Effect of Cyclin A2 on the proliferation of rat primary cardiomyocytes

doi:10.16098/j.issn.0529-1356.2015.01.010

Abstract

Abstract:

Objective To investigate the effect of Cyclin A2 on the proliferation of cardiomyocytes. Methods Twelve SD neonatal rats were used, cardiomyocytes were isolated from the neonatal rat heart, cultured in dish and identified by immunofluorescence. The primary cultured cardiomyocytes were divided into three groups: group 1 which cells were transduced with adenovirus expressing Cyclin A2 and eGFP, group 2 which cells were transduced with andenovirus expressing only eGFP and group 3 in which, untransduced cells were used as negative control. eGFP was employed to determine transduction efficiency. The expression of Cyclin A2, phosphorylated histone H3 (H3P), cardiac troponin -T (cTnT) were detected by immunofluorescence at day 3 after cardiomyocytes were transduced with andenovirus. Results 1. Analysis of eGFP expression showed that the transduction efficiency was (97±0.74)%. 2. cTnT, a specific marker protein for myocardial cells, was mainly found in the cytoplasm in eGFP-expressing and untransduced cells, suggesting that adenovirus/eGFP had no toxicity in the cultured cardiomyocytes. Immuno-staining results indicated that the percentage of isolated cTnT positive cells was (95±0.62)%. CyclinA2 was expressed mainly in the nucleus. H3P was also localized to the nucleus.3. In the CyclinA2 -overexpressing cells, the percentage of H3P positive cells were significantly increased compared to those untransduced or only eGFP-expressing cells (P<0.05). The experimental group showed a large number of multinucleated cells, in which the dual-core cell-based, followed by three nuclear cells. Conclusion Adenovirus vector transfection of primary cardiomyocytes has a good infection efficiency. Cyclin A2-overexpression in primary cardiomyocytes promotes to form a dual-core.

Key words: Cyclin A2, Cardiomyocyte, Proliferation, Gene transfection, Immunofluorescence, Rat

LU Xing ZHANG Jin-zhong GUO Zhi-kun CHANG Yu-qiao JIA Yin-ming XU Zhen-ping*. Effect of Cyclin A2 on the proliferation of rat primary cardiomyocytes[J]. AAS, 2015, 46(1): 57-62.

References

［1］Jopling C, Sleep E, Raya M, et al. Zebrafish heart regeneration occurs by cardiomyocyte dedifferentiation and proliferation ［J］.Nature, 2010, 464(7288):606-609.
［2］Porrello ER, Mahmoud AI, Simpson E, et al. Transient regenerative potential of the neonatal mouse heart ［J］. Science, 2011, 331(6020): 1078-1080.
［3］Gong D, Ferrell JE Jr. The roles of cyclin A2, B1, and B2 in early and late mitotic events ［J］. Mol Biol Cell, 2010, 21(18): 3149-3161. 
［4］Pagano M, Pepperkok R, Verde F, et al. Cyclin A is required at two points in the human cell cycle ［J］. EMBO, 1992, 11(3):961-971.
［5］Cheng RK, Asai T, Tang H, et al. Cyclin A2 induces cardiac regeneration after myocardial infarction and prevents heart failure［J］. Circ Res, 2007, 100(12): 1741-1748. 
［6］Chaudhry HW, Dashoush NH, Tang H, et al. Cyclin A2 mediates cardiomyocyte mitosis in the postmitotic myocardium ［J］. Biol Chem, 2004, 279(34): 35858-35866.
［7］Di J, Zhang LZh, Bai ShL. Phospho-histone H3-An approach to detect the mitotic index of cardiomyocytes［J］. Acta Anatomica Sinica, 2008, 39(2): 272-274. (in Chinese)
邸菁，张凌志，柏树令. 磷酸化组蛋白H3-检测心肌细胞有丝分裂指数的可靠指标［J］.解剖学报，2008，39(2): 272-274.
［8］Bolli P, Vardabasso C, Bernstein E, et al. Chromatin immunoprecipitation of adult murine cardiomyocytes ［J］. Curr Protoc Cell Biol, 2013, Chapter 17: Unit 1714
［9］Nurse P. A long twentieth century of the cell cycle and beyond ［J］. Cell, 2000, 100(1):71-78.
［10］Sherr CJ, Roberts JM, Inhibitors of mammalian G1cyclin-dependent kinases ［J］. Genes Dev, 1995, 9(10), 1149-1163.
［11］Castanedo G, Clark K, Wang S, et al. CDK2/cyclinA inhibitors: targeting the cyclinA recruitment site with small molecules derived from peptide leads ［J］. Bioorg Med Chem Lett, 2006, 16(6): 1716-1720.
［12］Chaudhry HW, Dashoush NH, Tang H, et al. Cyclin A2 mediates cardiomyocyte mitosis in the postmitotic myocardium ［J］. Biol Chem, 2004, 279(34):35858-35866.
［13］Shapiro SD, Ranjan AK, Kawase Y, et al. Cyclin A2 induces cardiac regeneration after myocardial infarction through cytokinesis of adult cardiomyocytes ［J］. Sci Transl Med, 2014, 6(224): 224-227.
［14］Cheng RK, Asai T, Tang H, et al. Cyclin A2 induces cardiac regeneration after myocardial infarction and prevents heart failure［J］. Circ Res, 2007, 100(12):1741-1748.
［15］Kabeche L, Compton DA. Cyclin A regulates kinetochore microtubules to promote faithful chromosome segregation ［J］. Nature, 2013, 502(7469): 110-113.

[1]	YANG Meng-yi NIU Shi-bo ZHANG Jing DANG Jie MA Zhan-bing LU Hong HUO Zheng-hao. Association between index finger and ring finger length ratios and polymorphism of homeobox A11 gene locus among Ningxia college students [J]. Acta Anatomica Sinica, 2024, 55(1): 62-66.
[2]	XIE Hui-lan FANG Fang LIN Yi. Mechanism of Chir99021 regulating Wnt/β-catenin signaling pathway inhibiting osteogenic differentiation of rat dental pulp stem cells [J]. Acta Anatomica Sinica, 2024, 55(1): 67-72.
[3]	WEI Xi-xi LI Chao-hong ZHAO Chen-lu TANG Jia-ping LIU Yu-zhen. Characterization of group Ⅰ metabotropic glutamate receptors in rat superior cervical ganglion and their changes following chronic intermittent hypoxia [J]. Acta Anatomica Sinica, 2024, 55(1): 3-9.
[4]	MA Ting-ting CHEN Jian-hong LIU Ai-cui LI Hai-ning. Role of inhibiting lncRNA TUG1 to down-regulate nucleotide binding oligomerization domain like receptor protein 1 inflammasome in delaying the progression of Alzheimer’s disease [J]. Acta Anatomica Sinica, 2024, 55(1): 32-42.
[5]	SU Peng WANG Xing-yi LIANG Jing-yan XIONG Tian-qing. Optimization and identification of a low density and high purity method for primary hippocampal neuron culture from fetal rats [J]. Acta Anatomica Sinica, 2024, 55(1): 113-119.
[6]	HAO Yong-ming GUO Lei ZHOU Cheng-hui PEI Han-jun LI Li ZHAO Zhe . Establishment of a rat model of myocardial hypertrophy by a modified abdominal aortic coarctation method [J]. Acta Anatomica Sinica, 2024, 55(1): 120-124.
[7]	XU Ya-ping YU Yue-xin CHEN Jin-fu WANG Ke-ke GUO Zhikun . Structural distribution and mechanism of elastic fibers and collagen fibers in ventricular interstitium of aged rats [J]. Acta Anatomica Sinica, 2023, 54(6): 716-721.
[8]	LIU Li-ping ZHU Meng-ni XU Hui . Effect of interleukin-6 on nucleated erythrocytes in lipopolysaccharide induced preeclampsia rats [J]. Acta Anatomica Sinica, 2023, 54(6): 722-729.
[9]	ZHAO Wei-ming LI Xiao YU Guo-ying WANG Gai-ping CHANG Cui-fang . Effect of serine protease inhibitor Kazaltype 1 on the proliferation of hepatocellular carcinoma cell and its mechanism [J]. Acta Anatomica Sinica, 2023, 54(6): 695-702.
[10]	WANG Wen-juan DING Yu-tong SU Hao REN Jie SUN Yan-rong WANG Han-fei LU Jia-li ZHANG Lin-qian BAI Yu QIN Li-hua. Changes of Nogo-A expression in hippocampus and striatum of rats under low estrogen condition [J]. Acta Anatomica Sinica, 2023, 54(6): 620-627.
[11]	LIANG Pan-pan YU Ai-mei DU Jing KOU Wen-hui WANG Huan-huan SONG Ai-xia. Inhibitory effect of sodium ferulate on inflammatory response in migraine rats based on c-Jun N-terminal kinase/p38 mitogen-activated protein kinase signaling pathway [J]. Acta Anatomica Sinica, 2023, 54(6): 652-659.
[12]	YAO Su-hua YAN Jun XIE Fang LIN Chun-hua HUANG Zheng-chun. Nerve repair effect of olanzapine-mediated cyclic AMP response element binding protein/brain-derived neurotrophic factor/receptor tyrosine kinase receptors B pathway on schizophrenic rats [J]. Acta Anatomica Sinica, 2023, 54(6): 660-667.
[13]	XU Ying LIU Bin ZHENG Xing HE Zong-zhao. Role and mechanism of complement C3a receptor in the cognitive impairment of septic rats induced by cecal ligation and perforation [J]. Acta Anatomica Sinica, 2023, 54(6): 668-675.
[14]	LIU Zhe-chuan LI Kun MA Shuai-nan MENG Jia-qi WANG Yan-qin. Effects of liraglutide on inflammation and mitochondrial fusion/division in Parkinson’s disease model of mice induced by paraquat [J]. Acta Anatomica Sinica, 2023, 54(6): 676-681.
[15]	LI Xia ZHU Shi-jie TANG Zhong-sheng LUO Ya-fei FAN Rui-juan XIE Gao-yu KOU Yun-fang LU Ying. Mechanism of electroacupuncture regulating the ligand pathway of tyrosine kinase receptor in improving vascular dementia [J]. Acta Anatomica Sinica, 2023, 54(6): 689-694.