Cyclic stretch promotes osteogenesis-related gene expression in osteoblast-like cells through a cofilin-associated mechanism

Gao,Jie; Fu,Shanmin; Zeng,Zhaobin; Li,Feifei; Niu,Qiannan; Jing,Da; Feng,Xue

doi:10.3892/mmr.2016.5239

Cyclic stretch promotes osteogenesis-related gene expression in osteoblast-like cells through a cofilin-associated mechanism

Authors:
- Jie Gao
- Shanmin Fu
- Zhaobin Zeng
- Feifei Li
- Qiannan Niu
- Da Jing
- Xue Feng
View Affiliations

Affiliations: State Key Laboratory of Military Stomatology, Department of Orthodontics, School of Stomatology, Fourth Military Medical University, Xi'an, Shaanxi 710032, P.R. China, Department of Stomatology, General Hospital of Shenyang Military Area Command, Shenyang, Liaoning 110084, P.R. China, Department of Biomedical Engineering, Fourth Military Medical University, Xi'an, Shaanxi 710032, P.R. China
Published online on: May 10, 2016 https://doi.org/10.3892/mmr.2016.5239
Pages: 218-224
Copyright: © Gao et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )

Metrics: Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )

Cited By (CrossRef): 0 citations Loading Articles...

Abstract

Osteoblasts have the capacity to perceive and transduce mechanical signals, and thus, regulate the mRNA and protein expression of a variety of genes associated with osteogenesis. Cytoskeletal reconstruction, as one of the earliest perception events for external mechanical stimulation, has previously been demonstrated to be essential for mechanotransduction in bone cells. However, the mechanism by which mechanical signals induce cytoskeletal deformation remains poorly understood. The actin‑binding protein, cofilin, promotes the depolymerization of actin and is understood to be important in the regulation of activities in various cell types, including endothelial, neuronal and muscle cells. However, to the best of our knowledge, the importance of cofilin in osteoblastic mechanotransduction has not been previously investigated. In the present study, osteoblast‑like MG‑63 cells were subjected to physiological cyclic stretch stimulation (12% elongation) for 1, 4, 8, 12 and 24 h, and the expression levels of cofilin and osteogenesis-associated genes were quantified with reverse transcription‑quantitative polymerase chain reaction, immunofluorescence staining and western blotting analyses. Additionally, knockdown of cofilin using RNA interference was conducted, and the mRNA levels of osteogenesis‑associated genes were compared between osteoblast‑like cells in the presence and absence of cofilin gene knockdown. The results of the present study demonstrated that cyclic stretch stimulates the expression of genes associated with osteoblastic activities in MG‑63 cells, including alkaline phosphatase (ALP), osteocalcin (OCN), runt‑related transcription factor 2 (Runx2) and collagen‑1 (COL‑1). Cyclic stretch also regulates the mRNA and protein expression of cofilin in MG‑63 cells. Furthermore, stretch‑induced increases in the levels of osteogenesis-associated genes, including ALP, OCN, Runx2 and COL‑1, were reduced following cofilin gene knockdown. Together, these results demonstrate that cofilin is involved in the regulation of mechanical load‑induced osteogenesis and, to the best of our knowledge, provides the first evidence demonstrating the importance of cofilin in osteoblastic mechanotransduction.

View Figures

View References

1	Harter LV, Hruska KA and Duncan RL: Human osteoblast-like cells respond to mechanical strain with increased bone matrix protein production independent of hormonal regulation. Endocrinology. 136:528–535. 1995.PubMed/NCBI
2	Frost HM: Bone's mechanostat: A 2003 update. Anat Rec A Discov Mol Cell Evol Biol. 275:1081–1101. 2003. View Article : Google Scholar : PubMed/NCBI
3	Tan H, Biechler S, Junor L, Yost MJ, Dean D, Li J, Potts JD and Goodwin RL: Fluid flow forces and rhoA regulate fibrous development of the atrioventricular valves. Dev Biol. 374:345–356. 2013. View Article : Google Scholar
4	Yano Y, Geibel J and Sumpio BE: Cyclic strain induces reorga-nization of integrin alpha 5 beta 1 and alpha 2 beta 1 in human umbilical vein endothelial cells. J Cell Biochem. 64:505–513. 1997. View Article : Google Scholar : PubMed/NCBI
5	Archambault J, Tsuzaki M, Herzog W and Banes AJ: Stretch and interleukin-1beta induce matrix metalloproteinases in rabbit tendon cells in vitro. J Orthop Res. 20:36–39. 2002. View Article : Google Scholar : PubMed/NCBI
6	Mayr M, Hu Y, Hainaut H and Xu Q: Mechanical stress-induced DNA damage and rac-p38MAPK signal pathways mediate p53-dependent apoptosis in vascular smooth muscle cells. FASEB J. 16:1423–1425. 2002.PubMed/NCBI
7	Wang JH and Thampatty BP: An introductory review of cell mechanobiology. Biomech Model Mechanobiol. 5:1–16. 2006. View Article : Google Scholar : PubMed/NCBI
8	Pan J, Wang T, Wang L, Chen W and Song M: Cyclic strain-induced cytoskeletal rearrangement of human periodontal ligament cells via the Rho signaling pathway. PLoS One. 9:e915802014. View Article : Google Scholar : PubMed/NCBI
9	Qi YX, Qu MJ, Long DK, Liu B, Yao QP, Chien S and Jiang ZL: Rho-GDP dissociation inhibitor alpha downregulated by low shear stress promotes vascular smooth muscle cell migration and apoptosis: A proteomic analysis. Cardiovasc Res. 80:114–122. 2008. View Article : Google Scholar : PubMed/NCBI
10	Sit ST and Manser E: Rho GTPases and their role in organizing the actin cytoskeleton. J Cell Sci. 124:679–683. 2011. View Article : Google Scholar : PubMed/NCBI
11	Bamburg JR and Bernstein BW: ADF/cofilin. Curr Biol. 18:R273–R275. 2008. View Article : Google Scholar : PubMed/NCBI
12	Bamburg JR and Wiggan OP: ADF/cofilin and actin dynamics in disease. Trends Cell Biol. 12:598–605. 2002. View Article : Google Scholar : PubMed/NCBI
13	Bamburg JR: Proteins of the ADF/cofilin family: Essential regulators of actin dynamics. Annu Rev Cell Dev Biol. 15:185–230. 1999. View Article : Google Scholar : PubMed/NCBI
14	Maekawa M, Ishizaki T, Boku S, Watanabe N, Fujita A, Iwamatsu A, Obinata T, Ohashi K, Mizuno K and Narumiya S: Signaling from Rho to the actin cytoskeleton through protein kinases ROCK and LIM-kinase. Science. 285:895–898. 1999. View Article : Google Scholar : PubMed/NCBI
15	Edwards DC, Sanders LC, Bokoch GM and Gill GN: Activation of LIM-kinase by Pak1 couples Rac/Cdc42 GTPase signalling to actin cytoskeletal dynamics. Nat Cell Biol. 1:253–259. 1999. View Article : Google Scholar : PubMed/NCBI
16	Bernstein BW and Bamburg JR: ADF/cofilin: A functional node in cell biology. Trends Cell Biol. 20:187–195. 2010. View Article : Google Scholar : PubMed/NCBI
17	Dumas V, Ducharne B, Perrier A, Fournier C, Guignandon A, Thomas M, Peyroche S, Guyomar D, Vico L and Rattner A: Extracellular matrix produced by osteoblasts cultured under low-magnitude, high-frequency stimulation is favourable to osteogenic differentiation of mesenchymal stem cells. Calcif Tissue Int. 87:351–364. 2010. View Article : Google Scholar : PubMed/NCBI
18	Livak KJ and Schmittgen TD: Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method. Methods. 25:402–408. 2001. View Article : Google Scholar
19	Torcasio A, van Lenthe GH and Van Oosterwyck H: The importance of loading frequency, rate and vibration for enhancing bone adaptation and implant osseointegration. Eur Cell Mater. 16:56–68. 2008.PubMed/NCBI
20	Zeichen J, van Griensven M and Bosch U: The proliferative response of isolated human tendon fibroblasts to cyclic biaxial mechanical strain. Am J Sports Med. 28:888–892. 2000.PubMed/NCBI
21	Watson PA: Function follows form: Generation of intracellular signals by cell deformation. FASEB J. 5:2013–2019. 1991.PubMed/NCBI
22	Mayr M, Hu Y, Hainaut H and Xu Q: Mechanical stress-induced DNA damage and rac-p38MAPK signal pathways mediate p53-dependent apoptosis in vascular smooth muscle cells. FASEB J. 16:1423–1425. 2002.PubMed/NCBI
23	Pan J, Wang T, Wang L, Chen W and Song M: Cyclic strain-induced cytoskeletal rearrangement of human periodontal ligament cells via the Rho signaling pathway. PLoS One. 9:e915802014. View Article : Google Scholar : PubMed/NCBI
24	Van Troys M, Huyck L, Leyman S, Dhaese S, Vandekerkhove J and Ampe C: Ins and outs of ADF/cofilin activity and regulation. Eur J Cell Biol. 87:649–667. 2008. View Article : Google Scholar : PubMed/NCBI
25	Nishida E, Iida K, Yonezawa N, Koyasu S, Yahara I and Sakai H: Cofilin is a component of intranuclear and cytoplasmic actin rods induced in cultured cells. Proc Natl Acad Sci USA. 84:5262–5266. 1987. View Article : Google Scholar : PubMed/NCBI
26	Lin MC, Galletta BJ, Sept D and Cooper JA: Overlapping and distinct functions for cofilin, coronin and Aip1 in actin dynamics in vivo. J Cell Sci. 123:1329–1342. 2010. View Article : Google Scholar : PubMed/NCBI
27	Pan JS, Han Y, Chen DP, Xu L, Qi YX and Yan ZQ: Cyclic strain promotes migration of human periodontal ligament cell via extracellular signal-regulated kinase (ERK) signaling pathway. Zhonghua Kou Qiang Yi Xue Za Zhi. 45:80–84. 2010.In Chinese. PubMed/NCBI
28	Liu X, Zhang X and Luo ZP: Strain-related collagen gene expression in human osteoblast-like cells. Cell Tissue Res. 322:331–334. 2005. View Article : Google Scholar : PubMed/NCBI
29	Larsson T, Aspden RM and Heinegård D: Effects of mechanical load on cartilage matrix biosynthesis in vitro. Matrix. 11:388–394. 1991. View Article : Google Scholar : PubMed/NCBI
30	Jing D, Cai J, Wu Y, Shen G, Li F, Xu Q, Xie K, Tang C, Liu J, Guo W, et al: Pulsed electromagnetic fields partially preserve bone mass, microarchitecture and strength by promoting bone formation in hindlimb-suspended rats. J Bone Miner Res. 29:2250–2261. 2014. View Article : Google Scholar : PubMed/NCBI
31	Lin T, Zeng L, Liu Y, DeFea K, Schwartz MA, Chien S and Shyy JY: Rho-ROCK-LIMK-cofilin pathway regulates shear stress activation of sterol regulatory element binding proteins. Circ Res. 92:1296–1304. 2003. View Article : Google Scholar : PubMed/NCBI
32	Campbell JJ, Blain EJ, Chowdhury TT and Knight MM: Loading alters actin dynamics and up-regulates cofilin gene expression in chondrocytes. Biochem Biophys Res Commun. 361:329–334. 2007. View Article : Google Scholar : PubMed/NCBI
33	Pederson T: As functional nuclear actin comes into view, is it globular, filamentous, or both? J Cell Biol. 180:1061–1064. 2008. View Article : Google Scholar : PubMed/NCBI
34	Zheng B, Han M, Bernier M and Wen JK: Nuclear actin and actin-binding proteins in the regulation of transcription and gene expression. FEBS J. 276:2669–2685. 2009. View Article : Google Scholar : PubMed/NCBI
35	Iida K, Matsumoto S and Yahara I: The KKRKK sequence is involved in heat shock-induced nuclear translocation of the 18-kDa actin-binding protein, cofilin. Cell Struct Funct. 17:39–46. 1992. View Article : Google Scholar : PubMed/NCBI
36	Burgos-Rivera B, Ruzicka DR, Deal RB, McKinney EC, King-Reid L and Meagher RB: ACTIN DEPOLYMERIZING FACTOR9 controls development and gene expression in Arabidopsis. Plant Mol Biol. 68:619–632. 2008. View Article : Google Scholar : PubMed/NCBI
37	Pendleton A, Pope B, Weeds A and Koffer A: Latrunculin B or ATP depletion induces cofilin-dependent translocation of actin into nuclei of mast cells. J Biol Chem. 278:14394–14400. 2003. View Article : Google Scholar : PubMed/NCBI