Small GTPases of the Rho family (RhoA and Rac-1) are responsible for cytoskeleton dynamics (particularly actin polymerisation) and control cellular tension. For these reasons, they are implicated in the commitment of multipotent cells (MCs). In one hand, increased tension (important RhoA activity) is commonly associated with osteogenesis (OS), in the other hand, a reduced one (low RhoA activity) is associated with adipogenesis (AD). Nevertheless, precise RhoGTPases regulations during stem cell commitment are not known. We and others observed that strontium (an anti-osteoporotic drug) promotes OS and prevents AD of MCs. We hypothesize that strontium (Sr) effects may be explained by its ability to modulate RhoGTPases activities.
To address this question, we cultivated C3H10T1/2 on microspheres in a dynamic bioreactor in conditions where OS (apatite coated beads) and AD (rosiglitazone) were promoted. 3D cultures were treated or not with 5 mM of Sr up to 6 days. OS, AD specific markers were evaluated by quantitative RT-PCR, RhoA and Rac1 activities were quantified by specific G-LISA. In our hands, OS is associated with sustained RhoA and increasing Rac1 activities whereas AD is characterized by a Rac1 activity reduction. We observe, as expected, that Sr inhibits AD (PPARγ2, C/EBPα, FABP4 & Adiponectin) and promote OS (ALP, OSX, BSP & OPN). During MC commitment, Sr increases significantly both RhoA and Rac1 activities from day 2 to 6 in OS and AD conditions. Dual activation of RhoA and Rac1 in MCs may explain why AD is blocked by strontium and fits with the promotion of fibrillogenesis and Wnt/β catenin signalling respectively controlled by RhoA and Rac1 in OS conditions.
As RhoA and Rac1 are mainly regulated by integrins and VEGFR dependent pathways, we measure various integrins, VEGF isoforms and receptors expressions. We identify matrix-bound VEGF188 and Flk1 as potential targets of Sr and ongoing studies will help understanding ability of VEGF188 to regulate RhoGTPases.
In conclusion, the ability of Sr to stimulate both RhoA and Rac1 in MCs may explain its particular action on osteoblasto- and adipo-genesis.
17 May 2014 - 20 May 2014