ABSTRACT: Endometrial carcinoma is one of the most common gynecological malignancies in women. The diagnosis of the disease at early or premalignant stages is crucial for the patient's prognosis. To date, diagnosis and follow-up of endometrial carcinoma and hyperplasia require invasive procedures. Therefore, there is considerable demand for the identification of biomarkers to allow non-invasive detection of these conditions.
In this study, we performed a quantitative proteomics analysis on serum samples from simple endometrial hyperplasia, complex endometrial hyperplasia, atypical endometrial hyperplasia, and endometrial carcinoma patients, as well as healthy women. Serum samples were first depleted of high-abundance proteins, labeled with isobaric tags (iTRAQ), and then analyzed via two-dimensional liquid chromatography and tandem mass spectrometry. Protein identification and quantitation information were acquired by comparing the mass spectrometry data against the International Protein Index Database using ProteinPilot software. Bioinformatics annotation of identified proteins was performed by searching against the PANTHER database.
In total, 74 proteins were identified and quantified in serum samples from endometrial lesion patients and healthy women. Using a 1.6-fold change as the benchmark, 12 proteins showed significantly altered expression levels in at least one disease group compared with healthy women. Among them, 7 proteins were found, for the first time, to be differentially expressed in atypical endometrial hyperplasia. These proteins are orosomucoid 1, haptoglobin, SERPINC 1, alpha-1-antichymotrypsin, apolipoprotein A-IV, inter-alpha-trypsin inhibitor heavy chain H4, and histidine-rich glycoprotein.
The differentially expressed proteins we discovered in this study may serve as biomarkers in the diagnosis and follow-up of endometrial hyperplasia and endometrial carcinoma.
Journal of Hematology & Oncology 01/2011; 4:15. · 3.99 Impact Factor
ABSTRACT: To investigate the expression of exogenous gene transferred by piggyBac (PB) transposon in various gynecological malignant cell lines and reveal its potential application of gene therapy in gynecological cancer.
Amplified herpes simplex virus thymidine kinase (HSV-tk) gene coding region by PCR and integrated it into PB expression vector, PB[Act-RFP]DS, for reconstructing PB[Act-RFP, HSV-tk]DS (pPB/TK). By using different transfection reagents: FuGENE HD, jetPEI, lipofectamine 2000, pPB/TK together with helper plasmid Act-PBase were cotransfected into four mostly common gynecological malignant tumor cell lines HeLa, JEG-3, SKOV3 and HEC-1B. The mRFP1 report gene expressions was observed and detected by fluorescence microscope and flow cytometry to analyze transfection efficiency. The expressions of HSV-tk and mRFP1 gene were detected by reverse transcription PCR (RT-PCR). The cytotoxic effect of various concentration of pro-drug ganciclovir (GCV) for transfected cells was detected by methyl thiazole tetrazolium assay. The transfected cells were positive sorted by flow cytometry and limiting diluted to obtain the stable transfected cell line. The insertion sites of foreign gene transferred by PB transposon in genome were analyzed by inverse PCR.
(1) Double digests analysis and sequences test demonstrated that pPB/TK vector was reconstructed successfully. (2) Using three different transfective reagents, PB transposon transferred HSV-tk gene and mRFP1 gene into HeLa, HEC-1B, SKOV3 and JEG-3 cell efficiently, and the transfection efficiency of pPB/TK for the same cell was different by using different transfective reagents; in Hela cell, the transfection efficiency of FuGENE HD [(78.7+/-9.2)%] was higher than that of lipofectamine 2000 [(54.1+/-11.4)%] and jetPEI [(46.5+/-7.4)%, all P<0.05]; using the same transfective reagent, the transfection efficiency of pPB/TK was also different on various cell lines, using FuGENE HD, the transfection efficiency of pPB/TK on HeLa, JEG-3 and SKOV3 cell was (78.7+/-9.2)%, (74.4+/-8.9)% and (83.2+/-9.7)% respectively, which all were higher than that on HEC-1B [(39.5+/-8.7)%, P<0.05]. (3) RT-PCR showed that there were the mRNA expression of HSV-tk and mRFP1 in all cell lines. (4) 50% inhibitory concentration of GCV for transfected cells, HeLa, JEG-3, SKOV3 and HEC-1B, was 1.29, 3.35, 0.09 and 13.28 microg/ml respectively. Inhibitory effect of GCV (10 microg/ml) on SKOV3 transfected with pPB/TK was (86+/-9)%, which was superior to that transfected with pORF-HSVtk alone [(52+/-12)%, P<0.05]. (5) The insertion sites of PB transposon in the target cells genome were located at TTAA sites. mRFP1 expression still could be detected in three months after transfected.
PB transposon could transfer exogenous gene into various gynecological malignant cells, which could integrated into genome and obtain a long-term and stable expression. It is expected that PB transposon may supply a more efficient and safer transgene technology platform for gene therapy in gynecological cancer.
Zhonghua fu chan ke za zhi 04/2010; 45(4):292-7.
ABSTRACT: To prepare follicle stimulating hormone (FSH) polypeptide modified nanoparticles (NP) in order to achieve specific ovarian tumor targeting.
Expression of FSH receptor protein in human liver cancer and ovarian cancer cell lines BEL-7402, SKOV-3 and Caov-3 was detected by immunocytochemistry. The polypeptide fragment of FSH beta 81-95 amino acids (FSHL81-95) was synthesized and covalently coupled to NP. The specific binding of FSHL81-95 and FSHL81-95-NP was examined by fluorescence microscopy and flow cytometry.
BEL-7402 and SKOV-3 cells were negative for FSH receptor staining, while Caov-3 cells were positive. The diameters of NP were about 100 nm, with a Zeta potential of -25 mV or so. Caov-3 cells showed a more specific interaction with FSHL81-95-NP than SKOV-3 cells (4.17 +/- 0.86 and 2.30 +/- 0.21; P < 0.05). The uptake of FSHL81-95-NP was more than NP in Caov-3 cells (4.17 +/- 0.86 and 0.41 +/- 0.32; P < 0.05). FSHL81-95-NP showed a selective targeting at Caov-3 cells compared with control NP.
FSH polypeptide modified NP could selectively target ovarian cancer cells expressing FSH receptor, which might contribute to specific endocytosis mediated by FSH receptor.
Zhonghua fu chan ke za zhi 07/2008; 43(7):533-7.
ABSTRACT: The objective was to study the role of PI3K signaling in the development of cervical cancer and the antitumor effect of PI3K inhibitors.
PI3K protein and mRNA expression of cervical cancer and non-neoplastic tissues were analyzed by Western blotting and RT-PCR. PI3K/Akt/mTOR pathway components in HeLa cells were assessed by immunocytochemistry and Western blotting. The inhibitive effect of LY294002 on HeLa cells was studied using MTT assay and flow cytometry.
PI3K protein expression was detected in 25 out of 31 tumor specimens. Compared with non-neoplastic tissues, significant overexpression was observed in tumor tissues. For PI3K overexpression with all clinicopathological features, a decreasing trend in adenocarcinoma, advanced stage, and grade was observed. PI3K inhibitor LY294002 efficiently inhibited HeLa cell growth with IC50 of 20.77 microM, and induced apoptosis. The apoptotic rate was 36% at 3h after LY294002 treatment. These pharmacological roles of LY294002 might be played through the PI3K/Akt/mTOR signaling pathway.
The PI3K signaling pathway was implicated in the development of cervical cancer. The activation of its signaling molecules might have clinical implications. Novel targeted therapies for the PI3K/Akt/mTOR signaling pathway components could provide a useful adjuvant therapeutic strategy for cervical cancer.
European Journal of Obstetrics & Gynecology and Reproductive Biology 05/2008; 139(2):237-44. · 1.97 Impact Factor