Endogenous Wnt Signaling Promotes Proliferation and Suppresses Osteogenic Differentiation in Human Adipose Derived Stromal Cells

Abstract

Multipotential adult mesenchymal stem cells (MSC) are able to differentiate along several known lineages, and lineage commitment is tightly regulated through specific cellular mediators and interactions. Human adipose tissues contain cell populations that have similar characteristics to bone marrow stromal cells. Wnt proteins have been reported to be involved in proliferation and differentiation of stem cells. RNA interference (RNAi) has recently emerged as a specific and efficient method to silence gene expression in mammalian cells. To analyze the role of beta-catenin signaling in human adipose stromal cells (hADSC), the effects of beta-catenin short hairpin RNAs (shRNA) expression and Wnt3a conditioned media on the growth and differentiation properties of hADSC were examined. Expression of an RNAi molecule to beta-catenin from a lentivirus vector decreased beta-catenin expression in hADSC, as indicated by Western blot and immunohistochemistry. Cells transduced with sibeta-catenin lentivirus had decreased CFU and lower numbers of cells per colony than transduced control cells, but this outcome did not result from altered attachment efficiency of hADSC. The inhibition of beta-catenin signal by RNAi expression increased osteogenic differentiation. The treatment of Wnt3a conditioned media increased cellular beta-catenin levels and the rate of cellular proliferation, but inhibited osteogenic differentiation. Transduction of beta-catenin RNAi lentivirus blocked the effect of Wnt3a on proliferation of hADSC. Taken together, these findings indicate that endogenous Wnt3a plays an important role in the regulation of proliferation and differentiation of hADSC.

Opposite Spectrum of Activity of Canonical Wnt Signaling in the Osteogenic Context of Undifferentiated and Differentiated Mesenchymal Cells: Implications for Tissue Engineering

“…Our observation that Wnt3a treatment inhibits osteogenic differentiation in mesenchymal cells (e.g., ASCs and E16) is consistent with previous in vitro studies performed on other multipotent mesenchymal cells. [41][42][43][44] In addition, our study provides also in vivo evidence complementing in vitro results. It must be pointed out that in vitro Wnt3a treatment represents a constant and continuous treatment of cells, compared to in vivo treatment where calvarial bones are initially exposed to doses of Wnt3a protein, which gradually decreased overtime.…”

Glycogen synthase kinase-3α/β inhibition promotes in vivo amplification of endogenous mesenchymal progenitors with osteogenic and adipogenic potential and their differentiation to the osteogenic lineage

“…Elucidating how Wnts act as instructive cues for the recruitment, maintenance, and maturation of MSCs and their differentiated progenies is of primary interest given the potential use of these cells in regenerative medicine. Prior studies have implicated canonical Wnt signaling in the regulation of bone metabolism at several levels, in amplifying the undifferentiated MSC pool, (7,8) commitment of undifferentiated MSCs to the osteoblast lineage, (13,14) and stimulation of their differentiation. (14) However, differences in cell preparations, use of models that did not take into account the cellular context, or use of nonphysiologic loss-and gain-of-function approaches are some of the reasons for the emergence of contrasting data.…”

“…(7,8) Moreover, the overexpression of LRP5, a key coreceptor specifically involved in canonical Wnt signaling, was reported to increase proliferation of MSCs. (9) Activation of Wnt signaling by lithium chloride, a nonspecific GSK-3 inhibitor, was shown to suppress dexamethasone-induced osteogenesis.…”

Diverse Effects of Cyclic AMP Variants on Osteogenic and Adipogenic Differentiation of Human Mesenchymal Stromal Cells

“…We and others have demonstrated that activation of Wnt signaling in hMSCs inhibits osteogenic differentiation [34][35][36][37], and the high number of Wnt-regulated genes in 8-br-cAMP possibly negatively correlates with osteogenic differentiation. Insulin-like growth factor (IGF) signaling and in particular the IGF binding proteins (IGFBPs), were also differentially expressed between the two types of cultures.…”