TWIST Family of Basic Helix-Loop-Helix Transcription Factors Mediate Human Mesenchymal Stem Cell Growth and Commitment

Abstract

The TWIST family of basic helix-loop-helix transcription factors, Twist-1 and Dermo-1 are known mediators of mesodermal tissue development and contribute to correct patterning of the skeleton. In this study, we demonstrate that freshly purified human bone marrow-derived mesenchymal stromal/stem cells (MSC) express high levels of Twist-1 and Dermo-1 which are downregulated following ex vivo expansion. Enforced expression of Twist-1 or Dermo-1 in human MSC cultures increased expression of the MSC marker, STRO-1, and the early osteogenic transcription factors, Runx2 and Msx2. Conversely, overexpression of Twist-1 and Dermo-1 was associated with a decrease in the gene expression of osteoblast-associated markers, bone morphogenic protein-2, bone sialoprotein, osteopontin, alkaline phosphatase and osteocalcin. High expressing Twist-1 or Dermo-1 MSC lines exhibited an enhanced proliferative potential of approximately 2.5-fold compared with control MSC populations that were associated with elevated levels of Id-1 and Id-2 gene expression. Functional studies demonstrated that high expressing Twist-1 and Dermo-1 MSC displayed a decreased capacity for osteo/chondrogenic differentiation and an enhanced capacity to undergo adipogenesis. These findings implicate the TWIST gene family members as potential mediators of MSC self-renewal and lineage commitment in postnatal skeletal tissues by exerting their effects on genes involved in the early stages of bone development.

Identification of Novel EZH2 Targets Regulating Osteogenic Differentiation in Mesenchymal Stem Cells

“…MSC were isolated and cultured using plastic adherence [34]. Culture media: Alpha modified Eagle's medium (a-MEM) (Sigma Aldrich, Inc., St Louis, MO) supplemented with 10% (v/v) fetal calf serum (SAFC Biosciences, Melbourne, VIC), 50 U/mL, 5 mg/mL penicillin, streptomycin (Sigma Aldrich, Inc.), 1 mM sodium pyruvate (Sigma Aldrich, Inc.), 100 mM L-ascorbate-2-phosphate (Wako Pure Chemical industries, Richmond, VA), 2 mM l-glutamine ( JRH Biosciences/Sigma, Lenexa, KS), and 10 mM HEPES (Sigma Aldrich, Inc.), and cultured at 37°C, >90% humidity and 5% CO2.…”

“…In vitro osteogenic differentiation assay siRNA-treated MSC in 96-well plates were cultured in either noninductive culture media or osteogenic inductive media (a-MEM supplemented with 5% (v/v) FCS, 2 mM l-Glutamine, 1 mM sodium pyruvate, 10 mM HEPES buffer and 50 U/mL penicillin-streptomycin, 100 mM L-ascorbate-2-phosphate, 0.1 mM dexamethasone, and 2.6 mM KH2PO4) [34] In vitro adipogenic differentiation assay siRNA-treated MSC in 96-well plates were cultured in either noninductive culture media or adipogenic inductive media (a-MEM supplemented with 10% [v/v] FCS, 2 mM lGlutamine, 1 mM sodium pyruvate, 10 mM HEPES buffer and 50 U/mL penicillin-streptomycin, 100 mM l-ascorbate-2-phosphate, 0.5 mM methyl isobutyl methylxanthine, 0.5 mM hydrocortisone, and 60 mM indomethacin) [34] for 14 days with media changes twice weekly. Oil Red O (Sigma Aldrich, Inc.) identified lipid formed by human MSC under adipogenic differentiation.…”

Twist2 contributes to breast cancer progression by promoting an epithelial–mesenchymal transition and cancer stem-like cell self-renewal

“…Twist2, similar to Twist1, is a potential mediator of mesenchymal stem cell self-renewal and lineage commitment. In addition, Twist2 may function to regulate critical transcription factors and osteo-/chondrogenic-inductive factors in the early stages of cell fate determination in human mesenchymal stem cells (Isenmann et al, 2009). However, the functional significance of Twist2 in human breast cancer and in regulating CSC stemness as well as the underlying cellular and molecular mechanisms are poorly studied.…”

TWISTing an embryonic transcription factor into an oncoprotein

“…In Drosophila pupals, Twi was found to be expressed within the adult muscle precursor cells, which invade the larval oblique muscles and was determined as essential for their splitting into dorsal longitudinal muscles found in the thorax of the adult fly (Cripps and Olson, 1998). Ectopic expression of TWIST1 or TWIST2 in human bone marrow-derived mesenchymal stromal/stem cells (MSC) was found to maintain an immature stromal phenotype, to inhibit their osteo-chondrogenic potential and conversely to increase their adipocyte potential, strengthening their role in cell fate determination (Isenmann et al, 2009). TWIST2 was additionally found to be expressed in granulocyte-macrophage progenitors, preventing their proliferation and differentiation into macrophages, neutrophils and basophils, through the inactivation of both RUNX1 and C/EBPa transcription factors (Sharabi et al, 2008).…”