The Roles And Underlying Mechanisms Of TASK-3 Channel In The Different Effects Of Anesthetics On Growth And Metastasis Of Ovarian Cancer Cells

Part1.Effects of anesthetics on cell growth and metastasis of ovarian cancerObjective :To examine the effects of anesthetics on cell proliferation,cell cycle and apoptosis rate,cell migration and invasion of SKOV-3 in vitro.And further to examine the effects of anesthetics on ovarian cancer growth and lung metastasis in vivo.Methods: CCK-8 and flow cytometry were used to examine the effects of anesthetics(lidocaine,sevoflurane and propofol)on cell proliferation,cell cycle and apoptosis.The effects of local anesthetics and inhalation anesthetics on cell migration and invasion ability of SKOV-3 were examined by Transwell assay.The effects of intraperitoneal injection of lidocaine and inhalation of sevoflurane on tumor growth after 27 days were observed.Lung metastasis model of ovarian cancer was established by injecting SKOV-3 via the tail vein of nude mice,and the lungs were taken for HE staining after 27 days of treatment with lidocaine and sevoflurane,and the effects of drugs on lung metastasis were observed under the microscope.Results:Lidocaine treatment significantly decreased cell proliferation,caused G2 phase arrest and increased the apoptosis rate of SKOV-3 in a concentration-dependent manner.However,after sevoflurane treatment,cell viability and the G2 phase rate of SKOV-3 were increased,and the apoptosis rate of SKOV-3 was decreased.The cell viability of SKOV-3 only decreased when the concentration of propofol was above 100?M.TASK-3 channel is upregulated in ovarian cancer compared with normal ovarian tissue,and TASK-3 channel is highly expressed in SKOV-3.Lidocaine treatment significantly decreased cell proliferation,caused G2 phase arrest and increased the apoptosis rate of SKOV-3 in a concentration-dependent manner.However,after treatment of sevoflurane,cell viability and The G2 phase rate of SKOV-3 were increased,and the apoptosis rate of SKOV-3 was decreased.The cell viability of SKOV-3 only decreased when the concentration of propofol was above 100?M.Lidocaine markedly decreased cell migration and invasion of SKOV-3,however,treatment of sevoflurane can significantly increased cell invasion and migration ofSKOV-3.Similarly,lidocaine can significantly inhibited ovarian cancer growth rate and lung metastasis in vivo,sevoflurane can increased ovarian cancer growth and lung metastasis.Conclusions: lidocaine decreased cell viability,caused G2 phase arrest and increased the apoptosis rate of SKOV-3 in a concentration-dependent manner.And lidocaine can markedly increased cell migration and invasion of SKOV-3.Treatment of sevoflurane can increased cell viability and the G2 phase rate and decreased apoptosis rate,and significantly increased cell invasion and migration of SKOV-3.Lidocaine can significantly inhibited ovarian cancer growth and lung metastasis in vivo,sevoflurane can increased ovarian cancer growth and lung metastasis.Part2.TASK-3 channel contributes to the effects of anesthetics on cell growth and metastasis of ovarian cancer cellObjectives: To examine the expression of TASK-3 channel in normal ovarian tissue and ovarian cancer and SKOV-3 cells,and to unveil the role of TASK-3 channel in effects of lidocaine and sevoflurane on apoptosis,cell migration,invasion of SKOV-3 in vitro.And to study the role of TASK-3 channel in effects of lidocaine and sevoflurane on ovarian cancer growth and metastasis in vivo.Methods: Samples of normal ovarian tissues and ovarian cancer were collected from the Department of gynecology and obstetrics of Union hospital.And culturing Human normal ovarian cell line IOSE-80 and Human ovarian cancer cell line SKOV-3.The mRNA and protein expressions of TASK-3 channel were detected by RT-PCR and Western blot.TASK-3 shRNA lentivirus were constructed and transfected in SKOV-3,and TASK-3 shRNA lentiviru transfected stably SKOV-3 was screened.Effects of lidocaine and sevoflurane on apoptosis rate were examined by flow cytometer.Effects of these drugs on cell migration and invasion were demonstrated by Transwell assay.To establish xenograft model of ovarian cancer in female nude mice,TASK-3 shRNA lentivirus transfected stably SKOV-3 and SKOV-3 cells were subcutaneously injected into the right side of nude mice,and assess the tumor growth after treatment of lidocaine and sevoflurane(about 27 days after treatment).And TASK-3 shRNA lentivirus transfected stably SKOV-3 and SKOV-3 cells were intravenously injected via tail vein.After 27 days,the lung was H&E staining to study the effects of lidocaine and sevoflurane on tumor lung metastasis in vivo.Results: TASK-3 channel is upregulated in ovarian cancer compared with normal ovarian tissue.Similarly,TASK-3 channel is highly expressed in human ovarian cancer cell line SKOV-3.The expression of TASK-3 channel was significantly decreased in TASK-3 shRNA lentivirus transfected stably SKOV-3.TASK-3 shRNA caused significant decrease in cell viability.Treatment of lidocaine and TASK-3 shRNA can increased the apoptosis rate and decreased cell migration and invasion of SKOV-3.TASK-3 shRNA pretreatment can attenuated these effects of lidocaine.Treatment of sevoflurane can decreased the apoptosis rate and increased cell migration and invasion of SKOV-3.And TASK-3 shRNA pretreatment can attenuated the effects of sevoflurane.By compared with non-treated SKOV-3,treatment of TASK-3 shRNA can decreased the growth rate and lung metastasis number and metastasis area of ovarian cancer cell.Similarly,lidocaine can significantly inhibited ovarian cancer growth rate and lung metastasis in vivo,sevoflurane can increased ovarian cancer growth and lung metastasis,and TASK-3 shRNA pretreatment can attenuated the effects of lidocaine and sevoflurane on ovarian cancer growth and lung metastasis.Conclusions: TASK-3 channel is upregulated in ovarian cancer.In vitro,treatment of lidocaine and TASK-3 shRNA can increased the apoptosis rate and decreased cell migration and invasion of SKOV-3.Treatment of sevoflurane can decreased the apoptosis rate and increased cell migration and invasion of SKOV-3.TASK-3 shRNA pretreatment can attenuated these effects of lidocaine and sevoflurane.In vivo,lidocaine can significantly inhibited ovarian cancer growth rate and lung metastasis sevoflurane can increased ovarian cancer growth and lung metastasis,and TASK-3 shRNA pretreatment can attenuated the effects of lidocaine and sevoflurane.Part3.TASK-3 channels contribute to the effects of anesthetics on apoptosis of ovarian cancer cellObjectives: To examine the subcellular location of TASK-3 channel in SKOV-3,and to unveil the role of TASK-3 channel in the effects of anesthetics on mitochondrial membrane potential and the downstream apoptosis signaling of SKOV-3Methods: The subcellular expression of TASK-3 channel was examined by immunofluorescence.Effects of lidocaine and sevoflurane on mitochondrial membrane potential,cytochrome C were examined by flow cytometer and western blot.Results: TASK-3 channel is expressed in cell membrane,mitochondrion and nucleus,highly expressed in mitochondrion.Treatment of lidocaine and TASK-3 shRNA can decreased mitochondrial membrane potential,caused cytochrome c leak in cytoplasm,Thus decreased the expression of cytochrome c in mitochondrial,increased the expression of cytochrome c in cytoplasm.Lidocaine markedly decreased cell migration and invasion of SKOV-3.TASK-3 shRNA pretreatment can attenuated these effects of lidocaine.Conclusions:TASK-3 channel is highly expressed in mitochondrion of SKOV-3.Lidocaine and TASK-3 shRNA can decreased mitochondrial membrane potential,caused cytochrome c leak in cytoplasm,Thus decreased the expression of cytochrome C in mitochondrial,increased the expression of cytochrome c in cytoplasm.