Fibroblast growth factor receptor signaling promotes radial glial identity and interacts with Notch1 signaling in telencephalic progenitors

The Notch and fibroblast growth factor (FGF) pathways both regulate cell fate specification during mammalian neural development. We have shown previously that Notch1 activation in the murine forebrain promotes radial glial identity. This result, together with recent evidence that radial glia can be progenitors, suggested that Notch1 signaling might promote progenitor and radial glial character simultaneously. Consistent with this idea, we found that in addition to promoting radial glial character in vivo, activated Notch 1 (ActN1) increased the frequency of embryonic day 14.5 (E14.5) ganglionic eminence (GE) progenitors that grew into neurospheres in FGF2. Constitutive activation of C-promoter binding factor (CBF1), a Notch pathway effector, also increased neurosphere frequency in FGF2, suggesting that the effect of Notch1 on FGF responsiveness is mediated by CBF1. The observation that ActN1 promoted FGF responsiveness in telencephalic progenitors prompted us to examine the effect of FGF pathway activation in vivo. We focused on FGFR2 because it is expressed in radial glia in the GEs where ActN1 increases FGF2 neurosphere frequency, but not in the septum where it does not. Like ActN1, activated FGFR2 (ActFGFR2) promoted radial glial character in vivo. However, unlike ActN1, ActFGFR2 did not enhance neurosphere frequency at E14.5. Additional analysis demonstrated that, unexpectedly, neither ActFGFR2 nor ActFGFR1 could replace the need for ligand in promoting neurosphere proliferation. This study suggests that telencephalic progenitors with radial glial morphology are maintained by interactions between the Notch and FGF pathways, and that the mechanisms by which FGF signaling promotes radial glial character in vivo and progenitor proliferation in vitro can be uncoupled

pH-responsive high-density lipoprotein-like nanoparticles to release paclitaxel at acidic pH in cancer chemotherapy

Jae-Yoon Shin,1,* Yoosoo Yang,1,* Paul Heo,1 Ji-Chun Lee,1 ByoungJae Kong,1 Jae Youl Cho,1 Keejung Yoon,1 Cheol-Su Shin,2 Jin-Ho Seo,3 Sung-Gun Kim,4 Dae-Hyuk Kweon11Department of Genetic Engineering, College of Biotechnology and Bioengineering, and Center for Human Interface Nano Technology, Sungkyunkwan University, 2APTech Research Center, Suwon, 3Department of Agricultural Biotechnology, Seoul National University, Seoul, 4Department of Biomedical Science, Youngdong University, Chungbuk, South Korea*These authors contributed equally to this workBackground: Nanoparticles undergoing physicochemical changes to release enclosed drugs at acidic pH conditions are promising vehicles for antitumor drug delivery. Among the various drug carriers, high-density lipoprotein (HDL)-like nanoparticles have been shown to be beneficial for cancer chemotherapy, but have not yet been designed to be pH-responsive.Methods and results: In this study, we developed a pH-responsive HDL-like nanoparticle that selectively releases paclitaxel, a model antitumor drug, at acidic pH. While the well known HDL-like nanoparticle containing phospholipids, phosphatidylcholine, and apolipoprotein A-I, as well as paclitaxel (PTX-PL-NP) was structurally robust at a wide range of pH values (3.8–10.0), the paclitaxel nanoparticle that only contained paclitaxel and apoA-I selectively released paclitaxel into the medium at low pH. The paclitaxel nanoparticle was stable at physiological and basic pH values, and over a wide range of temperatures, which is a required feature for efficient cancer chemotherapy. The homogeneous assembly enabled high paclitaxel loading per nanoparticle, which was 62.2% (w/w). The molar ratio of apolipoprotein A-I and paclitaxel was 1:55, suggesting that a single nanoparticle contained approximately 110 paclitaxel particles in a spherical structure with a 9.2 nm diameter. Among the several reconstitution methods applied, simple dilution following sonication enhanced the reconstitution yield of soluble paclitaxel nanoparticles, which was 0.66. As a result of the pH responsiveness, the anticancer effect of paclitaxel nanoparticles was much more potent than free paclitaxel or PTX-PL-NP.Conclusion: The anticancer efficacy of both paclitaxel nanoparticles and PTX-PL-NP was dependent on the expression of scavenger receptor class B type I, while the killing efficacy of free paclitaxel was independent of this receptor. We speculate that the pH responsiveness of paclitaxel nanoparticles enabled efficient endosomal escape of paclitaxel before lysosomal break down. This is the first report on pH-responsive nanoparticles that do not contain any synthetic polymer.Keywords: pH responsiveness, nanoparticle, apolipoprotein A-I, paclitaxe

Mind bomb 1 in the lymphopoietic niches is essential for T and marginal zone B cell development

Notch signaling regulates lineage decisions at multiple stages of lymphocyte development, and Notch activation requires the endocytosis of Notch ligands in the signal-sending cells. Four E3 ubiquitin ligases, Mind bomb (Mib) 1, Mib2, Neuralized (Neur) 1, and Neur2, regulate the Notch ligands to activate Notch signaling, but their roles in lymphocyte development have not been defined. We show that Mib1 regulates T and marginal zone B (MZB) cell development in the lymphopoietic niches. Inactivation of the Mib1 gene, but not the other E3 ligases, Mib2, Neur1, and Neur2, abrogated T and MZB cell development. Reciprocal bone marrow (BM) transplantation experiments revealed that Mib1 in the thymic and splenic niches is essential for T and MZB cell development. Interestingly, when BM cells from transgenic Notch reporter mice were transplanted into Mib1-null mice, the Notch signaling was abolished in the double-negative thymocytes. In addition, the endocytosis of Dll1 was impaired in the Mib1-null microenvironment. Moreover, the block in T cell development and the failure of Dll1 endocytosis were also observed in coculture system by Mib1 knockdown. Our study reveals that Mib1 is the essential E3 ligase in T and MZB cell development, through the regulation of Notch ligands in the thymic and splenic microenvironments

Src Is a Prime Target Inhibited by Celtis choseniana

Celtis choseniana is the traditional plant used at Korea as a herbal medicine to ameliorate inflammatory responses. Although Celtis choseniana has been traditionally used as a herbal medicine at Korea, no systemic research has been conducted on its anti-inflammatory activity. Therefore, the present study explored an anti-inflammatory effect and its underlying molecular mechanism using Celtis choseniana methanol extract (Cc-ME) in macrophage-mediated inflammatory responses. In vitro anti-inflammatory activity of Cc-ME was evaluated using RAW264.7 cells and peritoneal macrophages stimulated by lipopolysaccharide (LPS), pam3CSK4 (Pam3), or poly(I:C). In vivo anti-inflammatory activity of Cc-ME was investigated using acute inflammatory disease mouse models, such as LPS-induced peritonitis and HCl/EtOH-induced gastritis. The molecular mechanism of Cc-ME-mediated anti-inflammatory activity was examined by Western blot analysis and immunoprecipitation using whole cell and nuclear fraction prepared from the LPS-stimulated RAW264.7 cells and HEK293 cells. Cc-ME inhibited NO production and mRNA expression of inducible nitric oxide synthase (iNOS), cyclooxygenase (COX-2), and tumor necrosis factor-alpha (TNF-α) in the RAW264.7 cells and peritoneal macrophages induced by LPS, pam3, or poly(I:C) without cytotoxicity. High-performance liquid chromatography (HPLC) analysis showed that Cc-ME contained anti-inflammatory flavonoids quercetin, luteolin, and kaempferol. Among those, the content of luteolin, which showed an inhibitory effect on NO production, was highest. Cc-ME suppressed the NF-κB signaling pathway by targeting Src and interrupting molecular interactions between Src and p85, its downstream kinase. Moreover, Cc-ME ameliorated the morphological finding of peritonitis and gastritis in the mouse disease models. Therefore, these results suggest that Cc-ME exerted in vitro and in vivo anti-inflammatory activity in LPS-stimulated macrophages and mouse models of acute inflammatory diseases. This anti-inflammatory activity of Cc-ME was dominantly mediated by targeting Src in NF-κB signaling pathway during macrophage-mediated inflammatory responses

AP-1-Targeting Anti-Inflammatory Activity of the Methanolic Extract of Persicaria chinensis

In traditional Chinese medicine, Persicaria chinensis L. has been prescribed to cure numerous inflammatory disorders. We previously analyzed the bioactivity of the methanol extract of this plant (Pc-ME) against LPS-induced NO and PGE2 in RAW264.7 macrophages and found that it prevented HCl/EtOH-induced gastric ulcers in mice. The purpose of the current study was to explore the molecular mechanism by which Pc-ME inhibits activator protein- (AP-) 1 activation pathway and mediates its hepatoprotective activity. To investigate the putative therapeutic properties of Pc-ME against AP-1-mediated inflammation and hepatotoxicity, lipopolysaccharide- (LPS-) stimulated RAW264.7 and U937 cells, a monocyte-like human cell line, and an LPS/D-galactosamine- (D-GalN-) induced acute hepatitis mouse model were employed. The expression of LPS-induced proinflammatory cytokines including interleukin- (IL-) 1β, IL-6, and tumor necrosis factor-α (TNF-α) was significantly diminished by Pc-ME. Moreover, Pc-ME reduced AP-1 activation and mitogen-activated protein kinase (MAPK) phosphorylation in both LPS-stimulated RAW264.7 cells and differentiated U937 cells. Additionally, we highlighted the hepatoprotective and curative effects of Pc-ME pretreated orally in a mouse model of LPS/D-GalN-intoxicated acute liver injury by demonstrating the significant reduction in elevated serum AST and ALT levels and histological damage. Therefore, these results strongly suggest that Pc-ME could function as an antihepatitis remedy suppressing MAPK/AP-1-mediated inflammatory events

Lunatic Fringe Deficiency Cooperates with the Met/Caveolin Gene Amplicon to Induce Basal-like Breast Cancer

Basal-like breast cancers (BLBC) express a luminal progenitor gene signature. Notch receptor signaling promotes luminal cell fate specification in the mammary gland, while suppressing stem cell self-renewal. Here we show that deletion of Lfng, a sugar transferase that prevents Notch activation by Jagged ligands, enhances stem/progenitor cell proliferation. Mammary-specific deletion of Lfng induces basal-like and claudin-low tumors with accumulation of Notch intracellular domain fragments, increased expression of proliferation-associated Notch targets, amplification of the Met/Caveolin locus, and elevated Met and Igf-1R signaling. Human BL breast tumors, commonly associated with JAGGED expression, elevated MET signaling, and CAVEOLIN accumulation, express low levels of LFNG. Thus, reduced LFNG expression facilitates JAG/NOTCH luminal progenitor signaling and cooperates with MET/CAVEOLIN basal-type signaling to promote BLBC

Lunatic Fringe Deficiency Cooperates with the Met/Caveolin Gene Amplicon to Induce Basal-Like Breast Cancer

Basal-like breast cancers (BLBC) express a luminal progenitor gene signature. Notch receptor signaling promotes luminal cell fate specification in the mammary gland, while suppressing stem cell self-renewal. Here we show that deletion of Lfng, a sugar transferase that prevents Notch activation by Jagged ligands, enhances stem/progenitor cell proliferation. Mammary-specific deletion of Lfng induces basal-like and claudin-low tumors with accumulation of Notch intracellular domain fragments, increased expression of proliferation-associated Notch targets, amplification of the Met/Caveolin locus, and elevated Met and Igf-1R signaling. Human BL breast tumors, commonly associated with JAGGED expression, elevated MET signaling, and CAVEOLIN accumulation, express low levels of LFNG. Thus, reduced LFNG expression facilitates JAG/NOTCH luminal progenitor signaling and cooperates with MET/CAVEOLIN basal-type signaling to promote BLBC

Cigarette smoke extracts and cadmium induce COX-2 expression through γ-secretase-mediated p38 MAPK activation in C6 astroglia cells.

Exposure to cigarette smoke has been implicated in the progression of cerebrovascular and neurological disorders like stroke through inflammation and blood-brain barrier disruption. In this study, we investigated the signaling cascade activated by cigarette smoke extracts (CSE) and cadmium (Cd) resulting in the COX-2 induction in C6 rat astroglia cells. CSE or Cd induced Notch1 cleavage and activated p38 MAPK and CREB signaling pathways in C6 astroglia cells. Knockdown of nicastrin using siRNA or γ-secretase inhibitors, DAPT and L-685,486, reduced Notch1 cleavage and phosphorylation of p38 MAPK and CREB, while phosphorylation of ERK and JNK remained unaffected. Additionally, the blockage of γ-secretase activity did not show any effect on the phosphorylation of AKT, another upstream activator of CREB, indicating that γ-secretase-mediated CREB activation occurs via p38 MAPK. γ-secretase inhibitor also inhibited the CSE and Cd-mediated increase in the expression of COX-2. Furthermore, recombinant overexpression of Notch1 intracellular domain resulted in an increase in the expression of COX-2. Notch signaling induced by CSE and Cd induced apoptosis in C6 cells. Our results demonstrate that CSE exposure activated the p38 MAPK and CREB-mediated induction in COX-2 expression in astrocytes via γ-secretase-mediated Notch1 signaling. Our data provides novel insights into the potential mechanism of pro-inflammatory response activated by exposure to cigarette smoke

Non-cell autonomous promotion of astrogenesis at late embryonic stages by constitutive YAP activation.

Although astrocytes have gained increased recognition as an important regulator in normal brain function and pathology, the mechanisms underlying their genesis are not well understood. In this study, we show that constitutive YAP activation by in utero introduction of a non-degradable form of the YAP gene (YAP 5SA) causes productive GFAP+ cell generation at late embryonic periods, and this activity is nuclear localization- and TEAD transcription factor-dependent. Moreover, we found that the GFAP+ cells were not YAP 5SA-expressing cells themselves but cells in the vicinity in vivo. Conditioned medium prepared from YAP 5SA-expressing cells induced GFAP+ cell production in vitro, suggesting that a soluble factor(s) was mediating the astrogenic activity of YAP 5SA. Indeed, YAP 5SA expression greatly increased CNTF and BMP4 transcription in neural progenitor cells, and a neutralizing antibody against CNTF reduced the astrogenic effects of YAP 5SA-conditioned medium. Furthermore, the YAP 5SA-expressing cells were identified as FN1+ mesenchymal cells which are responsible for the precocious astrogenesis. These results suggest a novel molecular mechanism by which YAP activation can induce astrogenesis in a non-cell autonomous manner

Suppressor of Fused Is Critical for Maintenance of Neuronal Progenitor Identity during Corticogenesis

Proper lineage progression and diversification of neural progenitor cells (NPCs) ensures the generation of projection neuron (PN) subtypes in the mammalian neocortex. Here, we show that Suppressor of Fused (Sufu) controls PN specification by maintaining the identity of NPCs in the embryonic neocortex. Deletion of Sufu in NPCs of the E10.5 mouse neocortex led to improper specification of progenitors and a reduction in intermediate progenitors (IPs) during corticogenesis. We found that Sufu deletion resulted in unstable Gli2 and Gli3 activity, leading to the ectopic activation of Sonic hedgehog (Shh) signaling. The role of Sufu in maintaining progenitor identity is critical at early stages of corticogenesis, since deletion of Sufu at E13.5 did not cause similar abnormalities. Our studies revealed that Sufu critically modulates Shh signaling at early stages of neurogenesis for proper specification and maintenance of cortical NPCs to ensure the appropriate generation of cortical PN lineages