PURPOSE: The molecular mechanisms that are responsible for the initiation and progression of breast cancers are largely unknown. This study was to analyze the cyclin B1, cdc2, p53 and p16 tumor suppressor genes in human breast cancer. MATERIALS AND METHODS: To investigate the role of cyclin B1, cdc2, p53 and p16 in the pathogenesis and progression of breast carcinomas, 98 cases of breast cancers were examined by immunohistochemical method. The correlations of cyclin B1, cdc2, p53 and p16 expression with various clinico-pathologic findings were analysed. RESULTS: In the normal breast tissues, cyclin B1, cdc2 and p16 were weakly expressed, while p53 was not expressed. On the other hand, cyclin B1, cdc2, p53 and p16 were overexpressed in breast cancer, showing correlation between the expression of cyclin B1 and cdc2 and breast cancers (p=0.00). The overexpressions of cdc2 and p16 were correlated with an infiltrative tumor border pattern and this was statistically significant (p<0.05). In addition, the overexpression of cdc2 was correlated with histologic high grade carcinomas (p=0.00). CONCLUSION: Cyclin B1 and cdc2 appeared to be involved in the genesis or progression of breast cancers. In addition, the overexpressions of p16 and p53 may play important roles in more aggressive tumor and the overexpression of cdc2 is associated with progression of tumor to a higher grade of breast carcinomas. The deranged overexpressions of cyclin B1, cdc2, p16 and p53 may play an important role in human breast carcinogenesis.
Adult ; Aged ; Breast Neoplasms/*genetics/metabolism/pathology ; Cyclin B/*genetics/metabolism ; Cyclin B1/*genetics/metabolism ; Cyclin-Dependent Kinase Inhibitor p16/*genetics/metabolism ; Female ; Gene Expression Regulation, Neoplastic ; Humans ; Immunohistochemistry ; Middle Aged ; Tumor Suppressor Protein p53/*genetics/metabolism

<b>AIMb>To analyze the functional interactions of Cyclin with p53 and Atm in spermatogenesis and DNA double-strand break repair.

<b>METHODSb>Two lines of double knockout mice were generated. Spermatogenesis and double strand break repair mechanisms were analyzed in Cyclin A1 (Ccna1); p53- and Ccna1; Atm-double knockout mice.

<b>RESULTSb>The block in spermatogenesis observed in Cyclin A1-/- (Ccna1-/-) testes at the mid-diplotene stage is associated with polynucleated giant cells. We found that Ccna1-deficient testes and especially the giant cells accumulate unrepaired DNA double-strand breaks, as detected by immunohistochemistry for phosphorylated H2AX. In addition, the giant cells escape from apoptosis. The development of giant cells occurred in meiotic prophase I, because testes lacking ATM, which are known to develop spermatogenic arrest earlier than prophase I, do not develop giant cells in the absence of cyclin A1. Cyclin A1 interacted with p53 and phosphorylated p53 in complex with CDK2. Interestingly, p53-deficiency significantly increased the number of giant cells in Ccna1-deficient testes. Gene expression analyses of a panel of DNA repair genes in the mutant testes revealed that none of the genes examined were consistently misregulated in the absence of cyclin A1.

<b>CONCLUSIONb>Ccna1-deficiency in spermatogenesis is associated with defects in DNA double-strand break repair, which is enhanced by loss of p53.

Animals ; Ataxia Telangiectasia Mutated Proteins ; Cell Cycle ; genetics ; physiology ; Cell Cycle Proteins ; genetics ; metabolism ; Cyclin A ; genetics ; metabolism ; Cyclin A1 ; Cyclin B ; Cyclin B2 ; DNA ; genetics ; DNA Repair ; genetics ; physiology ; DNA-Binding Proteins ; genetics ; metabolism ; Gene Expression Regulation ; genetics ; physiology ; Male ; Mice ; Mice, Knockout ; Protein-Serine-Threonine Kinases ; genetics ; metabolism ; Spermatogenesis ; genetics ; physiology ; Testis ; cytology ; metabolism ; Tumor Suppressor Protein p53 ; genetics ; metabolism ; Tumor Suppressor Proteins ; genetics ; metabolism

To identify the integration sites in the host genome for the hepatitis B virus (HBV)-encoded X protein (HBx) in hepatocellular carcinoma (HCC) biopsies that are positive for hepatitis B surface antigen (HBsAg). HCC biopsies were obtained from six patients that were HBV carriers, as demonstrated by the presence of HBsAg in their serum and sero-negativity for antibody to HBsAg. DNA was extracted from the tissue, fractionated, and circularized. Primers were designed according to the HBx sequence and used to amplify the circularized DNA templates by inverse polymerase chain reaction (IPCR). The amplified DNA fragments were checked by electrophoresis, cloned into the PMD18-T expression vector, and sequenced. Sequence alignment was performed by the Blast algorithms. Seven electrophoresis bands yielded 22 sequencing results, which represented a total of three HBx integration sites in the host genome: 19q12, 2q32.2, 22q12. The 19q12 integration site encompasses the CCNE1 gene, which encodes a G1/S-specific cyclin-E1. HBx-related integration sites exist in HBsAg-positive HCC biopsies. The CCNE1 gene may play a role in the development of HBx-related HCC.
Carcinoma, Hepatocellular ; blood ; genetics ; Cyclin E ; genetics ; DNA Primers ; DNA, Viral ; genetics ; Hepatitis B Surface Antigens ; metabolism ; Hepatitis B virus ; genetics ; physiology ; Humans ; Liver Neoplasms ; blood ; genetics ; Oncogene Proteins ; genetics ; Trans-Activators ; genetics ; Virus Integration

<b>OBJECTIVEb>To explore the relationship among HBV-associated histopathological indexes, x gene mutations and the methylation status of p16INK4A promoter in liver with chronic hepatitis B virus infection, in order to illustrate their role in p16INK4A hypermethylation and HCC progression.

<b>METHODSb>Twenty-three cases of surgically resected HBV-associated hepatocellular carcinoma and twenty-five fine needle aspiration biopsy cases of chronic hepatitis B were chosen for this study. The methylation status of the p16INK4A promoter in tumors, their corresponding peritumoral samples and chronic hepatitis B cases was determined by methylation-specific polymerase chain reaction (MSP). EnVision two-step immunohistochemical staining showed the expression of viral antigens in situ. Tissue HBV DNA levels were determined by real-time fluorescence quantitative PCR. Polymerase chain reaction and the direct sequencing method was used for mutation analysis of HBV x gene.

<b>RESULTSb>In peritumoral samples (P = 0. 025) and chronic hepatitis B cases (P = 0.029), the expression of HBx protein in methylated groups was all significantly higher than that in unmethylated groups of p16INK4A gene. But in tumors, there was no such significant difference. Other HBV antigens including HBsAg and HBcAg, tissue HBV DNA levels and point mutations of HBV x gene did not show a relationship with the methylation status of p16INK4A gene.

<b>CONCLUSIONb>The data suggest that p16INK4A hypermethylation correlated closely with higher HBx expression in precancerous lesions. HBx may play an important role in the early stage of HBV-associated hepatocarcinogenesis via induction of hepermethylation of p16INK4A promoter.

Adult ; Carcinoma, Hepatocellular ; genetics ; metabolism ; virology ; Cyclin-Dependent Kinase Inhibitor p16 ; genetics ; DNA Methylation ; DNA, Viral ; genetics ; metabolism ; Female ; Hepatitis B Core Antigens ; metabolism ; Hepatitis B Surface Antigens ; metabolism ; Hepatitis B virus ; genetics ; immunology ; metabolism ; Hepatitis B, Chronic ; genetics ; metabolism ; virology ; Humans ; Liver ; metabolism ; pathology ; virology ; Liver Cirrhosis ; genetics ; metabolism ; virology ; Liver Neoplasms ; genetics ; metabolism ; virology ; Male ; Middle Aged ; Point Mutation ; Polymerase Chain Reaction ; methods ; Promoter Regions, Genetic ; genetics ; Trans-Activators ; genetics ; metabolism

<b>OBJECTIVEb>To investigate the feasibility of detecting cyclin D1 mRNA in paraffin-embedded tissues by reverse transcriptase polymerase chain reaction (RT-PCR) and competitive RT-PCR and its diagnostic and differential diagnostic significance for mantle cell lymphoma (MCL).

<b>METHODSb>Paraffin-embedded samples of 36 cases of MCL, 71 cases of other small B-cell lymphomas and 20 cases of lymphoid reactive hyperplasia as control group were retrieved from archival materials. Cyclin D1 protein and its mRNA was detected by EnVision and RT-PCR and competitive RT-PCR in all samples. House-keeping gene PGK was choosen as internal control.

<b>RESULTSb>(1) Cyclin D1 protein was expressed in 27 of the 38 MCL (71.1%). No cyclin D1 expression was found in the control group. (2) PGK was detected in 103 of the 116 cases (88.8%) and also detected in 34 of 36 MCL cases (94.7%). (3) cyclin D1 mRNA was detected in 34 nodal mantle cell lymphoma cases by RT-PCR in paraffin-embedded tissues. The positive rate of cyclin D1 mRNA was 94.4% in mantle cell lymphomas after exclusion of the 2 cases which were negative for both cyclin D1 mRNA and PGK. cyclin D1 mRNA was not detected in other nodal small B-cell lymphomas or lymphoid reactive hyperplasia, except 1 case of B-SLL. Sequencing analysis showed that sequences were identical to cyclin D1. (4) Cyclin D1 mRNA overexpression was detected in 27 cases of nodal mantle cell lymphoma by competitive RT-PCR in paraffin-embedded tissues. The positive rate of cyclin D1 mRNA overexpression was 75.0% in mantle cell lymphomas after exclusion of 2 cases which were negative for both cyclin D1 mRNA and PGK. cyclin D1 mRNA overexpression was not detected in other nodal small B-cell lymphomas or lymphoid reactive hyperplasia.

<b>CONCLUSIONb>RT-PCR and competitive RT-PCR detection of cyclin D1 mRNA overexpression could be used for the diagnosis and differential diagnosis of mantle cell lymphoma in paraffin-embedded blocks.

Cyclin D1 ; biosynthesis ; genetics ; Diagnosis, Differential ; Humans ; Leukemia, Lymphocytic, Chronic, B-Cell ; genetics ; metabolism ; pathology ; Lymphoma, Follicular ; genetics ; metabolism ; Lymphoma, Mantle-Cell ; genetics ; metabolism ; pathology ; Paraffin Embedding ; RNA, Messenger ; biosynthesis ; genetics ; Reverse Transcriptase Polymerase Chain Reaction ; methods

<b>OBJECTIVEb>To investigate the frequency of cyclin A overexpression in hepatocellular carcinoma (HCC) and its relationship with clinical significance and HBx gene integration.

<b>METHODSb>PCR, RT-PCR and Western blot were used to detect the gene, mRNA and protein level of cyclin A in the tumor and nontumorous tissue. PCR and Southern blot were used to detect the integration of HBx gene in HCC.

<b>RESULTSb>Amplification of cyclin A gene was found in 1 of 35 patients; overexpression of cyclin A mRNA and protein was found in 16 of 35, 21 of 35 patients, respectively. Overexpression of cyclin A protein was correlated with patient's age, tumor size and HBx integration.

<b>CONCLUSIONb>Overexpression of cyclin A occurs in the early stage of HCC carcinogenesis. It may be one of the important approaches by which HBV affects the normal cell cycle of hepatocyte.

Adult ; Aged ; Carcinoma, Hepatocellular ; genetics ; metabolism ; physiopathology ; virology ; Cyclin A ; genetics ; Female ; Gene Expression ; Genes, Viral ; Hepatitis B virus ; genetics ; Humans ; Liver Neoplasms ; genetics ; metabolism ; physiopathology ; virology ; Male ; Middle Aged ; RNA, Messenger ; Trans-Activators ; genetics ; Virus Integration

Cyclin B1, a positive regulator, controls mitosis occurrence, plays an important role in cell proliferation. To investigate the clinical significance of cyclin B1, the expression of cyclin B1 in acute leukemia (AL) patients was measured; the expression of cyclin B1 and p21(cipl), and their cell cycle distribution were assayed by flow cytometry in 136 adult patients with newly diagnosed AL, 10 continuous complete remission (CCR) AL and 17 normal controls; the mRNA of cyclin B1 and p21(cipl), and the proliferation cell nuclear antigen (PCNA) in patients and normal controls were detected with semi-quantitative reverse transcription polymerase chain reaction (RT-PCR). The results showed that the expression of cyclin B1 in newly diagnosed AL patients was significantly higher than that in normal controls. For the relapsed AL patients, the cyclin B1 expression was also higher than that in normal controls, but lower than that in newly diagnosed cases, there was no significant difference between the remission cases and normal controls, nor difference between CCR AL patients and normal controls. All patients with high cyclin B1 expression had an unscheduled expression manner, that cyclin B1 protein appeared in G(1) phase, and in some case it even higher than that of G(2) phase. The response rate (partial remission + complete remission) and survival rate in the cyclin B1 high expressed patients were higher than that of cyclin B1 low expressed patients. The relapse rate in cyclin B1 high expressed patients was higher than that in cyclin B1 normally expressed patients. The survival rate in cyclin B1 high expressed patients was higher than that in cyclin B1 low expression patients. A negative correlation between the expression of cyclin B1 and p21(cipl) was observed. Additionally, cyclin B1 protein expression was generally correlated with proliferation index (PI) and proliferation cell nuclear antigen (PCNA). It is concluded that this study demonstrates for the first time cyclin B1 overexpression and abnormally distribution in cell cyclin of newly diagnosed AL patients. It was considered that cyclin B1 may play an important role in leukemic pathogeneses and can be one of the factors influencing the prognosis of AL patients.
Acute Disease ; Adolescent ; Adult ; Aged ; Cell Proliferation ; Cyclin B ; genetics ; Cyclin B1 ; Cyclin-Dependent Kinase Inhibitor p21 ; genetics ; Female ; HL-60 Cells ; Humans ; Kaplan-Meier Estimate ; Leukemia ; drug therapy ; genetics ; pathology ; Male ; Middle Aged ; Neoplasm Recurrence, Local ; Prognosis ; Proliferating Cell Nuclear Antigen ; genetics ; RNA, Messenger ; biosynthesis ; genetics ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction

Astrocytoma ; genetics ; metabolism ; pathology ; Brain Neoplasms ; classification ; genetics ; metabolism ; pathology ; Child ; Child, Preschool ; Chromosomal Proteins, Non-Histone ; genetics ; metabolism ; Chromosome Deletion ; Cyclin-Dependent Kinase Inhibitor p16 ; genetics ; metabolism ; DNA-Binding Proteins ; genetics ; metabolism ; Ependymoma ; genetics ; metabolism ; pathology ; Hedgehog Proteins ; genetics ; metabolism ; Humans ; Medulloblastoma ; classification ; genetics ; metabolism ; pathology ; Proto-Oncogene Proteins B-raf ; genetics ; metabolism ; Rhabdoid Tumor ; genetics ; metabolism ; pathology ; SMARCB1 Protein ; Signal Transduction ; Transcription Factors ; genetics ; metabolism ; Wnt Proteins ; metabolism ; beta Catenin ; genetics ; metabolism

This study aimed to observe the effect of terpinen-4-ol(T4O) on the proliferation of vascular smooth muscle cells(VSMCs) exposed to high glucose(HG) and reveal the mechanism via the Krüppel-like factor 4(KLF4)/nuclear factor kappaB(NF-κB) signaling pathway. The VSMCs were first incubated with T4O for 2 h and then cultured with HG for 48 h to establish the model of inflammatory injury. The proliferation, cell cycle, and migration rate of VSMCs were examined by MTT method, flow cytometry, and wound healing assay, respectively. The content of inflammatory cytokines including interleukin(IL)-6 and tumor necrosis factor-alpha(TNF-α) in the supernatant of VSMCs was measured by enzyme-linked immunosorbent assay(ELISA). Western blot was employed to determine the protein levels of proliferating cell nuclear antigen(PCNA), Cyclin D1, KLF4, NF-κB p-p65/NF-κB p65, IL-1β, and IL-18. The KLF4 expression in VSMCs was silenced by the siRNA technology, and then the effects of T4O on the cell cycle and protein expression of the HG-induced VSMCs were observed. The results showed that different doses of T4O inhibited the HG-induced proliferation and migration of VSMCs, increased the percentage of cells in G_1 phase, and decreased the percentage of cells in S phase, and down-regulated the protein levels of PCNA and Cyclin D1. In addition, T4O reduced the HG-induced secretion and release of the inflammatory cytokines IL-6 and TNF-α and down-regulated the expression of KLF4, NF-κB p-p65/NF-κB p65, IL-1β, and IL-18. Compared with si-NC+HG, siKLF4+HG increased the percentage of cells in G_1 phase, decreased the percentage of cells in S phase, down-regulated the expression of PCNA, Cyclin D1, and KLF4, and inhibited the activation of NF-κB signaling pathway. Notably, the combination of silencing KLF4 with T4O treatment further promoted the changes in the above indicators. The results indicate that T4O may inhibit the HG-induced proliferation and migration of VSMCs by down-regulating the level of KLF4 and inhibiting the activation of NF-κB signaling pathway.
NF-kappa B/metabolism* ; Interleukin-18/metabolism* ; Proliferating Cell Nuclear Antigen/genetics* ; Cyclin D1/metabolism* ; Tumor Necrosis Factor-alpha/metabolism* ; Muscle, Smooth, Vascular ; Cell Proliferation ; Signal Transduction ; Cytokines/metabolism* ; Glucose/metabolism*

The human Gadd45 protein family plays critical roles in DNA repair, negative growth control, genomic stability, cell cycle checkpoints and apoptosis. Here we report the crystal structure of human Gadd45γ [corrected], revealing a unique dimer formed via a bundle of four parallel helices, involving the most conserved residues among the Gadd45 isoforms. Mutational analysis of human Gadd45γ [corrected] identified a conserved, highly acidic patch in the central region of the dimer for interaction with the proliferating cell nuclear antigen (PCNA), p21 and cdc2, suggesting that the parallel dimer is the active form for the interaction. Cellular assays indicate that: (1) dimerization of Gadd45γ [corrected] is necessary for apoptosis as well as growth inhibition, and that cell growth inhibition is caused by both cell cycle arrest and apoptosis; (2) a conserved and highly acidic patch on the dimer surface, including the important residues Glu87 and Asp89, is a putative interface for binding proteins related to the cell cycle, DNA repair and apoptosis. These results reveal the mechanism of self-association by Gadd45 proteins and the importance of this self-association for their biological function.
Amino Acid Motifs ; Animals ; Apoptosis ; radiation effects ; CDC2 Protein Kinase ; Cell Cycle ; Cell Survival ; Crystallography, X-Ray ; Cyclin B ; metabolism ; Cyclin-Dependent Kinase Inhibitor p21 ; metabolism ; Cyclin-Dependent Kinases ; HeLa Cells ; Humans ; Intracellular Signaling Peptides and Proteins ; chemistry ; genetics ; metabolism ; Mice ; Mutagenesis, Site-Directed ; Mutation, Missense ; Proliferating Cell Nuclear Antigen ; metabolism ; Protein Binding ; Protein Interaction Domains and Motifs ; Protein Multimerization ; Protein Structure, Quaternary ; Ultraviolet Rays