OBJECTIVETo investigate the expressions of cyclin E, cyclin dependent kinase 2 (CDK-2) and cyclin-dependent kinase inhibitor p57(KIP2) in human gastric cancer, and to evaluate the relationships between protein levels and clinicopathological parameters.

METHODSWestern blot was used to measure the expressions of cyclin E, CDK-2 and p57(KIP2) proteins in the surgically resected gastric carcinoma, adjacent normal mucosa and metastatic lymph nodes from 36 patients.

RESULTSCyclin E and CDK-2 protein levels were higher in gastric cancer tissues in comparison with normal tissues (P < 0.05). Overexpression of cyclin E was correlated with lymph node involvement, poor histological grade and serosa invasion (P < 0.05). Overexpression of CDK-2 was correlated with lymph nodes involvement (P < 0.05). No statistically significant difference between cyclin E and CDK-2 expression was found when samples were stratified according to tumor size (P > 0.05). Expression of cyclin E and CDK-2 showed a positive linear correlation (r = 0.451, P = 0.01). Protein levels of p57(KIP2) were lower in gastric cancer tissues than in the normal mucosa (P < 0.05). Decreased expression of p57(KIP2) was correlated with lymph node involvement (P < 0.05). No statistically significant difference in p57(KIP2) expression was found when sample were stratified according to tumor size, histological grade or serosa invasion (P > 0.05). In metastatic lymph nodes, expression of cyclin E was increased and the expression of p57(KIP2) decreased.

CONCLUSIONOverexpressions of cyclin E, CDK-2 and downregulated expression of p57(KIP2) may play important roles in tumorigenesis and metastatic potential of gastric cancer.

Blotting, Western ; CDC2-CDC28 Kinases ; Cyclin E ; analysis ; physiology ; Cyclin-Dependent Kinase 2 ; Cyclin-Dependent Kinase Inhibitor p57 ; Cyclin-Dependent Kinases ; analysis ; physiology ; Humans ; Lymphatic Metastasis ; Nuclear Proteins ; analysis ; physiology ; Protein-Serine-Threonine Kinases ; analysis ; physiology ; Stomach Neoplasms ; chemistry ; pathology

Carcinoma of the uterine cervix is one of the most common malignancies among women worldwide. Human papillomaviruses (HPV) have been identified as the major etiological factor in cervical carcinogenesis. However, the time lag between HPV infection and the diagnosis of cancer indicates that multiple steps, as well as multiple factors, may be necessary for the development of cervical cancer. The development and progression of cervical carcinoma have been shown to be dependent on various genetic and epigenetic events, especially alterations in the cell cycle checkpoint machinery. In mammalian cells, control of the cell cycle is regulated by the activity of cyclin-dependent kinases (CDKs) and their essential activating coenzymes, the cyclins. Generally, CDKs, cyclins, and CDK inhibitors function within several pathways, including the p16INK4A-cyclin D1-CDK4/6-pRb-E2F, p21WAF1-p27KIP1-cyclinE-CDK2, and p14ARF-MDM2-p53 pathways. The results from several studies showed aberrant regulation of several cell cycle proteins, such as cyclin D, cyclin E, p16 INK4A, p21WAF1, and p27KIP1, as characteristic features of HPV- infected and HPV E6/E7 oncogene-expressing cervical carcinomas and their precursors. These data suggested further that interactions of viral proteins with host cellular proteins, particularly cell cycle proteins, are involved in the activation or repression of cell cycle progression in cervical carcinogenesis.
Uterine Cervical Neoplasms/*pathology ; Tumor Suppressor Protein p53/physiology ; Tumor Suppressor Protein p14ARF/physiology ; Retinoblastoma Protein/physiology ; Proto-Oncogene Proteins c-mdm2/physiology ; Humans ; Female ; E2F Transcription Factors/physiology ; Cyclin-Dependent Kinase Inhibitor p27/physiology ; Cyclin-Dependent Kinase Inhibitor p21/physiology ; Cyclin-Dependent Kinase Inhibitor p16/physiology ; Cyclin-Dependent Kinase 4/physiology ; Cyclin-Dependent Kinase 2/physiology ; Cyclin E/physiology ; Cyclin D1/physiology ; Cell Cycle/*physiology

Cyclin-dependent protein kinase (CDK) plays an important role in the replication of herpes simplex virus (HSV) and other important human disease viruses. But which kinds of CDK are required in the replication of HSV is still not clear. In this study, we infected dominant negative CDK2 cell line with different multiplicity of infection (MOI) of HSV-1-KOS strain (abbreviated as HSV below), and the results showed that the yield of HSV depended on the MOI; the replication of HSV delayed about 3 h as compared with that of the control in the one-step growth curve replication experiments; the CDK2 activity was induced 6 h post HSV infection and reached the highest 9 h post infection; the HSV went into rapid productive replication after the CDK2 was induced. We propose herein that the CDK2 is required in the initiation of replication of HSV.
Animals ; Cell Proliferation ; Cercopithecus aethiops ; Cyclin-Dependent Kinase 2 ; physiology ; HT29 Cells ; Humans ; Simplexvirus ; physiology ; Vero Cells ; Virus Replication

Cell cycle plays a crucial role in cell development. Cell cycle progression is mainly regulated by cyclin dependent kinase (CDK), cyclin and endogenous CDK inhibitor (CKI). Among these, CDK is the main cell cycle regulator, binding to cyclin to form the cyclin-CDK complex, which phosphorylates hundreds of substrates and regulates interphase and mitotic progression. Abnormal activity of various cell cycle proteins can cause uncontrolled proliferation of cancer cells, which leads to cancer development. Therefore, understanding the changes in CDK activity, cyclin-CDK assembly and the role of CDK inhibitors will help to understand the underlying regulatory processes in cell cycle progression, as well as provide a basis for the treatment of cancer and disease and the development of CDK inhibitor-based therapeutic agents. This review focuses on the key events of CDK activation or inactivation, and summarizes the regulatory processes of cyclin-CDK at specific times and locations, as well as the progress of research on relevant CDK inhibitor therapeutics in cancer and disease. The review concludes with a brief description of the current challenges of the cell cycle process, with the aim to provide scientific references and new ideas for further research on cell cycle process.
Cyclin-Dependent Kinases/metabolism* ; Cyclins/metabolism* ; Protein Serine-Threonine Kinases ; Cell Cycle Proteins/metabolism* ; Cell Cycle/physiology* ; Cyclin-Dependent Kinase 2

OBJECTIVETo explore the influence of different nutritional support routes on the intestinal mucosal epithelial cell cycle in burned rats.

METHODSSixty-six Wistar rats inflicted with 30% TBSA III degree burns on the back were employed as the model and were randomly divided into enteral feeding group (EF) and intravenously parenteral nutrition group (PN). Equal volume of nutritional support fluid containing predetermined equal amount of calories and nitrogen was applied via feeding or intravenously infusion through external jugular vein. The indices were observed on 6, 12, 24, 48 and 72 postburn hours (PBHs) with the reference to those in 6 normal rats. The intestinal epithelial cell cycle in jejunal and ileal mucous membrane was analyzed by flow cytometry. Western blotting method was employed in the examination of the expression of cyclin D1, E and that of cyclin dependent kinase (CDK)2 and CDK4.

RESULTS(1) lntestinal mucosal epithelial G0/G1 ratio in jejunum in EF group was significantly lower than that in PN group at 72 PBHs (P < 0.05). While the ratio in ileum in EF was obviously higher than that in PN groups at 6, 12, 48 and 72 PBHs (P < 0.05). (2) The cell percentage of S phase in EF group was evidently higher than that in PN group (P < 0.05 - 0.01) at 48 and 72 PBHs. (3) Intestinal mucosal cyclin D1 expression increased significantly in EF group at 24 PBHs and in PN group at 48 PBHs (P < 0.05) and which in EF group was obviously higher than that in PN group at 72 PBHs (P < 0.05). (4) The expression of the intestinal mucosal cyclin E in EF group at 72 PBHs was evidently higher than the control value and that in PN group (P < 0.05). (5) The expression of CDK2 exhibited no obvious difference among PN,EF and control group (P < 0.05). The CDK4 expression in EF group increased obviously at 72 PBHs (P < 0.05).

CONCLUSIONEarly postburn enteral feeding was beneficial to the progression of intestinal mucosal epithelial cell cycle and to the repairing and renovation of injured intestinal mucosal membrane. Cyclin and CDK might be important in the modulation of the intestinal mucosal epithelial cell cycle.

Animals ; Burns ; metabolism ; pathology ; CDC2-CDC28 Kinases ; Cell Cycle ; physiology ; Cyclin D1 ; metabolism ; Cyclin-Dependent Kinase 2 ; Cyclin-Dependent Kinase 4 ; Cyclin-Dependent Kinases ; metabolism ; Disease Models, Animal ; Enteral Nutrition ; Female ; G1 Phase ; physiology ; Intestinal Mucosa ; metabolism ; pathology ; Male ; Protein-Serine-Threonine Kinases ; metabolism ; Proto-Oncogene Proteins ; Rats ; Rats, Wistar ; Resting Phase, Cell Cycle ; physiology ; S Phase ; physiology

OBJECTIVESTo unravel the molecular mechanism of proliferation inhibition induced by transfection of pemt2-cDNA into rat CBRH-7919 hepatoma cells.

METHODSWe started with the highly expressed PEMT2 clone. Cell culture and Western blotting techniques were used to examine the expression of cyclinD1/CDK4, cyclinE/CDK2, phospho-Rb, caspase-3, c-jun and caveolins.

RESULTSOur results showed that CDK4, CDK2, phospho-Rb and c-jun were down regulated in the pemt2 highly expressed cell clone. The high expression clone of pemt2-transfected cells also showed over expression of caspase-3.

CONCLUSIONThe reductions of proliferation and apoptosis of pemt2 transfected cells could be related to the G1 phase arrest induced by down-regulation of the cell cycle-associated proteins.

Animals ; Apoptosis ; physiology ; Caspase 3 ; biosynthesis ; genetics ; Cyclin-Dependent Kinase 2 ; biosynthesis ; genetics ; Cyclin-Dependent Kinase 4 ; biosynthesis ; genetics ; Cyclin-Dependent Kinases ; biosynthesis ; genetics ; Down-Regulation ; Liver Neoplasms, Experimental ; genetics ; metabolism ; Phosphatidylethanolamine N-Methyltransferase ; genetics ; metabolism ; Rats ; Transfection ; Tumor Cells, Cultured

BACKGROUNDRhoA/Rho kinase (ROCK) pathway is involved in pulmonary arterial hypertension (PAH) and pulmonary artery smooth muscle cell (PASMC) proliferation. Inhibition of ROCK has been proposed as a treatment for PAH. But the mechanism of RhoA/ROCK pathway and its downstream signaling in proliferation of human PASMCs is unclear. We investigated the effect of fasudil, a selective ROCK inhibitor, on platelet-derived growth factor (PDGF) induced human PASMC proliferation, and the possible association between RhoA/ROCK and extracellular signal-regulated kinase (ERK), p27(Kip1).

METHODSHuman PASMCs were cultured with the stimulation of 10 ng/ml PDGF, and different concentrations of fasudil were added before the addition of mitogen. Cell viability and cell cycle were determined with MTT and flow cytometry respectively. ROCK activity, ERK activity and protein expression of proliferating cell nuclear angigen (PCNA) and p27(Kip1) were measured by immunoblotting.

RESULTSBy MTT assay, PDGF significantly increased the OD value that represented human PASMC proliferation, and pretreatment with fasudil significantly reversed this effect in a dose-dependent manner. After PDGF stimulation, the percentage of cells in S phase increased dramatically from 15.6% to 24.3%, while the percentage in G(0)/G(1) phase was reduced from 80.6% to 59%. And pretreatment with fasudil reversed the cell cycle effect of PDGF significantly in a dose-dependent manner. PDGF markedly induced ROCK activity and ERK activity with a peak at 15 minutes, which were significantly inhibited by fasudil. In addition, fasudil significantly inhibited PDGF-induced PCNA expression and fasudil also upregulated p27(Kip1) expression in human PASMCs, which decreased after PDGF stimulation.

CONCLUSIONRhoA/ROCK is vital for PDFG-induced human PASMC proliferation, and fasudil effectively inhibited PDGF-induced human PASMC proliferation by up-regulation of p27(Kip1), which may be associated with inhibition of ERK activity.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine ; analogs & derivatives ; pharmacology ; Cell Proliferation ; drug effects ; Cells, Cultured ; Cyclin-Dependent Kinase Inhibitor p27 ; metabolism ; Humans ; MAP Kinase Signaling System ; physiology ; Muscle, Smooth, Vascular ; cytology ; Platelet-Derived Growth Factor ; pharmacology ; Protein Kinase Inhibitors ; pharmacology ; Pulmonary Artery ; cytology ; Up-Regulation

OBJECTIVETo investigate the effect of Notch1 on cell cycle, apoptosis and migration of laryngeal squamous cell carcinoma cell line Hep-2 and explore its possible molecular mechanism.

METHODSHep-2 cells were divided into three groups: untreated group, empty vector group and Notch1 group. The effects of upregulation of Notch1 expression on cell proliferation, cell cycle and apoptosis were assessed by CCK-8 staining and flow cytometry, respectively, and effect of upregulation of Notch1 expression on cell migration of Hep-2 cells was studied using Boyden chamber assay, and further expression changes of genes related to cell proliferation, cell cycle, apoptosis and migration were detected by semi-quantitative RT-PCR and Western blotting.

RESULTSCompared with the untreated group and empty vector group, cell proliferation of Hep-2 in the Notch1 group was significantly inhibited (P < 0.05). The results of flow cytometry revealed that the percentage of cells at G0/G1 phase in the Notch1 group was (70.43 +/- 1.36)%, significantly higher than the (46.39 +/- 1.19)% in the untreated group or (48.41 +/- 1.18)% in the empty vector group, and there was a significant difference among the three groups (P = 0.000). In addition, the percentage of apoptotic cells in the Notch1 group was (22.46 +/- 0.90)%, significantly higher than that in the untreated group [(5.77 +/- 0.37)%] or empty vector group [(6.09 +/- 0.20)%], with a significant difference among the three groups (P = 0.000). The results of Boyden chamber assay demonstrated that the number of cells migrated through membrane in the Notch1 group was evidently lower than that in the untreated group and empty vector group. Moreover, the results of semi-quantitative RT-PCR and Western blotting showed that cell proliferation inhibition and cell cycle arrest were closely associated with downregulation of cyclin D1, cyclin E and CDK2 expressions and upregulation of p53 expression. In addition, upregulation of Notch1 expression mediated cell apoptosis was tightly related to upregulation of caspase 3/9 expressions and downregulation of Bcl-2, while the decrease of Hep-2 cell migration ability was obviously correlated with downregulation of MMP-2/-9 expressions.

CONCLUSIONSNotch1 signaling pathway may play a pivotal role in the occurrence and development of laryngeal squamous cell carcinoma. Further study may elucidate that Notchl signaling pathway may become a molecular target for therapy of laryngeal squamous cell carcinoma.

Apoptosis ; Carcinoma, Squamous Cell ; metabolism ; pathology ; Caspase 3 ; metabolism ; Caspase 9 ; metabolism ; Cell Cycle ; Cell Line, Tumor ; Cell Movement ; Cell Proliferation ; Cyclin D1 ; metabolism ; Cyclin E ; metabolism ; Cyclin-Dependent Kinase 2 ; metabolism ; Humans ; Laryngeal Neoplasms ; metabolism ; pathology ; Matrix Metalloproteinase 2 ; metabolism ; Matrix Metalloproteinase 9 ; metabolism ; Oncogene Proteins ; metabolism ; Proto-Oncogene Proteins c-bcl-2 ; metabolism ; Receptor, Notch1 ; metabolism ; physiology ; Signal Transduction ; Tumor Suppressor Protein p53 ; metabolism

BACKGROUNDPulmonary artery smooth muscle cell (PASMC) proliferation plays an important role in pulmonary vessel structural remodelling. At present, the mechanisms related to proliferation of PASMCs are not clear. Focal adhesion kinase (FAK) is a widely expressed nonreceptor protein tyrosine kinase. Recent research indicates that FAK is implicated in signalling pathways which regulate cytoskeletal organization, adhesion, migration, survival and proliferation of cells. Furthermore, there are no reports about the role of FAK in human pulmonary artery smooth muscle cells (HPASMCs). We investigated whether FAK takes part in the intracellular signalling pathway involved in HPASMCs proliferation and apoptosis, by using antisense oligodeoxynucleotides (ODNs) to selectively suppress the expression of FAK protein.

METHODSCultured HPASMCs stimulated by fibronectin (40 microg/ml) were passively transfected with ODNs, sense FAK, mismatch sense and antisense-FAK respectively. Expression of FAK, Jun NH2-terminal kinase (JNK), cyclin-dependent kinase 2 (CDK 2) and caspase-3 proteins were detected by immunoprecipitation and Western blots. Cell cycle and cell apoptosis were analysed by flow cytometry. In addition, cytoplasmic FAK expression was detected by immunocytochemical staining.

RESULTSWhen compared with mismatch sense group, the protein expressions of FAK, JNK and CDK 2 in HPASMCs decreased in antisense-FAK ODNs group and increased in sense-FAK ODNs group significantly. Caspase-3 expression upregulated in HPASMCs when treated with antisense ODNs and downregulated when treated with sense ODNs. When compared with mismatch sense ODNs group, the proportion of cells at G1 phase decreased significantly in sense ODNs group, while the proportion of cells at S phase increased significantly. In contrast, compared with mismatch sense ODNs group, the proportion of cells at G1 phase was increased significantly in antisense-FAK ODNs group. The level of cell apoptosis in antisense-FAK group was higher than in the mismatch sense group and the latter was higher than sense-FAK group. In addition, the sense-FAK ODNs group was strongly stained by immunocytochemistry, whereas the antisense-FAK ODNs group was weakly stained.

CONCLUSIONSThe results suggest that FAK relates to the proliferation of HPASMCs. Antisense-FAK ODNs inhibit HPASMCs proliferation and facilitate their apoptosis. It is possible that FAK via JNK, CDK 2 signalling pathways enhances HPASMCs proliferation and via caspase-3 inhibits HPASMCs apoptosis.

Apoptosis ; CDC2-CDC28 Kinases ; analysis ; Caspase 3 ; Caspases ; analysis ; Cell Cycle ; Cell Proliferation ; drug effects ; Cells, Cultured ; Cyclin-Dependent Kinase 2 ; Focal Adhesion Kinase 1 ; Focal Adhesion Protein-Tyrosine Kinases ; Humans ; Immunohistochemistry ; JNK Mitogen-Activated Protein Kinases ; analysis ; Muscle, Smooth, Vascular ; cytology ; Myocytes, Smooth Muscle ; physiology ; Oligodeoxyribonucleotides, Antisense ; pharmacology ; Protein-Tyrosine Kinases ; analysis ; physiology ; Pulmonary Artery ; cytology

In order to study the potential effects of exogenous WT1 gene isoform on apoptosis in leukemia cell line NB4 and its possible molecular mechanisms, the eukaryotic expression recombinant vector (pCB6(+)/WTA) containing full-length human WT1 isoform (WTA: -17aa/-KTS) cDNA and the vacant vector-alone were introduced into the leukemia cell line NB4 respectively by electroporation. The WTA mRNA and protein in cells were detected by RT-PCR and Western blot. Binding of Annexin V were tested by flow cytometry and agarose gel electrophoresis to verify whether exogenous WTA could induce apoptosis of NB4 cells. Expressions of p21, p53, bcl-2, bcl-XL and c-myc genes were determined by semi-quantitative RT-PCR after introducing recombinant vectors into the NB4 cells. The results showed that in exposure to As(2)O(3) at 0.8 micromol/L for 48 hours, the NB4/WTA cells exhibited the morphological hallmarks of apoptosis, the marked DNA ladder shown by gel electrophoresis, and the enhanced apoptosis rate marked by Annexin V. RT-PCR showed an increase in p21 and c-myc genes expression, a decrease in bcl-2 and a relative constant expression of p53, bcl-XL in NB4/WTA cells. It is concluded that the introduction and expression of exogenous WTA gene can lead to apoptosis of NB4/WTA cells by down-regulating the Bcl-2 gene expression and up-regulating the p21 and c-myc genes expression.
Apoptosis ; genetics ; physiology ; Blotting, Western ; Cell Line, Tumor ; Cyclin-Dependent Kinase Inhibitor p21 ; genetics ; Gene Expression Regulation, Neoplastic ; Humans ; Leukemia ; genetics ; metabolism ; pathology ; Proto-Oncogene Proteins c-bcl-2 ; genetics ; Proto-Oncogene Proteins c-myc ; genetics ; RNA, Messenger ; biosynthesis ; genetics ; Reverse Transcriptase Polymerase Chain Reaction ; Transfection ; Tumor Suppressor Protein p53 ; genetics ; WT1 Proteins ; genetics ; metabolism ; physiology ; bcl-X Protein ; genetics