Lentiviral‑mediated Shh reverses the adverse effects of high glucose on osteoblast function and promotes bone formation via Sonic hedgehog signaling

Jiang,Zhu‑Ling; Jin,Han; Liu,Zhong‑Shuang; Liu,Ming‑Yue; Cao,Xiao‑Fang; Jiang,Yang‑Yang; Bai,Hong‑Dan; Zhang,Bin; Li,Ying

doi:10.3892/mmr.2019.10540

Lentiviral‑mediated Shh reverses the adverse effects of high glucose on osteoblast function and promotes bone formation via Sonic hedgehog signaling

Authors:
- Zhu‑Ling Jiang
- Han Jin
- Zhong‑Shuang Liu
- Ming‑Yue Liu
- Xiao‑Fang Cao
- Yang‑Yang Jiang
- Hong‑Dan Bai
- Bin Zhang
- Ying Li
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Affiliations: Department of Implantology, The Second Affiliated Hospital, Harbin Medical University, Harbin, Heilongjiang 150001, P.R. China, Institute of Hard Tissue Development and Regeneration, The Second Affiliated Hospital, Harbin Medical University, Harbin, Heilongjiang 150001, P.R. China, Department of Dentistry, The Second Affiliated Hospital, Harbin Medical University, Harbin, Heilongjiang 150001, P.R. China, Department of Dentistry, The Affiliated Hospital, Harbin Institute of Technology, Harbin, Heilongjiang 150001, P.R. China, Feiyang Dental Clinic, Heihe, Heilongjiang 164300, P.R. China
Published online on: July 31, 2019 https://doi.org/10.3892/mmr.2019.10540
Pages: 3265-3275
Copyright: © Jiang et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

Patients with diabetes tend to have an increased incidence of osteoporosis, which may be associated with hyperglycemia; however, the pathogenic mechanisms governing this interaction remain unknown. The present study sought to investigate whether elevated extracellular glucose levels of bone mesenchymal stem cells (BMSCs) could influence osteoblastic differentiation and whether the intracellular Sonic hedgehog (Shh) pathway could adjust the effects. Furthermore, to verify the results in vivo, a rat tooth extraction model was constructed. BMSCs were incubated in eight types of culture medium, including low glucose (LG), LG + lentivirus (Lenti), LG + Lenti‑small interfering RNA (Lenti‑siRNA), LG + Lenti‑Shh, high glucose (HG), HG + Lenti, HG + Lenti‑siRNA and HG + Lenti‑Shh. The lentiviral transfection efficiency was observed using a fluorescence microscope; protein and mRNA expression was detected by western blotting and reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR). The matrix mineralization and alkaline phosphatase (ALP) activity of BMSCs were examined by Alizarin red staining and ALP activity assays, respectively. The expression of osteogenesis‑related genes in BMSCs were quantified by RT‑qPCR. The alveolar ridge reduction was measured and histological sections were used to evaluate new bone formation in the tooth socket. With high concentrations of glucose, Shh expression, matrix mineralization nodules formation, ALP activity and the levels of bone morphogenic protein 4 (BMP4), bone sialoprotein (BSP) and osteopontin (OPN) expression were greatly reduced compared with LG and corresponding control groups. Whereas activated Shh signaling via Lenti‑Shh could increase the number of matrix mineralization nodules, ALP activity, and the expression levels of BMP4, BSP and OPN in BMSCs. Additionally, in vivo assays demonstrated that Lenti‑Shh induced additional bone formation. Collectively, the results of the present study indicated that HG inhibited the Shh pathway in osteoblasts and resulted in patterning defects during osteoblastic differentiation and bone formation, while the activation of Shh signaling could suppress these deleterious effects.

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