Concept: Sonic hedgehog
BACKGROUND: Thuja orientalis has been traditionally used to treat patients who suffer from baldness and hair loss in East Asia. The present study sought to investigate the hair growth-promoting activity of T. orientalis hot water extract and the underlying mechanism of action. METHODS: After T. orientalis extract was topically applied to the shaved dorsal skin of telogenic C57BL/6 N mice, the histomorphometric analysis was employed to study induction of the hair follicle cycle. To determine the effect of T. orientalis extract on the telogen to anagen transition, the protein expression levels of beta-catenin and Sonic hedgehog (Shh) in hair follicles were determined by immunohistochemistry. RESULTS: We observed that T. orientalis extract promoted hair growth by inducing the anagen phase in telogenic C57BL/6 N mice. Specifically, the histomorphometric analysis data indicates that topical application of T. orientalis extract induced an earlier anagen phase and prolonged the mature anagen phase, in contrast to either the control or 1% minoxidil-treated group. We also observed increases in both the number and size of hair follicles of the T. orientalis extract-treated group. Moreover, the immunohistochemical analysis reveals earlier induction of beta-catenin and Shh proteins in hair follicles of the T. orientalis extract-treated group, compared to the control or 1% minoxidil-treated group. CONCLUSION: These results suggest that T. orientalis extract promotes hair growth by inducing the anagen phase in resting hair follicles and might therefore be a potential hair growth-promoting agent.
Axons must switch responsiveness to guidance cues during development for correct pathfinding. Sonic Hedgehog (Shh) attracts spinal cord commissural axons ventrally toward the floorplate. We show that after crossing the floorplate, commissural axons switch their response to Shh from attraction to repulsion, so that they are repelled anteriorly by a posterior-high/anterior-low Shh gradient along the longitudinal axis. This switch is recapitulated in vitro with dissociated commissural neurons as they age, indicating that the switch is intrinsic and time dependent. 14-3-3 protein inhibition converted Shh-mediated repulsion of aged dissociated neurons to attraction and prevented the correct anterior turn of postcrossing commissural axons in vivo, an effect mediated through PKA. Conversely, overexpression of 14-3-3 proteins was sufficient to drive the switch from Shh-mediated attraction to repulsion both in vitro and in vivo. Therefore, we identify a 14-3-3 protein-dependent mechanism for a cell-intrinsic temporal switch in the polarity of axon turning responses.
Administration of cadmium (Cd) after 60 h (H) incubation induces ventral body wall defect (VBWD) similar to the omphalocele phenotype in the chick embryo. In this model, the earliest histological changes have been observed in somites commencing at 4-h post-treatment (4H). The molecular mechanism by which Cd acts in this critical period of embryogenesis still remains unclear. Sonic hedgehog (SHH) signalling plays an important role in vertebrate development, including somitogenesis and thus ventral body wall formation. Patched (PTCH), a cell membrane receptor for SHH, is expressed in somites and Patched knockout mice display somite dysfunction. Another transmembrane receptor, Smoothened (SMO), is also expressed in somites and transduces the SHH signal regulated by PTCH. We designed this study to test the hypothesis that SHH signalling is downregulated during the critical period of early embryogenesis in the Cd-induced omphalocele chick model.
Activation of sonic hedgehog (HH) signaling pathway has been implicated in aggressiveness and progression of gastrointestinal tumors. We planned this study to identify a subgroup of gastric cancer (GC) patients with HH activation and to assess the effect of a HH inhibitor in HH activated GC in vitro. We surveyed HH pathway activation among 512 GC specimens for protein expression of various target genes involved in HH pathway: Indian hedgehog (IHH), patched-1 (PTCH1), smoothened (SMO), GLI2, and FOXA2. We analyzed the correlations between the expression of these factors and clinicopathological features and prognosis. In vitro, ten gastric cancer cell lines were screened for anti-tumoractivity of an HH inhibitor, GDC-0449. Among the 512 specimens, 105 (20.0 %), 83 (16.3 %), 130 (25.5 %), 61 (12.0 %), and 206 (40.8 %) were positive for IHH, PTCH1, GLI2, SMO, and FOXA2 expression, respectively. PTCH1 expression was more frequently observed in well- or moderately differentiated tubular adenocarcinoma, intestinal type and low stage GC. GLI2 was correlated with lymphovascular invasion and intestinal type GC. A high-stage and negative PTCH1 staining were identified as unfavorable independent risk factors for overall survival in multivariate analysis (P < 0.001, 0.045, respectively). For IHH, SMO, and FOXA2, there was no statistical difference in clinicopathologic variables and survival outcome. An HH inhibitor had particularly potent effects on GC cell lines with SMO mRNA overexpression. This is the largest report to analyze the hedgehog pathway in GC. PTCH1 overexpression was an independent prognostic factor for survival and SMO overexpression which was found in 12.0 % of GC patients might be the potential predictive marker of HH inhibitor.
Limb reduction and loss are hallmarks of snake evolution. Although advanced snakes are completely limbless, basal and intermediate snakes retain pelvic girdles and small rudiments of the femur. Moreover, legs may have re-emerged in extinct snake lineages [1-5], suggesting that the mechanisms of limb development were not completely lost in snakes. Here we report that hindlimb development arrests in python embryos as a result of mutations that abolish essential transcription factor binding sites in the limb-specific enhancer of Sonic hedgehog (SHH). Consequently, SHH transcription is weak and transient in python hindlimb buds, leading to early termination of a genetic circuit that drives limb outgrowth. Our results suggest that degenerate evolution of the SHH limb enhancer played a role in reduction of hindlimbs during snake evolution. By contrast, HOXD digit enhancers are conserved in pythons, and HOXD gene expression in the hindlimb buds progresses to the distal phase, forming an autopodial (digit) domain. Python hindlimb buds then develop transitory pre-chondrogenic condensations of the tibia, fibula, and footplate, raising the possibility that re-emergence of hindlimbs during snake evolution did not require de novo re-evolution of lost structures but instead could have resulted from persistence of embryonic legs. VIDEO ABSTRACT.
The evolution of body shape is thought to be tightly coupled to changes in regulatory sequences, but specific molecular events associated with major morphological transitions in vertebrates have remained elusive. We identified snake-specific sequence changes within an otherwise highly conserved long-range limb enhancer of Sonic hedgehog (Shh). Transgenic mouse reporter assays revealed that the in vivo activity pattern of the enhancer is conserved across a wide range of vertebrates, including fish, but not in snakes. Genomic substitution of the mouse enhancer with its human or fish ortholog results in normal limb development. In contrast, replacement with snake orthologs caused severe limb reduction. Synthetic restoration of a single transcription factor binding site lost in the snake lineage reinstated full in vivo function to the snake enhancer. Our results demonstrate changes in a regulatory sequence associated with a major body plan transition and highlight the role of enhancers in morphological evolution. PAPERCLIP.
Dysregulation of the sonic hedgehog (Shh) signaling pathway has been associated with cancer stem cells (CSC) and implicated in the initiation of pancreatic cancer. Pancreatic CSCs are rare tumor cells characterized by their ability to self-renew, and are responsible for tumor recurrence accompanied by resistance to current therapies. The lethality of these incurable, aggressive and invasive pancreatic tumors remains a daunting clinical challenge. Thus, the objective of this study was to investigate the role of Shh pathway in pancreatic cancer and to examine the molecular mechanisms by which sulforaphane (SFN), an active compound in cruciferous vegetables, inhibits self-renewal capacity of human pancreatic CSCs. Interestingly, we demonstrate here that Shh pathway is highly activated in pancreatic CSCs and plays important role in maintaining stemness by regulating the expression of stemness genes. Given the requirement for Hedgehog in pancreatic cancer, we investigated whether hedgehog blockade by SFN could target the stem cell population in pancreatic cancer. In an in vitro model, human pancreatic CSCs derived spheres were significantly inhibited on treatment with SFN, suggesting the clonogenic depletion of the CSCs. Interestingly, SFN inhibited the components of Shh pathway and Gli transcriptional activity. Interference of Shh-Gli signaling significantly blocked SFN-induced inhibitory effects demonstrating the requirement of an active pathway for the growth of pancreatic CSCs. SFN also inhibited downstream targets of Gli transcription by suppressing the expression of pluripotency maintaining factors (Nanog and Oct-4) as well as PDGFRα and Cyclin D1. Furthermore, SFN induced apoptosis by inhibition of BCL-2 and activation of caspases. Our data reveal the essential role of Shh-Gli signaling in controlling the characteristics of pancreatic CSCs. We propose that pancreatic cancer preventative effects of SFN may result from inhibition of the Shh pathway. Thus Sulforaphane potentially represents an inexpensive, safe and effective alternative for the management of pancreatic cancer.
The Bone Morphogenetic Protein (BMP) family reiteratively signals to direct disparate cellular fates throughout embryogenesis. In the developing dorsal spinal cord, multiple BMPs are required to specify sensory interneurons (INs). Previous studies suggested that the BMPs act as concentration-dependent morphogens to direct IN identity, analogous to the manner in which sonic hedgehog patterns the ventral spinal cord. However, it remains unresolved how multiple BMPs would cooperate to establish a unified morphogen gradient. Our studies support an alternative model: BMPs have signal-specific activities directing particular IN fates. Using chicken and mouse models, we show that the identity, not concentration, of the BMP ligand directs distinct dorsal identities. Individual BMPs promote progenitor patterning or neuronal differentiation by their activation of different type I BMP receptors and distinct modulations of the cell cycle. Together, this study shows that a ‘mix and match’ code of BMP signaling results in distinct classes of sensory INs.
The G protein-coupled receptor Smoothened (Smo) is the signal transducer of the Sonic Hedgehog (Shh) pathway. Smo signals through G protein-dependent and -independent routes, with G protein-independent canonical signaling to Gli effectors requiring Smo accumulation in the primary cilium. The mechanisms controlling Smo activation and trafficking are not yet clear but likely entail small-molecule binding to pockets in its extracellular cysteine-rich domain (CRD) and/or transmembrane bundle. Here, we demonstrate that the cytosolic phospholipase cPLA2α is activated through Gβγ downstream of Smo to release arachidonic acid. Arachidonic acid binds Smo and synergizes with CRD-binding agonists, promoting Smo ciliary trafficking and high-level signaling. Chemical or genetic cPLA2α inhibition dampens Smo signaling to Gli, revealing an unexpected contribution of G protein-dependent signaling to canonical pathway activity. Arachidonic acid displaces the Smo transmembrane domain inhibitor cyclopamine to rescue CRD agonist-induced signaling, suggesting that arachidonic acid may target the transmembrane bundle to allosterically enhance signaling by CRD agonist-bound Smo.
Sonic hedgehog (Shh) determines cerebellar granule cell (GC) progenitor proliferation and medulloblastoma pathogenesis. However, the pathways regulating GC progenitors during embryogenesis before Shh production by Purkinje neurons and their roles in tumorigenesis remain unclear. The cilium-localized G-protein-coupled receptor Gpr161 suppresses Shh-mediated signaling in the neural tube. Here, by deleting Gpr161 in mouse neural stem cells or GC progenitors, we establish Gpr161 as a tumor suppressor in Shh subtype medulloblastoma. Irrespective of Shh production in the cerebellum, Gpr161 deletion increased downstream activity of the Shh pathway by restricting Gli3-mediated repression, causing more extensive generation and proliferation of GC progenitors. Moreover, earlier deletion of Gpr161 during embryogenesis increased tumor incidence and severity. GC progenitor overproduction during embryogenesis from Gpr161 deletion was cilium dependent, unlike normal development. Low GPR161 expression correlated with poor survival of SHH subtype medulloblastoma patients. Gpr161 restricts GC progenitor production by preventing premature and Shh-dependent pathway activity, highlighting the importance of basal pathway suppression in tumorigenesis.