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Objective To explore the role and mechanism of keratin 19 (KRT19) in breast cancer.Methods Quantitative real-time polymerase chain reaction (qRT-PCR) assay was used to determine KRT19 levles in 35 cases of breast cancer tissues and normal tissues.The correlation between KRT19 levels and clinical property of breast cancer was analyzed.Meanwhile,the expression levels of KRT19 in several breast cancer cells and mammary epithelial cell Michigan cancer foundation (MCF)-10A were evaluated by qRT-PCR assay.Over-expressed and knocked-down KRT19 in breast cancer ceils,3-(4,5-dimenthylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) assay was performed to detect the proliferation and chemosensitivity of these cells.The ability of forming colon of these breast cancer cells with treated KRT19 was studied via colony-forming unit assays.Western blot assay was performed to determine expression levels of ceil cycler related proteins.Results KRT19 was upregulated in breast cancer tissues comparing to normal tissues.KRT19 was positively related to tumor node metastasis (TNM) stage and distant metastasis of breast cancer.Similarly,KRT19 was highly expressed in breast cancer cells compared to mammary epithelial cell MCF-10A.The proliferation and colony-forming ability was significantly enhanced in MCF-7 cell with o,verexpressed KRT19.MTS assay showed that chemosensitivity of MCF-7 cells with overexpression of KRT19 was much more remarkably reduced than the control group.However,knocking down KRT19 in breast cancer cells MDA-MB-231 got the opposite results.Western blot assay suggested that KRT19 could obviously upregulated cyclin-dependent kinase 1 (CDK1) but not p27 expression levels.Conclusions KRT19 was upregulated in breast cancer tissues and was positively related to TNM stage and distant metastasis of breast cancer.KRT19 can significantly enhance proliferation and chemoresistance of breast cancer cells via upregulating CDK1.
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Objective To investigate the effect of artesunate on the cell cycle of human multiple myeloma RPMI8226 cells and explore the related molecularmechanisms. Methods RPMI8226 cellswere treated with 0, 25, and 50 pg/mL artesunate. The morphology change of cells was observed under transmission electron microscope, the cell cycle distribution was detected by flow cytometry, and the expression of cyclin B1 and p34cdc2 protein was examined by Western blotting analysis. Results After treated with 50 pg/mL artesunate for 48 h, most RPMI8226 cells showed characteristic morphology of apoptosis. With the increase of artesuate concentration, the proportion of RPMI8226 cells in G0/G1 phase was significantly decreased (P<0. 05) and that of cells in G2/M phase was significantly increased (P<0.05), suggesting that artesunate induced noticeable G2/M arrest. Cyclin B1 level was increased and the p34cdc2 level was decreased with the increase of artesuate concentration. Conclusion Artesunate can induce G2/M arrests in RPMI8226 cells, which may be related to the increasedcyclin B1 expression and decreased p34cdc2 expression.
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Objective Proliferative cholangitis (PC) is responsible for stone recurrence and biliary restenosis, this study was to investigate the and-proliferative effect of CDC2 kinase shRNA on PC. Methods The common bile duct of PC rat model was given an intralumenal administration of 0. 5 ml of CDC2 kinase shRNA. Results CDC2 kinase shRNA treatment effectively inhibited the expression of CDC2 kinase,PCNA, and procollagen I , resulting in the inhibition of hyperplasia of biliary epithelium, submucosal gland, and collagen fibers. Also, the lithogenic potentiality of PC decreased due to the inhibition of endogenous β-glucuronidase secretion. Conclusion The anti-proliferative effect of CDC2 kinase shRNA on PC may prevent biliary restenosis and stone recurrence.
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Objective To describe the spatial and temporal characteristics of neurofibrillary tangles (NFT) in brains of patients with Niemann-Pick disease type C (NPC). Methods In order to analyze formation of NFT in NPC, the brains collected from 17 patients with NPC aged from 7 months to 55 years old were investigated using antibodies against the protein tau and the specific proteins in mitotic phase. Results Typical NFT could be detected in the parahippocampns of a NPC patient as early as at 4 years old. The number of NFT were increasing along with the time. Gradually, the hippocampus and other regions of the temporal lobes and the frontal lobes would be affected with the time. Immunohistochemically, the NFT formed the shape similar to NFT seen in AD brains, without the presence of senile plagues. Interestingly,mitosis-phase markers appeared in the degenerating NPC neurons prior to hyperphesphorylation of tau and formation of NFT. Conclusions The formation of NFT does not result from aging, for there is no close relation between the presence of senile plagues and the formation of NFT. Ectopic activation of cdc2/cyclinB1 kinase complex might be an early event leading to NFT formation. Antagonist of the kinase complex may potentially slow down the formation of NFT.
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PURPOSE: Recent studies have suggested that p53 regulates the G2 checkpoint in the cell cycle and this function is required for the maintenance of genomic integrity. In this study, we addressed a role of p53 in escaping from cell cycle G2 arrest following DNA damage. MATERIALS AND METHODS: Cell cycle checkpoint arrest in the human colon cancer cell line HCT116 and its derivatives carry p53 or p21 deletions, were examined by FACS analysis, immunoprecipitation, Western blot and IP-kinase assay. RESULTS: While the cells with functional p53 were arrested at both the G1 and G2 checkpoints, the p53-deficient cells failed to arrest at G1, but they were arrested at G2. However, the p53-deficient cells failed to sustain G2 checkpoint arrest and they entered mitosis earlier than did the p53-positive cells and so this resulted in extensive cell death. Cdc2 kinase becomes reactivated in p53-deficient cells in association with entry into mitosis, but not in the p53-positive cells. Upon DNA damage, the p21-deficient cells, like the p53-negative cells, not only failed to repress cdk2- dependent NF-Y phosphorylation, but they also failed to repress the expression of such cell cycle G2-regulatory genes as cdc2, cyclin B, RNR-R2 and cdc25C, which have all been previously reported as targets of NF-Y transcription factor. CONCLUSION: p53 is essential to prevent immature escaping from cell cycle G2 checkpoint arrest through p21-mediated cdk2 inactivation, and this leads to inhibition of cdk2-dependent NF-Y phosphorylation and NF-Y dependent transcription of the cell cycle G2-rgulatory genes, including cdc2 and cyclin B.
Subject(s)
Humans , Blotting, Western , CCAAT-Binding Factor , CDC2 Protein Kinase , Cell Cycle Checkpoints , Cell Cycle , Cell Death , Cell Line , Colonic Neoplasms , Cyclin B , DNA Damage , G2 Phase , Immunoprecipitation , Mitosis , Phosphorylation , Phosphotransferases , Transcription Factors , Tumor Suppressor Protein p53 , United NationsABSTRACT
Objective To investigate the role of expressions of pituitary tumor transforming gene(PTTG) and p27 in glioma,and explore the relationship between the expressions of them and cell cycle regulation.Methods Specimens of 40 patient with gliomas were divided into gradeⅠ:8 cases,gradeⅡ:10 cases,gradeⅢ:13 cases, gradeⅣ:9 cases,PTTGmRNA and p27mRNA were detected by reverse transcription polymerase chain reaction (RT-PCR),and their expressions of PTTG,P27,CDK4 protein were detected by immunostaining assay using strep- tavidin-peroxidase(SP) method.Results The expressions of PTTGmRNA in gradeⅠ～Ⅳwere (0.907?0.065),(1.109?0.083),(1.312?0.089),(1.499?0.215) respectively,there was significant difference among the groups(P