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Optogenetic control of mesenchymal cell fate towards precise bone regeneration.

ABSTRACT: Rationale: Spatial-temporal control of cell fate in vivo is of great importance for regenerative medicine. Currently, there remain no practical strategies to tune cell-fate spatial-temporally. Optogenetics is a biological technique that widely used to control cell activity in genetically defined neurons in a spatiotemporal-specific manner by light. In this study, optogenetics was repurposed for precise bone tissue regeneration. Methods: Lhx8 and BMP2 genes, which are considered as the master genes for mesenchymal stem cell proliferation and differentiation respectively, were recombined into a customized optogenetic control system. In the system, Lhx8 was constitutively expressed, while BMP2 together with shLhx8 expression was driven by blue light. Results: As expected, blue light induced BMP2 expression and inactivated Lhx8 expression in cells infected with the optogenetic control system. Optogenetic control of BMP2 and Lhx8 expression inversely regulates MSC fate in vitro. By animal study, we found that blue light could fine-tune the regeneration in vivo. Blue light illumination significantly promotes bone regeneration when the scaffold was loaded with MSCs infected with adeno-Lhx8, GI-Gal4DBD, LOV-VP16, and BMP2-shLhx8. Conclusions: Together, our study revealed that optogenetic control of the master genes for mesenchymal stem cell proliferation and differentiation would be such a candidate strategy for precise regenerative medicine.

SUBMITTER: Wang W

PROVIDER: S-EPMC6857041 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Optogenetic control of mesenchymal cell fate towards precise bone regeneration.

Wang Weicai W Huang Delan D Ren Jianhan J Li Runze R Feng Zhicai Z Guan Chenyu C Bao Baicheng B Cai Bin B Ling Junqi J Zhou Chen C

Theranostics 20191018 26

Rationale: Spatial-temporal control of cell fate in vivo is of great importance for regenerative medicine. Currently, there remain no practical strategies to tune cell-fate spatial-temporally. Optogenetics is a biological technique that widely used to control cell activity in genetically defined neurons in a spatiotemporal-specific manner by light. In this study, optogenetics was repurposed for precise bone tissue regeneration. Methods: Lhx8 and BMP2 genes, which are conside ...[more]

PMID: 31754390

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Project description:BackgroundThe spatiotemporal regulation of essential genes is crucial for controlling the growth and differentiation of cells in a precise manner during regeneration. Recently, optogenetics was considered as a potent technology for sophisticated regulation of target genes, which might be a promising tool for regenerative medicine. In this study, we used an optogenetic control system to precisely regulate the expression of Lhx8 to promote efficient bone regeneration.MethodsQuantitative real-time PCR and western blotting were used to detect the expression of Lhx8 and osteogenic marker genes. Alkaline phosphatase staining and alizarin red staining were used to detect alkaline phosphatase activity and calcium nodules. A customized optogenetic expression system was constructed to regulate Lhx8, of which the expression was activated in blue light but not in dark. We also used a critical calvarial defect model for the analysis of bone regeneration in vivo. Moreover, micro-computed tomography (micro-CT), three-dimensional reconstruction, quantitative bone measurement, and histological and immunohistochemistry analysis were performed to investigate the formation of new bone in vivo.ResultsDuring the osteogenic differentiation of BMSCs, the expression levels of Lhx8 increased initially but then decreased thereafter. Lhx8 promoted the early proliferation of BMSCs but inhibited subsequent osteogenic differentiation. The optogenetic activation of Lhx8 in BMSCs in the early stages of differentiation by blue light stimulation led to a significant increase in cell proliferation, thus allowing a sufficient number of differentiating BMSCs to enter the later osteogenic differentiation stage. Analysis of the critical calvarial defect model revealed that the pulsed optogenetic activation of Lhx8 in transplanted BMSCs over a 5-day period led to a significant increase in the generation of bone in vivo.ConclusionsLhx8 plays a critical role in balancing proliferation and osteogenic differentiation in BMSCs. The optogenetic activation of Lhx8 expression at early stage of BMSCs differentiation led to better osteogenesis, which would be a promising strategy for precise bone regeneration.

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Optogenetic control of mesenchymal cell fate towards precise bone regeneration.

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Optogenetic control of mesenchymal cell fate towards precise bone regeneration.

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