Effect of fluid shear force from a fluid flow device simulating mechanical motion on RANKL expression in myeloma cells, osteocytes and osteoclasts

doi:10.3969/j.issn.2095-4344.0608

Abstract

Abstract:

BACKGROUND: Myeloma patients have less movement because of bone pain and osteolytic destruction, which result in bone loss and disease aggravation.

OBJECTIVE: To explore the effect of myeloma cells (U266 cells) on osteocytes (Y4 cells), osteoblasts and osteoclasts as well as RANKL secreted by osteocytes under fluid shear stress simulating human mechanical movement by a fluid flow devise.

METHODS: A flow and no-flow model was established. U266 cell culture fluid was used in the experimental group and Y4 standard culture fluid was used in the flow and control groups. Myeloma cell line U266 and mouse osteocytes cell line Y4 were cultured and passaged in vitro. Y4 cells were allotted and cultured for 48 hours, and the morphology of osteocytes and osteoclasts was observed and counted under microscope. The level of RANKL was quantitatively determined by ELISA and western blot assay. The RANKL, sample and standard culture system was established, RAW264.7 cells were cultured in vitro, and induced by RANKL. Number and morphology of RAW264.7 cells differentiating into osteoclasts were observed by TRAP staining.

RESULTS AND CONCLUSION: Compared with the control group, Y4 cells in U266 culture supernatant exhibited morphological changes and their number was significantly decreased. The number of TRAP-positive osteoclasts was increased and the expression of RANKL protein in Y4 cells was significantly up-regulated (P < 0.05). Comparison between the flow model and the no-flow model showed that the number of Y4 cells was significantly increased, the expression of RANKL protein decreased and the number of TRAP-positive osteoclasts was significantly reduced (P < 0.05). Our study indicates that myeloma cells can inhibit the growth of normal osteocyte and might promote the expression of RANKL protein and the growth of osteoclasts. Compared with static state, fluid shear stress may promote the proliferation of osteocytes and might inhibit the expression of RANKL protein and the growth of osteoclasts. So, we hypothesize that mild exercise may prevent the progression of myeloma bone disease.

中国组织工程研究杂志出版内容重点：生物材料；骨生物材料; 口腔生物材料; 纳米材料; 缓释材料; 材料相容性；组织工程

Key words: Multiple Myeloma, Osteoclasts, RANK Ligand, Tissue Engineering

CLC Number:

R455

R318

Wang Xiaotao, Tian Shen, He Yuchan, Wu Chunye, Zhang Junyan, Yang Shaofang, Wang Hangfei, You Lidan. Effect of fluid shear force from a fluid flow device simulating mechanical motion on RANKL expression in myeloma cells, osteocytes and osteoclasts[J]. Chinese Journal of Tissue Engineering Research, 2019, 23(2): 251-256.

Figures/Tables 4

References

[1] Hameed A,Brady JJ,Dowling P,et al.Bone disease in multiple myeloma: pathophysiology and management. Cancer Growth Metastasis. 2014;7:33-42.

[2] Palma BD,Guasco D,Pedrazzoni M,et al.Osteolytic lesions, cytogenetic features and bone marrow levels of cytokines and chemokines in multiple myeloma patients: Role of chemokine (C-C motif) ligand 20.Leukemia. 2016;30(2): 409-416.

[3] Minarik J,Hermanova Z,Petrova P,et al.Prospective study of signalling pathways in myeloma bone disease with regard to activity of the disease, extent of skeletal involvement and correlation to bone turnover markers.Eur J Haematol.2016; 97(2):201-207.

[4] Bolzoni M, Storti P, Bonomini S, et al.Immunomodulatory drugs lenalidomide and pomalidomide inhibit multiple myeloma-induced osteoclast formation and the RANKL/OPG ratio in the myeloma microenvironment targeting the expression of adhesion molecules. Exp Hematol.2013;41(4): 387-397.e1

[5] Graves DT,Alshabab A,Albiero ML,et al.Osteocytes play an important role in experimental periodontitis in healthy and diabetic mice through expression of RANKL.J Clin Periodontol. 2018;45(3):285-292.

[6] Delgado-Calle J,Anderson J,Cregor MD,et al.Bidirectional Notch Signaling and Osteocyte-Derived Factors in the Bone Marrow Microenvironment Promote Tumor Cell Proliferation and Bone Destruction in Multiple Myeloma.Cancer Res.2016; 76(5):1089-1100.

[7] Vallet S, Pozzi S, Patel K, et al.A novel role for CCL3 (MIP-1a) in myeloma-induced bone disease via osteocalcin downregulation and inhibition of osteoblast function. Leukemia.2011;25(7):1174-1181.

[8] Wang XT, He YC, Zhou SY, et al.Bone marrow plasma macrophage inflammatory protein-1 alpha(MIP-1 alpha) and sclerostin in multiple myeloma: Relationship with bone disease and clinical characteristics.2014;38(5):525-531.

[9] Walker RE, Lawson MA, Buckle CH.et al. Myeloma bone disease: pathogenesis, current treatments and future targets.Br Med Bull.2014;111(1):117-138.

[10] Colucci S, Brunetti G, Oranger A, et al.Myeloma cells suppress osteoblasts through sclerostin secretion.Blood Cancer J.2011;22(1):1-7.

[11] Higgs JT, Lee JH, Wang H.et al. Mesenchymal stem cells expressing osteoprotegerin variants inhibit osteolysis in a murine model of multiple myeloma.Blood Adv.2017;1(25): 2375-2385.

[12] Palma BD,Guasco D,Pedrazzoni M,et al.Osteolytic lesions, cytogenetic features and bone marrow levels of cytokines and chemokines in multiple myeloma patients: Role of chemokine (C-C motif) ligand 20.Leukemia.2016;30(2):409-416.

[13] Groeneveldt L,Mein G,Garrod R,et al.A mixed exercise training programme is feasible and safe and may improve quality of life and muscle strength in multiple myeloma survivors. BMC Cancer.2013;13(31):1-10.

[14] Miceli TS, Colson K, Faiman BM,et al.Maintaining bone health in patients with multiple myeloma: survivorship care plan of the International Myeloma Foundation Nurse Leadership Board.Clin J Oncol Nurs.2011;15 Suppl:9-23.

[15] Pagnotti GM,Chan ME,Adler BJ,et al.Low intensity vibration mitigates tumor progression and protects bone quantity and quality in a murine model of myeloma.Bone.2016;90:69-79.

[16] Li X, Niu QT, Warmington KS,et al.Progressive increases in bone mass and bone strength in an ovariectomized rat model of osteoporosis after 26 weeks of treatment with a sclerostin antibody.Endocrinology.2014;155(12):4785-4797.

[17] McGrath RP,Kraemer WJ,Vincent BM.et al.Muscle-Strength Is Protective Against Osteoporosis in an Ethnically Diverse Sample of adults.J Strength Cond Res.2017;31(9): 2586-2589.

[18] Middleton K,Al-Dujaili S,Mei X,et al.Microfluidic co-culture platform for investigating osteocyte-osteoclast signalling during fluid shearstress mechanostimulation.J Biomech. 2017;59:35-42.

[19] Shah KM, Orton P, Mani N,et al. Osteocyte physiology and response to fluid shear stress are impaired following exposure to cobalt and chromium: Implications for bone health following joint replacement.J Orthop Res.2017;35(8):1716-1723.

[20] Sartawi Z,Schipani E,Ryan KB, et al.Sphingosine 1-phosphate (S1P) signalling: Role in bone biology and potential therapeutic target for bone repair.Pharmacol Res. 2017;125(Pt B):232-245.

[21] Meshcheryakova A,Mechtcheriakova D,Pietschmann P.Sphingosine 1-phosphate signaling in bone remodeling: multifaceted roles and therapeutic potential.Expert Opin Ther Targets. 2017;21(7):725-737.

[22] Zhang J,Ding C,Meng X,et al.Nitric oxide modulates the responses of osteoclast formation to static magnetic fields. Electromagn Biol Med.2017;13:1-12.

[23] Xiao L,Zhu L,Yang S,et al.Different correlation of sphingosine-1-phosphate receptor 1 with receptor activator of nuclear factor kappa B ligand and regulatory T cells in rat periapical lesions.J Endod.2015;41(4):479-486.

[24] Kim BJ, Shin KO, Kim H ,et al.The effect of sphingosine-1-phosphate on bone metabolism in humans depends on its plasma/bone marrow gradient.J Endocrinol Invest.2016;39(3):297-303.

[25] Yuan M,Chen J,Zeng Z.Knockdown of macrophage inhibitory cytokine-1 in RPMI-8226 human multiple myeloma cells inhibits osteoclastic differentiation through inhibiting the RANKL-Erk1/2 signaling pathway.Mol Med Rep.2016;14(6): 5199-5204.

[26] Pitari MR,Rossi M,Amodio N,et al.Inhibition of miR-21 restores RANKL/OPG ratio in multiple myeloma-derived bone marrow stromal cells and impairs the resorbing activity of mature osteoclasts.Oncotarget.2015;6(29):27343-27358.

[27] Kristensen IB,Christensen JH,Lyng MB,et al.Expression of osteoblast and osteoclast regulatory genes in the bone marrow microenvironment in multiple myeloma: only up-regulation of Wnt inhibitors SFRP3 and DKK1 is associated with lytic bone disease.Leuk Lymphoma.2014; 55(4):911-919.

[28] Delgado-Calle J,Arozamena J,Pérez-López J,et al.Role of BMPs in the regulation of sclerostin as revealed by an epigenetic modifier of human bone cells.Mol Cell Endocrinol. 2013;369(1-2):27-34.

[29] 陈少龙,赵良功,滕元君,等.周期性和持续性流体剪切力对成骨细胞OPG、RANKL蛋白表达的影响[J].中国医学物理学杂志, 2015, 32(1):120-123,133.