Caspase-9 in myoblasts is involved in mechanical signal transdunction under cyclic stretch

doi:10.3969/j.issn.2095-4344.2014.15.015

Abstract

Abstract:

BACKGROUND: The adaptive reconstruction of maxillofacial muscles would happen when functional orthopedic treatment is done to cure micromaxillary deformity. The myoblast is the main responder in the process of adaptive reconstruction, and cyclic stretch can induce apoptosis of myoblasts. Caspase-9 is an important factor in the mitochondrial apoptosis pathway.
OBJECTIVE: To investigate the expression of Caspase-9 in different cyclic stretch.
METHODS: Based on myoblasts cultured in vitro-mechanical stimulation model, the rat L6 myoblasts were loaded stretch for 1, 6, 12 and 24 hours through multi-channel cell stress loading system, while the control group received no stretch. The morphological change and growth of myoblasts were observed under inverted phase contrast microscope; the expression of the mRNA and protein of Caspase-9 were detected by RT-PCR and western blot analysis, respectively.
RESULTS AND CONCLUSION: Under inverted phase contrast microscope, the rat L6 myoblasts at cyclic stretch maintained a good growth state and biological characteristics; there was no cell degeneration; and the loss rate was extremely low, which could demonstrate that myoblast in vitro-mechanical stimulation model was established successfully. The results of RT-PCR and western blot analysis showed that, the expression of Caspase-9 mRNA and Cleaved Caspase-9 protein was significantly increased as the loading time prolonged, and the expression of Procaspase-9 protein was significantly decreased as the time. We can conclude that Caspase-9 is involved in the mechanical signal transduction of cyclic stretch.

中国组织工程研究杂志出版内容重点：组织构建；骨细胞；软骨细胞；细胞培养；成纤维细胞；血管内皮细胞；骨质疏松；组织工程

全文链接：

Key words: Caspase-9, myoblasts, apoptosis, mitochondria, reverse transcription polymerase chain reaction

CLC Number:

R318

Wang Shuang-yu, Wang Hong-ling, Xia Chen-lei, Ding Xian, Sun Xian-rui, Zhang Qiang, Li Jian-ping, Yan Xiao, Liu Wen, Zhang Yue, Yao Ru-yong, Yuan Xiao. Caspase-9 in myoblasts is involved in mechanical signal transdunction under cyclic stretch[J]. Chinese Journal of Tissue Engineering Research, 2014, 18(15): 2383-2389.

Figures/Tables 7

References

[1]Matsunaga K, Usui A, Yamaguchi K, et al. An anatomical study of the muscles that attach to the articular disc of the temporomandibular joint. Clin Anat. 2009;22(8):932-940.
[2]Tanaka E, Hirose M, Inubushi T, et al. Effect of hyperactivity of the lateral pterygoid muscle on the temporomandibular joint disk. J Biomech Eng. 2007;129(6):890-897.
[3]Wang L, Pan J, Wang T, et al. Pathological cyclic strain-induced apoptosis in human periodontal ligament cells through the RhoGDIalpha/caspase-3/PARP pathway. PLoS One. 2013;8(10):e75973.
[4]Rust C, Gores GJ. Apoptosis and liver disease. Am J Med. 2000;108(7):567-574.
[5]Liu J, Wang Y, Yuan X, et al. Cyclic-stretch induces the apoptosis of myoblast by activation of Caspase-3 protease in a magnitude-dependent manner. Int J Biochem Cell Biol. 2010;42(12):2004-2011.
[6]唐惠玲.线粒体介导的细胞凋亡的研究进展[J].安徽医药,2012, 16(11):1696-1699.
[7]Jiang X, Wang X. Cytochrome C-mediated apoptosis. Annu Rev Biochem. 2004;73:87-106.
[8]Huttemann M, Helling S, Sanderson TH, et al. Regulation of mitochondrial respiration and apoptosis through cell signaling: cytochrome c oxidase and cytochrome c in ischemia/reperfusion injury and inflammation. Biochim Biophys Acta. 2012;1817(4):598-609.
[9]Gu S, Zhao Y, Guo J, et al. High expression of APAF-1 elevates erythroid apoptosis in iron overload myelodysplastic syndrome. Tumour Biol. 2014;35(3): 2211-2218.
[10]Sun KW, Ma YY, Guan TP, et al. Oridonin induces apoptosis in gastric cancer through Apaf-1, cytochrome c and caspase-3 signaling pathway. World J Gastroenterol. 2012;18(48):7166-7174.
[11]Shoshan-Barmatz V, Keinan N, Abu-Hamad S, et al. Apoptosis is regulated by the VDAC1 N-terminal region and by VDAC oligomerization: release of cytochrome c, AIF and Smac/Diablo. Biochim Biophys Acta. 2010;1797(6-7): 1281-1291.
[12]王辉,张琳,沈树娜. Caspases家族在妇科疾病中的研究进展[J]. 医学综述,2012,18(9):1314-1316.
[13]Porter AG, Janicke RU. Emerging roles of caspase-3 in apoptosis. Cell Death Differ. 1999;6(2):99-104.
[14]Salakou S, Kardamakis D, Tsamandas AC, et al. Increased Bax/Bcl-2 ratio up-regulates caspase-3 and increases apoptosis in the thymus of patients with myasthenia gravis. In Vivo. 2007;21(1):123-132.
[15]Renatus M, Stennicke HR, Scott FL, et al. Dimer formation drives the activation of the cell death protease caspase 9. Proc Natl Acad Sci U S A. 2001;98(25):14250-14255.
[16]Acehan D, Jiang X, Morgan DG, et al. Three-dimensional structure of the apoptosome: implications for assembly, procaspase-9 binding, and activation. Mol Cell. 2002; 9(2): 423-432.
[17]田臻,杨竹丽,贾文敏,等. p38MAPK信号通路与应力介导成肌细胞的凋亡(英文)[J].中国组织工程研究与临床康复,2011, 15(15):2751-2754.
[18]阎潇. Caspase-3在应力介导的成肌细胞凋亡中的作用及其机制[D].青岛:青岛大学,2012.
[19]Khan A, Farah CS, Savage NW, et al. Th1 cytokines in oral lichen planus. J Oral Pathol Med. 2003;32(2):77-83.
[20]胡锴,刘好朋,万婷,等. 线粒体与细胞凋亡的关系[J].中国畜牧兽医,2010,37(12):86-88.
[21]Czerniczyniec A, Bustamante J, Lores-Arnaiz S. Improvement of mouse brain mitochondrial function after deprenyl treatment. Neuroscience. 2007;144(2):685-693.
[22]Bellido T, Huening M, Raval-Pandya M, et al. Calbindin-D28k is expressed in osteoblastic cells and suppresses their apoptosis by inhibiting caspase-3 activity. J Biol Chem. 2000;275(34):26328-26332.
[23]朱炎杰,高维娟. Cyt-C与caspase在细胞凋亡中的作用及相互关系[J]. 中国老年学杂志,2011,31(15):2990-2992.
[24]刘晓婷,王延让,张明. 线粒体介导细胞凋亡的研究进展[J]. 环境与健康杂志,2013,28(2):182-185.
[25]Cande C, Cohen I, Daugas E, et al. Apoptosis-inducing factor (AIF): a novel caspase-independent death effector released from mitochondria. Biochimie. 2002;84(2-3): 215-222.
[26]Zou H, Li Y, Liu X, et al. An APAF-1.cytochrome c multimeric complex is a functional apoptosome that activates procaspase-9. J Biol Chem. 1999;274(17): 11549-11556.
[27]Hu Y, Benedict MA, Ding L, et al. Role of cytochrome c and dATP/ATP hydrolysis in Apaf-1-mediated caspase-9 activation and apoptosis. EMBO J. 1999;18(13):3586-3595.
[28]Saleh A, Srinivasula SM, Acharya S, et al. Cytochrome c and dATP-mediated oligomerization of Apaf-1 is a prerequisite for procaspase-9 activation. J Biol Chem. 1999;274(25):17941-17945.
[29]Brentnall M, Rodriguez-Menocal L, De Guevara RL, et al. Caspase-9, caspase-3 and caspase-7 have distinct roles during intrinsic apoptosis. BMC Cell Biol. 2013;14:32.
[30]Hirota M, Ito T, Okudela K, et al. Cell proliferation activity and the expression of cell cycle regulatory proteins in oral lichen planus. J Oral Pathol Med. 2002;31(4):204-212.
[31]Sutton VR, Davis JE, Cancilla M, et al. Initiation of apoptosis by granzyme B requires direct cleavage of bid, but not direct granzyme B-mediated caspase activation. J Exp Med. 2000; 192(10):1403-1414.
[32]Sitailo LA, Tibudan SS, Denning MF. Activation of caspase-9 is required for UV-induced apoptosis of human keratinocytes. J Biol Chem. 2002;277(22):19346-19352.
[33]Wurstle ML, Laussmann MA, Rehm M. The central role of initiator caspase-9 in apoptosis signal transduction and the regulation of its activation and activity on the apoptosome. Exp Cell Res. 2012;318(11):1213-1220.
[34]李志恒. 大鼠颅脑损伤后Caspase-9、MMP-3和NGB表达变化的研究[D]. 乌鲁木齐:新疆医科大学,2012.
[35]Guerrero AD, Schmitz I, Chen M, et al. Promotion of Caspase Activation by Caspase-9-mediated Feedback Amplification of Mitochondrial Damage. J Clin Cell Immunol. 2012;3(3). pii: 1000126.
[36]Chavez-Galan L, Sada-Ovalle I, Baez-Saldana R, et al. Monocytes from tuberculosis patients that exhibit cleaved caspase 9 and denaturalized cytochrome c are more susceptible to death mediated by Toll-like receptor 2. Immunology. 2012;135(4):299-311.
[37]Wang YJ, Niu XP, Yang L, et al. Effects of celecoxib on cycle kinetics of gastric cancer cells and protein expression of cytochrome C and caspase-9. Asian Pac J Cancer Prev. 2013; 14(4):2343-2347.
[38]Fang F, Gong PS, Zhao HG, et al. Mitochondrial Modulation of Apoptosis Induced by Low-dose Radiation in Mouse Testicular Cells. Biomed Environ Sci. 2013;26(10):820-830.
[39]刘江涛,郭雄,吴翠艳,等. 线粒体细胞色素c介导的caspase-9激活通路参与大骨节病关节软骨细胞凋亡的发生机制[J]. 西安交通大学学报:医学版,2011,32(5):580-584.
[40]曹军军,杨茂伟,郭宝磊,等. 高糖通过线粒体途径诱导成骨细胞凋亡[J]. 中国生物化学与分子生物学报,2012,28(12): 1109-1114.