Hedgehog (Hh) signaling plays a major role in multiple aspects of embryonic development, which involves both short-and long-range signaling from localized Hh sources. One unusual aspect of Hh signaling is the autoproteolytic processing of Hh followed by lipid modification. As a consequence, the N-terminal fragment of Hh becomes membrane anchored on the cell surface of Hh-producing cells. A key issue in Hh signaling is to understand the molecular mechanisms by which lipid-modified Hh protein is transported from its sites of synthesis and subsequently moves through the morphogenetic field. The dispatched gene, which encodes a putative multipass membrane protein, was initially identified in Drosophila and is required in Hh-producing cells, where it facilitates the transport of cholesterol-modified Hh. We report the identification of the mouse dispatched (Disp) gene and a phenotypic analysis of Disp mutant mice. Disp-null mice phenocopy mice deficient in the smoothened gene, an essential component for Hh reception, suggesting that Disp is essential for Hh signaling. This conclusion was further supported by a detailed molecular analysis of Disp knockout mice, which exhibit defects characteristic of loss of Hh signaling. We also provide evidence that Disp is not required for Hh protein synthesis or processing, but rather for the movement of Hh protein from its sites of synthesis in mice. Taken together, our results reveal a conserved mechanism of Hh protein movement in Hh-producing cells that is essential for proper Hh signaling.

In this study we examined the expression of RyR subtypes and the role of RyRs in neurotransmitter- and hypoxia-induced Ca2+ release and contraction in pulmonary artery smooth muscle cells (PASMCs). Under perforated patch clamp conditions, maximal activation of RyRs with caffeine or inositol triphosphate receptors (IP3Rs) with noradrenaline induced equivalent increases in [Ca2+]i and Ca2+-activated Cl− currents in freshly isolated rat PASMCs. Following maximal IP3-induced Ca2+ release, neither caffeine nor chloro-m-cresol induced a response, whereas prior application of caffeine or chloro-m-cresol blocked IP3-induced Ca2+ release. In cultured human PASMCs, which lack functional expression of RyRs, caffeine failed to affect ATP-induced increases in [Ca2+]i in the presence and absence of extracellular Ca2+. The RyR antagonists ruthenium red, ryanodine, tetracaine, and dantrolene greatly inhibited submaximal noradrenaline– and hypoxia-induced Ca2+ release and contraction in freshly isolated rat PASMCs, but did not affect ATP-induced Ca2+ release in cultured human PASMCs. Real-time quantitative RT-PCR and immunofluorescence staining indicated similar expression of all three RyR subtypes (RyR1, RyR2, and RyR3) in freshly isolated rat PASMCs. In freshly isolated PASMCs from RyR3 knockout (RyR3−/−) mice, hypoxia-induced, but not submaximal noradrenaline–induced, Ca2+ release and contraction were significantly reduced. Ruthenium red and tetracaine can further inhibit hypoxic increase in [Ca2+]i in RyR3−/− mouse PASMCs. Collectively, our data suggest that (a) RyRs play an important role in submaximal noradrenaline– and hypoxia-induced Ca2+ release and contraction; (b) all three subtype RyRs are expressed; and (c) RyR3 gene knockout significantly inhibits hypoxia-, but not submaximal noradrenaline–induced Ca2+ and contractile responses in PASMCs.

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Nineteen soil samples were collected in and around Songshan coking plant in Guangdong province of China and analyzed for eighteen polycyclic aromatic hydrocarbons (PAHs) by gas chromatographymass spectrometry (GC-MS). The total concentration of PAHs ranged from 2.36 to 1146.39 mg kg À1 dry weight, varying significantly among the sampling sites, most individual PAHs were correlated with each other. A cluster analysis was performed to examine the correlation of PAH distribution, five groups were observed with sample types in the coking plant. 2-3 ring PAHs were predominant in group I and II, while 4-5 ring PAHs showed great abundance in group III, IV and V, which contributed to the distance from the emission sources in the coking plant and the behaviors of particle-bound and gaseous PAHs. The ratios of Flu : (Flu + Pyr), BaA : (BaA + Chr), InP : (InP + BgP) and Ant : (Ant + Phen) ratios were 0.51-0.87, 0.16-0.89, 0.47-0.68 and 0.03-0.60, respectively. The total index of all studied soils was > 6, indicating that the source of the PAHs in coking plant soils were from the pyrolysis processes. Health risk assessments were carried out by dermal PAH exposure data to quantify cancer risk. The resultant lifetime exposure levels due to TEQ BaP desorbed onto skin for workers ranged from 2.25 Â 10 À7 to 7.86 Â 10 À5 mg kg À1 per day, and the estimated cancer risks were between 8.45 Â 10 À6 and 2.94 Â 10 À3 , indicating that the dermal exposures of PAHs to coking workers might be acceptable in most soil sites.