Effect Of Piper Nigrum’s Active Constituent (Dmf) On Self-renewal Capability And Expression Of Twist In Ocslcs Derived From Ovarian Cancer Skov3Cell Line

Objective:To investigate the effect of the piper nigrum’s active constituent5,7-dimethoxyflavone(DMF) on self-renewal capability, proliferation and expression of Twist protein in ovarian cancer sphere forming cells (OCSLCs) derived from human ovarian cancer SKOV3cell line, and explore the mechanisms underlying piper nigrum’s active constituent (DMF) inhibits ovarian cancer stem cells characteristics.Methods:Human ovarian cancer was cultured in vitro. Used the suspension culture with stem cell-conditioned medium to enrich and gather the Ovarian cancer sphere-forming cells and identified as OCSLCs. Self-renewal capability of OCSLCs was detected by pneumonosphere formation assay. The expression of stem biomarkers CD133and CD44in ovarian cancer SKOV3shpere-forming cells and nonsorted ovarian cancer cell were analyzed by Western blot. We established ovarian cancer transplantation tumor model in nude mice to estimate the tumorigensis capacity of human ovarian cancer sphere-forming cells and parental SKOV3cell line;and the transplation tumor tissues were Preserved, routine hematoxylin-eosin staning, pathological and morphological were detected by optical microscope. The MTT assay was used to measure the effect of piper nigrum’s active constituent (DMF) on proliferation of OCSLCs. We used the Western blot to compare the expression of Twist protein in SKOV3sphere-forming cells and parental SKOV3cells, and the effect of piper nigrum’s active constituent (DMF) on expression of Twist protein in OCSLCs.Results:1. The ovarian cancer sphere cells derived from human ovarian cancer SKOV3cell line which was cultured in stem cell-conditioned medium condition growing into a sphere.2. Pneumonosp here formation assay showed sphere-forming cells of human ovarian cancer SKOV3cells has self-renewal capability.3. The expression level of stem biomarkers CD133and CD44protein were higher in ovarian cancer sphere-forming cells derived from SKOV3cells than parental SKOV3cells.4. The results of ovarian cancer transplantation tumor experiment in nude mice demonstrated ovarian cancer sphere-forming cells derived from SKOV3cells have high tumorigensis capacity.5. MTT assay proved that piper nigrum’s active constituent (DMF) could inhibit cell proliferation activity of OCSLCs.6. Piper nigrum’s active constituent (DMF) could inhibit tumorigenic ability of OCSLC.7. Piper nigrum’s active constituent (DMF) could inhibit self-renewal capability of OCSLCs.8. Piper nigrum’s active constituent (DMF) could inhibit expression of Twist protein with the dose-dependent manner. Conclusion:1. Ovarian cancer sphere-forming cells derived from ovarian cancer SKOV3cells which was cultured in stem cell-conditioned medium have cancer stem cell characteristics.2. Piper nigrum’s active constituent (DMF) can inhibit self-renewal capability, proliferation activity and tumorigenic ability of OCSLCs.3. The self-renewal inhibited role of piper nigrum’s active constituent (DMF) of OCSLCs was associated with down-regulating expression cancer signaling molecule Twist.4. Piper nigrum’s active constituent (DMF) regulated the expression level of Twist protein in SKOV3ovarian cancer stem like cells which can affect its cancer stem cells property; it maybe a potential approach for curing the ovarian cancer focusing on the key pathological factors "phlegm" and "hemostasis".