Combined Treatment with Low-Level Laser and rhBMP-2 Promotes Differentiation and Mineralization of Osteoblastic Cells under Hypoxic Stress.
10.1007/s13770-018-0167-1
- Author:
Jin Ho HEO
1
;
Jeong Hun CHOI
;
In Ryoung KIM
;
Bong Soo PARK
;
Yong Deok KIM
Author Information
1. Department of Oral and Maxillofacial Surgery, Pusan National University, 49 Busandaehak-ro, Mulgeum-eup, Yangsan-si, Gyeongsangnam-do 50612, Republic of Korea. ydkimdds@pusan.ac.kr
- Publication Type:Original Article
- Keywords:
Hypoxia;
Osteoblast;
Laser;
p38;
PKD
- MeSH:
Alkaline Phosphatase;
Anoxia;
Blotting, Western;
Cell Culture Techniques;
Collagen Type I;
Humans;
In Vitro Techniques;
Low-Level Light Therapy;
Miners*;
Osteoblasts*;
Oxygen;
Phosphorylation;
Phosphotransferases;
Transcription Factors
- From:
Tissue Engineering and Regenerative Medicine
2018;15(6):793-801
- CountryRepublic of Korea
- Language:English
-
Abstract:
BACKGROUND: The aim of this study was to evaluate the combined effect of low-level laser treatment (LLLT) and recombinant human bone morphological protein-2 (rhBMP-2) applied to hypoxic-cultured MC3T3-E1 osteoblastic cells and to determine possible signaling pathways underlying differentiation and mineralization of osteoblasts under hypoxia. METHODS: MC3T3-E1 cells were cultured under 1% oxygen tension for 72 h. Cell cultures were divided into four groups: normoxia control, low-level laser (LLL) alone, rhBMP-2 combined with LLLT, and rhBMP-2 under hypoxia. Laser irradiation was applied at 0, 24, and 48 h. Cells were treated with rhBMP-2 at 50 ng/mL. Alkaline phosphatase activity was measured at 3, 7, and 14 days to evaluate osteoblastic differentiation. Cell mineralization was determined with Alizarin red S staining at 7 and 14 days. Western blot assays were performed to evaluate whether p38/protein kinase D (PKD) signaling was involved. RESULTS: The results indicate that LLLT and rhBMP-2 synergistically increased alkaline phosphatase (ALP) activity and mineralization. Western blot analyses showed that expression of type I collagen, runt-related transcription factor 2 (RUNX2), and Osterix (Osx), increased and expression of hypoxia-inducible factor 1-alpha (HIF-1α), decreased more in the LLLT and rhBMP-2 combined group than in the rhBMP-2 or LLL alone groups. Moreover, LLLT and rhBMP-2 stimulated p38 phosphorylation and rhBMP-2 and LLLT increased Prkd1 phosphorylation. CONCLUSION: Combined treatment with rhBMP-2 and LLL induced differentiation and mineralization of hypoxiccultured MC3T3-E1 osteoblasts by activating p38/PKD signaling in vitro.
