[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.

Inhibitory effects of methotrexate on spontaneous motility and Cajal-like type of tubal interstitial cells in rabbit oviduct.

OBJECTIVE: To study the adverse biomechanical effects of methotrexate (MTX) on spontaneous tubal motility and on a widely distributed Cajal-like type of tubal interstitial cells (t-ICC) in rabbits. In our previous study, MTX was confirmed to cause acute endosalpingitis, and ultrastructural and steroid receptor damage in rat's endosalpinx in a dose-dependent manner. DESIGN: Differences in spontaneous tubal contractions and cellular distribution of t-ICC in isthmus were evaluated in response to MTX. SETTING: Medical school research laboratory. ANIMAL(S): Twenty nonpregnant female New Zealand albino rabbits in estrus stage were divided equally into four groups. INTERVENTION(S): Rabbits received IM MTX (1, 5, 10 mg/kg body weight) and controls received physiological saline. MAIN OUTCOME MEASURE(S): On day 7, in vitro motility studies measuring spontaneous tubal contractions were performed, and cellular distribution of t-ICC was determined by immunohistochemistry. RESULT(S): Methotrexate produced a concentration-dependent inhibition of spontaneous isthmus contractions (frequency in 5, 10 mg/kg groups, and amplitude in 1, 5, 10 mg/kg MTX groups). It decreased significantly compared with the control group. Meanwhile, MTX at 5, 10 mg/kg decreased the population of c-kit immunoreactive t-ICC significantly. CONCLUSION(S): The decreased t-ICC may contribute to the diminished tubal smooth muscle contractility caused by MTX as observed. Tubal interstitial cells might be new potential targets for a variety of dysfunctional tubal motility diseases.

Protective effect of calcium folinate against methotrexate-induced endosalpinx damage in rats.

The aim of this study was to evaluate the protective effect of calcium folinate (CF) applied in 10% of the methotrexate (MTX) dosage against morphologic and steroid-receptor damage induced by MTX in rat endosalpinx. The result indicated that endosalpingitis, the ultrastructural damage of endosalpinx, and a change in estrogen and P receptor expression induced by low- and high-dose MTX in endosalpinx can be reversed completely and partly (B1, B2) by combined treatment with CF, suggesting that CF combined with MTX protects against the side effects induced by MTX.

[Proteomic study of total protein of Pinellia pedatisecta Schott effect on human ovarian cancer SKOV3 cells].

OBJECTIVE: To study the proteomics of Pinellia pedatisecta Schott total protein on human ovarian cancer SKOV3 cells. METHODS: SKOV3 cells were in vitro cultured. The growth inhibition of SKOV3 cells in the logarithmic phase with different concentrations of Pinellia pedatisecta Schott (0, 0.05, 0.10, 0.20, 0.30, 0.40, and 0.50 mg/mL) at different time points (24, 48, 72, and 96 h) was analyzed by CCK-8 colorimetry. The total protein was extracted after adding 0.296 mg/mL Pinellia pedatisecta Schott protein for 48 h. The protein with differential expressions was screened out using two-dimensional gel electrophoresis (2-DE) and matrix-assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF-MS). RESULTS: The protein of Pinellia pedatisecta Schott (at 0.10 - 0.50 mg/mL) could significantly inhibit the growth of SKOV3 cells in a time- and dose-dependent manner (P < 0.05, P < 0.01). After analyzed by 2-DE and MALDI-TOF-MS, 43 differential protein dots were successfully identified (21 up-regulated and 22 down-regulated), including alpha-enolase 1, eukaryotic initiation factor 3alpha, cyclophilin B, and so on. CONCLUSIONS: Protein of Pinellia pedatisecta Schott could significantly inhibit the growth of SKOV3 cells, and lead to the proteomics changes of SKOV3 cell strain. They might be correlated with its anti-tumor mechanisms.