Photobiomodulation therapy's impact on angiogenesis and osteogenesis in orthodontic tooth movement: in vitro and in vivo study

Abstract

Background: This study explores the effectiveness of Photobiomodulation Therapy (PBMT) in enhancing orthodontic tooth movement (OTM), osteogenesis, and angiogenesis through a comprehensive series of in vitro and in vivo investigations. The in vitro experiments involved co-culturing MC3T3-E1 and HUVEC cells to assess PBMT's impact on cell proliferation, osteogenesis, angiogenesis, and associated gene expression. Simultaneously, an in vivo experiment utilized an OTM rat model subjected to laser irradiation at specific energy densities.

Methods: In vitro experiments involved co-culturing MC3T3-E1 and HUVEC cells treated with PBMT, enabling a comprehensive assessment of cell proliferation, osteogenesis, angiogenesis, and gene expression. In vivo, an OTM rat model was subjected to laser irradiation at specified energy densities. Statistical analyses were performed to evaluate the significance of observed differences.

Results: The results revealed a significant increase in blood vessel formation and new bone generation within the PBMT-treated group compared to the control group. In vitro, PBMT demonstrated positive effects on cell proliferation, osteogenesis, angiogenesis, and gene expression in the co-culture model. In vivo, laser irradiation at specific energy densities significantly enhanced OTM, angiogenesis, and osteogenesis.

Conclusions: This study highlights the substantial potential of PBMT in improving post-orthodontic bone quality. The observed enhancements in angiogenesis and osteogenesis suggest a pivotal role for PBMT in optimizing treatment outcomes in orthodontic practices. The findings position PBMT as a promising therapeutic intervention that could be seamlessly integrated into orthodontic protocols, offering a novel dimension to enhance overall treatment efficacy. Beyond the laboratory, these results suggest practical significance for PBMT in clinical scenarios, emphasizing its potential to contribute to the advancement of orthodontic treatments. Further exploration of PBMT in orthodontic practices is warranted to unlock its full therapeutic potential.

Keywords: Angiogenesis; Bone formation; Co-culture; Low-level laser therapy; Orthodontic treatment.

Fig. 1

Impact of Varied Energy Density of the laser on Co-Culture Systems Over Different Time Durations. A Co-culture system with varying energy density levels for 12 hours. B Co-culture system with varying energy density levels for 24 hours. C Co-culture system with varying energy density levels for 48 hours. D Co-culture system with varying energy density levels for 72 hours. (**: P < 0.01, ***: P < 0.001; compared to the control group)

Fig. 2

Impact of Photobiomodulation Therapy (PBMT) on Osteogenic Differentiation in Co-Culture Systems. A ALP staining for assessing osteogenic differentiation. B Evaluation of osteogenic differentiation through Alizarin Red S staining. C Alkaline Phosphatase Activity Assay results (**: P < 0.001, compared to the MC3T3-E1 group; ###: P < 0.001, compared to the MC3T3 + HUVEC group). D Measurement of Runx2, OCN, and COL-1α1 expression levels by qRT-PCR (: P < 0.05, **: P < 0.01; compared to the M group; #: P < 0.05; compared to the MH group)

Fig. 3

The Impact of PBMT on Angiogenesis in Co-Culture Systems. A Representative images of tube formation in MC3T3-E1, HUVEC, and MC3T3-E1/HUVEC co-culture systems under light microscopy (100x magnification, scale bars represent 100 μm). B Quantification of the number of tubular branches in each experimental group (**: P < 0.01, **: P < 0.001; compared to the MC3T3-E1 group; #: P < 0.05; compared to the HUVEC group). C Statistical analysis of the total length of tubular branches in each experimental group (: P < 0.05, **: P < 0.001; compared to the MC3T3-E1 group; #: P < 0.05; compared to the HUVEC group). D-E Measurement of HIF-1α and VEGF expression levels by qRT-PCR (: P < 0.05, **: P < 0.01, ***: P < 0.001; compared to the HUVEC group; #: P < 0.05, ##: P < 0.01; compared to the MC3T3-E1 + HUVEC group)

Fig. 4

Animal Model and H&E Staining. C represents the control group, while 0–5 correspond to various levels of laser radiation characterized by energy densities of 0 J/cm², 1.7 J/cm², 3.5 J/cm², 5.3 J/cm², 7.1 J/cm², and 8.8 J/cm². A Establishment of the Orthodontic Tooth Movement Rat Model. (B) Statistical Analysis of Mesial Movement Distance of the Left Maxillary First Molar in Each Group (*: P < 0.05, compared to the OTM group). C-D Hematoxylin and Eosin (H&E) stained sections (40×, upper row, scale bar = 500 μm; 100×, lower row, scale bar = 200 μm). P denotes the Periodontal Ligament; Ab signifies Alveolar Bone; R represents Tooth Root

Fig. 5

Immunohistochemical Analysis of VEGF and CD31 Expression. C represents the control group, while 0–5 correspond to various levels of laser radiation characterized by energy densities of 0 J/cm², 1.7 J/cm², 3.5 J/cm², 5.3 J/cm², 7.1 J/cm², and 8.8 J/cm². A Immunohistochemical staining results of VEGF in various groups, showing OTM lateral maxilla and first molar sagittal tissue sections at 40x magnification, as well as local periodontal tissue of the distal root of the first molar at 200x magnification. B Comparison of VEGF expression in the periodontal ligament on the tension side between the control group and the experimental group(*: P < 0.05; compared to the control group; #: P < 0.05,; compared to the OTM group). C Immunohistochemical staining results of CD31 in different groups, presenting OTM lateral maxilla and first molar sagittal tissue sections at 40x magnification, and local periodontal tissue of the distal root of the first molar at 200x magnification. D Comparison of the expression levels of CD31 in the periodontal ligament on the tension side between the control group and the experimental group(*: P < 0.05; compared to the control group; #: P < 0.05,; compared to the OTM group)