Recombinant Human Bone Morphogenic Protein-2 Induces the Differentiation and Mineralization of Osteoblastic Cells Under Hypoxic Conditions Via Activation of Protein Kinase D and P38 Mitogen-Activated Protein Kinase Signaling Pathways

Overview

Journal Tissue Eng Regen Med

Date 2019 Jan 4

PMID 30603499

Citations 2

Authors

Woo-Hun Ha

Hwa-Sik Seong

Na-Rae Choi

Bong-Soo Park

Yong-Deok Kim

Affiliations

Soon will be listed here.

Abstract

Hypoxia suppresses osteoblastic differentiation and the bone-forming capacity. As the leading osteoinductive growth factor used clinically in bone-related regenerative medicine, recombinant human bone morphogenic protein-2 (rhBMP-2) has yielded promising results in unfavorable hypoxic clinical situations. Although many studies have examined the effects of rhBMP-2 on osteoblastic differentiation, mineralization and the related signaling pathways, those of rhBMP-2 on osteoblastic cells remain unknown, particularly under hypoxic conditions. Therefore, this study was conducted under a 1% oxygen tension to examine the differentiating effects of rhBMP-2 on osteoblastic cells under hypoxia. rhBMP-2 could also induce the differentiation and mineralization of Osteoblastic (MC3T3-E1) cells under 1% hypoxic conditions. rhBMP-2 could also induce the differentiation and mineralization of MC3T3-E1 cells under 1% hypoxic conditions. rhBMP-2 increased the alkaline phosphatase {ALP} activity in a time dependent manner, and expression of ALP, collagen type-1 (Col-1) and osteocalcin (OC) mRNA were up-regulated significantly in a time- and concentration-dependent manner. In addition, the area of the mineralized nodules increased gradually in a concentration-dependent manner. Western blot analysis, which was performed to identify the signaling pathways underlying rhBMP-2-induced osteoblastic differentiation under hypoxic conditions, showed that rhBMP-2 significantly promoted the phosphorylation of the p38 mitogen-activated protein kinase (MAPK) in a time-dependent manner. A pretreatment with SB203580, a p38 MAPK inhibitor, inhibited the rhBMP-2-mediated differentiation and mineralization. Moreover, the phosphorylation of p38 induced by rhBMP-2 was inhibited in response to a pretreatment of the cells with Go6976, a protein kinase D {PKD) inhibitor. These findings suggest that rhBMP-2 induces the differentiation and mineralization of MC3T3-E1 cells under hypoxic conditions via activation of the PKD and p38 MAPK signaling pathways.

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References

Cochran D, Schenk R, Buser D, Wozney J, Jones A . Recombinant human bone morphogenetic protein-2 stimulation of bone formation around endosseous dental implants. J Periodontol. 1999; 70(2):139-50. DOI: 10.1902/jop.1999.70.2.139. View

Boyne P, Marx R, Nevins M, Triplett G, Lazaro E, Lilly L . A feasibility study evaluating rhBMP-2/absorbable collagen sponge for maxillary sinus floor augmentation. Int J Periodontics Restorative Dent. 1997; 17(1):11-25. View

Higuchi T, Kinoshita A, Takahashi K, Oda S, Ishikawa I . Bone regeneration by recombinant human bone morphogenetic protein-2 in rat mandibular defects. An experimental model of defect filling. J Periodontol. 1999; 70(9):1026-31. DOI: 10.1902/jop.1999.70.9.1026. View

Kobayashi M, Takiguchi T, Suzuki R, Yamaguchi A, Deguchi K, Shionome M . Recombinant human bone morphogenetic protein-2 stimulates osteoblastic differentiation in cells isolated from human periodontal ligament. J Dent Res. 1999; 78(10):1624-33. DOI: 10.1177/00220345990780100701. View

Lewis J, Lee J, Underwood J, Harris A, Lewis C . Macrophage responses to hypoxia: relevance to disease mechanisms. J Leukoc Biol. 1999; 66(6):889-900. DOI: 10.1002/jlb.66.6.889. View

Sciadini M, Johnson K . Evaluation of recombinant human bone morphogenetic protein-2 as a bone-graft substitute in a canine segmental defect model. J Orthop Res. 2000; 18(2):289-302. DOI: 10.1002/jor.1100180218. View

Kaneko H, Arakawa T, Mano H, Kaneda T, Ogasawara A, Nakagawa M . Direct stimulation of osteoclastic bone resorption by bone morphogenetic protein (BMP)-2 and expression of BMP receptors in mature osteoclasts. Bone. 2000; 27(4):479-86. DOI: 10.1016/s8756-3282(00)00358-6. View

Spector J, Mehrara B, GREENWALD J, Saadeh P, Steinbrech D, Bouletreau P . Osteoblast expression of vascular endothelial growth factor is modulated by the extracellular microenvironment. Am J Physiol Cell Physiol. 2000; 280(1):C72-80. DOI: 10.1152/ajpcell.2001.280.1.C72. View

Dinenno F, Seals D, DeSouza C, Tanaka H . Age-related decreases in basal limb blood flow in humans: time course, determinants and habitual exercise effects. J Physiol. 2001; 531(Pt 2):573-9. PMC: 2278480. DOI: 10.1111/j.1469-7793.2001.0573i.x. View

10.

Gallea S, Lallemand F, Atfi A, Rawadi G, Ramez V, Spinella-Jaegle S . Activation of mitogen-activated protein kinase cascades is involved in regulation of bone morphogenetic protein-2-induced osteoblast differentiation in pluripotent C2C12 cells. Bone. 2001; 28(5):491-8. DOI: 10.1016/s8756-3282(01)00415-x. View

11.

Chen T, Shen W, Kraemer F . Human BMP-7/OP-1 induces the growth and differentiation of adipocytes and osteoblasts in bone marrow stromal cell cultures. J Cell Biochem. 2001; 82(2):187-99. DOI: 10.1002/jcb.1145. View

12.

Kozawa O, Hatakeyama D, Uematsu T . Divergent regulation by p44/p42 MAP kinase and p38 MAP kinase of bone morphogenetic protein-4-stimulated osteocalcin synthesis in osteoblasts. J Cell Biochem. 2002; 84(3):583-9. View

13.

Park J, Park B, Kim H, Park T, Baek H . Hypoxia decreases Runx2/Cbfa1 expression in human osteoblast-like cells. Mol Cell Endocrinol. 2002; 192(1-2):197-203. DOI: 10.1016/s0303-7207(02)00036-9. View

14.

Saito Y, Yoshizawa T, Takizawa F, Ikegame M, Ishibashi O, Okuda K . A cell line with characteristics of the periodontal ligament fibroblasts is negatively regulated for mineralization and Runx2/Cbfa1/Osf2 activity, part of which can be overcome by bone morphogenetic protein-2. J Cell Sci. 2002; 115(Pt 21):4191-200. DOI: 10.1242/jcs.00098. View

15.

Hu Y, Chan E, Wang S, Li B . Activation of p38 mitogen-activated protein kinase is required for osteoblast differentiation. Endocrinology. 2003; 144(5):2068-74. DOI: 10.1210/en.2002-220863. View

16.

Hassel S, Schmitt S, Hartung A, Roth M, Nohe A, Petersen N . Initiation of Smad-dependent and Smad-independent signaling via distinct BMP-receptor complexes. J Bone Joint Surg Am. 2003; 85-A Suppl 3:44-51. DOI: 10.2106/00004623-200300003-00009. View

17.

Gordillo G, Sen C . Revisiting the essential role of oxygen in wound healing. Am J Surg. 2003; 186(3):259-63. DOI: 10.1016/s0002-9610(03)00211-3. View

18.

Lemonnier J, Ghayor C, Guicheux J, Caverzasio J . Protein kinase C-independent activation of protein kinase D is involved in BMP-2-induced activation of stress mitogen-activated protein kinases JNK and p38 and osteoblastic cell differentiation. J Biol Chem. 2003; 279(1):259-64. DOI: 10.1074/jbc.M308665200. View

19.

Guicheux J, Lemonnier J, Ghayor C, Suzuki A, Palmer G, Caverzasio J . Activation of p38 mitogen-activated protein kinase and c-Jun-NH2-terminal kinase by BMP-2 and their implication in the stimulation of osteoblastic cell differentiation. J Bone Miner Res. 2003; 18(11):2060-8. DOI: 10.1359/jbmr.2003.18.11.2060. View

20.

Termaat M, den Boer F, Bakker F, Patka P, Haarman H . Bone morphogenetic proteins. Development and clinical efficacy in the treatment of fractures and bone defects. J Bone Joint Surg Am. 2005; 87(6):1367-78. DOI: 10.2106/JBJS.D.02585. View