Citation
- Authors: Chen M. et al.
- Year: 2022
- Journal: Cell Mol Life Sci 79 107
- Applications: in vitro / in vivo / DNA, siRNA / in vivo-jetPEI, jetPRIME
- Cell type: ST2
Method
in vitro: The expression construct of MTSS1 was purchased from Origene (Rockville, MD, USA). Each construct or the vector was transfected into ST2 cells at the confluence of 70-80% using JetPRIME transfection reagent. After 4 hrs of transfection, the transfection medium was removed and replaced with complete culture medium. The cells were induced at appropriate confluence to allow osteogenic differentiation.
in vivo: To deliver the siRNAs to the bone marrow of mice, 2’-Ome modified MTSS1 siRNA or control siRNA (Genepharma, Shanghai, China) was formulated with in vivo-jetPEI (Polyplus, Illkirch, France) following the supplier’s instructions.
Briefly, for one mouse, 11 μg siRNA and 1.5 μL in vivo-jetPEI were diluted, respectively, with 12.5 μl of 5% glucose and then mixed. Afterwards, the formulated mixture was injected to the marrow cavity of the mouse tibia. The in vivo efficacy of the MTSS1 siRNA in silencing the endogenous mRNA expresssion was tested in mice 48 h after receiving intra-tibial transfection, with the mice receiving control siRNA as control. The mice received the transfections twice in total, with an interval of 3 weeks
Abstract
Metastasis suppressor 1 (MTSS1) plays an inhibitory role in tumorigenesis and metastasis of a variety of cancers. To date, the function of MTSS1 in the differentiation of marrow stromal progenitor cells remains to be explored. In the current study, we investigated whether and how MTSS1 has a role in osteoblast differentiation and bone homeostasis. Our data showed that MTSS1 mRNA was upregulated during osteoblast differentiation and downregulated in the osteoblastic lineage cells of ovariectomized and aged mice. Functional studies revealed that MTSS1 promoted the osteogenic differentiation from marrow stromal progenitor cells. Mechanistic explorations uncovered that the inactivation of Src and afterward activation of canonical Wnt signaling were involved in osteoblast differentiation induced by MTSS1. The enhanced osteogenic differentiation induced by MTSS1 overexpression was attenuated when Src was simultaneously overexpressed, and conversely, the inhibition of osteogenic differentiation by MTSS1 siRNA was rescued when the Src inhibitor was supplemented to the culture. Finally, the in vivo transfection of MTSS1 siRNA to the marrow of mice significantly reduced the trabecular bone mass, along with the reduction of trabecular osteoblasts, the accumulation of marrow adipocytes, and the increase of phospho-Src-positive cells on the trabeculae. No change in the number of osteoclasts was observed. This study has unraveled that MTSS1 contributes to osteoblast differentiation and bone homeostasis through regulating Src-Wnt/β-catenin signaling. It also suggests the potential of MTSS1 as a new target for the treatment of osteoporosis.