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2.
Asian Pac J Cancer Prev ; 16(6): 2495-9, 2015.
Article in English | MEDLINE | ID: mdl-25824786

ABSTRACT

Mitogen-activated protein kinase/extracellular signal-regulated kinase kinase kinase 3 (MEKK3) is an important serine/threonine protein kinase and a member of the MAPK family. MEKK3 can effectively activate the MEK/ERK signaling pathway and promote an autocrine growth loop critical for tumor genesis, cell proliferation, terminal differentiation, apoptosis and survival. To explore the relationship between MEKK3 and cell apoptosis, clinicopathology and prognosis, we characterize the expression of MEKK3, pERK and FoxP3 in the renal clear cell carcinoma (RCCC). Protein expression was detected by tissue microarray and immunochemistry in 46 cases of RCCC and 28 control cases. Expression levels of CD3+ ,CD3+CD4+,CD3+CD8+,CD4+CD25+, CD4+CD25+ FoxP3+ were assessed by flow cytometry and analyzed for their association with pathological factors, correlation and prognosis in RCCC. Expression of MEKK3, pERK and FoxP3 was significantly up-regulated in RCCC as compared to control levels (p<0.01), associated with pathological grade (p<0.05)and clinical stage (p<0.05). CD4+CD25+ Foxp3+ Treg cells were also significantly increased in RCCC patients (p<0.05). Cox multivariate regression analysis showed that MEKK3, pERK expression and patholigical stage were independent prognostic factors in patients with RCCC (p<0.05). MEKK3 can be used as an important marker of early diagnosis and prognostic evaluation in RCCC. It may be associated with imbalance of anti-tumor immunity and overexpression of pERK. Expression of MEKK3 and pERK are significantly increased in RCCC, with protein expression and clinical stage acting as independent prognostic factors.


Subject(s)
Biomarkers, Tumor/metabolism , Carcinoma, Renal Cell/metabolism , Extracellular Signal-Regulated MAP Kinases/metabolism , Forkhead Transcription Factors/metabolism , Kidney Neoplasms/metabolism , MAP Kinase Kinase Kinase 3/metabolism , Adult , Aged , Aged, 80 and over , Antibodies, Monoclonal/immunology , Carcinoma, Renal Cell/mortality , Carcinoma, Renal Cell/pathology , Case-Control Studies , Female , Flow Cytometry , Follow-Up Studies , Humans , Immunoenzyme Techniques , Kidney Neoplasms/mortality , Kidney Neoplasms/pathology , Male , Middle Aged , Neoplasm Staging , Phosphorylation , Prognosis , Survival Rate , T-Lymphocyte Subsets/metabolism , T-Lymphocyte Subsets/pathology , Tissue Array Analysis , Tumor Cells, Cultured
3.
Asian Pac J Cancer Prev ; 15(13): 5271-6, 2014.
Article in English | MEDLINE | ID: mdl-25040987

ABSTRACT

Mitogen-activated protein kinase/extracellular signal-regulated kinase kinase kinase 3 (MEKK3) is an important protein kinase and a member of the MAPK family, which regulates cellular responses to environmental stress and serves as key integration points along the signal transduction cascade that not only link diverse extracellular stimuli to subsequent signaling molecules but also amplify the initiating signals to ultimately activate effector molecules and induce cell proliferation, differentiation and survival. To explore the relationship between MEKK3 and cell apoptosis, clinicopathology and prognosis, we characterize the expression of MEKK3 and survivin in cervical cancer. MEKK3 and survivin expression was measured by RT-PCR and Western blotting of fresh surgical resections from 30 cases of cervical cancer and 25 cases of chronic cervicitis. Protein expression was detected by tissue microarray and immunochemistry (En Vision) in 107 cases of cervical cancer, 86 cases of cervical intraepithelial neoplasia (CIN), and 35 cases of chronic cervicitis. Expression patterns were analyzed for their association with clinicopathological factors and prognosis in cervical cancer. Expression of MEKK3 and survivin mRNA was significantly higher in cervical cancer than in the controls (p<0.05). MEKK3 and survivin expression differed significantly between cervical carcinoma, CIN, and cervicitis (p<0.05) and correlated with clinical stage, infiltration depth, and lymph node metastasis (p<0.05). MEKK3 expression was positively correlated with survivin (p<0.05). Kaplan-Meier survival analysis showed that MEKK3 and survivin expression, lymph node metastasis, depth of invasion, and FIGO stage reduce cumulative survival. Cox multivariate regression analysis showed that MEKK3, survivin, and clinical staging are independent prognostic factors in cervical cancer (p<0.05). Expression of MEKK3 and survivin are significantly increased in cervical cancer, their overexpression participating in the occurrence and development of cervical cancer, with protein expression and clinical staging acting as independent prognostic factors for patients with cervical cancer.


Subject(s)
Inhibitor of Apoptosis Proteins/genetics , MAP Kinase Kinase Kinase 3/genetics , Uterine Cervical Neoplasms/genetics , Uterine Cervical Neoplasms/pathology , Adult , Aged , Apoptosis/genetics , Female , Humans , Kaplan-Meier Estimate , Lymphatic Metastasis/genetics , Lymphatic Metastasis/pathology , Middle Aged , Prognosis , RNA, Messenger/genetics , Survivin , Uterine Cervicitis/genetics , Uterine Cervicitis/pathology , Uterine Cervical Dysplasia/genetics , Uterine Cervical Dysplasia/pathology
4.
PLoS One ; 9(6): e97739, 2014.
Article in English | MEDLINE | ID: mdl-24905462

ABSTRACT

The mitotic spindle checkpoint (SAC) genes have been considered targets of anticancer therapies. Here, we sought to identify the attractive mitotic spindle checkpoint genes appropriate for human hepatocellular carcinoma (HCC) therapies. Through expression profile analysis of 137 selected mitotic spindle checkpoint genes in the publicly available microarray datasets, we showed that 13 genes were dramatically up-regulated in HCC tissues compared to normal livers and adjacent non-tumor tissues. A role of the 13 genes in proliferation was evaluated by knocking them down via small interfering RNA (siRNA) in HCC cells. As a result, several mitotic spindle checkpoint genes were required for maintaining the proliferation of HCC cells, demonstrated by cell viability assay and soft agar colony formation assay. Then we established sorafenib-resistant sublines of HCC cell lines Huh7 and HepG2. Intriguingly, increased TTK expression was significantly associated with acquired sorafenib-resistance in Huh7, HepG2 cells. More importantly, TTK was observably up-regulated in 46 (86.8%) of 53 HCC specimens. A series of in vitro and in vivo functional experiment assays showed that TTK overexpression promoted cell proliferation, anchor-dependent colony formation and resistance to sorafenib of HCC cells; TTK knockdown restrained cell growth, soft agar colony formation and resistance to sorafenib of HCC cells. Collectively, TTK plays an important role in proliferation and sorafenib resistance and could act as a potential therapeutic target for human hepatocellular carcinoma.


Subject(s)
Carcinoma, Hepatocellular/genetics , Cell Cycle Proteins/genetics , Genes, cdc , Liver Neoplasms/genetics , Protein Serine-Threonine Kinases/genetics , Protein-Tyrosine Kinases/genetics , Carcinoma, Hepatocellular/metabolism , Cell Cycle Proteins/metabolism , Cell Proliferation , Drug Resistance, Neoplasm , Gene Expression Regulation, Neoplastic , Hep G2 Cells , Hepatocytes/drug effects , Hepatocytes/metabolism , Humans , Liver Neoplasms/metabolism , Niacinamide/analogs & derivatives , Niacinamide/pharmacology , Phenylurea Compounds/pharmacology , Protein Serine-Threonine Kinases/metabolism , Protein-Tyrosine Kinases/metabolism , Sorafenib
5.
Zhonghua Gan Zang Bing Za Zhi ; 15(3): 196-8, 2007 Mar.
Article in Chinese | MEDLINE | ID: mdl-17407710

ABSTRACT

OBJECTIVE: To construct TIMP-1 siRNA eukaryotic expression vectors and evaluate their effect on TIMP-1 mRNA expression in hepatic stellate cells. METHODS: The combinant lone DNA with cutting sites of BamH I and Xho I enzyme according to the sequences of 447-465, 552-540 TIMP-1 of rats and nonspecific sequence were selected and cloned to pGEM-T vector and sub-cloned to pRNAT-U6.2. They were then identified by double enzyme digestion analysis and DNA sequencing. Three plasmids were transfected into T6 separately through an oligofectamine package. TIMP-1 mRNA expression was evaluated by RT-PCR. RESULTS: Targeting sequences of TIMP-1 siRNA eukaryotic expression vectors were correct. TIMP-1 mRNA expression was significantly reduced by transfecting them into the T6. CONCLUSION: We successfully constructed two TIMP-1 siRNA eukaryotic expression vectors and the transfected cells can significantly suppress the TIMP-1 expression.


Subject(s)
Genetic Vectors , Hepatic Stellate Cells , RNA, Small Interfering , Tissue Inhibitor of Metalloproteinase-1/genetics , Animals , Cell Line , Gene Silencing , Plasmids , Rats , Transfection
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