Abstract 76: Meox1 is a Cell-cycle Oscillator Required for Mitotic Transition and Proliferation in Cardiac Fibroblasts

2014 ◽  
Vol 115 (suppl_1) ◽  
Author(s):  
Shuichiro Higo ◽  
Yoshihiro Asano ◽  
Yuki Masumura ◽  
Yasushi Sakata ◽  
Masafumi Kitakaze ◽  
...  
Keyword(s):  
Heart Failure ◽  
Cell Cycle ◽  
Cardiac Fibroblasts ◽  
Tissue Fibrosis ◽  
Cell Cycle Synchronization ◽  
M Phase ◽  
A Cell ◽  
End Stage

Background: Tissue fibrosis plays important roles in the pathogenesis of chronic diseases, including heart failure. The mechanism underlying interstitial fibroblast proliferation is a promising analytical target for therapeutic applications. Here we developed quantitative epigenome profiling to identify a critical regulator in interstitial cell populations that emerges during the progression of heart failure. Methods and Results: We subjected pressure-overloaded hearts of mice to trimethylated histone H3 lysine 4 (H3K4me3) ChIP-sequence and RNA-sequence. Expression analysis followed by quantitative H3K4me3 profiling identified 45 fibrosis-related genes with significant H3K4me3 enrichment in failing hearts, including Meox1 transcription factor. Meox1 emerged in the interstitial fibrotic region in failing heart, and intriguingly Meox1 was expressed in the limited population of cardiac fibroblasts both in vivo and in vitro. Meox1-positive fibroblasts were increased in response to a paracrine signal from cardiomyocytes, and knockdown of Meox1 completely inhibited the reactive proliferation of cardiac fibroblasts stimulated by conditioned medium from cardiomyocytes. Gene expression profiling combined with siRNAs clarified that Meox1 depletion resulted in down regulation in the mitosis-related genes including Aurora B kinase. Indeed, Meox1 depletion decreased the cells under mitosis, but conversely increased the proportion of DNA synthesizing cells, thereby inhibited mitotic transition. The cell-cycle synchronization analysis and promoter analysis using live-cell imaging clarified that Meox1 oscillated throughout the cell-cycle and specifically emerged in G2/M phase. Finally, we revealed that Meox1 heterogenously expressed in the interstitial fibrotic are of human ventricular heart tissues from patients with end-stage heart failure. Notably, Meox1 expression was significantly correlated with the fibrosis-related genes in diseased ventricular heart tissues (n=15), suggesting the pathological relevance in clinical settings. Conclusion: Our findings identify a novel cell-cycle regulator and propose that Meox1 is a potential target for therapies aimed at preventing tissue fibrosis.

scholarly journals Cardiomyocytes fuse with surrounding noncardiomyocytes and reenter the cell cycle

2004 ◽  
Vol 167 (2) ◽  
pp. 351-363 ◽  
Author(s):  
Katsuhisa Matsuura ◽  
Hiroshi Wada ◽  
Toshio Nagai ◽  
Yoshihiro Iijima ◽  
Tohru Minamino ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Endothelial Cells ◽  
Adult Stem Cells ◽  
Bone Marrow Cells ◽  
Cardiac Fibroblasts ◽  
Neonatal Cardiomyocytes ◽  
Marker Proteins ◽  
M Phase

The concept of the plasticity or transdifferentiation of adult stem cells has been challenged by the phenomenon of cell fusion. In this work, we examined whether neonatal cardiomyocytes fuse with various somatic cells including endothelial cells, cardiac fibroblasts, bone marrow cells, and endothelial progenitor cells spontaneously in vitro. When cardiomyocytes were cocultured with endothelial cells or cardiac fibroblasts, they fused and showed phenotypes of cardiomyocytes. Furthermore, cardiomyocytes reentered the G2-M phase in the cell cycle after fusing with proliferative noncardiomyocytes. Transplanted endothelial cells or skeletal muscle–derived cells fused with adult cardiomyocytes in vivo. In the cryoinjured heart, there were Ki67-positive cells that expressed both cardiac and endothelial lineage marker proteins. These results suggest that cardiomyocytes fuse with other cells and enter the cell cycle by maintaining their phenotypes.

scholarly journals Romidepsin Induces G2/M Phase Arrest and Apoptosis in Cholangiocarcinoma Cells

2020 ◽  
Vol 19 ◽  
pp. 153303382096075
Author(s):  
Pihong Li ◽  
Luguang Liu ◽  
Xiangguo Dang ◽  
Xingsong Tian
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Arrest ◽  
Antitumor Effect ◽  
Caspase 3 ◽  
M Cell ◽  
Cycle Arrest ◽  
Phase Arrest ◽  
M Phase

Background: Cholangiocarcinoma (CCA) is an extremely intractable malignancy since most patients are already in an advanced stage when firstly discovered. CCA needs more effective treatment, especially for advanced cases. Our study aimed to evaluate the effect of romidepsin on CCA cells in vitro and in vivo and explore the underlying mechanisms. Methods: The antitumor effect was determined by cell viability, cell cycle and apoptosis assays. A CCK-8 assay was performed to measure the cytotoxicity of romidepsin on CCA cells, and flow cytometry was used to evaluate the effects of romidepsin on the cell cycle and apoptosis. Moreover, the in vivo effects of romidepsin were measured in a CCA xenograft model. Results: Romidepsin could reduce the viability of CCA cells and induce G2/M cell cycle arrest and apoptosis, indicating that romidepsin has a significant antitumor effect on CCA cells in vitro. Mechanistically, the antitumor effect of romidepsin on the CCA cell lines was mediated by the induction of G2/M cell cycle arrest and promotion of cell apoptosis. The G2/M phase arrest of the CCA cells was associated with the downregulation of cyclinB and upregulation of the p-cdc2 protein, resulting in cell cycle arrest. The apoptosis of the CCA cells induced by romidepsin was attributed to the activation of caspase-3. Furthermore, romidepsin significantly inhibited the growth of the tumor volume of the CCLP-1 xenograft, indicating that romidepsin significantly inhibited the proliferation of CCA cells in vivo. Conclusions: Romidepsin suppressed the proliferation of CCA cells by inducing cell cycle arrest through cdc2/cyclinB and cell apoptosis by targeting caspase-3/PARP both in vitro and in vivo, indicating that romidepsin is a potential therapeutic agent for CCA.

Protein kinase D inhibitor CRT0066101 suppresses bladder cancer growth in vitro and in vivo through induction of cell cycle arrest at the G2-M phase.

2017 ◽  
Vol 35 (15_suppl) ◽  
pp. e16024-e16024
Author(s):  
Qingdi Quentin Li ◽  
Iawen Hsu ◽  
Thomas Sanford ◽  
Reema S. Railkar ◽  
Piyush K. Agarwal
Keyword(s):  
Cell Cycle ◽  
Flow Cytometry ◽  
Bladder Cancer ◽  
Cyclin B1 ◽  
Cancer Growth ◽  
Protein Kinase D ◽  
Bladder Cancer Cells ◽  
M Phase

e16024 Background: Protein Kinase D (PKD) is implicated in tumor growth, death, invasion, and progression. CRT0066101 is an inhibitor of PKD and has antitumor activity in several types of carcinomas. However, the effect and mechanism of CRT0066101 in bladder cancer remain unknown. Methods: The MTS assay was used to evaluate the ability of CRT0066101 to inhibit cellular proliferation in bladder cancer cells. Cell cycle was analyzed by flow cytometry. Protein expression and phosphorylation were assessed by western blotting. Results: We showed that CRT0066101 suppressed the proliferation and migration of 4 bladder cancer cell lines in vitro. We also demonstrated that CRT0066101 inhibited tumor growth in an in vivo mouse model of bladder cancer. To verify the role of PKD in bladder tumor, we found that PKD2 was highly expressed in 8 bladder cancer lines and that RNA interference-mediated silencing of the PKD2 gene dramatically reduced bladder cancer growth in vitro and in vivo, suggesting that the effect of the compound in bladder cancer is mediated through inhibition of PKD2. This notion was confirmed by demonstrating that the levels of PKD2 and phospho-PKD2 (Ser-876) were markedly decreased in CRT0066101-treated bladder cancer. In addition, our cell cycle analysis by flow cytometry revealed that CRT0066101 arrested bladder cancer cells at the G2-M phase. We further validated these data by immunoblotting showing that treatment of bladder carcinoma cells with CRT0066101 downregulated the expression of cyclin B1, cdc2 and cdc25C, but elevated the levels of p27kip1, gadd45a, chk1/2, and wee1. Finally, CRT0066101 was found to increase the phosphorylation of cdc2 and cdc25C, which lead to reduction in cdc2-cyclin B1 activity. Conclusions: These novel findings suggest that CRT0066101 inhibits bladder cancer growth through modulating the cell cycle G2 checkpoint and inducing cell cycle G2-M arrest, which lead to blockade of cell cycle progression. QQL and IH contributed equally to this work.

scholarly journals Th1 effector T cells selectively orchestrate cardiac fibrosis in nonischemic heart failure

2017 ◽  
Vol 214 (11) ◽  
pp. 3311-3329 ◽  
Author(s):  
Tania Nevers ◽  
Ane M. Salvador ◽  
Francisco Velazquez ◽  
Njabulo Ngwenyama ◽  
Francisco J. Carrillo-Salinas ◽  
...  
Keyword(s):  
Heart Failure ◽  
T Cells ◽  
Cardiac Fibrosis ◽  
Cardiac Fibroblasts ◽  
Th1 Cells ◽  
Dependent Manner ◽  
Effector Cells ◽  
Ifn Γ

Despite emerging data indicating a role for T cells in profibrotic cardiac repair and healing after ischemia, little is known about whether T cells directly impact cardiac fibroblasts (CFBs) to promote cardiac fibrosis (CF) in nonischemic heart failure (HF). Recently, we reported increased T cell infiltration in the fibrotic myocardium of nonischemic HF patients, as well as the protection from CF and HF in TCR-α−/− mice. Here, we report that T cells activated in such a context are mainly IFN-γ+, adhere to CFB, and induce their transition into myofibroblasts. Th1 effector cells selectively drive CF both in vitro and in vivo, whereas adoptive transfer of Th1 cells, opposite to activated IFN-γ−/− Th cells, partially reconstituted CF and HF in TCR-α−/− recipient mice. Mechanistically, Th1 cells use integrin α4 to adhere to and induce TGF-β in CFB in an IFN-γ–dependent manner. Our findings identify a previously unrecognized role for Th1 cells as integrators of perivascular CF and cardiac dysfunction in nonischemic HF.

In vivo and in vitro phosphorylation studies of numatrin, a cell cycle regulated nuclear protein, in insulin-stimulated NIH 3T3 HIR cells

1991 ◽  
Vol 194 (2) ◽  
pp. 289-296 ◽  
Author(s):  
Nili Feuerstein ◽  
Paul A. Randazzo
Keyword(s):  
Cell Cycle ◽  
Nuclear Protein ◽  
A Cell ◽  
Nih 3T3

scholarly journals 1,2:5,6-dianhydrogalactitol inhibits human glioma cell growth in vivo and in vitro by arresting the cell cycle at G2/M phase

2017 ◽  
Vol 38 (4) ◽  
pp. 561-570 ◽  
Author(s):  
Chun Peng ◽  
Xin-ming Qi ◽  
Ling-ling Miao ◽  
Jin Ren
Keyword(s):  
Cell Cycle ◽  
Cell Growth ◽  
Glioma Cell ◽  
Human Glioma ◽  
Human Glioma Cell ◽  
M Phase

scholarly journals The Cak1p Protein Kinase Is Required at G1/S and G2/M in the Budding Yeast Cell Cycle

Genetics ◽  
1997 ◽  
Vol 147 (1) ◽  
pp. 57-71 ◽  
Author(s):  
Ann Sutton ◽  
Richard Freiman
Keyword(s):  
Cell Cycle ◽  
Protein Kinase ◽  
Cell Cycle Progression ◽  
Budding Yeast ◽  
Protein Kinase Activity ◽  
Synthetic Lethal ◽  
Protein Levels ◽  
M Phase

Abstract The CAK1 gene encodes the major CDK-activating kinase (CAK) in budding yeast and is required for activation of Cdc28p for cell cycle progression from G2 to M phase. Here we describe the isolation of a mutant allele of CAK1 in a synthetic lethal screen with the Sit4 protein phosphatase. Analysis of several different cak1 mutants shows that although the G2 to M transition appears most sensitive to loss of Cak1p function, Cak1p is also required for activation of Cdc28p for progression from G1 into S phase. Further characterization of these mutants suggests that, unlike the CAK identified from higher eukaryotes, Cak1p of budding yeast may not play a role in general transcription. Finally, although Cak1 protein levels and in vitro protein kinase activity do not fluctuate during the cell cycle, at least a fraction of Cak1p associates with higher molecular weight proteins, which may be important for its in vivo function.

Abstract 331: Mir-433 Controls Cardiac Fibrosis

2015 ◽  
Vol 117 (suppl_1) ◽  
Author(s):  
Lichan Tao ◽  
Xiaoting Wu ◽  
Ping Chen ◽  
Shanshan Li ◽  
Xiaomin Zhang ◽  
...  
Keyword(s):  
Myocardial Infarction ◽  
Cardiac Fibrosis ◽  
Cardiac Fibroblasts ◽  
Heart Diseases ◽  
Mice Model ◽  
Ki 67 ◽  
End Stage ◽  
Common Manifestation

Background: Cardiac fibrosis, a result of multiple injurious insults in heart, is a final common manifestation of chronic heart diseases and can lead to end-stage cardiac failure. MicroRNAs (miRNAs, miRs) participate in many essential biological processes and their dysfunction has been implicated in a variety of cardiovascular diseases including fibrosis. miR-433 has recently been implicated in renal fibrosis, however, its role in cardiac fibrosis is unclear. Methods and results: miR-433 was increased in heart samples from dilated cardiomyopathy patients as determined by qRT-PCRs. In addition, miR-433 was also consistently upregulated in mice model of cardiac fibrosis after myocardial infarction or heart failure. Additionally, miR-433 was found to be enriched in fibroblasts compared to cardiomyocytes. In neonatal cardiac fibroblasts, forced expression of miR-433 promoted cell proliferation as indicated by EdU and Ki-67 staining. Moreover, miR-433 overexpression promoted the transdifferentiation of fibroblasts into myofibroblasts as determined by qRT-PCR and western blot for α-SMA and collagen whether in the presence of TGF-β or not, indicating that miR-433 is sufficient to induce fibrosis. In addition, knockdown of miR-433 inhibited proliferation and the transdifferentiation into myofibroblasts, indicating that miR-433 is required for cardiac fibrosis. Interestingly, miR-433 did not affect the migration of cardiac fibroblast. Importantly, miR-433 antagomir could partially attenuate cardiac fibrosis induced by myocardial infarction in mice. Conclusion: both in vitro and in vivo. Inhibition of miR-433 represents a novel therapeutic strategy for cardiac fibrosis.

scholarly journals BRCA1 Phosphorylation by Aurora-A in the Regulation of G2to M Transition

2004 ◽  
Vol 279 (19) ◽  
pp. 19643-19648 ◽  
Author(s):  
Mutsuko Ouchi ◽  
Nobuko Fujiuchi ◽  
Kaori Sasai ◽  
Hiroshi Katayama ◽  
Yohji A. Minamishima ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Phosphorylation Site ◽  
Site Directed Mutagenesis ◽  
Aurora A ◽  
M Phase ◽  
Cancer Tumor ◽  
A Site ◽  
Interfering Rna

Aurora-A/BTAK/STK15 localizes to the centrosome in the G2-M phase, and its kinase activity regulates the G2to M transition of the cell cycle. Previous studies have shown that the BRCA1 breast cancer tumor suppressor also localizes to the centrosome and that BRCA1 inactivation results in loss of the G2-M checkpoint. We demonstrate here that Aurora-A physically binds to and phosphorylates BRCA1. Biochemical analysis showed that BRCA1 amino acids 1314–1863 binds to Aurora-A. Site-directed mutagenesis indicated that Ser308of BRCA1 is phosphorylated by Aurora-Ain vitro. Anti-phospho-specific antibodies against Ser308of BRCA1 demonstrated that Ser308is phosphorylatedin vivo. Phosphorylation of Ser308increased in the early M phase when Aurora-A activity also increases; these effects could be abolished by ionizing radiation. Consistent with these observations, acute loss of Aurora-A by small interfering RNA resulted in reduced phosphorylation of BRCA1 Ser308, and transient infection of adenovirus Aurora-A increased Ser308phosphorylation. Mutation of a single phosphorylation site of BRCA1 (S308N), when expressed in BRCA1-deficient mouse embryo fibroblasts, decreased the number of cells in the M phase to a degree similar to that with wild type BRCA1-mediated G2arrest induced by DNA damage. We propose that BRCA1 phosphorylation by Aurora-A plays a role in G2to M transition of cell cycle.

scholarly journals Kallistatin/ serpina3c inhibits fibrosis after myocardial infarction by regulating glycolytic pathway

2021 ◽  
Vol 42 (Supplement_1) ◽  
Author(s):  
J.J Ji ◽  
Y.U.Y.U Yao
Keyword(s):  
Myocardial Infarction ◽  
Heart Failure ◽  
Transcriptional Activation ◽  
Cardiac Fibroblasts ◽  
Rna Seq ◽  
Funding Sources ◽  
Proliferation And Differentiation ◽  
Key Enzyme

Abstract Background Fibrotic remodeling is the main pathological mechanism of heart failure after infarction. Studies have shown that the serine protease inhibitor Kallistatin/Serpina3c could inhibit the progression of myocardial hypertrophy and fibrosis after myocardial infarction, but the specific mechanism is unknown. Methods and results We used Serpina3c−/− and C57BL/6 mice to construct a MI model in vivo. TGF-β and hypoxia interfered with cardiac fibroblasts (CFs) to construct a fibrosis model in vitro. RNA-seq assessed the effect of Serpina3c knockout on the transcriptome. CCK-8 and BrdU were used to assess cell proliferation. Western blot and qPCR were used to verify the changes of glycolytic enzymes in the sequencing results. Co-IP was used to verify the interaction between Serpina3c and NR4A1. The results showed that Serpina3c expression decreased in the fibrotic tissue of MI heart and in the fibrosis model in vitro. After Serpina3c knockout, the degree of myocardial infarction fibrosis aggravated, and the proliferation and differentiation of CFs increased. Up-regulation of Serpina3c expression in CFs had the opposite effect, suggesting that Serpina3c had the ability to resist myocardial fibrosis. RNA-Seq results showed that glycolysis-related genes were significantly increased after Serpina3c knockout. Among them, the key enzyme of glycolysis, enolase (ENO1), had the largest change, and at the same time, the glycolysis reaction was enhanced. Inhibition of ENO1 can antagonize the promotion of Serpina3c knockout on the proliferation and differentiation of CFs. Activating the activity of NR4A1 could negatively regulate the expression of ENO1 and then inhibited the proliferation and differentiation of CFs. Conclusions Serpina3c inhibited the transcriptional activation of ENO1 by binding to NR4A1, and then reduced the fibrosis after myocardial infarction by inhibiting glycolysis. In short, Serpina3c provides a potential intervention target for the prevention and treatment of heart failure after infarction. FUNDunding Acknowledgement Type of funding sources: Foundation. Main funding source(s): National Nature Science Foundation of China (No. 81770452) Serpina3c is a key protein in MI.

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