SMAC exhibits anti-tumor effects in ECA109 cells by regulating expression of inhibitor of apoptosis protein family

BACKGROUND The poor prognosis and rising incidence of esophageal cancer highlight the need for improved therapeutics that are essential prior to treatment.LCL161 is an SMAC(second mitochondrial activator of caspases)mimic and inhibitor of apoptosis protein(IAP)antagonist which exhibits anti-tumor effects and improves the chemical sensitivity of many cancers.AIM To ascertain the effects and mechanisms of the SMAC analog LCL161 on esophageal cancer cells.METHODS MTT assay and TUNEL assay were used to detect cell proliferation and apoptosis,respectively.Western blot analysis was used to study the molecular mechanisms of LCL161-induced death of ECA109 cells.RESULTS LCL161 decreased ECA109 cell proliferation in dose-and time-dependent manner and induced apoptosis of ECA109 cells in a dose-dependent manner.Also,LCL161 induced a significant decrease in the expression of the XIAP and significant increase in the expression of Caspase-3.In addition,Bax increased significantly with increasing concentrations of LCL161,and the relative expression of Bax was significantly different between groups.CONCLUSION These findings support the hypothesis that LCL161 can inhibit proliferation and induce apoptosis in esophageal cancer cells by regulating the expression of IAP family members,suggesting that it has potential to be an effective treatment for esophageal squamous cell carcinoma.

Tonglian Decoction(通莲汤)Arrests the Cell Cycle in S-phase by Targeting the Nuclear Factor-Kappa B Signal Pathway in Esophageal Carcinoma Eca109 Cells

Objective： To investigate the anti-tumor activity and molecular mechanism of Tonglian Decoction （通莲汤, TLD） on esophageal carcinoma Eca109 cells. Methods： Ecal09 cells were treated with TLD and its separated formulae, including the clearing-heat and detoxification formula （Q）, activating-blood and promoting-qi formula （H） and nourishing-yin and blood formula （Z）. Cell proliferation was measured using the 3-（4,5-dimethyl-2- thiazolyl）-2,5-diphenyl-2-H-tetrazolium bromide assay, cell morphology was observed using a microscope, the cell cycle was measured using flow cytometry and the activity of the nuclear factor-kappa B （NF- K B） signal pathway was detected by Western blot. Results： The half maximal inhibitory concentrations of TLD, Q and H were 386, 771 and 729 rag/L, respectively. TLD, Q and H significantly inhibited cell proliferation, with 69.43%, 60.84% and 61.90% of treated cells in the G1 phase of the cell cycle. The percentage of cells in S phase increased significantly after treatment with TLD, Q, and H compared with the control group （P〈0.05）, and TLD showed the strongest effect. Z had no influence on the cell cycle compared with the control group （P〉0.05）. Western blot detection indicated slight differences in the inhibition of the NF- K B pathway by the different formulae. TLD formula strongly inhibited IKK β, NF- K B, interleukin-6 and tumor necrosis factor-α expression compared with the control group. Conclusions： TLD inhibited Eca109 cell proliferation by arresting cells in S phase. The possible mechanism might be related to inhibiting the NF- K B transduction cascade. The combination of the herbs found in the three separate formulae, H, Q and Z, work synergistically in TLD to produce the inhibitory effects of TLD treatment on Eca109 proliferation.

Wilfoside C3N Promotes Tumor Cell Death by Activating Gammadelta T Cells-Mediated Anti-tumor Immunity

Vδ1^+γδ T lymphocytes are known to play important roles in anti-tumor immunity.We recently reported an anti-tumor activity of wilfoside C3 N,an active component extracted from Chinese medicinal herbs.In the current study,we evaluated the role of Vδ1^+γδ T cells in C3 N anti-tumor activity using an in vitro cell co-culture model.We found that C3 N induced the ECA109 tumor cells to undergo apoptosis in the presence of Vδ1^+γδ T cells.The level of ECA109 apoptosis maximized when both C3 N and Vδ1 + γδ T cells were present,which correlated with the increased expression of Fas on ECA109 and Fas ligand on Vδ1^+γδ T cells induced by C3 N.In addition,C3 N also enhanced secretion of cytokines,perforin and granzymes by Vδ1^+γδ T cells.These observations suggest that activation of Vδ1^+ γδ T cells may play a critical role in C3N-mediated anti-tumor activity.

Impact of resveratrol on the expression of apoptosis related gene survivin and bax in human cancer cells

Objective： We explored the mechanism of apoptosis in human esophageal cancer Ecal09 cells by resveratrol. Methods： The suppressive ratio of resveratrol on Ecal09 cells proliferation was evaluated by MTT colorimetric assay and morphology was observed by transmission electron microscope. The expression of survivin and bax was analyzed by RT-PCR and Flow Cytometry （FCM）. Results： Resveratrol inhibited the growth of Ecal09 calls in a dose-and time-dependent man- ner, and the suppressive ratio arrived at 76.42%. Morphological apoptosis could be observed after treated with resveratrol.The bulk of some drug-treated cells turned small and the nuclear chromatin became condensed and rnarginated. The results determined by RT-PCR and FCM showed that resveratrol could down-regulate surviving, while up-regulate bax. Conclusion： Resveratrol could induce the apoptosis of human esophageal cancer Ecal09 cells, and its possible molecular mechanisms might be related to modulation the expression of survivin and bax.

Role of stress-activated MAP kinase P38 in cisplatin-and DTT-induced apoptosis of the esophageal carcinoma cell line Eca109

AIM： To study the role of P38 kinase in esophageal cancer cell apoptosis induced by genotoxin, cisplatin and the unfolded protein response （UPR） inducer, dithiothreitol （DTT）. METHODS： Esophageal carcinoma cell line Eca109 was cultured in RPMI 1640 medium to 70% confluency and treated with either cisplatin, DTT, or cisplatin plus DTT in the presence or absence of P38 inhibitor, SB203580. The untreated cells served as the control. The esophageal carcinoma cell apoptosis was detected by agarose gel DNA ladder analysis and quantified by flow cytometry. The P38 phosphorylation was detected by immunohistochemistry using antibodies specific to phosphorylated P38 protein. RESULTS： （1） Both cisplatin and DTF induced apoptosis in the esophageal cancer cell line Eca109 as shown by DNA ladder formation; （2） As detected by antibodies specific for the phosphorylated P38 protein （p-P38）, both cisplatin and DTT treatments activated the stress-activated enzyme, MAP kinase P38. The number of positive cells was about 50% for the treatment groups, comparing to that of 10% for untreated group. DTF treatment, but not cisplatin treatment, induces nuclear localization of p-P38; （3） As measured by flow cytometry, inhibition of P38 activity by SB203580 blocks DTT- and cisplatin-induced apoptosis. The rates for DTT, cisplatin, and DTT plus cisplatin-induced apoptosis were 16.8%, 17.1%, and 21.4%, respectively. Addition of the SB compound during the incubation reduced the apoptotic rate to about 7.6% for all the treatment groups, suggesting that P38 activation is essential for cisplatin- and DTT-induced apoptosis in Eca109 cells. CONCLUSION： （1） Both DTT and cisplatin were able to induce apoptosis in esophageal cancer cell line Eca109; （2） P38 MAP kinase is essential for DTT- and cisplatininduced apoptosis in Eca109 cells; （3） P38 activation may be the common signaling component relaying the multiple upstream signaling events to the downstream cell death program.

EXPRESSION OF PLACENTAL ALKALINE PHOSPHATASE IN ESOPHAGEAL CANCER CELL LINE Eca109

The expression and properties of alkaline phosphatase (ALP) in Eca109 cells, a cell line derived fromhuman esophageal cancer were studied with specific inhibition assay and polyacrylamide gel electrophoresis.The results showed that ALP of Eca109 cells was heat stable and was strongly inhibited by L-pheuylalanine, but slightly inhibited by urea. Preduisolone could causedramatic increase in activity of ALP, but no change in ALP isozyme and concomitant increase in lactic dehydrogenase activity were found after prednisolone treatment. The results suggested that placental alkaline phosphatase as an oncodevelopmental gene product could be expressed ectopically by Eca109 cells and prednisolone could specifically induce increase in its activity.