Accelerated Bone Loss in Transgenic Mice Expressing Constitutively Active TGF-ß Receptor Type I.

Transforming growth factor beta (TGF-ß) is a key factor mediating the intercellular crosstalk between the hematopoietic stem cells and their microenvironment. Here, we investigated the skeletal phenotype of transgenic mice expressing constitutively active TGF-ß receptor type I under the control of Mx1-Cre (Mx1;TßRICA mice). µCT analysis showed decreased cortical thickness, and cancellous bone volume in both femurs and mandibles. Histomorphometric analysis confirmed a decrease in cancellous bone volume due to increased osteoclast number and decreased osteoblast number. Primary osteoblasts showed decreased ALP and mineralization. Constitutive TßRI activation increased osteoclast differentiation. qPCR analysis showed that Tnfsf11/Tnfrsf11b ratio, Ctsk, Sufu, and Csf1 were increased whereas Runx2, Ptch1, and Ptch2 were decreased in Mx1;TßRICA femurs. Interestingly, Gli1, Wnt3a, Sp7, Alpl, Ptch1, Ptch2, and Shh mRNA expression were reduced whereas Tnfsf11/Tnfrsf11b ratio was increased in Mx1;TßRICA mandibles. Similarly, osteoclast-related genes were increased in Mx1;TßRICA osteoclasts whereas osteoblast-related genes were reduced in Mx1;TßRICA osteoblasts. Western blot analysis indicated that SMAD2 and SMAD3 phosphorylation was increased in Mx1;TßRICA osteoblasts, and SMAD3 phosphorylation was increased in Mx1;TßRICA osteoclasts. CTSK was increased while RUNX2 and PTCH1 was decreased in Mx1;TßRICA mice. Microindentation analysis indicated decreased hardness in Mx1;TßRICA mice. Our study indicated that Mx1;TßRICA mice were osteopenic by increasing osteoclast number and decreasing osteoblast number, possibly by suppressing Hedgehog signaling pathways.

Distinctions in bone matrix nanostructure, composition, and formation between osteoblast-like cells, MG-63, and human mesenchymal stem cells, UE7T-13.

Osteoblast-like cells and human mesenchymal stem cells (hMSCs) are frequently employed as osteoprogenitor cell models for evaluating novel biomaterials in bone healing and tissue engineering. In this study, the characterization of UE7T-13 hMSCs and MG-63 human osteoblast-like cells was examined. Both cells can undergo osteogenesis and produce calcium extracellular matrix; however, calcium nodules produced by MG-63 lacked a central mass and appeared flatter than UE7T-13. The absence of growing calcium nodules in MG-63 was discovered by SEM-EDX to be associated with the formation of alternating layers of cells and calcium extracellular matrix. The nanostructure and composition analysis showed that UE7T-13 had a finer nanostructure of calcium nodules with a higher calcium/phosphate ratio than MG-63. Both cells expressed high intrinsic levels of collagen type I alpha 1 chain, while only UE7T-13 expressed high levels of alkaline phosphatase, biomineralization associated (ALPL). High ALP activity in UE7T-13 was not further enhanced by osteogenic induction, but in MG-63, low intrinsic ALP activity was greatly induced by osteogenic induction. These findings highlight the differences between the two immortal osteoprogenitor cell lines, along with some technical notes that should be considered while selecting and interpreting the pertinent in vitro model.