[Effects of Brucea javanica oil emulsion on human papilloma virus type 16 infected cells and mechanisms research].
OBJECTIVE
To explore the in vitro inhibitive effect and underlying mechanisms of Brucea Javanica oil emulsion (BJOE) on human papilloma virus (HPV) type 16 infected cells.


METHODS
The HPV16 E61E7 immortalized human ectocervical Ect1/E6E7 cell line and the CaSki cell line were selected as the in vitro models of premalignant cervical lesion and cervical cancer respectively. After treated with BJOE at different concentrations (5, 10, 20, and 40 microg/mL) at the operation time points (24, 48, and 72 h), the effects of BJOE on proliferative activities were measured by MTT assay. The morphologic changes of cell apoptosis stained with Hochest 33,258 were observed by fluorescence microscope. The effect on the cell apoptosis rate was analyzed by Annexin V-FITC/PI double-labeled flow cytometry. The mRNA expressions of HPV16 E6 and E7 were determined by semi-quantitative RT-PCR. The protein expressions of HPV16 E6, E7 oncogene, and specifically interacted p53, Rb antioncogene were stained by immunocytochemical staining (Elivison two-step procedure).


RESULTS
(1) The proliferative activities of the Ect1/E6E7 cell and the CaSki cell treated with BJOE at different concentrations (5, 10, 20, and 40 p g/mL) at the operation time points (24, 48, and 72 h) were obviously inhibited, showing dose- and time-dependent manners (P <0.05). (2) Typical changes of apoptosis were observed in both HPV 16 positive cell lines after treated with BJOE. The cell apoptosis rates increased markedly after being cultured with BJOE at different concentrations (5, 10, and 20 microg/mL) for 48 h (P < 0.05). (3) After treated with BJOE at different concentrations (5, 10, and 20 microg/mL) for 48 h, the HPV16 E6 and E7 mRNA relative expressions in both HPV 16 positive cell lines decreased significantly (P < 0.05). (4) After treated with BJOE at different concentrations (5, 10, and 20 microg/ mL), the expressions of HPV16 E6, E7, and mutant p53 protein decreased gradually (P < 0.05), while the Rb protein expression increased gradually (P < 0.05).


CONCLUSIONS
BJOE showed obvious in vitro inhibitory effects on HPV type 16 infected cells. Its underlying mechanisms might be possibly associated with down-regulating expressions of HPV16 E6 and E7 oncogenes.
Department of Gynecology, First Affiliated Hospital of Wenzhou Medical College, Zhejiang (325000), China. 627830566@qq.com
Yan
Y
Hu
Department of Gynecology, First Affiliated Hospital of Wenzhou Medical College, Zhejiang (325000), China.
Xiao-jie
XJ
Wan
Department of Gynecology, First Affiliated Hospital of Wenzhou Medical College, Zhejiang (325000), China.
Liu-liu
LL
Pan
Institute of Medical Sciences, First Affiliated Hospital of Wenzhou Medical College, Zhejiang (325000), China.
Sheng-hui
SH
Zhang
Department of Gynecology, First Affiliated Hospital of Wenzhou Medical College, Zhejiang (325000), China.
Fei-yun
FY
Zheng
D017209:Apoptosis / Q000187:drug effects; D032269:Brucea / Q000737:chemistry*; D045744:Cell Line, Tumor; D004365:Drugs, Chinese Herbal / Q000494:pharmacology*; D005260:Female; D052162:Human papillomavirus 16 / Q000187:drug effects* / Q000472:pathogenicity*; D006801:Humans; D009856:Oncogene Proteins, Viral / Q000378:metabolism; D050725:Papillomavirus E7 Proteins / Q000378:metabolism; D030361:Papillomavirus Infections; D010938:Plant Oils / Q000494:pharmacology*; D012097:Repressor Proteins / Q000378:metabolism
D004365:Drugs, Chinese Herbal; C068651:E6 protein, Human papillomavirus type 16; D009856:Oncogene Proteins, Viral; D050725:Papillomavirus E7 Proteins; D010938:Plant Oils; D012097:Repressor Proteins; C059731:oncogene protein E7, Human papillomavirus type 16