Primary cilia/intraflagellar transport mediates mechanics-responsive signaling pathway and promotes osteogenic differentiation of bone marrow stromal stem cells

doi:10.12307/2024.192

Abstract

Abstract: BACKGROUND: Mechanical stimulation has been confirmed to promote osteogenic differentiation of bone marrow stromal stem cells, but the mechanism is unknown. Primary cilia are important mechanoreceptors and regulate various signaling pathways such as TGF-β1/BMP-2/SMAD. They are likely to be important targets for mechanical regulation of bone marrow stromal stem cells.
OBJECTIVE: To investigate the effect and mechanism of fluid shear stress on osteogenic differentiation of bone marrow stromal stem cells.
METHODS: Rat bone marrow stromal stem cells were divided into control group, mechanical stimulation group (fluid shear mechanics intervention by shaking table), mechanical stimulation + IFT88 silencing group (mechanical stimulation + silencing IFT88 expression with siRNA). After 24 hours of intervention, qRT-PCR was utilized to determine the expression of transforming growth factor β1 and bone morphogenetic protein 2. Western blot assay was used to detect the expression of phosphorylated SMAD2/3 protein. Immunofluorescent staining of primary cilia was conducted and morphology was analyzed.
RESULTS AND CONCLUSION: Shear stress stimulation could promote the transcriptional activity of transforming growth factor β1 and bone morphogenetic protein 2 genes, and increase the expression of phosphorylated SMAD2/3 protein. After siRNA interfered with primary cilia, this mechanical response effect was significantly reduced. There was a Spearman correlation between the change ratio of the primary cilium area of bone marrow stromal stem cells and the increased ratio of transforming growth factor β1 and bone morphogenetic protein 2 gene transcription. These findings indicate that primary cilia/intraflagellar transport mediates the activation of fluid shear stress-responsive transforming growth factor β1/bone morphogenetic protein 2/SMAD signaling pathway and promotes osteogenic differentiation of bone marrow stromal stem cells.

Key words: bone marrow stromal stem cell, transforming growth factor β1, bone morphogenetic protein 2, primary cilia, intraflagellar transport, shear stress stimulation

CLC Number:

Ma Zhanhua, Yan Xu, Jiang Yan, Cao Zhengming, Wang Yongkui, Li Dongzhe, Yang Tengyue, Jin Yikai, Fu Su, Zhang Chunlin. Primary cilia/intraflagellar transport mediates mechanics-responsive signaling pathway and promotes osteogenic differentiation of bone marrow stromal stem cells[J]. Chinese Journal of Tissue Engineering Research, 2024, 28(25): 3937-3941.

Figures/Tables 4

References

[1] MUSUMECI G. The effect of mechanical loading on articular cartilage. J Funct Morphol Kinesiol. 2016;1(2):154-161.
[2] METZGER TA, KREIPKE TC, VAUGHAN TJ, et al. The in situ mechanics of trabecular bone marrow: the potential for mechanobiological response. J Biomech Eng. 2015;137(1). doi: 10.1115/1.4028985.
[3] 黎润光,邵景范,魏明发.机械力学对体外骨髓间充质干细胞的影响[J].中国组织工程研究与临床康复,2007,11(3):551-554.
[4] CORRIGAN MA, JOHNSON GP, STAVENSCHI E, et al. TRPV4-mediates oscillatory fluid shear mechanotransduction in mesenchymal stem cells in part via the primary cilium. Sci Rep. 2018;8(1):3824.
[5] WHEWAY G, NAZLAMOVA L, HANCOCK JT. Signaling through the Primary Cilium. Front Cell Dev Biol. 2018;6:8.
[6] MCMURRAY RJ, WANN AK, THOMPSON CL, et al. Surface topography regulates wnt signaling through control of primary cilia structure in mesenchymal stem cells. Sci Rep. 2013;3:3545.
[7] YAN JL, ZHOU J, MA HP, et al. Pulsed electromagnetic fields promote osteoblast mineralization and maturation needing the existence of primary cilia. Mol Cell Endocrinol. 2015;404:132-140.
[8] 刘勇,裴国献,江汕,等.筋膜瓣包裹组织工程骨修复兔大段骨缺损的动态变化[J].中国组织工程研究与临床康复,2008,12(24): 4672-4676.
[9] 廖欣宇,王福科,王国梁.骨组织工程支架的进展与挑战[J].中国组织工程研究,2021,25(28):4553-4560.
[10] ANDRZEJEWSKA A, LUKOMSKA B, JANOWSKI M. Concise Review: Mesenchymal Stem Cells: From Roots to Boost. Stem Cells. 2019;37(7): 855-864.
[11] SUWITTAYARAK R, KLINCUMHOM N, NGAOKRAJANG U, et al. Shear Stress Enhances the Paracrine-Mediated Immunoregulatory Function of Human Periodontal Ligament Stem Cells via the ERK Signalling Pathway. Int J Mol Sci. 2022;23(13):7119.
[12] RUI YF, LUI PP, NI M, et al. Mechanical loading increased BMP-2 expression which promoted osteogenic differentiation of tendon-derived stem cells. J Orthop Res. 2011;29(3):390-396.
[13] YOUREK G, MCCORMICK SM, MAO JJ, et al. Shear stress induces osteogenic differentiation of human mesenchymal stem cells. Regen Med. 2010;5(5):713-724.
[14] SONAM S, SATHE SR, YIM EK, et al. Cell contractility arising from topography and shear flow determines human mesenchymal stem cell fate. Sci Rep. 2016;6:20415.
[15] STAVENSCHI E, LABOUR MN, HOEY DA. Oscillatory fluid flow induces the osteogenic lineage commitment of mesenchymal stem cells: The effect of shear stress magnitude, frequency, and duration. J Biomech. 2017;55:99-106.
[16] MILL P, CHRISTENSEN ST, PEDERSEN LB. Primary cilia as dynamic and diverse signalling hubs in development and disease. Nat Rev Genet. 2023;24(7):421-441.
[17] WANG W, JACK BM, WANG HH, et al. Intraflagellar Transport Proteins as Regulators of Primary Cilia Length. Front Cell Dev Biol. 2021;9:661350.
[18] RYS JP, DUFORT CC, MONTEIRO DA, et al. Discrete spatial organization of TGFβ receptors couples receptor multimerization and signaling to cellular tension. Elife. 2015;4:e09300.
[19] CLEMENT CA, AJBRO KD, KOEFOED K, et al. TGF-β signaling is associated with endocytosis at the pocket region of the primary cilium. Cell Rep. 2013;3(6):1806-1814.
[20] LABOUR MN, RIFFAULT M, CHRISTENSEN ST, et al. TGFβ1 - induced recruitment of human bone mesenchymal stem cells is mediated by the primary cilium in a SMAD3-dependent manner. Sci Rep. 2016;6:35542.
[21] CHEN G, DENG C, LI YP. TGF-β and BMP signaling in osteoblast differentiation and bone formation. Int J Biol Sci. 2012;8(2):272-288.
[22] LIU M, ALHARBI M, GRAVES D, et al. IFT80 Is Required for Fracture Healing Through Controlling the Regulation of TGF-β Signaling in Chondrocyte Differentiation and Function. J Bone Miner Res. 2020; 35(3):571-582.