Fluid flow modulates the expression of genes involved in the Wnt signaling pathway in osteoblasts in 3D culture conditions

Jia,Yuan-Yuan; Li,Feng; Geng,Ning; Gong,Ping; Huang,Shi-Jie; Meng,Ling-Xian; Lan,Jing; Ban,Yu

doi:10.3892/ijmm.2014.1694

Fluid flow modulates the expression of genes involved in the Wnt signaling pathway in osteoblasts in 3D culture conditions

Authors:
- Yuan-Yuan Jia
- Feng Li
- Ning Geng
- Ping Gong
- Shi-Jie Huang
- Ling-Xian Meng
- Jing Lan
- Yu Ban
View Affiliations

Affiliations: State Key Laboratory of Oral Diseases, West China School of Stomatology, Sichuan University, Chengdu, Sichuan 610041, P.R. China, Department of Stomatology, The People's Hospital of Deyang City, Deyang, Sichuan 618000, P.R. China
Published online on: March 11, 2014 https://doi.org/10.3892/ijmm.2014.1694
Pages: 1282-1288

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

The balance between osteoclastic bone resorption and osteoblastic bone formation maintains bone mass, while mechanical loads stimulate bone formation and suppress resorption. The molecular mechanisms responsible for this process have not yet been fully elucidated. In the present study, we assessed whether mechanical stimulation by pulsating fluid flow (PFF) leads to functional Wnt production and affects the function of osteoblasts. ROS17/2.8 osteoblasts were submitted to 1-4 h PFF (0.8 Pa) by three-dimensional (3D) cell culture system with fluid flow. PFF upregulated the gene expression levels of adenomatous polyposis coli, alkaline phosphatase, low density lipoprotein receptor-related protein 5 (LRP5), Wnt3a and β-catenin [catenin beta 1 (CTNNB1)] in all the groups of osteoblasts. Our results suggest that mechanical stimulation by PFF induces the differentiation of osteoblasts and the activation of the Wnt/β-catenin signaling pathway in a 3D cell culture system. Furthermore, mechanical stress plays an important role in the Wnt/β-catenin signaling pathway and is involved in bone formation.

View Figures

View References

1	Harada S and Rodan GA: Control of osteoblast function and regulation of bone mass. Nature. 423:349–355. 2003. View Article : Google Scholar : PubMed/NCBI
2	Carter DR, Van der Meulen MC and Beaupre GS: Mechanical factors in bone growth and development. Bone. 18(Suppl 1): 5S–10S. 1996. View Article : Google Scholar : PubMed/NCBI
3	You J, Yellowley CE, Donahue HJ, Zhang Y, Chen Q and Jacobs CR: Substrate deformation levels associated with routine physical activity are less stimulatory to bone cells relative to loading-induced oscillatory fluid flow. J Biomech Eng. 122:387–393. 2000. View Article : Google Scholar
4	Ehrlich PJ and Lanyon LE: Mechanical strain and bone cell function: a review. Osteoporos Int. 13:688–700. 2002. View Article : Google Scholar : PubMed/NCBI
5	Bodine PVN: Wnt signaling in bone development. Bone and Development. Springer; London: pp. 137–152. 2010, View Article : Google Scholar
6	Zhang R, Oyajobi BO, Harris S, Chen D, Tsao C, Deng HW and Zhao M: Wnt/β-catenin signaling activates bone morphogenetic protein 2 expression in osteoblasts. Bone. 52:145–156. 2013.
7	Guo Y, Li PF, Shu XC, Deng H, Ma HL and Sun L: Involvement of Wnt/β-catenin signaling in the osteogenesis of bone marrow mesenchymal stem cells induced by drynaria total flavonoids. Zhonghua Yi Xue Za Zhi. 92:2288–2291. 2012.(In Chinece).
8	Fischbach C, Kong HJ, Hsiong SX, Evangelista MB, Yuen W and Mooney DJ: Cancer cell angiogenic capability is regulated by 3D culture and integrin engagement. Proc Natl Acad Sci USA. 106:399–404. 2009. View Article : Google Scholar : PubMed/NCBI
9	Byrne EM, Farrell E, McMahon LA, Haugh MG, O’Brien FJ, Campbel VA, Prendergast PJ and O’Connell BC: Gene expression by marrow stromal cells in a porous collagen-glycosaminoglycan scaffold is affected by pore size and mechanical stimulation. J Mater Sci Mater Med. 19:3455–3463. 2008. View Article : Google Scholar
10	Helmke CD: Factors affecting bone cell growth and differentiation under differing culture conditions. Masters Thesis. Rice University, ETD. http://hdl.handle.net/1911/17342. 2000
11	Jarrahy R, Huang W, Rudkin GH, Lee JM, Ishida K, Berry MD, Sukkarieh M, Wu BM, Yamaguchi DT and Miller TA: Osteogenic differentiation is inhibited and angiogenic expression is enhanced in MC3T3-E1 cells cultured on three-dimensional scaffolds. Am J Physiol Cell Physiol. 289:C408–C414. 2005. View Article : Google Scholar : PubMed/NCBI
12	Barron MJ, Tsai CJ and Donahue SW: Mechanical stimulation mediates gene expression in MC3T3 osteoblastic cells differently in 2D and 3D environments. J Biomech Eng. 132:0410052010. View Article : Google Scholar : PubMed/NCBI
13	Wu YY, Ban Y, Geng N, Wang YY, Liu XG, Yu T and Gong P: Evaluation of different culture techniques of osteoblasts on 3D scaffolds. Cent Eur J Biol. 5:456–465. 2010. View Article : Google Scholar
14	Fu Q, Liu YH, Xu YJ, Guo L and Wu CJ: Design and use of flow chamber for fluid shear stress on cells in vitro. Journal of Sun Yat-Sen University. S12009.(In Chinese).
15	Engler AJ, Sen S, Sweeney HL and Discher DE: Matrix elasticity directs stem cell lineage specification. Cell. 126:677–689. 2006. View Article : Google Scholar : PubMed/NCBI
16	Evans ND, Minelli C, Gentleman E, LaPointe V, Patankar SN, Kallivretaki M, Chen X, Roberts CJ and Stevens MM: Substrate stiffness affects early differentiation events in embryonic stem cells. Eur Cell Mater. 18:1–14. 2009.PubMed/NCBI
17	Ghosh K and Ingber DE: Micromechanical control of cell and tissue development: implications for tissue engineering. Adv Drug Deliv Rev. 59:1306–1318. 2007. View Article : Google Scholar : PubMed/NCBI
18	Karamichos D, Skinner J, Brown R and Mudera V: Matrix stiffness and serum concentration effects matrix remodelling and ECM regulatory genes of human bone marrow stem cells. J Tissue Eng Regen Med. 2:97–105. 2008. View Article : Google Scholar : PubMed/NCBI
19	Orr AW, Helmke BP, Blackman BR and Schwartz MA: Mechanisms of mechanotransduction. Dev Cell. 10:11–20. 2006. View Article : Google Scholar
20	Weinbaum S, Cowin SC and Zeng Y: A model for the excitation of osteocytes by mechanical loading-induced bone fluid shear stresses. J Biomech. 27:339–360. 1994. View Article : Google Scholar : PubMed/NCBI
21	Bakker AD, Soejima K, Klein-Nulend J and Burger EH: The production of nitric oxide and prostaglandin E2 by primary bone cells is shear stress dependent. J Biomech. 34:671–677. 2001. View Article : Google Scholar : PubMed/NCBI
22	Riancho JA, Olmos JM, Pineda B, García-Ibarbia C, Pérez-Núñez MI, Nan DN, Velasco J, Cano A, García-Pérez MA, Zarrabeitia MT and González-Macías J: Wnt receptors, bone mass, and fractures: gene-wide association analysis of LRP5 and LRP6 polymorphisms with replication. Eur J Endocrinol. 164:123–131. 2011. View Article : Google Scholar : PubMed/NCBI
23	Bonewald LF and Johnson ML: Osteocytes, mechanosensing and Wnt signaling. Bone. 42:606–615. 2008. View Article : Google Scholar : PubMed/NCBI
24	Clevers H and Nusse R: Wnt/β-catenin signaling and disease. Cell. 149:1192–1205. 2012.
25	Kobayashi Y, Maeda K and Takahashi N: Roles of Wnt signalingin bone formation and resorption. Jpn Dent Sci Rev. 44:76–82. 2008. View Article : Google Scholar
26	Hill TP, Später D, Taketo MM, Birchmeier W and Hartmann C: Canonical Wnt/beta-catenin signaling prevents osteoblasts from differentiating into chondrocytes. Dev Cell. 8:727–738. 2005. View Article : Google Scholar : PubMed/NCBI
27	Day TF, Guo X, Garrett-Beal L and Yang Y: Wnt/β-catenin signaling in mesenchymal progenitors controls osteoblast and chondrocyte differentiation during vertebrate skeletogenesis. Dev Cell. 8:739–750. 2005.
28	Tüysüz B, Bursalı A, Alp Z, Suyugül N, Laine CM and Mäkitie O: Osteoporosis-pseudoglioma syndrome: three novel mutations in the LRP5 gene and response to bisphosphonate treatment. Horm Res Paediatr. 77:115–120. 2012.PubMed/NCBI
29	Wodarz A and Nusse R: Mechanisms of Wnt signaling in development. Annu Rev Cell Dev Biol. 14:59–88. 1998. View Article : Google Scholar
30	Johnson ML and Kamel MA: The Wnt signaling pathway and bone metabolism. Curr Opin Rheumatol. 19:376–382. 2007. View Article : Google Scholar : PubMed/NCBI
31	Kato M1, Patel MS, Levasseur R, Lobov I, Chang BH, Glass DA II, Hartmann C, Li L, Hwang TH, Brayton CF, Lang RA, Karsenty G and Chan L: Cbfa1-independent decrease in osteoblast proliferation, osteopenia, and persistent embryonic eye vascularization in mice deficient in Lrp5, a Wnt coreceptor. J Cell Biol. 157:303–314. 2002. View Article : Google Scholar : PubMed/NCBI
32	Owen TA, Aronow M, Shalhoub V, et al: Progressive development of the rat osteoblast phenotype in vitro: reciprocal relationships in expression of genes associated with osteoblast proliferation and differentiation during formation of the bone extracellular matrix. J Cell Physiol. 143:420–430. 1990. View Article : Google Scholar
33	Rawadi G, Vayssière B, Dunn F, Baron R and Roman-Roman S: BMP-2 controls alkaline phosphatase expression and osteoblast mineralization by a Wnt autocrine loop. J Bone Miner Res. 18:1842–1853. 2003. View Article : Google Scholar : PubMed/NCBI
34	Olivares-Navarrete R, Hyzy SL, Hutton DL, Dunn GR, Appert C, Boyan BD and Schwartz Z: Role of non-canonical Wnt signaling enhances osteoblast maturation on microstructured titanium surfaces. Acta Biomater. 7:2740–2750. 2011. View Article : Google Scholar : PubMed/NCBI
35	Jullien N, Maudinet A, Leloutre B, Ringe J, Häupl T and Marie PJ: Downregulation of ErbB3 by Wnt3a contributes to wnt-induced osteoblast differentiation in mesenchymal cells. J Cell Biochem. 113:2047–2056. 2012. View Article : Google Scholar : PubMed/NCBI
36	Natsume H, Tokuda H, Matsushima-Nishiwaki R, Kato K, Yamakawa K, Otsuka T and Kozawa O: Wnt3a upregulates transforming growth factor-β-stimulated VEGF synthesis in osteoblasts. Cell Biochem Funct. 29:371–377. 2011.
37	Sakai A: Space flight/bedrest immobilization and bone. Osteocyte as a sensor of mechanical stress and Wnt signal. Clin Calcium. 22:1829–1835. 2012.(In Japanese).
38	Sakai A: Mechanical stress and Wnt signal. Clin Calcium. 23:839–845. 2013.(In Japanese).