein kinase receptor B4 (EphB4) and myeloid cell leukemia protein 1 (Mcl-1) protein levels in bone marrow mononuclear cells, K562 cell line and transplanted tumor tissues of CML patients. Results:The expressions of EphB4 protein in A2-E2 cells of 5 patients with CML were higher than those in A1-E1 cells (all P < 0.01). The IC50 of K562-W, K562-R and CPT-K562-R cells for imatinib were (0.160±0.015) mg/L, (5.450±0.460) mg/L, (0.300±0.035) mg/L, and the difference was statistically significant ( F = 390.65, P < 0.01). In cells of K562-W group, EphB4 and Mcl-1 proteins were expressed at low levels (0.54±0.02 and 0.70±0.08); in cells of K562-R group, the expressions of EphB4 and Mcl-1 proteins were enhanced (3.04±0.11 and 2.88±0.04); in cells of CPT-K562-R group, the expressions of EphB4 and Mcl-1 proteins decreased (0.57±0.03 and 0.38±0.04). Before imatinib treatment, there was no statistically significant difference in the tumor volumes of nude mice among the K562-W, K562-R and CPT-K562-R xenograft groups ( F = 0.39, P = 0.68), suggesting the transplanted tumors formed in nude mice were balanced; after imatinib treatment, the difference in the tumor volumes among the three groups were statistically significant ( F = 26.16, P < 0.01). The survival time of nude mice in the K562-W, K562-R and CPT-K562-R xenograft groups was (18.5±3.3) d, (10.0±2.4) d and (17.5±1.6) d, and the difference was statistically significant ( F = 20.45, P < 0.01). In K562-W xenograft group, both EphB4 and Mcl-1 proteins were expressed at low levels (0.55±0.06 and 0.67±0.06); in K562-R xenograft group, the expressions of EphB4 and Mcl-1 proteins were enhanced (1.95±0.08 and 6.21±0.53); the expressions of EphB4 and Mcl-1 in CPT-K562-R xenograft group decreased (0.59±0.04 and 0.37±0.04) and were close to their expressions in K562-W xenograft group. Conclusion:CPT may enhance the sensitivity of CML to imatinib by inhibiting the expressions of EphB4 and Mcl-1, and this may be a targeted pathway for imatinib therapy.
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Objective: To investigate whether piperlongumine can sensitize prostate cancer cells to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and trigger apoptosis in prostate cells.Methods: Human prostate cancer cell lines PC3, LNCaP, and VCaP were cultured with piperlongumine and TRAIL. Then, cell proliferation, migration, caspase activation, apoptotic protein expressions, and death receptor expressions were measured. Results: Piperlongumine inhibited cell proliferation at low doses (<10 μM) alone and in combination with TRAIL (25 ng/mL), induced apoptosis, and suppressed cyclooxygenase activation. Additionally, piperlongumine induced expression of death receptors which potentiated TRAIL-induced apoptosis in cancer cells but did not affect decoy receptors. Piperlongumine also downregulated tumor cell-survival pathways, inhibited colony formation and migration of cancer cells alone or in combination with TRAIL. The combination of piperlongumine with TRAIL was found to be synergistic. Conclusions: Our findings indicate that piperlongumine can sensitize cancer cells to TRAIL through the upregulation of death receptors and can trigger apoptosis with the downregulation of anti-apoptotic proteins.
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Objective: To investigate whether piperlongumine can sensitize prostate cancer cells to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and trigger apoptosis in prostate cells. Methods: Human prostate cancer cell lines PC3, LNCaP, and VCaP were cultured with piperlongumine and TRAIL. Then, cell proliferation, migration, caspase activation, apoptotic protein expressions, and death receptor expressions were measured. Results: Piperlongumine inhibited cell proliferation at low doses (<10 μM) alone and in combination with TRAIL (25 ng/mL), induced apoptosis, and suppressed cyclooxygenase activation. Additionally, piperlongumine induced expression of death receptors which potentiated TRAIL-induced apoptosis in cancer cells but did not affect decoy receptors. Piperlongumine also downregulated tumor cell-survival pathways, inhibited colony formation and migration of cancer cells alone or in combination with TRAIL. The combination of piperlongumine with TRAIL was found to be synergistic. Conclusions: Our findings indicate that piperlongumine can sensitize cancer cells to TRAIL through the upregulation of death receptors and can trigger apoptosis with the downregulation of anti-apoptotic proteins.
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Objective: To explore the possibility of promoting tumor necrosis factor-related apoptosis inducing ligand (TRAIL)-induced apoptosis in prostate cancer PC-3 cell by inhibiting Krüppel-like factor 5 (KLF5). Methods: MTT assay, flow cytometry, Western blot assay and qRT-PCR assay were deployed to detect the cell viability, apoptosis and apoptotic markers in KLF5-inhibited and TRAIL-induced PC-3 cells. Results: After KLF5 was inhibited in TRAIL-induced PC-3 cells, cell viability reduced, apoptosis enhanced, the expressions of DR4 and DR5 increased while the expression of cellular fas-associated death domain-like interleukin-1β converting enzyme inhibitory protein (c-FLIP) decreased. Conclusion: Inhibiting KLF5 suppresses cell viability by promoting TRAIL-induced apoptosis in prostate cancer cell PC-3. It may be a potential means to treat hormone-insensitive prostate cancer.
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Objective To explore the plasma level change of soluble tumor necrosis factor related apoptosis inducing ligand (sTRAIL) in patients with systemic lupus erythematosus (SLE) and its clinical significance.Methods Quantitative real-time reverse transcription-polymerase chain reaction (RT-PCR) was used to detect the mRNA expressions of TRAIL and TRAIL receptors-1 (TRAIL-R1) and TRAIL-R2 in the peripheral blood mononuclear cells (PBMCs) derived from active SLE patients (n =26) and healthy controls.Enzyme linked immuno sorbent assay (ELISA) was used to detect the plasma level of sTRAIL in the active SLE patients (n=42),healthy controls (n=21).Pearson correlation analysis was used to analyze the correlation of sTRAIL with clinical and laboratory parameters.Results The plasma levels of sTRAIL [(82±5) pg/ml] in SLE were significantly higher than that in healthy controls [(49 ±3) pg/ml],the difference was statistically significant (t=4.10,P＜0.01).The plasma levels of sTRAIL in SLE with inactive disease [(92±14) pg/ml],mild active disease [(80±9) pg/ml],moderate active disease [(74±12) pg/ml] and severe active disease [(83±8) pg/ml] were higher than healthy controls,the difference was statistically significant (H=18.07,P＜0.01).The mRNA levels of TRAIL and TRAIL-R2 in PBMCs derived from SLE patients were significantly higher than those in healthy controls [(1.04±0.08) vs (1.80±0.25),t=2.10,P＜0.05 and (1.07±0.12) vs (2.08±0.21),t=3.27,P＜0.01].In SLE with moderate and severe active disease,plasma sTRAIL levels were associated with the 24 hours urine protein.Conclusion Plasma sTRAIL may be predictors of SLE disease activity and TRAIL pathway may be a new potential target of SLE treatment.
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Objective@#To explore the plasma level change of soluble tumor necrosis factor related apoptosis inducing ligand (sTRAIL) in patients with systemic lupus erythematosus (SLE) and its clinical significance.@*Methods@#Quantitative real-time reverse transcription-polymerase chain reaction (RT-PCR) was used to detect the mRNA expressions of TRAIL and TRAIL receptors-1 (TRAIL-R1) and TRAIL-R2 in the peripheral blood mononuclear cells (PBMCs) derived from active SLE patients (n=26) and healthy controls. Enzyme linked immuno sorbent assay (ELISA) was used to detect the plasma level of sTRAIL in the active SLE patients (n=42), healthy controls (n=21). Pearson correlation analysis was used to analyze the correlation of sTRAIL with clinical and laboratory parameters.@*Results@#The plasma levels of sTRAIL [(82±5) pg/ml] in SLE were significantly higher than that in healthy controls [(49±3) pg/ml], the difference was statistically significant (t=4.10, P<0.01). The plasma levels of sTRAIL in SLE with inactive disease [(92±14) pg/ml], mild active disease [(80±9) pg/ml], moderate active disease [(74±12) pg/ml] and severe active disease [(83±8) pg/ml] were higher than healthy controls, the difference was statistically significant (H=18.07, P<0.01). The mRNA levels of TRAIL and TRAIL-R2 in PBMCs derived from SLE patients were significantly higher than those in healthy controls [(1.04±0.08) vs (1.80±0.25), t=2.10, P<0.05 and (1.07±0.12) vs (2.08±0.21), t=3.27, P<0.01]. In SLE with moderate and severe active disease, plasma sTRAIL levels were associated with the 24 hours urine protein.@*Conclusion@#Plasma sTRAIL may be predictors of SLE disease activity and TRAIL pathway may be a new potential target of SLE treatment.
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Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a promising agent for anticancer therapy. The identification of small molecules that can establish the sensitivity of prostate cancer (PCa) cells to TRAIL-induced apoptosis is crucial for the targeted treatment of PCa. PC3, DU145, JAC-1, TsuPr1, and LNCaP cells were treated with Andrographolide (Andro) and TRAIL, and the apoptosis was measured using the Annexin V/PI double staining method. Real time-polymerase chain reaction (PCR) and Western blot analysis were performed to measure the expression levels of target molecules. RNA interference technique was used to down-regulate the expression of the target protein. We established a nude mouse xenograft model of PCa, which was used to measure the caspase-3 activity in the tumor cells using flow cytometry. In this research study, our results demonstrated that Andro preferentially increased the sensitivity of PCa cells to TRAIL-induced apoptosis at subtoxic concentrations, and the regulation mechanism was related to the up-regulation of DR4. In addition, it also increased the p53 expression and led to the generation of reactive oxygen species (ROS) in the cells. Further research revealed that the DR4 inhibition, p53 expression, and ROS generation can significantly reduce the apoptosis induced by the combination of TRAIL and Andro in PCa cells. In conclusion, Andro increases the sensitivity of PCa cells to TRAIL-induced apoptosis through the generation of ROS and up-regulation of p53 and then promotes PCa cell apoptosis associated with the activation of DR4.
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Animals , Humans , Male , Mice , Antineoplastic Agents/pharmacology , Apoptosis/drug effects , Cell Line, Tumor , Diterpenes/pharmacology , Drug Synergism , Mice, Nude , Neoplasm Transplantation , PC-3 Cells , Prostatic Neoplasms/metabolism , Reactive Oxygen Species/metabolism , Receptors, TNF-Related Apoptosis-Inducing Ligand/metabolism , TNF-Related Apoptosis-Inducing Ligand/pharmacology , Tumor Suppressor Protein p53/metabolism , Xenograft Model Antitumor AssaysABSTRACT
Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a promising agent for anticancer therapy. The identification of small molecules that can establish the sensitivity of prostate cancer (PCa) cells to TRAIL-induced apoptosis is crucial for the targeted treatment of PCa. PC3, DU145, JAC-1, TsuPr1, and LNCaP cells were treated with Andrographolide (Andro) and TRAIL, and the apoptosis was measured using the Annexin V/PI double staining method. Real time-polymerase chain reaction (PCR) and Western blot analysis were performed to measure the expression levels of target molecules. RNA interference technique was used to down-regulate the expression of the target protein. We established a nude mouse xenograft model of PCa, which was used to measure the caspase-3 activity in the tumor cells using flow cytometry. In this research study, our results demonstrated that Andro preferentially increased the sensitivity of PCa cells to TRAIL-induced apoptosis at subtoxic concentrations, and the regulation mechanism was related to the up-regulation of DR4. In addition, it also increased the p53 expression and led to the generation of reactive oxygen species (ROS) in the cells. Further research revealed that the DR4 inhibition, p53 expression, and ROS generation can significantly reduce the apoptosis induced by the combination of TRAIL and Andro in PCa cells. In conclusion, Andro increases the sensitivity of PCa cells to TRAIL-induced apoptosis through the generation of ROS and up-regulation of p53 and then promotes PCa cell apoptosis associated with the activation of DR4.
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Objective@#To investigate the effect and intrinsic mechanism of Honokiol (HNK) enhanced tumor necrosis factor-related apoptosis inducing ligand (TRAIL) on hepatocellular carcinoma HepG2 cells.@*Methods@#HepG2 cells were routinely cultured. Firstly, a concentration gradient of HNK was given to observe its effect on the vitality of tumor cells. Western blot were used to detect change in the expression levels of c-jun N-terminal kinase (JNK), death receptor 4 (DR4), and DR5.Secondly, we observed the effect of combined drugs (HNK and TRAIL) on the vitality of tumor cells. Apoptosis-related protein expression levels were detected to determine the apoptosis condition. Thirdly, JNK inhibitor SP600125 was used to block the JNK pathway, and it was evaluated whether JNK signaling pathway mediated the DR4 and DR5 levels and finally, the subcutaneous tumor model of nude mice was constructed, and enhancement effect of HNK on TRAIL was confirmed in vivo.@*Results@#Cell vitality was decreased (P < 0.05) in a dose-dependent manner after treatment with gradient HNK. Combined effect of TRAIL and HNK was superior to that of single drug administration (P < 0.05). Western blot analysis showed that pJNK level increased after HNK treatment and TRAIL receptor DR4 and DR5 expression were up-regulated. Combined application of HNK and TRAIL, B lymphocyte tumor factor 2 (BCL2) decreased significantly while Bcl2 related X protein (Bax) increased significantly. Blocking JNK pathway by SP600125, the expression of DR4 and DR5 decreased (P < 0.05), Bax expression decreased and Bcl2 expression increased compared with HNK+TRAIL group. In vivo results showed that TRAIL inhibited the growth of subcutaneous tumors, and enhanced inhibition effect in combination with TRAIL and HNK.@*Conclusion@#HNK may enhance the inhibitory effect of TRAIL on HepG2 cells by activating JNK pathway and then upregulating the expression of DR4 and DR5.
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AIM:To investigate the effects of evodiamine on the growth and apoptosis of human hepatocellular carcinoma Huh7 cells,and to illustrate the molecular mechanism that evodiamine enhances antitumor activity of tumors nec -rosis factor-related apoptosis-inducing ligand(TRAIL)in Huh7 cells.METHODS: The cell viability was measured by MTT assay.The cell cycle distribution was analyzed by flow cytometry.The apoptosis rate was determined by TUNEL stai-ning.The protein levels of cell cycle-and apoptosis-related proteins were detected by Western blot analysis.RESULTS:Treatment of Huh7 cells with evodiamine reduced the cell viability(P<0.05).Evodiamine induced cell cycle arrest in G2/M phase by upregulation of p27,cyclin B1, cell division cycle protein 2(Cdc2)and p-Cdc2.Evodiamine triggered apoptosis accompanied by cleavage of caspase-3 and poly(ADP-ribose)polymerase(PARP).Combination of evodiamine with TRAIL significantly reduced the cell viability and increased cleavage of caspase -3 and PARP as compared with the use of each agent alone.Moreover,evodiamine increased the expression of death receptor 5(DR5)in the Huh7 cells.CON-CLUSION:Evodiamine inhibits the cell growth by reducing the cell viability and inducing cell cycle arrest.Evodiamine also triggers cell apoptosis and enhances the sensitivity of Huh 7 cells to TRAIL by upregulating the expression of DR5.
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Objective To explore the effect of chloroquine on death receptor 5 (DR5) expression of hepatocellular carcinoma Huh7 cells and cell proliferation and apoptosis induced by tumor necrosis factor related apoptosis-inducing ligand (TRAIL).Methods Huh7 cells were divided into four groups:the control group (1∶1 000 dimethyl sulfoxide),TRAIL group (50 μg/L),chloroquine group (10 μmol/L) and TRAIL +chloroquine group (TRAIL 50 μg/L + chloroquine 10 μmol/L).Thiazolyl blue tetrazolium bromide (MTT) assay was used to determine the proliferation activity of cells,immunofluorescence was used to detect the expression of DR5,4',6-diamidino-2-phenylindole (DAPI) staining was used to observe cell apoptosis and Western blot was used to detect the expression of cleaved poly ADP-ribose polymerase (PARP).Results TRAIL treatment could decrease Huh7 cells proliferation activity;when compared with the cell viability in the control group,the cell proliferation inhibition rate of chloroquine group,TRAIL group and TRAIL+ chloroquine group was (89±8) ％,(53±10) ％ and (27±7) ％,respectively;compared with TRAIL group alone,cell proliferation activity was decreased in TRAIL+ chloroquine group (t =3.922,P =0.017).The expression of DR5 was upregulated in chloroquine group,and the cell apoptosis signaling was activated in TRAIL + chloroquine group.The cell apoptosis rate of TRAIL group and TRAIL + chloroquine group was (10.0±2.3) ％ and (20.4±4.0) ％,respectively,and there was a statistical difference (t =3.894,P =0.018).Conclusion Chloroquine can enhance the cell chemosensitivity to TRAIL treatment by upregulating the expression of DR5 in Huh7 cells.
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BACKGROUND: Bile acids have anti-cancer properties in a certain types of cancers. We determined anticancer activity and its underlying molecular mechanism of ursodeoxycholic acid (UDCA) in human DU145 prostate cancer cells. METHODS: Cell viability was measured with an MTT assay. UDCA-induced apoptosis was determined with flow cytometric analysis. The expression levels of apoptosis-related signaling proteins were examined with Western blotting. RESULTS: UDCA treatment significantly inhibited cell growth of DU145 in a dose-dependent manner. It induced cellular shrinkage and cytoplasmic blebs and accumulated the cells with sub-G1 DNA contents. Moreover, UDCA activated caspase 8, suggesting that UDCA-induced apoptosis is associated with extrinsic pathway. Consistent to this finding, UDCA increased the expressions of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) receptor, death receptor 4 (DR4) and death receptor 5 (DR5), and TRAIL augmented the UDCA-induced cell death in DU145 cells. In addition, UDCA also increased the expressions of Bax and cytochrome c and decreased the expression of Bcl-xL in DU145 cells. This finding suggests that UDCA-induced apoptosis may be involved in intrinsic pathway. CONCLUSIONS: UDCA induces apoptosis via extrinsic pathway as well as intrinsic pathway in DU145 prostate cancer cells. UDCA may be a promising anti-cancer agent against prostate cancer.
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Humans , Apoptosis , Bile Acids and Salts , Blister , Blotting, Western , Caspase 8 , Cell Death , Cell Survival , Cytochromes c , Cytoplasm , DNA , Necrosis , Prostate , Prostatic Neoplasms , Receptors, TNF-Related Apoptosis-Inducing Ligand , Ursodeoxycholic AcidABSTRACT
Objective To explore the effect of inhibiting Akt phosphorylation on tumor necrosis factor related apoptosis inducing ligand (TRAIL)-induced apoptosis of human non-small cell lung cancer (NSCLC) H1299 cells with wild type EGFR and KRAS.Methods The TRAIL-induced apoptosis was examined by the Annexin V-FITC/PI.The expressions of TRAIL-activated Akt phosphorylation and p-Akt were measured by Western blot.After cells were treated with LY294002, an inhibitor of PI3K-Akt pathway, Annexin V-FITC/PI and Western blot were used to analyze the alteration of TRAIL-induced apoptosis and Akt phosphorylation, respectively.Results H1299 cells were not sensitive to TRAIL-induced apoptosis.When TRAIL concentration was 100 ng/ml, the apoptosis rate of the test group was significantly higher than that of the control group [(15.06±1.29) ％ vs (3.56±0.50) ％, t =66.953, P =0.000].When TRAIL concentration was 500 ng/ml, the difference was not statistically significant compared with apoptosis rate of 100 ng/ml TRAIL group [(18.65±2.09) ％ vs (15.06±1.29) ％, t =2.423, P =0.136].The expression level of Akt phosphorylation in H1299 cells was increased by TRAIL in a time-dependent way.When cells were pretreated with LY294002, TRAIL-induced Akt phosphorylation was suppressed to baseline level.At the same time, the apoptosis rate in LY294002-treated group was significantly higher than that in TRAIL group [(41.65±4.62) ％ vs (15.82±0.61) ％, t =39.028, P =0.001].Conclusions TRAIL-induced Akt phosphorylation can antagonize TRAIL-induced apoptosis.Inhibition of Akt phosphorylation can significantly enhance the sensitivity of NSCLC H1299 cells with wild type EGFR and KRAS to TRAIL-induced apoptosis.
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Background and purpose:The study hasfound that tumornecrosis factor-related apoptosis inducing ligand (TRAIL) can enhance the cytotoxic effect of chemotherapeutic drugs on tumor cells. The aim of our study was to investigate the effects of the tumor TRAIL and cisplatin combined application on the growth and apoptosis of human ovarian cancer cell lines SKOV3 and OVCAR3, and the possible mechanism.Methods:Under the combined application of cisplatin and TRAIL, MTT method and flow cytometry were used to detect the proliferation and apoptosis of SKOV3 of OVCAR3 cells; the mRNA expression levels of death receptors, DR4 and DR5, were detected by real-time lfuorescent quantitative polymerase chain reaction (RTFQ-PCR). At the same time, the protein expression levels of DR4 and DR5 were detected by Western blot.Results:SKOV3 and OVCAR3 cells were sensitive to TRAIL protein, and with the increasing of TRAIL protein concentration, cell growth inhibitory rate up to 64%. When the combination application of TRAIL and cisplatin, the inhibition rate of the two cells reached more than 92%, and the two drugs showed high synergistic effect, compared with the single drug group (P<0.05). Flowcytometry analysis indicated that the synergistic killing effect of TRAIL and cisplatin was mainly due to the cell apoptosis. RTFQ-PCR and Western blot detection results showed that the DR4 and DR5 were up-regulated under the combined application of TRAIL and cisplatin.Conclusion:In vitro, TRAIL and cisplatin combined application can signiifcantly inhibit the proliferation of human ovarian cancer cells and induce tumor cell apoptosis. TRAIL can obviously enhance the sensitivity of cisplatin to tumor cells. The mechanism may be related to the increased death receptor DR4 and DR5 expression level.
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Cancer is nowadays one of the main causes of death worldwide. The numbers have shown that one in three people will develop cancer at some point in their lives. Cancer is a major issue for the whole humanity, and therefore a lot of research has been running for the past years in order to understand the mechanisms that underlie tumorigenesis and eventually cancer formation, with the perspective to discover new approaches for effective treatment. Gene therapy strategies that intended to tackle cancer systemically are often impaired by inefficient delivery of the vector to the tumor site. Several studies have shown the possibility of using mesenchymal stem cells (MSCs) as a future therapeutic mechanism against cancer, since they possess important features such as the ability to home to and target cancer cells. The engineering of MSCs to produce and deliver an apoptotic factor, calledtumor necrosis factor-related apoptosis-inducing ligand (TRAIL) has been studied excessively. TRAIL is a transmembrane protein that causes selective apoptosis of tumor cells but does not have any harmful effects on the normal neighboring cells. Experiments have shown that this approach has significant results in mouse models and has now proceeded for clinical trials.
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AIM:To determine whether caudatin , a C21 steroidal aglycone , enhances tumor necrosis factor-re-lated apoptosis-inducing ligand ( TRAIL)-associated HepG2 cell apoptosis .METHODS:Cell growth inhibition was deter-mined by MTT assay and cell colony formation assay .The TUNEL apoptosis detection kit was used to analyze cell apopto-sis, and the protein expression was examined by Western blotting .RESULTS:Combination of caudatin with TRAIL signi-ficantly reduced cell proliferation and increased the apoptotic rate of HepG 2 cells compared with the use of each agent alone.This was evidenced by marked increases in caspase-3, caspase-7, caspase-9 and PARP cleavages in the cells treated with caudatin and TRAIL-compared with control group .Combination of caudatin with TRAIL also led to the strong suppres-sion of survivin .CONCLUSION:Caudatin synergizes HepG 2 cells to TRAIL-induced apoptosis by promoting the cleava-ges of caspase-3, caspase-7, caspase-9 and PARP and inhibiting the expression of survivin .
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[ ABSTRACT] AIM:To construct a prokaryotic expression plasmid to produce recombinant human tumor necrosis factor-related apoptosis-inducing ligand ( TRAIL) and to verify the biological activity of TRAIL.METHODS: The pro-karyotic expression plasmid pET-28a (+)-TRAIL114-281 was constructed.Human soluble TRAIL was obtained through opti-mized inducing protein expression and purification conditions.The biological activity of TRAIL was verified by CCK-8 as-say.The apoptosis-inducing effect of TRAIL alone and/or in combination with proteasome inhibitor bortezomib ( Velcade, PS-341) on the tumor cell lines H460 ( TRAIL-sensitive) and K562 ( TRAIL-resistance) for 24 h was determined.The ap-optotic rates of the cells were analyzed by flow cytometry with Annexin V-FITC/PI staining.The activities of caspase-8,-9 and -3 in the cells were detected by colorimetric method.The protein expression of Bax, Bcl-2 and cFLIP was measured by Western blot.The expression of DR4 and DR5 in the H460 cells and K562 cells after treated with bortezomib for 24 h was detected by flow cytometry.RESULTS:The recombinant human soluble TRAIL protein with stable bioactivity was success-fully acquired, which induced apoptosis in H460 cells and K562 cells.After treatment with different concentrations of TRAIL, the apoptotic rate of H460 cells was significantly increased with the increase in the concentration of TRAIL ( P<0.05), but the apoptotic rate of K562 cells was not affected by the increasing TRAIL concentration.Apoptotic rate in com-bination group was obviously higher than that in single group ( P<0.05 ) .In the process of apoptosis, the activities of caspase-8,-9 and -3 in H460 cells and K562 cells were both increased.The expression of Bcl-2 and cFLIP in treatment groups ( especially the combination group) was decreased compared with control group.No significant change of the Bax expression level was observed.The expression of DR4 and DR5 in the H460 cells and K562 cells was significantly up-regu-lated after treated with bortezomib ( P<0.05 ) .CONCLUSION: Bortezomib combined with recombinant human soluble TRAIL synergistically induces apoptosis in tumor cell lines H460 and K562 through initiating intrinsic apoptotic pathways by up-regulating death receptors DR4 and DR5, and reducing the expression of antiapoptotic proteins Bcl-2 and cFLIP.
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Objective To study the correlation between individual gene polymorphisms of transforming growth factor (TGF)-β1 + 869 T/C,tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) + 1525 G/A genes and nodular thyroid disease.Methods From September 2007 to December 2009,a total of 544 patients with nodular thyroid disease diagnosed in the Department of Endocrinology,The First Affiliated Hospital of Baotou Medical College,Inner Mongolia University of Science and Technology were selected,including 136 cases of nodular goiter patients (node group),132 cases of thyroid tumor (adenoma group),146 cases of Graves patients (GD group),and 130 cases of Hashimoto's thyroiditis (HT group).One hundred and thirty-five healthy subjects were enrolled as control group.Two milliliters of fasting venous blood of all subjects were collected.Polymorphisms of the TGF-β1 + 869 T/C and the TRAIL 1525 A/G genes were identified by the polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) and the restriction fragment length polymorphism (PCR-RFLP) methods.Results TGF-β1 + 869 T/C:The CC genotypes and C allele frequencies of nodular goiter group [47.0％(64/136),63.2％(172/ 272)] were significantly higher than those of normal control group [18.0％(22/135),45.2％ (122/270); x2 =30.76,17.79,all P ＜ 0.05].The genotypes and allele frequencies of adenoma group[42.4％ (56/132),59.1％ (156/264)] were significantly higher than those of the normal control group (x2 =24.40,10.34,all P ＜ 0.05).The risk of population carrying the C allele suffering from nodular goiter was 2.086 times of those carrying the T allele (OR =2.086; 95％ CI:1.480-2.943).The risk of population carrying the C allele suffering from adenoma was 1.752 times of those carrying the T allele (OR =1.752,95％ CI:1.244-2.469).TRAIL + 1525 G/A:the genotypes and allele frequencies of nodular goiter group [40.4％ (55/136),62.9％ (171/272)] were significantly higher than those of normal control group [12.0％ (16/135),48.5％ (131/270); x2 =9.176,11.307,all P ＜ 0.05].The genotypes and allele frequencies of adenoma group[53.3％ (70/132),73.1％ (193/264)] were significantly higher than those of the normal control group (x2 =9.806,33.82,all P ＜ 0.05).The risk of population carrying the G allele suffering from nodular goiter was 1.796 times of those carrying the A allele (OR =1.796,95％ CI:1.275-2.531).The risk of population carrying the G allele suffering from adenoma was 2.884 times of those carrying the A allele (OR =2.884,95％ CI:2.009-4.142).Conclusions TGF-β1 + 869 T/C and TRAIL + 1525 G/A gene polymorphisms may be related to the incidence of nodular thyroid diseases; G allele of TRAIL and C allele of TGF-β1 may be predisposing genes of patients with nodular goiter.