Antioxidant activity of ethanol extract of Lycium barbarum's leaf with removal of chlorophyll / 한국영양학회지

PURPOSE: The aim of this study was to estimate the antioxidant activities of 50%, 70%, and 100% ethanol extracts of Lycium barbarum leaf and chlorophyll removal extract. METHODS: The antioxidant activities were estimated by measuring total polyphenol content and by assays of 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2′-azino-bis (3-ethylbenzothiazoline-6-sulfate) (ABTS) radical scavenging activities and ferric reducing antioxidant power (FRAP). In addition, reactive oxygen species (ROS) production, DNA fragmentation, and antioxidant enzyme (superoxide dismutase and catalase) activities of the extracts were measured in hydrogen peroxide (H2O2)-stressed HepG2 cells. RESULTS: The total polyphenol content, DPPH and ABTS radical scavenging activities, and FRAP value of the extracts increased in an ethanol concentration-dependent manner. The antioxidant activities of the chlorophyll-removal extracts were much higher than those of the chlorophyll-containing extracts. Cytotoxicity was not observed in HepG2 cells with extracts up to 1,000 µg/mL. All extracts inhibited ROS production in a concentration-dependent manner from 31.3 µg/mL and inhibited DNA damage at 250 µg/mL. The SOD and catalase activities of cell lines treated with the extracts and H2O2 were similar to those of normal cells, indicating a strong protective effect. CONCLUSION: Lycium barbarum leaf extracts had high antioxidant activities and protected H2O2-stressed HepG2 cells. Since the chlorophyll-removal extract exhibited higher antioxidant activities than the chlorophyll-containing ones and the cytoprotective effect was similar, chlorophyll removal extract of Lycium barbarum leaf could be developed as ingredients of functional food and cosmetics.

Time-course Transcriptional Profiling of Human Amniotic Fluid-derived Stem Cells Using Microarray / Journal of the Korean Cancer Association, 대한암학회지

PURPOSE: To maintain the homeostasis of stem cells and prevent their ability to initiate tumorigenesis, it is important to identify and modify factors that prevent or accelerate stem cell senescence. We used microarrays to attempt to identify such factors in human amniotic fluid (HAF)-derived stem cells. MATERIALS AND METHODS: To identify gene expression changes over a time course, we compared gene expression profiles of HAF-derived stem cells in different passages (1st, 2nd, 4th, 6th, 8th, and 10th) using a Sentrix Human illumina microarray. RESULTS: Of the 25,804 genes in the microarray chip, 1,970 showed an over 2-fold change relative to the control (the 1st passage)-either upregulated or downregulated. Quantitative real-time PCR validated the microarray data for selected genes: markedly increased genes were CXCL12, cadherin 6 (CDH6), and folate receptor 3 (FOLR3). Downregulated genes included cyclin D2, keratin 8, insulin-like growth factor 2 (IGF2), natriuretic peptide precursor B (NPPB) and cellular retinoic acid binding protein 2 (CRABP2). The expression pattern of the selected genes was consistent with the microarray data except for CXCL12 and IGF2. Interestingly, the expression of NPPB was dramatically downregulated along the time course; it was almost completely shut-down by the 10th passage. In contrast, FOLR3 mRNA expression was dramatically increased. CONCLUSION: Taken together, although a function for NPPB and FOLR3 in stem cell senescence has not been reported, our results strongly suggest that NPPB and/or FOLR3 play a significant role in the regulation of stem cell senescence.

Establishment of Leukemia Mouse Model Using Mouse-Derived A20 Leukemic Cells, and Detection of Tumor Cells in Bone Marrow / 한국실험동물학회지

Objectives of this study were to establish a leukemia mouse model in the Balb/c mouse based upon the A20 cell line (murine B-lymphoma/leukemia cell line, H-2d). Here we demonstrate for the first time that A20 cells were infiltrated into tissue and bone marrow, thereby evaluate the feasibility of using A20 leukemic cells as a leukemia model. In the study, changes of behavior, survival rate and histological changes of major organs after intravenous injection of A20 cells (1x105, 1x106 or 1x107) into Balb/c mice were observed. After inoculation of 1x106 cells, animals survived up to 38.3 days, although there were no significant correlation between the number of injected cells and life-span. At 21 and 28 days post-injection, both hematoxylin-eosin and CD45R immunohistochemical stains showed diffuse large B-cell lymphoma in the liver. FACS analysis was performed after injection of fluorescent nanomaterial (MNPs@SiO2 RITC)-labeled A20 cells. The labeled A20 cells were detected in bone marrow from 6 hours post-inoculation, indicative of the cellular infiltration. This is the first study that demonstrated the invasion of A20 cells into the bone marrow of Balb/c model using A20 cells. With the occurrence of systemic lesions following metastasis of the cells into lymph nodes and neighboring tissues via bone marrow infiltration, it is suggested that the A20 cell-inoculated Balb/c miouse could be an animal model of acute lymphocytic leukemia.

Comparison of cell growth suppression in SiHa cervical carcinoma cell line by human papillomavirus type 16 E6/E7 siRNAs / 대한산부인과학회지

OBJECTIVE: Human cervical cancer is caused by the high-risk types of human papillomavirus (HPV) such as HPV16, which possess the E6 and E7 oncogenes, whose expressions are a prerequisite for cancer development. We performed this study to compare the efficacy of antitumor activity by HPV siRNA which silences only E6 or both E6/E7. METHODS: We transfected siRNA 377 (HPV16 E6 siRNA), siRNA 3 (HPV16 E6 siRNA), and siRNA 198 (HPV16 E7 siRNA) into SiHa cell line (siRNA 377 silences only E6, and siRNA 3 and siRNA 198 silence both E6 and E7). We experimented cell counts and morphologic changes 24 and 48 hours after transfection and expressions of HPV16 E6/E7 mRNA by RT-PCR. RESULTS: siRNA 377, siRNA 3, and siRNA 198 suppressed the cell growth. siRNA 3 and siRNA 198 were more potent than siRNA 377 in cell growth suppression. siRNA 377 knocked down the expression of E6 mRNA, and both siRNA 3 and siRNA 198 knocked down the expression of E6/E7 mRNA. CONCLUSION: Our results suggest that simultaneous suppression of E6 and E7 was more potent than E6-specific suppression in cancer cell growth.

Vascular shutdown effects by tetraarsenic oxide in TC-1 cells implanted C57BL/6 mice / 대한산부인과학회지

OBJECTIVE: Arsenic trioxide (As2O3) is known to have potent anti-vascular activity and significantly suppress solid tumor growth. The present study was conducted to investigate the vascular shutdown effects of a novel arsenic compound, tetraasrsenic oxide (As4O6), in comparison with As2O3 using cervical cancer animal model. METHODS: Mice tumor challenge model was used C57BL/6 mice transplanted with TC-1 cells. After the growth of tumors was reached up 200~250 mm3, mice were divided into 3 groups randomly for control and treatment of either As2O3 or As4O6. As2O3 and As4O6 was treated by i.p. injection. The tumor size was caliperated in twice for weeks and anti-vascular effect were assessed by Evans blue extraction assay and Hoechst 33342 staining. In tumor tissue, histopathological feaure was obserevd by hematoxylin and eosin (H&E) staining. RESULTS: In mice treated with either As2O3 and As4O6 (i.p.), both of As2O3 and As4O6 was significantly suppressed the tumor growth compared with control group. Moreover, effect of As4O6 is more pronounced. These tumor growth inhibition is led to the massive necrosis and vacular shutdown in tumor tissue. CONCLUSION: This study suggests that As4O6 may have potential anticancer activity via vascular shutdown in C57BL/6 mice transplanted with TC-1 cells.

Apigenin-induced apoptosis in cervical cancer cell lines / 대한산부인과학회지

OBJECTIVE: Apigenin is a widely distributed plant flavonoid and was proposed as a potent antitumor agent. In this study, we investigated the anticancer effects of apigenin on human cervical cancer cell lines. For this, the effects of apigenin on growth inhibition and apoptosis were examined and also mRNA expression of E6 and E7, the main causes of development of cervical cancer, was also evaluated. METHODS: To observe the anti-proliferative effects, cervical cancer cell lines, 5x103 cells/well (96 well plate) of Caski, HeLa and C33A were plated and 24 h later treated with apigenin for three days and then MTT assay was performed. For apoptosis analysis, Annexin V-FITC staining was performed. To examine the effect on anchorage-independent growth by apigenin, soft agar assay was performed. The mRNA expression of HPV E6/E7 was examined by quantitative RT-PCR. RESULTS: Apigenin inhibited the growth of all three kind of cervical cancer cell lines (CaSki, HeLa, and C33A) and induced apoptosis in these cell lines. Also, anchorage-independent growth of Caski cells in soft agar was inhibited approximately 3 folds by apigenin treatment. Unexpectedly, mRNA expression level of both E6 and E7 in HeLa cells was not significantly affected by apigenin. CONCLUSION: These studies showed that apigenin inhibits cervical cancer cell growth through the induction of apoptosis. However, mRNA expression of HPV E6/E7 genes was not affected by the treatment of apigenin, indicating that the anti-cancer effect of apigenin in cervical cancer might be mediated via other pathway. Taken together, these results suggest that apigenin may provide a new therapeutic approach for cervical cancer.

Retraction: Comparison of As2O3 and As4O6 in the Detection of SiHa Cervical Cancer Cell Growth Inhibition Pathway / Journal of the Korean Cancer Association, 대한암학회지

No abstract available.

Retraction: Tetraarsenic Oxide-mediated Apoptosis in a Cervical Cancer Cell Line, SiHa / Journal of the Korean Cancer Association, 대한암학회지

No abstract available.

Application of experion(TM) assay system for discovery of ovarian cancer serum biomarkers / 대한산부인과학회지

OBJECTIVE: The discovery of new biomarkers for ovarian cancer is clearly necessary for the detection and monitoring of the disease. Experion(TM) automated electrophoresis system can be employed in the identification of differentially expressed proteins in cancer cells. The objective of this study was to discover potential diagnostic serological biomarkers for ovarian cancer. METHODS: We performed protein expression difference analyses for 14 healthy women and 28 ovarian cancer patients with stage I, III and IV using Experion(TM) system. And then we checked the protein expression as silver staining after loading at 8~16% gradient gel for comparison with Experion(TM) gel image. The candidate biomarkers were purified and determined using MALDI-TOF mass spectrometer. RESULTS: The distinctive polypeptide peaks were detected at 115.40, 15.96, 14.8, 11.66, and 10.69 kDa and these five peaks were identified as ceruloplasmin, hemoglobin beta chain, hemoglobin sigma chain, serum amyloid A4, and amyloid related serum protein SAA, respectively. These proteins were significantly different between the sera of normal healthy women and ovarian cancer patients. CONCLUSIONS: Five proteins were found to be significantly different between the sera of normal healthy women and ovarian cancer patients. In addition, Experion(TM) assay system can provide high performance for analysis of ovarian cancer-related proteins by increasing the throughput while maintaining a high level of accuracy.

Protein Expression Profile using Two-Dimensional Gel Analysis in Squamous Cervical Cancer Patients / Journal of the Korean Cancer Association, 대한암학회지

PURPOSE: Screening in cervical cancer is now progressing to discover candidate genes and proteins that may serve as biological markers and that play a role in tumor progression. We examined the protein expression patterns of the squamous cell carcinoma (SCC) tissues from Korean women with using two-dimensional polyacrylamide gel electrophoresis (2-DE) and matrix assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometer. MATERIALS AND METHODS: Normal cervix and SCC tissues were solubilized and 2-DE was performed using pH 3~10 linear IPG strips of 17 cm length. The protein expression was evaluated using PDQuest 2-D software(TM). The differentially expressed protein spots were identified with a MALDI-TOF mass spectrometer, and the peptide mass spectra identifications were performed using the Mascot program and by searching the Swiss-prot or NCBInr databases. RESULTS: A total of 35 proteins were detected in SCC. 17 proteins were up-regulated and 18 proteins weredown-regulated. Among the proteins that were identified, 12 proteins (pigment epithelium derived factor, annexin A2 and A5, keratin 19 and 20, heat shock protein 27, smooth muscle protein 22 alpha, alpha-enolase, squamous cell carcinoma antigen 1 and 2, glutathione S-transferase and apolipoprotein a1) were protein previously known to be involved in tumor, and 21 proteins were newly identified in this study. CONCLUSION: 2-DE offers the total protein expression profiles of SCC tissues; further characterization of these differentially expressed proteins will give a chance to identify the badly needed tumor-specific diagnostic markers for SCC.

Cell Cycle Regulatory Protein Expression Profiles by Adenovirus p53 Infection in Human Papilloma Virus-associated Cervical Cancer Cells / Journal of the Korean Cancer Association, 대한암학회지

PURPOSE: The tumor suppressor gene, p53, has been established as an essential component for the suppression of tumor cell growth. In this study, we investigated the time-course anticancer effects of adenoviral p53 (Adp53) infection on human ovarian cancer cells to provide insight into the molecular-level understanding of the growth suppression mechanisms involved in Adp53-mediated apoptosis and cell cycle arrest. MATERIALS AND METHODS: Three human cervical cancer cell lines (SiHa, CaSki, HeLa and HT3) were used. The effect of Adp53 infection was studied via cell count assay, cell cycle analysis, FACS, Western blot and macroarray assay. RESULTS: Adp53 exerts a significant role in suppressing cervical cancer cell growth. Adp53 also showed growth inhibitory effects in each cell line, and it induced apoptosis and cell cycle arrest. Adp53 differentially regulated the expression of genes and proteins, and the gene expression profiles in the SiHa cells revealed that the p21, p53 and mdm2 expressions were significantly up-regulated at 24 and 48 hr. Western blot shows that the p21 and p53 expressionlevels were significantly increased after Adp53 infection. In addition, in all cell lines, both the CDK4 and PCNA protein expression levels were decreased 48 h after Adp53 infection. Cell cycle arrest at the G1 phase was induced only in the SiHa and HeLa cells, suggesting that exogenous infection of Adp53 in cancer cells was significantly different from the other HPV-associated cervical cancer cells. CONCLUSION: Adp53 can inhibit cervical cancer cell growth through induction of apoptosis and cell cycle arrest, as well as through the regulation of the cell cycle-related proteins. The Adp53-mediated apoptosis can be employed as an advanced strategy for developing preferential tumor cell-specific delivery.

Genomic gain and loss of cervical cancer using BAC Chip / 대한산부인과학회지

OBJECTIVE: Cervical cancer has long been linked to the sexually transmitted human papillomavirus (HPV), and the oncoproteins E6 and E7 disrupt the functions of tumour suppressor genes, resulting in genetic alteration. It was shown that loss of heterozygosity at 6p is a common genetic alteration in cervical cancer. However, the molecular genetics of cancer have only recently been understood, and for the development of cervical cancer additional genetic alterations in host cell genes are required. The present study has identified the differential changes of the cervical cancer-associated genetic alterations by a genome-wide array based comparative genomic hybridization (array-CGH). METHODS: We analyzed 15 cases of cervical cancer from St. Mary's hospital of The paraffin-fixed tissue samples were microdissected under microscope and DNA was extracted by the procedures of proteinase K digestion and chloroform extraction. Array-based CGH and genomic PCR were carried out with statistical analyses such as hierarchical clustering and Gene Ontology. The BAC array used in this study consisted of 1,440 human BACs, the space among the clones were approximately 2.08 megabase (Macrogen, Seoul, Korea). RESULTS: All of 15 cases of cervical cancer showed specific gains and losses. The analysis limit of average gains and losses was 53%. A significant positive correlation was found between 1p36.32, 3p14.2, 3q27.1, 7p21.1, 8q24.3 and 11q13.1 changes through the cervical carcinogenesis. The high-level of gain regions, BAC clones encoded GSDMDC1, RECQL4, TP73, ABCF3, ALG3, HDAC9, ESRRA and RPS6KA4 genes. Frequently gained BAC clones encoded genes were PRSS8, FUS, COL18A1, PCOLN3, MAFG and ASPSCR1. The genes encoded by frequently lost BAC clones were PTPRG, GRM7, ZDHHC3, EXOSC7, LRP1B and NR3C2. Also, hierarchical clustering of the expression data readily distinguished genomic alterations in cervical cancer. A subset of cellular processes from each gene was clustered by Gene Ontology database. CONCLUSION: Using Array-CGH, genomic alterations related to cervical cancer were identified to determine whether induction of chromosomal imbalances occurs prior to carcinogenesis. The high resolution of array-CGH combined with human genome database would give a chance to find out possible target genes present in the gained or lost clones.

Immunization with Adenoviral Vectors Carrying Recombinant IL-12 and E7 Enhanced the Antitumor Immunity against Human Papillomavirus 16-associated Tumor / Journal of the Korean Cancer Association, 대한암학회지

PURPOSE: Human papillomavirus (HPV) infection has a significant role in cervical carcinogenesis, and HPV oncoprotein E7 plays an important part in the formation and maintenance of cervical cancer. Interleukin-12 (IL-12) has been reported to induce a cellular immune response, and to suppress the tumor growth and the E7 production. Here we describe the use of adenoviral delivery of the HPV 16 E7 subunit (AdE7) along with adenoviral delivery of IL-12 (AdIL-12) in mice with HPV-associated tumors. MATERIALS AND METHODS: Mice were injected with TC-1 cells to establish TC-1 tumor, and then they were immunized with AdIL-12 and/or AdE7 intratumorally. The anti tumor effects induced by AdIL-12 and/or E7 were evaluated by measuring the size of the tumor. E7-specific antibody and INF-gamma production in sera, and the T-helper cell proliferative responses were then measured. Cytotoxic T-lymphocyte (CTL) and T cell subset depletion studies were also performed. RESULTS: Combined AdIL-12 and AdE7 infection at the tumor sites significantly enhanced the antitumor effects more than that of AdIL-12 or AdE7 single infection. This combined infection resulted in regression of the 9 mm sized tumors in 80% of animals as compare to the PBS group. E7-specific antibody and INF-gamma production in the sera, and the T-helper cell proliferative responses were significantly higher with coinfection of AdIL-12 and AdE7 than with AdIL-12 or AdE7 alone. CTL response induced by AdIL-12 and AdE7 in the coinjected group suggested that tumor suppression was mediated by mostly CD8+ and only a little by the CD4+ T cells. CONCLUSION: IL-12 and E7 application using adenovirus vector showed antitumor immunity effects against TC-1 tumor, and this system could be use in clinical applications for HPV-associated cancer.

Adenosine 5'-triphosphate induced NF-kappa B and intracellular calcium mobilization in cervical cancer cells / 대한산부인과학회지

OBJECTIVE: To know the effect of adenosine 5'-triphosphate (ATP) on intracellular calcium level and cell proliferation in cervical cancer cells. METHODS: Study design: Four different human cervical cancer cell lines (Caski, C33A, HeLaS3 and SiHa) were used in this study. The change of intracellular calcium level, cell proliferation and the activity of proliferation- and calcium-related transcription factors by extracellular ATP were examined in these cell lines. RESULTS: Extracellular ATP induced calcium mobilization, cell proliferation and the activation of NF-kappa B in all cell lines used. CONCLUSION: These results suggest that calcium mobilization and NF-kappa B dependent signaling pathway play an important role in the cell proliferation by ATP in cervical cancer.

Effects of cell growth inhibition on the combination of cisplatin with green tea extracts / 부인종양

OBJECTIVE: The chemotherapeutic agent cisplatin (cis-diamminedichloroplatinum (II)) is particularly effective against cervical cancer. The purpose of this study is to elucidate combination effect of cisplatin and green tea extracts on the growth inhibition of TC-1 cell. METHODS: To observe the anti-proliferative effects, we treated different doses of cisplatin (0.1, 0.5, 2.5 uM), GTP (1, 5, 25 ug/ml) and EGCG (25, 50, 100 uM). to TC-1 cells. Also, we treated 0.5 uM of cisplatin and different doses of GTP (1 and 5 ug/ml) or EGCG (25 and 50 uM). Cell viability was scored by use of MTT assay. In addition, E6 gene expression patterns in TC-1 cell were investigated by using RT-PCR. RESULTS: Cell growth inhibition in a dose dependent was observed at the different concentration of ciaplatin, GTP and EGCG. Also, in the groups treated by 0.5 uM of cisplatin and GTP (1 and 5 ug/ml) or EGCG (25 and 50 uM), the inhibition of cell growth showed with 12.2%, 6.9% and 63.4%, 72.2% as compared to the group treated by cisplatin only. In RT-PCR, down regulation of E6 was shown. CONCLUSION: Additive effect of the combination of cisplatin with GTP or EGCG on the inhibition of cell growth was observed. This effect suggests the possibility lowering the concentration of chemotherapeutic drugs, which alleviate the side effect of drugs.

Correlation of adenoassociated virus type 2 and high-risk human papillomavirus type 16 and 18 infection in the cervix of Korean women / 대한산부인과학회지

OBJECTIVE: Previous studies were showed that adenoassocited virus (AAV) infection was had negative effects on human papillomavirus (HPV) infection and that the cervical cancer cell growth is inhibited by AAV infection. We detected of AAV 2 and high-risk HPV infection and researched correlation with AAV 2 and HPV in cervical cell. METHODS: Cell of normal cervix (49 persons), infected HPV cervix (45 persons), cervical intraepithelial neoplasm (CIN) I (31 persons), II (20 persons), III (35 persons), and invasive cancer (30 persons) were investigated by PCR using AAV-2 and HPV type 16 and 18 specific primers. RESULTS: AAV 2 was detected in 8 out of 49 normal cervix (16.3%), 2 out of 45 infected HPV cervix (4.4%), 3 out of 31 CIN I (9.7%), 4 out of 20 CIN II (20%), 8 out of 35 CIN III (22.8%), and 3 out of 30 invasive cervical cancer cases (30%). However, HPV 16 was detected in 5 out of 49 normal cervix (10.2%), 20 out of 45 infected HPV cervix (44.4%), 13 out of 31 CIN I (42%), 11 out of 20 CIN II (55%), 19 out of 35 CIN III (54.3%), and 21 out of 30 invasive cervical cancer cases (70%). HPV 18 was detected in 6 out of 49 normal cervix (12.2%), 18 out of 45 infected HPV cervix (40%), 16 out of 31 CIN I (51.6%), 10 out of 20 CIN II (50%), 22 out of 35 CIN III (62.8%), and 13 out of 30 invasive cervical cancer cases (43.3%). CONCLUSION: AAV 2 was detected in normal and infected HPV cervix, CIN (I, II, III) and invasive cervical cancer. As compared to normal, CIN I and CIN II, suggesting significant correlation between AAV 2 and HPV type 16. Further, researches continue to be done relationship to AAV 2 and HPV infection in cervix.

Tetraarsenic Oxide-mediated Apoptosis in a Cervical Cancer Cell Line, SiHa / Journal of the Korean Cancer Association, 대한암학회지

PURPOSE: Diarsenic oxide, As2O3, has been reported to be effective in treating acute leukemia, and induce apoptosis in many tumor cells. In this study, the ability of a novel arsenical compound, As4O6 (tetraarsenic oxide), along with As2O3, for its ability to induce cell growth inhibition, as well as apoptosis, in human cervical cancer cells, SiHa cells, were evaluated in vitro. MATERIALS AND METHODS: To examine the levels of apoptosis, SiHa cells were given two sensitive doses, 0.5 and 1micrometer, of arsenical compounds, and a DNA fragmentation assay and FACS analysis were then conducted. In addition, a Western blotting assay was performed to identify target molecules for apoptosis. RESULTS: Both As2O3 and As4O6 induced dose-dependent inhibition of SiHa cell proliferation. In particular, As4O6 was more effective at suppressing SiHa cell growth than As2O3. In parallel with the inhibition of cell proliferation, As4O6 caused a significantly greater increase in the sub-G1 cell population than As2O3, as determined by propidium iodide DNA staining. This was confirmed by a DNA fragmentation assay and annexin V staining. The Western blotting analysis also showed that the expression of proliferating cell nuclear antigen (PCNA) was suppressed to a significantly greater extent by As4O6 than As2O3 at a dose of 0.5micrometer. However, the apoptosis-related protein, Bax, was expressed to a significantly greater extent due to As4O6 than As2O3. CONCLUSIONS: Taken together, these findings suggest that a novel arsenic compound, As4O6, possesses more potent anti-proliferative effects on human cervical cancer cells, with the induction of apoptosis also, at least via the activation of Bax protein in vitro.

Search for Gene Expression in Cervical Squamous Cell Carcinoma using GeneFishing(TM) DEG PCR Technique / 대한산부인과학회지

OBJECTIVE: The aim of this study was to investigate the gene expression profiles using GeneFishing(TM) DEG kit in Korean women with cervical squamous cell carcinoma. METHODS: Cervical cancer biopsies were obtained from patients at the Department of Obstetrics and Gynecology, St. Mary's hodpital. In this study, we used a common reference that was mixed with an equal amount of RNA extracted from non-cervical cancer patients. The profiles of expression genes between cervical normal and squamous cell carcinoma tissue were identified using GeneFishing(TM) DEG Kit and screened by BLAST search. RESULTS: Almost 100 differential expressed genes were identified in universal control and cervical squamous cell carcinoma, 53 of differential expressed genes, up-regulated expression of 32 and 21 down-regulated expression was sequenced. Up-regulated genes were calcylin, calgranulin A, TRK oncogene, HLC5, fibrillarin, collagene type I alpha1 etc. and down-regulated genes were galectin 1, PRP8 pre-mRNA precessing factor 8 homology, clusterin etc. CONCLUSION: We identified gene expression profile in cervical squamous cell carcinoma using GeneFishing(TM) Kit in Korean women. The functional genomics of these genes should be further studied.

Proteomic analyses of cervical cancer tissues by two-dimensional gel electrophoresis and mass spectrometry / 대한산부인과학회지

OBJECTIVE: Comparison of protein expressions by two-dimensional gel electrophoresis (2-DE) in normal cervix and squamous cell carcinoma tissues in Korean women. METHODS: Normal cervix and squamous cell carcinoma tissues were solubilized with 2-DE buffer and the first dimension of PROTEAN IEF CELL, isoelectric focusing (IEF), was performed using pH3-10 linear IPG strips of 17 cm. And then running 12% sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and sliver stain. Scanned image was analyzed using PDQuest 2-D softwareTM. Protein spot spectrum was identified by assisted laser desorption/ionization-time of fighting (MALDI-TOF) and the protein mass spectrum identifications were performed by searching protein databases of Swiss-prot/TrEMBL, Mascot and MS-FIT. RESULTS: We found 9 up-regulation proteins (Alpha enolase, Keratin 19 type I, Keratin 20 type I, Keratin 13 type I, beta-actin, Aflatoxin B1 aldehyde reductase 1, Annexin A2, Squamous cell carcinoma antigen 2, unknown), 7 down-reguation proteins (Annexin 1, Myosin regulatory light chain 2, 14-3-3 protein epsilon, Heat shock 27 kDa protein, Hypothetical protein (DKFZP434C1715), Tumor necrosis factor receptor superfamily member 13B, Smoth muscle protein 22-alpha) and 6 up and down-regulation proteins (Tropomyosin 1, Tropomyosin 2, Tropomyosin 3, Serine (or cysteine) proteinase inhibitor, Phosphatidylinositol transfer protein alpha isoform, Src homology 3 domain-containing protein HIP-55) between normal cervix and squamous cell carcinoma cell tissues. CONCLUSION: 2-DE offers total protein expressions between normal cervix and squamous cell carcinoma cell tissues, and searching of differently expressed protein for the diagnostic markers of squamous cell carcinoma tissue.

Expression Profiling of the Cellular Processes in Uterine Leiomyomas: Omic Approaches and IGF-2 Association with Leiomyosarcomas / Journal of the Korean Cancer Association, 대한암학회지

PURPOSE: This study utilized both cDNA microarray and 2D protein gel electrophoresis technology to investigate the multiple interactions of the genes and proteins involved in the pathophysiology of uterine leiomyomas. Also, Gene Ontology (GO) analysis was used to systematically characterize the global expression profiles, which were found to correlate with the leiomyosarcomas. MATERIALS AND METHODS: The uterine leiomyoma biopsies were obtained from patients in the Department of Obstetrics and Gynecology, The Catholic University of Korea. Differentially expressed transcriptome and proteome, in 6 paired leiomyoma and normal myometrium, were profiled. The total RNAs from the leiomyoma and normal myometrium were labeled with Cy5 and Cy3. All specimens were punch-biopsy-obtained, and frozen in liquid nitrogen. RESULTS: Screening of up to 17, 000 genes identified 71 that were either up-regulated or down-regulated (21 and 50, respectively). The gene expression profiles were classified into 420 mutually dependent functional sets, resulting in 611 cellular processes, according to the gene ontology. Also, the protein analysis, using 2D gel electrophoresis, identified 33 proteins (17 up-regulated and 16 down-regulated) with more than 500 total spots, which were classified into 302 cellular processes. Of these functional profilings, transcriptomes and proteoms down- regulations were shown in the cell adhesion, cell motility, organogenesis, enzyme regulator, structural molecule activity and responses to external stimulus functional activities, which are supposed to play important roles in the pathophysiology. In contrast, up-regulation was only shown in the nucleic acid binding activity. The CDKN2A, ADH1A, DCX, IGF2, CRABP2 and KIF5C were found to increase the reliability of this study, and correlate with the leiomyosarcomas. CONCLUSION: Potentially significant pathogenetic cellular processes showed that down-regulated functional profiling has an important impact on the discovery of the pathogenic pathways in leiomyomas and leiomyosarcomas. GO analysis can also overcome the complexity of the expression profiles of cDNA microarrays and 2D protein analyses, via a cellular process level approach. Thereby, a valuable prognostic candidate gene, with real relevance to disease-specific pathogenesis, can be found at cellular process levels.