[Apoptosis and growth arrest of human esophageal squamous cell carcinoma cell EC9706 induced by Fufangkushen injection].
OBJECTIVE
To investigate the inhibitory effect and apoptosis induction on human esophageal carcinoma EC9706 cell by Fufangkushen.


METHODS
The experiment of Fufangkushen was designed into three groups including 25.00 µl/ml group, 6.25 µl/ml group and control group in vitro. The method of MTT was used to evaluate the growth inhibition effects. Proliferating cell nuclear antigen (PCNA) was detected by immunohistochemistry (IHC) in vitro. The morphological changes of cells were observed under inverted microscope. FACS was used to analyze the distribution of cell cycle and apoptosis. The expressions of Bcl-2, Fas and caspase-3 in EC9706 cells were detected by Western blotting. The clone formation in plate was used to test the capacity of cell clone formation. Nude mice experiments were conducted to investigate the tumor inhibition of Fufangkushen in vivo. The mice were divided into 3 groups of 200 µl/d treatment, 25 µl/d treatment and saline control. PCNA and Bcl-2 were detected by IHC. And the apoptotic index was detected by terminal transferase dUTP nick end labeling (TUNEL) on xenograft of nude mice.


RESULTS
The proliferative capacities of 25.00 µl/ml group were lower than that of the control group at 48, 72, 96 h respectively (all P < 0.01). IHC showed the PCNA expressions, cell clone formation rate were both lower than that of control group (in 25.00 µl/ml treatment group both P < 0.05). Many apoptotic cells could be observed. And the apoptotic rate was higher in 25.00 µl/ml group than that in the control group ((25.2 ± 7.3)% vs (3.4 ± 1.5)%, P < 0.01). After a treatment of Fufangkushen, the activation of caspase-3 and the Fas were higher ((21.3 ± 4.4)% vs (1.8 ± 0.6)%, (30.2 ± 8.3)% vs (5.4 ± 1.6)%, both P < 0.01), the Bcl-2 were lower (P < 0.01) were observed in vitro. Comparing with the saline control group, the tumor weight in 200 µl/d treatment group were lower ((987 ± 386) vs (1935 ± 838) mg, P < 0.01) and the apoptotic index higher ((33.8 ± 8.7)% vs (5.3 ± 1.4)%, P < 0.01).


CONCLUSIONS
Fufangkushen can inhibit the proliferation of EC9706 cells and induce the cellular apoptosis. The mechanism of apoptosis is probably associated with the arrest of cell cycle, the up-regulation of Fas, the down-regulation of Bcl-2 and the activation of caspase-3 in ESCC EC9706 cells.
Department of Oncology, First Affiliated Hospital, Zhengzhou University, Zhengzhou, China.
Ai-lian
AL
Zhu
Feng
F
Wang
Qing-xia
QX
Fan
Wei
W
He
Liu-xing
LX
Wang
Pei-rong
PR
Zhao
D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D002294:Carcinoma, Squamous Cell / Q000473:pathology*; D053148:Caspase 3 / Q000378:metabolism; D045744:Cell Line, Tumor; D004365:Drugs, Chinese Herbal / Q000494:pharmacology*; D004938:Esophageal Neoplasms / Q000473:pathology*; D006801:Humans; D051379:Mice; D008819:Mice, Nude; D009368:Neoplasm Transplantation; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D019014:fas Receptor / Q000378:metabolism
D004365:Drugs, Chinese Herbal; C491901:FAS protein, human; D019253:Proto-Oncogene Proteins c-bcl-2; D019014:fas Receptor; C574916:fufangkushen; C505407:CASP3 protein, human; D053148:Caspase 3