No abstract

The Wnt/β-catenin signaling pathway appears to be particularly important for bone homeostasis, whereas nuclear accumulation of β-catenin requires the activation of Rac1, a member of the Rho small GTPase family. The aim of the present study was to investigate the role of RhoA/Rho kinase (Rock)-mediated Wnt/β-catenin signaling in the regulation of aging-associated bone loss. We find that Lrp5/6-dependent and Lrp5/6-independent RhoA/Rock activation by Wnt3a activates Jak1/2 to directly phosphorylate Gsk3β at Tyr216, resulting in Gsk3β activation and subsequent β-catenin destabilization. In line with these molecular events, RhoA loss- or gain-of-function in mouse embryonic limb bud ectoderms interacts genetically with Dkk1 gain-of-function to rescue the severe limb truncation phenotypes or to phenocopy the deletion of β-catenin, respectively. Likewise, RhoA loss-of-function in pre-osteoblasts robustly increases bone formation while gain-of-function decreases it. Importantly, high RhoA/Rock activity closely correlates with Jak and Gsk3β activities but inversely correlates with β-catenin signaling activity in bone marrow mesenchymal stromal cells from elderly male humans and mice, whereas systemic inhibition of Rock therefore activates the β-catenin signaling to antagonize aging-associated bone loss. Taken together, these results identify RhoA/Rock-dependent Gsk3β activation and subsequent β-catenin destabilization as a hitherto uncharacterized mechanism controlling limb outgrowth and bone homeostasis.

Rationale: The nuclear translocation of transcriptional factor Gli is indispensable for Hedgehog (Hh) signaling activation, whose deregulation causes cancer progressions; however, the mechanisms governing Gli nuclear translocation are poorly understood. Here, we report that the Gli translocation in response to Hh requires Rac1 activation. Methods: C3H10T1/2 cell line and mouse embryonic fibroblasts were used to explore the molecular mechanisms underlying Rac1 activity in regulation of Hh signaling transduction. Transgenic mouse strains and human medulloblastoma (MB) tissue samples were utilized to examine the role of Rac1 in Hh-directed limb bud development and MB progression. Results: We show that upon the binding of Hh to receptor Patched1 (Ptch1), receptor Smoothened (Smo) dissociates from Ptch1 and binds to Vav2, resulting in the increased phosphorylation levels of Vav2 at Y172, which further activates Rac1. The role of Rac1 is dependent on the regulation of phosphorylation levels of KIF3A at S689 and T694, which in turn affects IFT88 stability and subsequently dampens SuFu-Gli complex formation, leading to the release of Gli from the complex and the consequent translocation of Gli into the nucleus. Moreover, Vav2 phospho-Y172 levels are up-regulated in GFAP-Cre;SmoM2 +/- mouse cerebellum and human Shh type MB tissues, whereas deficiency of Rac1 in mouse embryonic limb bud ectoderm ( Prx1-Cre;Rac1 f/f ) impedes Hh activation by disruption of Gli nuclear translocation. Conclusion: Together, our results uncover the Rac1 activation and the subsequent Gli translocation as a hitherto uncharacterized mechanism controlling Hh signaling and may provide targets for therapeutic intervention of this signaling pathway.

Preeclampsia (PE), a pregnancy-specific syndrome with the major molecular determinants of placenta-borne oxidative stress and consequently impaired nitric oxide (NO) generation, has been considered to be one of the leading causes of maternal morbidity as well as mortality and preterm delivery worldwide. Several medical conditions have been found to be associated with increased PE risk, however, the treatment of PE remains unclear. Here, we report that Tianma Gouteng Decoction (TGD), which is used clinically for hypertension treatment, regulates oxidative stress and NO production in human extravillous trophoblast-derived TEV-1 cells. In human preeclamptic placental explants, reactive oxygen species (ROS) levels were elevated and NO production was inhibited, while TGD treatment at different periods effectively down-regulated the H2O2-induced ROS levels and significantly up-regulated the H2O2-suppressed NO production in human TEV-1 cells. Mechanistically, TGD enhanced the activity of total nitric oxide synthase (TNOS), which catalyze L-arginine oxidation into NO, and simultaneously, TGD promoted the expression of neuronal nitric oxide synthase (nNOS) and endothelial nitric oxide synthase (eNOS), two isoforms of nitric oxide synthetases (NOS) in human placenta, resulting in the increased NO generation. More importantly, TGD administration not only increased the weight gain during pregnancy and revealed a hypotensive effect, but also improved the placental weight gain and attenuated fetal growth restriction in an NG-nitro-L-arginine methyl ester (L-NAME)-induced mouse PE-like model. Our results thereby provide new insights into the role of TGD as a potentially novel treatment for PE.