Low intensity pulsed ultrasound (LIPUS) maintains osteogenic potency by the increased expression and stability of Nanog through spleen tyrosine kinase (Syk) activation

Cell Signal. 2019 Oct:62:109345. doi: 10.1016/j.cellsig.2019.109345. Epub 2019 Jun 19.

Abstract

Mesenchymal stem cells (MSCs) are a powerful tool for cell-based, clinical therapies like bone regeneration. Therapeutic use of cell transplantation requires many cells, however, the expansion process needed to produce large quantities of cells reduces the differentiation potential of MSCs. Here, we examined the protective effects of low intensity pulsed ultrasound (LIPUS) on the maintenance of osteogenic potency. Primary osteoblastic cells were serially passaged between 2 and 12 times with daily LIPUS treatment. We found that LIPUS stimulation maintains osteogenic differentiation capacity in serially passaged cells, as characterized by improved matrix mineralization and Osteocalcin mRNA expression. Decreased expression of Nanog, Sox2, and Msx2, and increased expression of Pparg2 from serial passaging was recovered in LIPUS-stimulated cells. We found that LIPUS stimulation not only increased but also sustained expression of Nanog in primary osteoblasts and ST2 cells, a mouse mesenchymal stromal cell line. Nanog overexpression in serially passaged cells mimicked the recuperative effects of LIPUS on osteogenic potency, highlighting the important role of Nanog in LIPUS stimulation. Additionally, we found that spleen tyrosine kinase (Syk) is an important signaling molecule to induce Nanog expression in LIPUS-stimulated cells. Syk activation was regulated by both Rho-associated kinase 1 (ROCK1) and extracellular ATP in a paracrine manner. Interestingly, the LIPUS-induced increase in Nanog mRNA expression was regulated by ATP-P2X4-Syk Y323 activation, while the improvement of Nanog protein stability was controlled by the ROCK1-Syk Y525/526 pathway. Taken together, these results indicate that LIPUS stimulation recovers and maintains the osteogenic potency of serially passaged cells through a Syk-Nanog axis.

Keywords: Differentiation; Mesenchymal stem cell; Osteoblast; Osteogenesis; Passage.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cell Differentiation / radiation effects
  • Gene Expression Regulation, Developmental / radiation effects
  • Mesenchymal Stem Cells / metabolism*
  • Mesenchymal Stem Cells / radiation effects
  • Mice
  • Nanog Homeobox Protein / genetics*
  • Osteoblasts / radiation effects
  • Osteogenesis / genetics*
  • Osteogenesis / radiation effects
  • SOXB1 Transcription Factors / genetics
  • Syk Kinase / genetics*
  • Ultrasonic Waves
  • rho-Associated Kinases / genetics*

Substances

  • Nanog Homeobox Protein
  • SOX2 protein, human
  • SOXB1 Transcription Factors
  • Syk Kinase
  • Rock1 protein, mouse
  • rho-Associated Kinases