The Mechanism Of Endoplasmic Reticulum Of Nanosecond Steep Pulse Induced Apoptosis In Human Ovarian Cancer Cell SKOV3

Following the reversible electrical breakdown are adjuvant to chemotherapy drug therapy for cancer and irreversible electrical breakdown directly kill cancer cells, The newly developed a foundation for treatment are nanosecond Steep pulse. It is not damaging cell membranes but to make the nucleus, mitochondria, endoplasmic reticulum etc organelles appear a series of functional change and change of signal transduction , inducing apoptosis of cell.At present, the study of nanosecond Steep pulse induce the apoptosis are simple, such as phosphatidyl serine (phosphatidylserine, PS) evaginate, the change of mitochondrial membrane potentialld and the intracellular free Ca²⁺, the evidences of the morphology etc, the apoptosis pathways never be systematic and in-deph searched,We find the best treatment parameters through the Annexin V/PI double dyeing technology testing cell apoptosis rate of different parameters group after steep pulse processing , determining the existence of apoptosis through DNA Ladde, proving Endoplasmic reticulum pathway of cell apoptosis Through the caspase12 protein, Chelating the increased intracellular free Ca²⁺ because of the effect of nanosecond steep pulse, then detect the rate of cell apoptosis and survival rate, the change of expression of caspase12 protein. Deeply discuss the endoplasmic reticulum mechanism of oarian cancer cell apoptosis inducted by nanosecond steep pulse, for the clinical use of nanosecond Steep pulse therapy of ovarian cancer provides new theoretical basis.part I:nanosecond Steep pulse induced apoptosis in human ovarian cancer cell SKOV3 and determine the best parametersObjective:To investigate the effect of nanosecond Steep pulse induced apoptosis in human ovarian cancer cell SKOV3and determined the best parameters.Methods:Annexin V / PI double staining flow cytometry detection of apoptosis of different parameters group treated with nanosecond steep pulse, and find the best treatment parameters.; gettingthe DNA bands though Agarose gel electrophoresis. cells were treated with the nanosecond steep pulse of the best treatment parameters .caspase12 protein level were detected by Western blotResults:Flow cytometry displayed that nanosecond steep pulse induced early apoptosis.the best treatment parameters: 90kV/cm,100ns,1Hz ,30s, Its early apoptotic rate was(64.01±5.07)% necrosis rat was (0.71±0.37)%. Agarose gel electrophoresis showed a multiple of 180-200bp DNA ladder, the expressions of caspase-12 were increased after 2,6,12hours (P<0.05),Conclusion:Steep nanosecond pulse significantly induced apoptosis in SKOV3 cells, and caused at least the endoplasmic reticulum pathwayPART II:The mechanism of endoplasmic reticulum of nanosecond Steep pulse induced apoptosis in human ovarian cancer cellObjective: Ca²⁺ is the role of ndoplasmic reticulum of apoptosis pathway in nanosecond steep pulse induced e human ovarian cancer cells.Methods:Different concentration of BAPTA-AM (0μ,25μ,50μ,100μmol/L )chelation of intracellular increase of free Ca²⁺,then cell viability were detected by MTT, Morphologic analysis of NSP-Induced cell by Hoechst 33342, Expression of caspase12 of NSP-Induced cell .Results:Cell viability were increased by BAPTA-AM chelation of intracellular Increase of free Ca²⁺(P<0.05), Apoptosis rate and the expressions of caspase—12 were decreased by BAPTA-AM chelation of intracellular Increase of free Ca²⁺(P<0.05).Conclusion: By antagonizing the intracellular free Ca²⁺ increase, inhibit human ovarian cancer cells SKOV3 apoptosis that induced by the nanosecond steep pulse. Increased intracellular Ca²⁺ and may mediate the endoplasmic reticulum apoptosis pathway is one of the mechanisms of ovarian cancer cells apoptosis induced by nanosecond pulses steep...