Activation of the Ca2+/calcineurin/NFAT2 pathway controls smooth muscle cell differentiation

International audienceCellular mechanisms controlling smooth muscle cells (SMCs) phenotypic modulation are largely unknown. Intracellular Ca2+ movements are essential to ensure SMC functions; one of the roles of Ca 2+ is to regulate calcineurin, which in turn induces nuclear localization of the nuclear factor of activated T-cell (NFAT). In order to investigate, during phenotypic differentiation of SMCs, the effect of calcineurin inhibition on NFAT2 nuclear translocation, we used a culture model of SMC differentiation in serum-free conditions. We show that the treatment of cultured SMC with the calcineurin inhibitor cyclosporine A induced their dedifferentiation while preventing their differentiation. These findings suggest that nuclear translocation of NFAT2 is dependent of calcineurin activity during the in vitro SMC differentiation kinetic and that the nuclear presence of NFAT2 is critical in the acquisition and maintenance of SMC differentiation

Mapping 3-dimensional neovessel organization steps using micro-computed tomography in a murine model of hindlimb ischemia-brief report.

International audienceOBJECTIVE: Studying the mechanisms of neovascularization and evaluating the effects of proangiogenic strategies require accurate analysis of the neovascular network. We sought to evaluate the contribution of the microcomputed tomography (mCT) providing high-resolution 3-dimensional (3D) structural data, to a better comprehension of the well-studied mouse hindlimb postischemic neovascularization. METHODS AND RESULTS: We showed a predominant arteriogenesis process in the thigh and a predominant angiogenesis-related process in the tibiofibular region, in response to ischemia during the first 15 days. After 15 days, mCT quantitative analysis reveals a remodeling of arterial neovessels and a regression depending on the restoration of the blood flow. We provided also new mCT data on the rapid and potent angiogenic effects of mesenchymal stem cell therapy on vessel formation and organization. We discussed the contribution of this technique compared with or in addition to data generated by the more conventional approaches. CONCLUSIONS: This study demonstrated that optimized mCT is a robust method for providing new insights into the 3D understanding of postischemic vessel formation

Sonic hedgehog mediates a novel pathway of PDGF-BB-dependent vessel maturation

International audienceRecruitment of mural cells, i.e. pericytes and smooth muscle cells (SMCs), is essential to improve the maturation of newly formed vessels. Sonic hedgehog (Shh) has been suggested to promote the formation of larger and more muscularized vessels, but the underlying mechanisms of this process have not yet been elucidated. We first identified Shh as a target of PDGF-BB and found that SMCs respond to Shh by upregulating ERK1/2 and Akt phosphorylation. We next showed that PDGF-BB-induced SMC migration was reduced after inhibition of Shh or its signaling pathway. Moreover, we found that PDGF-BB-induced SMC migration, involves Shh-mediated motility. In vivo, in the mouse model of corneal angiogenesis, Shh is expressed by mural cells of newly formed blood vessels. PDGF-BB inhibition reduced Shh expression, demonstrating that Shh is a target of PDGF-BB, confirming in vitro experiments. Finally, we found that in vivo inhibition of either PDGF-BB or Shh signaling reduces NG2+ mural cell recruitment into neovessels and subsequently reduces neo-vessel lifespan. Our findings demonstrate, for the first time, that Shh is involved in PDGF-BB-induced SMC migration and recruitment of mural cells into neo-vessels and elucidate the molecular signaling pathway involved in this process

Regulation of Endothelial Cell Cytoskeletal Reorganization by a Secreted Frizzled-Related Protein-1 and Frizzled 4- and Frizzled 7-Dependent Pathway : Role in Neovessel Formation

Consistent with findings of Wnt pathway members involved in vascular cells, a role for Wnt/Frizzled signaling has recently emerged in vascular cell development. Among the few Wnt family members implicated in vessel formation in adult, Wnt7b and Frizzled 4 have been shown as involved in vessel formation in the lung and in the retina, respectively. Our previous work has shown a role for secreted Frizzled-related protein-1 (sFRP-1), a proposed Wnt signaling inhibitor, in neovascularization after an ischemic event and demonstrated its role as a potent angiogenic factor. However the mechanisms involved have not been investigated. Here, we show that sFRP-1 treatment increases endothelial cell spreading on extracellular matrix as revealed by actin stress fiber reorganization in an integrin-dependent manner. We demonstrate that sFRP-1 can interact with Wnt receptors Frizzled 4 and 7 on endothelial cells to transduce downstream to cellular machineries requiring Rac-1 activity in cooperation with GSK-3β. sFRP-1 overexpression in endothelium specifically reversed the inactivation of GSK-3β and increased neovascularization in ischemia-induced angiogenesis in mouse hindlimb. This study illustrates a regulated pathway by sFRP-1 involving GSK-3β and Rac-1 in endothelial cell cytoskeletal reorganization and in neovessel formation