Meiotic recombination is carried out through a specialized pathway for the formation and repair of DNA double-strand breaks (DSBs) made by the Spo11 protein. The present study shed light on the functional role of cyclin, CYC2, in Tetrahymena thermophila which has transcriptionally high expression level during meiosis process. Knocking out the CYC2 gene results in arrest of meiotic conjugation process at 2.5-3.5 h after conjugation initiation, before the meiosis division starts, and in company with the absence of DSBs. To investigate the underlying mechanism of this phenomenon, a complete transcriptome profile was performed between wild-type strain and CYC2 knock-out strain. Functional analysis of RNA-Seq results identifies related differentially expressed genes (DEGs) including SPO11 and these DEGs are enriched in DNA repair/mismatch repair (MMR) terms in homologous recombination (HR), which indicates that CYC2 could play a crucial role in meiosis by regulating SPO11 and participating in HR.
Cell Cycle Checkpoints ; Cyclins ; genetics ; metabolism ; DNA Breaks, Double-Stranded ; DNA Mismatch Repair ; DNA Repair ; Endodeoxyribonucleases ; genetics ; metabolism ; Homologous Recombination ; Meiosis ; Microscopy, Fluorescence ; Phenotype ; Protozoan Proteins ; genetics ; metabolism ; Real-Time Polymerase Chain Reaction ; Sequence Analysis, RNA ; Tetrahymena thermophila ; genetics ; metabolism ; Transcriptome

OBJECTIVETo investigate c-myc and CCNE2 gene amplifications and their relationship in breast cancer.

METHODSSixty-six infiltrating ductal breast carcinomas with foci of ductal carcinoma in situ components collected from January 2005 to December 2007 were selected for tissue microarray and quantitative multi-gene FISH for c-myc and CCNE2 gene amplification, and the relationship with the clinicopathologic features was analyzed.

RESULTSOf the 66 cases, 18 (27.3%) showed c-myc amplification and 23 (34.8%) showed CCNE2 amplification. A strong correlation was found between c-myc and CCNE2 amplification (P < 0.01). The breast cancers showing c-myc and CCNE2 amplifications were all aneuploidy, and were HER2 positive (P < 0.05). Tumors with c-myc amplification also showed higher Ki-67 index (P < 0.05).

CONCLUSIONSC-myc and CCNE2 amplifications are common events in breast cancer, and they often coexist. C-myc and CCNE2 genes may play critical roles in the pathogenesis and development of breast cancer through unique and overlapping signaling pathways.

Aneuploidy ; Breast Neoplasms ; genetics ; Carcinoma in Situ ; genetics ; Carcinoma, Ductal, Breast ; genetics ; Carcinoma, Intraductal, Noninfiltrating ; genetics ; Cyclins ; genetics ; Female ; Gene Amplification ; Genes, myc ; Humans ; In Situ Hybridization, Fluorescence ; methods ; Tissue Array Analysis

PURPOSE: The effect of cyclin D1 overexpression on breast cancer outcomes and prognosis is controversial, even though amplification of the cyclin D1 gene, CCND1, has been shown to be associated with early relapse and poor prognosis. In this study, we examined the relationship between cyclin D1 overexpression and disease-specific survival (DSS). We also analyzed survival in patients who experienced recurrence. METHODS: We retrospectively analyzed data from patients diagnosed with ductal carcinoma between April 2005 and December 2010. We examined clinicopathologic factors associated with cyclin D1 overexpression and analyzed the influence of cyclin D1 on recurrence-free survival and DSS. RESULTS: We identified 236 patients diagnosed with primary breast cancer who completed all phases of their primary treatment. Cyclin D1 overexpression was significantly associated with longer DSS (5-year DSS, 89.9% in patients without cyclin D1 overexpression vs. 98.9% in patients with cyclin D1 overexpression; p=0.008). Multivariate analysis also found that patients with cyclin D1 overexpressing tumors had significantly longer disease-specific survival than patients whose tumors did not overexpress cyclin D1, with a hazard ratio for disease-specific mortality of 7.97 (1.17-54.22, p=0.034). However, in the group of patients who experienced recurrence, cyclin D1 overexpression was not significantly associated with recurrence-free survival. Cyclin D1 overexpression was significantly associated with increased survival after disease recurrence, indicating that cyclin D1 overexpression might be indicative of more indolent disease progression after metastasis. CONCLUSION: Cyclin D1 overexpression is associated with longer DSS, but not recurrence-free survival, in patients with breast cancer. Longer postrecurrence survival could explain the apparent inconsistency between DSS and recurrence-free survival. Patients with cyclin D1-overexpressing tumors survive longer, but with metastatic disease after recurrence. This information should spark the urgent development of tailored therapies to cure these patients.
Breast Neoplasms* ; Breast* ; Carcinoma, Ductal ; Cyclin D1* ; Cyclins* ; Disease Progression ; Genes, bcl-1 ; Humans ; Mortality ; Multivariate Analysis ; Neoplasm Metastasis ; Prognosis ; Recurrence* ; Retrospective Studies

To study the effect of micro (mi)RNA on cellular proliferation induced by hepatitis B x protein, HBx, in human liver cells and to investigate the underlying molecular mechanism of this cancer-related effect. The human L02 hepatocyte cell line was stably transfected with HBx (L02/HBx) or an HBx mutant (L02/HBx-d382) that induces higher levels of cellular proliferation. The differential miRNA expression profiles were determined by microarray analysis and confirmed by real-time PCR. Two miRNAs, miR-338-3p and miR-551b, that were found to be significantly down-regulated in the L02/HBx-d382 cells were selected for further study and transfected individually into cells using the lipofectamine procedure. The cell survival rate was analyzed by MTT assay, and cell cycles were assessed by flow cytometry. Expressions of cyclinD1, cyclinG1, and E2F1 were assessed by real-time PCR and Western blotting. Compared with the microarray miRNA profile of L02/pcDNA3.0 cells, six miRNAs were up-regulated and five miRNAs were down-regulated in the L02/HBx-d382 cells, while four miRNAs were up-regulated and 12 were down-regulated in the L02/HBx cells. The microarray results were consistent with real-time PCR results. Transfection of miR-338-3p and miR-551b significantly inhibited the cell survival rates (P less than 0.001) and induced G0/G1 phase cycle arrest. According to MTT results: for L02/HBx-d382 cells, compared with lipofectamine or non-transfected (NC) controls, the t value of miR-338-3p was 10.402, 9.133 and the t value of miR-551b was 8.763, 7.403; for L02/HBx cells, compared with lipofectamine or NC controls, the t value of miR-338-3p was 9.105, 8.074 and the t value of miR-551b was 7.673, 7.52. According to flow cytometry results: for L02/HBx-d382 cells, compared with lipofectamine or NC controls, the t value of miR-338-3p was 12.173, 11.107 and the t value of miR-551b was 15.364, 13.377; for L02/HBx cells, compared with lipofectamine or NC controls, the t value of miR-338-3p was 15.416, 13.378, and the t value of miR-551b was 13.276, 13.109. The protein levels of cyclinD1, cyclinG1, and E2F1 were significantly reduced by both miR-338-3p and miR-551b ( P less than 0.001). For L02/HBx-d382 cells, compared with lipofectamine or NC controls: E2F1 had t = 11.132, 10.031 and 12.017, 10.973, respectively; cyclinD1 had t = 15.654, 15.013 and 15.447, 14.733, respectively; cyclinG1 had t = 8.017, 7.661 and 7.402, 7.417, respectively. For L02/HBx cells, compared with lipofectamine or NC controls: E2F1 had t = 14.244, 13.331 and 15.022, 14.468, respectively; cyclinD1 had t = 8.695, 8.137 and 7.877, 7.503, respectively; cyclinG1 had t = 7.73, 7.471 and 7.596, 7.41, respectively. In contrast, the mRNA levels for E2F1, cyclinD1, and cylcinG1 showed no significant differences between the miRNA transfected cells and controls. Wild-type HBx and the high proliferation-inducing mutant HBx can influence the miRNA expression profile of L02 cells. HBx down-regulates miR-338-3p and miR-551b in L02 cells, and the high proliferation-inducing mutant has a more robust effect. The mechanism of miR-338-3p- or miR-551b-mediated cell growth inhibition appears to be related to the direct modulation of cyclinD1, cyclinG1, and E2F1.
Blotting, Western ; Carcinoma, Hepatocellular ; genetics ; metabolism ; pathology ; Cell Cycle ; Cell Line ; Cell Proliferation ; Cyclins ; genetics ; metabolism ; Gene Expression Regulation, Neoplastic ; Genes, Viral ; Hepatitis B virus ; genetics ; metabolism ; Hepatocytes ; metabolism ; pathology ; Humans ; Liver Neoplasms ; genetics ; metabolism ; pathology ; MicroRNAs ; genetics ; metabolism ; Mutation ; Oligonucleotide Array Sequence Analysis ; RNA, Messenger ; genetics ; Real-Time Polymerase Chain Reaction ; Trans-Activators ; genetics ; metabolism ; Transfection

PURPOSE: The aim of this study was to determine the expressions of Rb, p16, and cyclin D1 in soft tissue sarcomas, and we also wanted to identify the prognostic factors according to the clinicalpathologic features. MATERIALS AND METHODS: We reviewed the charts and radiographic films of 66 sarcoma patients. Tissue samples were collected from these patients. Immunochemistry was performed using formalin-fixed, paraffin-embedded tissue samples to examine the expressions of p16, Rb, and cyclin D1 proteins. RESULTS: The median duration of overall survival was 47.8 months (range, 20.0 to 70.7 months) and the 5 years survival rate was 39%. As for the correlation between the degree of immunohistochemical staining for Rb protein and the histological tumor grades, there was a significant difference with a p-value of 0.019. However, no significant correlation was shown for p16 and cyclin D1. The overall survival duration of the Rb negative group (staining cell <20%) and the heterogeneous group (cell staining 20 to 80%) was 53.5+/-6.6 months and the overall survival duration of the Rb homogeneous group was 18.3+/-6.4 months, and there was a significant difference with a p-value of 0.016. However, no significant difference was shown between the survival rate according to the p16 and cyclin D1 expressions. On the multivariate analysis that was done with Rb, p16, the tumor size, grade and site, and patient age, the Rb gene expression was the most significant independent prognostic factor with a risk ratio of 3.01 (p=0.04). CONCLUSION: The expression of Rb protein was correlated with the histologic grade and overall survival of patients with soft tissue sarcomas.
Cyclin D1 ; Cyclins ; Genes, Retinoblastoma ; Humans ; Immunochemistry ; Multivariate Analysis ; Odds Ratio ; Proteins ; Retinoblastoma Protein ; Sarcoma ; Survival Rate ; X-Ray Film

OBJECTIVETo investigate the mechanism of pilose antler polypeptides (PAP) resisting replicative senescence of rat chondrocyte serially subcultivated in vitro by means of PAP interfering and controlled experiment.

METHODSThe successive tert-generation (2nd passage, 3rd passage, 4th passage) chondrocytes and the 4th passage cells intervented by PAP were studied for senenscence mechanism. In this course, immunocytochemistry was applied for pl6, pRb, E2F, CyclinD, CDK4 and TRAP-ELISA (telomerase repeat amplification protocol assay-enzyme linked immunosorbent assay) was applied for telomerase activation to observe targets' changing regarding to senescence and the function of PAP.

RESULTSAlong with cell's replicative senescence, pl6, pRb and Cyclin D express significantly rised (P < 0.01), while E2F, CDK4 and telomerase express significantly lowerd (P < 0.01). Meanwhile, in PAP interfered group compared with which in 4th passage group, pl6, pRb and Cyclin D express significantly lowerd (P < 0.01l), while E2F, CDK4 and telomerase express significantly rised (P < 0.01).

CONCLUSIONPAP has function that it reversingly affect the express of factors which controlling cell life cycle and cell growth to postpone chondrocyte senenscence.

Animals ; Antlers ; chemistry ; Cellular Senescence ; drug effects ; Chondrocytes ; cytology ; drug effects ; Cyclin D ; Cyclin-Dependent Kinase 4 ; analysis ; Cyclin-Dependent Kinase Inhibitor p16 ; analysis ; Cyclins ; analysis ; E2F Transcription Factors ; analysis ; Peptides ; pharmacology ; Rats ; Rats, Sprague-Dawley ; Retinoblastoma Protein ; analysis

PURPOSE: Cyclin G2 has been reported to be a negative cell-cycle regulator in various cancer tissues. However, the pattern of cyclin G2 expression in gastric cancer is relatively unknown. We investigated the expression of cyclin G2 in gastric cancer tissues and evaluated the clinical significance of its expression. MATERIALS AND METHODS: Well-preserved gastric cancer tissues were consecutively obtained from 172 patients who underwent gastric cancer operations at Samsung Medical Center between November 1994 and December 1997. Cyclin G2 expression in the tissues was examined immunohistochemically, and the clinicopathological features and prognostic significance according to the expression were analyzed. RESULTS: Of the 172 gastric cancer tissues, cyclin G2 expression was positive in 43 tissues (25.0%). According to the stage, cyclin G2 expression was lower in more advanced stages (P<0.001). Negative expression of cyclin G2 was positively correlated with more advanced depth of tumor invasion (P<0.05), presence of lymph-node metastasis (P<0.05) and presence of lymphatic invasion (P<0.05). The prognosis of the cyclin G2 (+) group was significantly better than that of the cyclin G2 (-) group (P<0.001). Multivariate analysis revealed that T stage, lymph-node metastasis, distant metastasis, and lymphatic invasion were independent prognostic factors, but the expression of cyclin G2 was not. CONCLUSION: Cyclin G2 was expressed in 25% of the gastric cancer tissues, and negative expression of cyclin G2 was associated with more advanced tumor progression. Cyclin G2 may be a negative cell-cycle regulator in gastric cancer, and further studies are necessary to elucidate its exact role in the mechanism of carcinogenesis.
Carcinogenesis ; Cyclin G2* ; Cyclins* ; Humans ; Multivariate Analysis ; Neoplasm Metastasis ; Prognosis ; Stomach Neoplasms

PURPOSE: Cyclin G2 has been reported to be a negative cell-cycle regulator in various cancer tissues. However, the pattern of cyclin G2 expression in gastric cancer is relatively unknown. We investigated the expression of cyclin G2 in gastric cancer tissues and evaluated the clinical significance of its expression. MATERIALS AND METHODS: Well-preserved gastric cancer tissues were consecutively obtained from 172 patients who underwent gastric cancer operations at Samsung Medical Center between November 1994 and December 1997. Cyclin G2 expression in the tissues was examined immunohistochemically, and the clinicopathological features and prognostic significance according to the expression were analyzed. RESULTS: Of the 172 gastric cancer tissues, cyclin G2 expression was positive in 43 tissues (25.0%). According to the stage, cyclin G2 expression was lower in more advanced stages (P<0.001). Negative expression of cyclin G2 was positively correlated with more advanced depth of tumor invasion (P<0.05), presence of lymph-node metastasis (P<0.05) and presence of lymphatic invasion (P<0.05). The prognosis of the cyclin G2 (+) group was significantly better than that of the cyclin G2 (-) group (P<0.001). Multivariate analysis revealed that T stage, lymph-node metastasis, distant metastasis, and lymphatic invasion were independent prognostic factors, but the expression of cyclin G2 was not. CONCLUSION: Cyclin G2 was expressed in 25% of the gastric cancer tissues, and negative expression of cyclin G2 was associated with more advanced tumor progression. Cyclin G2 may be a negative cell-cycle regulator in gastric cancer, and further studies are necessary to elucidate its exact role in the mechanism of carcinogenesis.
Carcinogenesis ; Cyclin G2* ; Cyclins* ; Humans ; Multivariate Analysis ; Neoplasm Metastasis ; Prognosis ; Stomach Neoplasms

BACKGROUNDMutation and expression change of p21WAF1/CIP1 may play a role in the growth of osteosarcoma. This study was to investigate the expression of the p21WAF1/CIP1 gene in human osteosarcoma, p21WAF1/CIP1 gene DNA sequence change and their relationships with the phenotype and clinical prognosis.

METHODSp21WAF1/CIP1 gene in 10 normal people and the tumours of 45 osteosarcoma patients were examined using polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) with silver staining. The PCR product with an abnormal strand was sequenced directly. The p21WAF1/CIP1 gene mRNA and P21 protein of 45 cases of osteosarcoma were investigated by using in situ hybridization and immunohistochemistry, respectively.

RESULTSThe occurrence of P21 protein in osteosarcoma was 17.78% (8/45), and p21WAF1/CIP1 mRNA expression in osteosarcoma was 42.22% (19/45). The p21WAF1/CIP1 gene DNA sequencing of amplified production showed that in p21WAF1/CIP1 gene exon 3 of 36 cases of human osteosarcoma, there were 17 cases (47.22%) with C-->T at position 609; 10 normal blood samples' DNA sequence analysis yielded 8 cases (80.00%) with C-->T at the same position.

CONCLUSIONSAlong with the increase of malignancy, the expression of p21WAF1/CIP1mRNA and P21 protein in osteosarcoma tends to decrease. It is uncommon for the p21WAF1/CIP1 gene mutation to occur in human osteosarcoma. As a result, the possible existence of tumour subtypes of p21WAF1/CIP1 gene mutation should be investigated. Our research leads to the location of p21WAF1/CIP1 gene polymorphism of Chinese osteosarcoma patients, which can provide a basis for further research.

Cyclin-Dependent Kinase Inhibitor p21 ; Cyclins ; genetics ; Humans ; Osteosarcoma ; genetics ; Polymerase Chain Reaction ; Polymorphism, Single-Stranded Conformational ; RNA, Messenger ; analysis ; Sequence Analysis, DNA

OBJECTIVETo investigate the relationship between cyclin D2 and P210(BCR/ABL) tyrosine kinase in chronic myelogenous leukemia (CML).

METHODSRT-PCR, Western blot and flow cytometry were performed to detect the expression of cyclin D2 in K562 cells and in K562-ib-eGFP cells which express intracellular single-chain antibody (sFv, intrabody) against ABL tyrosine kinase domain.

RESULTSCyclin D2 expression in K562-ib-eGFP cells was 18.90% which was lower than that of control K562 cells (48.10%), and the number of S-phase cells in K562-ib-eGFP was 40.40% which was much lower than that in K562 cells (64.34%).

CONCLUSIONCyclin D2 is a potential down-stream signal molecule of the p210(BCR/ABL) tyrosine kinase in CML. The altered expression of cyclin D2 may contribute to the over proliferation of CML cells.

Blotting, Western ; Cyclin D2 ; Cyclins ; analysis ; genetics ; Flow Cytometry ; Genes, abl ; Humans ; K562 Cells ; Leukemia, Myelogenous, Chronic, BCR-ABL Positive ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction