Antitumor Effect And Mechanism Of GCPS On Tumor In Gastrointestinal Tract

OBJECTIVE: To extract and purify GCPs from GAE and to investigate theanti-tumor effects and the mechanisms of GCPs in vitro and in vivo.METHODS:（1）GCPs was analysised and purified by Dialysis assay, HPLC andTrincene-SDS-PAGE assays.（2）The morphological changes of esophageal EC109,liver cancer cells HepG2and stomach cancer cells MGC803were observed withinverted microscope in different concentration (3mg/mL,1.5mg/mL,1mg/mL、0.75mg/mL,0.375mg/mL) of the GCPs, and MTT assay was used to investigate theinhibitory effect of proliferation on the three gastrointestinal tract tumor cells line.Hoechst33258fluorescence staining was used to identify the nucleus apoptotic changeof HepG2treated with GCPs. The protein expression of bax and bcl-2were detectedwith western-blot assay.（3）A xenograft H22liver cancer model was established inKunming mice. The tumor-bearing mice were treated with daily intraperitonealinjections of normal saline (NS group) or GCPs (80,40or20mg/kg) for10days, oronce per two days with2mg/kg doxorubicin (ADR group, n=10each）. Serum tumornecrosis factor (TNF-α) and interleukin (IL)-6were quantified using ELISA assay.RESULTS:（1）Gecko Crude Peptides（GCPs）were analysised throughHPLC and Trincene-SDS-PAGE.（2）The some morphological changes of the threegastrointestinal tract tumor cells, treated with different concentration (3mg/mL,1.5mg/mL,1mg/mL、0.75mg/mL,0.375mg/mL) of the GCPs were shown withinverted microscope after24h,48h,72h. GCPs inhibited the proliferation of the tumorcells in a concentration and time-dependent manner. EC109cells, HepG2cells andMGC803cells were treated GCPs for48h, the IC50of GCPs was1.1mg/mL,1.2mg/mL and1.6mg/mL，respectively.（3）HepG2treated with GCPs showedapoptotic morphological changes that dense and strong blue fluorescence mass can beseen under fluorescence microscope；GCPs（1.6mg/mL，0.8mg/mL）decreasedthe expression of bcl-2protein, and increase the expression of bax protein.（4）The weight of the tumors (0.98±0.60g) were smaller (2.20±0.45g; P <0.01) while thewhite blood cell count, thymus index, spleen index were higher in the high dose GCPsgroup than the NS group (P <0.05), the VEGF expression in tumor tissue, TNF-α andIL-6levels in serum of the GCPs mice were lower than the NS mice (P <0.05).CONCLUSION: GCPs exert anti-tumor effects in vitro and in vivo by inducingapoptosis, increasing the bcl-2/bax ratio, downregulating VEGF and reducing serumlevels of the inflammatory factors TNF-α and IL-6.