In Vitro Antiproliferative Activity Of Propolis Flavonoids PB3A On Human Cancer Cell Lines

Objective: Natural product of Propolis Flavonoids exhibits a broad spectrum of biological activity, its extraction of Pinobanksin-3-acetate(PB3A) significantly inhibited the proliferation and induced apoptosis of colon cancer cells and leukemia cells. However, its effects on other tumor cells remain unclear so far. In this study, the effects of PB3 A on several tumor cell lines and normal liver L02 cell lines was tested. Meanwhile, screening the sensitive cell lines to PB3 A. Further find out key genes which PB3 A effected on. So that, we could provide experimental and theoretical basis for the development of anti-tumor drugs.Our study divided in to two parts 1. In vitro anti-proliferative activity of PB3 A against the six tested cancer cell lines and the and normal cell linesMorphological changes of lung cancer A549, Liver cancer HepG-2, cervical cancer Hela, breast cancer MDA-MB-231, eso phageal cancer EC-109, gastric carcinoma SGC-7901 cells and normal liverL02 cells were observed by inverted photo microscopy after treatment with different concentrations(20, 40, 80μg/mL) of PB3 A for different durations. the effect of different concentrations(10, 20, 40 and 80μg/mL) of PB3 A for different durations(24, 48, 72h) on proliferation of six cancer(A549, HepG-2, EC-109, Hela, MDA-MB-231, SGC 7901) cells and normal(liver L02) cells was analyzed by MTT assay. PB3 A provided the highest in vitro anti-proliferative activity against the six tested cancer cell lines and the low cytotoxicity against the normal cell lines in both time and dose-dependent manner. By contrast, PB3 A highly inhibited digestive system tumor cells, and revealed relatively high inhibition against lung cancer cells and cervical cancer cells. However, breast cancer cells MDA-MB-231 showed definitely high drug resistance of PB3 A. the 24 h apoptotic rate of six tumor cell lines and normal cell induced with PB3 A and cisplatinum were detected by FCM with Annexin V-FITC/PI double labeling.FCM analysis showed that(20, 40 and 80μg/mL) PB3 A could significantly increase the apoptotic rate of A549, HepG-2, EC-109, Hela, MDA-MB-231 and SGC-7901 cells, and suggesting a dose-dependent manner. The regression analysis of Six tumor cell lines. in apoptotic rate suggested that the inducing apoptosis of PB3 A on tumor cell lines of digestive system was obvious, PB3 A induced apoptosis of gastric cancer SGC-7901 cells was highly significant. 2.protein immunoblot(Western Blot)assay in gastric cancer SGC-7901 cells.by usingWestern Blot assay to detect the protein expression level of(heat shock 70 kDa protein,HSPA6), 45(growth arrest and DNA damage inducible protein 45,Gadd45),(Regulator of G-protein signaling 2,RGS2),(GTP binding protein over expressed in skeletal muscle,GEM) and FOS genes, β-actin as a loading control, to compare the differences of protein expression in the intervention groups and control group. The experimental results of western blottting showed that after intervention of gastric cancer SGC-7901 cells with 40μg/mL and 80μg/mL concentration of PB3 A, could induce up regulation of GADD45 G, HSPA6, GEM, RGSR and FOS protein expression. To comparing with the control group, in the 40μg/mL treatment group with PB3 A, GADD45 G, GEM and HSPA6 protein expression intensity were relatively significant(P<0.05). However, in the 80μg/mL treatment group, all the(GADD45G, HSPA6, GEM, RGSR and FOS) protein expression level had highly significant(P<0.01).