CERKL regulates autophagy via the NAD-dependent deacetylase SIRT1

<p>Macroautophagy/autophagy is an important intracellular mechanism for the maintenance of cellular homeostasis. Here we show that the <i>CERKL</i> (ceramide kinase like) gene, a retinal degeneration (RD) pathogenic gene, plays a critical role in regulating autophagy by stabilizing SIRT1. <i>In vitro</i> and <i>in vivo</i>, suppressing CERKL results in impaired autophagy. SIRT1 is one of the main regulators of acetylation/deacetylation in autophagy. In CERKL-depleted retinas and cells, SIRT1 is downregulated. ATG5 and ATG7, 2 essential components of autophagy, show a higher degree of acetylation in CERKL-depleted cells. Overexpression of SIRT1 rescues autophagy in CERKL-depleted cells, whereas CERKL loses its function of regulating autophagy in SIRT1-depleted cells, and overexpression of CERKL upregulates SIRT1. Finally, we show that CERKL directly interacts with SIRT1, and may regulate its phosphorylation at Ser27 to stabilize SIRT1. These results show that CERKL is an important regulator of autophagy and it plays this role by stabilizing the deacetylase SIRT1.</p

Proteomics and network pharmacology of Ganshu Nuodan capsules in the prevention of alcoholic liver disease

IntroductionGanshu Nuodan is a liver-protecting dietary supplement composed of Ganoderma lucidum (G. lucidum) spore powder, Pueraria montana (Lour.) Merr. (P. montana), Salvia miltiorrhiza Bunge (S. miltiorrhiza) and Astragalus membranaceus (Fisch.) Bunge. (A. membranaceus). However, its pharmacodynamic material basis and mechanism of action remain unknown.MethodsA mouse model of acute alcohol liver disease (ALD) induced by intragastric administration of 50% alcohol was used to evaluate the hepatoprotective effect of Ganshu Nuodan. The chemical constituents of Ganshu Nuodan were comprehensively identified by UPLC-QTOF/MS, and then its pharmacodynamic material basis and potential mechanism of action were explored by proteomics and network pharmacology.ResultsGanshu Nuodan could ameliorate acute ALD, which is mainly manifested in the significant reduction of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in serum and malondialdehyde (MDA) content in liver and the remarkably increase of glutathione (GSH) content and superoxide dismutase (SOD) activity in liver. Totally 76 chemical constituents were identified from Ganshu Nuodan by UPLC-QTOF/MS, including 21 quinones, 18 flavonoids, 11 organic acids, 7 terpenoids, 5 ketones, 4 sterols, 3 coumarins and 7 others. Three key signaling pathways were identified via proteomics studies, namely Arachidonic acid metabolism, Retinol metabolism, and HIF-1 signaling pathway respectively. Combined with network pharmacology and molecular docking, six key targets were subsequently obtained, including Ephx2, Lta4h, Map2k1, Stat3, Mtor and Dgat1. Finally, these six key targets and their related components were verified by molecular docking, which could explain the material basis of the hepatoprotective effect of Ganshu Nuodan.ConclusionGanshu Nuodan can protect acute alcohol-induced liver injury in mice by inhibiting oxidative stress, lipid accumulation and apoptosis. Our study provides a scientific basis for the hepatoprotective effect of Ganshu Nuodan in acute ALD mice and supports its traditional application

Fungi and cercozoa regulate methane-associated prokaryotes in wetland methane emissions

Wetlands are natural sources of methane (CH4) emissions, providing the largest contribution to the atmospheric CH4 pool. Changes in the ecohydrological environment of coastal salt marshes, especially the surface inundation level, cause instability in the CH4 emission levels of coastal ecosystems. Although soil methane-associated microorganisms play key roles in both CH4 generation and metabolism, how other microorganisms regulate methane emission and their responses to inundation has not been investigated. Here, we studied the responses of prokaryotic, fungal and cercozoan communities following 5 years of inundation treatments in a wetland experimental site, and molecular ecological networks analysis (MENs) was constructed to characterize the interdomain relationship. The result showed that the degree of inundation significantly altered the CH4 emissions, and the abundance of the pmoA gene for methanotrophs shifted more significantly than the mcrA gene for methanogens, and they both showed significant positive correlations to methane flux. Additionally, we found inundation significantly altered the diversity of the prokaryotic and fungal communities, as well as the composition of key species in interactions within prokaryotic, fungal, and cercozoan communities. Mantel tests indicated that the structure of the three communities showed significant correlations to methane emissions (p &lt; 0.05), suggesting that all three microbial communities directly or indirectly contributed to the methane emissions of this ecosystem. Correspondingly, the interdomain networks among microbial communities revealed that methane-associated prokaryotic and cercozoan OTUs were all keystone taxa. Methane-associated OTUs were more likely to interact in pairs and correlated negatively with the fungal and cercozoan communities. In addition, the modules significantly positively correlated with methane flux were affected by environmental stress (i.e., pH) and soil nutrients (i.e., total nitrogen, total phosphorus and organic matter), suggesting that these factors tend to positively regulate methane flux by regulating microbial relationships under inundation. Our findings demonstrated that the inundation altered microbial communities in coastal wetlands, and the fungal and cercozoan communities played vital roles in regulating methane emission through microbial interactions with the methane-associated community

RAGE Mediates Accelerated Diabetic Vein Graft Atherosclerosis Induced by Combined Mechanical Stress and AGEs via Synergistic ERK Activation

Aims/Hypothesis: Diabetes with hypertension rapidly accelerates vascular disease, but the underlying mechanism remains unclear. We evaluated the hypothesis that the receptor of advanced glycation end products (RAGE) might mediate combined signals initiated by diabetes-related AGEs and hypertension-induced mechanical stress as a common molecular sensor. Methods: In vivo surgical vein grafts created by grafting vena cava segments from C57BL/6J mice into the common carotid arteries of streptozotocin (STZ)-treated and untreated isogenic mice for 4 and 8 weeks were analyzed using morphometric and immunohistochemical techniques. In vitro quiescent mouse vascular smooth muscle cells (VSMCs) with either knockdown or overexpression of RAGE were subjected to cyclic stretching with or without AGEs. Extracellular signalregulated kinase (ERK) phosphorylation and Ki-67 expression were investigated. Results: Significant increases in neointimal formation, AGE deposition, Ki-67 expression, and RAGE were observed in the vein grafts of STZ-induced diabetic mice. The highest levels of ERK phosphorylation and Ki-67 expression in VSMCs were induced by simultaneous stretch stress and AGE exposure. The synergistic activation of ERKs and Ki-67 in VSMCs was significantly inhibited by siRNA-RAGE treatment and enhanced by over-expression of RAGE. Conclusion: RAGE may mediate synergistically increased ERK activation and VSMC proliferation induced by mechanica

Extraction optimization, structure features, and bioactivities of two polysaccharides from Corydalis decumbens

Two polysaccharides (CPS1 and CPW2) from Corydalis decumbens were obtained to develop insights into natural medical resources. Optimal extraction conditions of total sugars were researched using the method of response surface methodology, polysaccharides were purified using a combination of ethanol precipitation and anion-exchange chromatography, and structure features were analyzed by scanning electron microscopy, transmission electron microscopy, and Congo-red assay. The bioactivities were estimated in terms of antioxidant and anti-inflammatory effects. Total sugars were extracted with an experimental yield of 32.74% under optimum conditions. CPS1 and CPW2 were purified with yields of 12.01% and 8.23%, respectively. CPS1 was a unique polysaccharide with a molecular weight (Mw) of 360 kDa and consisted of glucose, galactose, mannose, and arabinose in a ratio of 4.9:2.0:1:1.9, and CPW2 was composed of glucose with the Mw of 550 kDa. CPS1 possessed a four-helix conformation, and CPW2 was identified as a linear molecule without branched and entangled chains. The mRNA expressions of TNF-α (71.80%), IL-1β (56.55%), IL-6 (43.98%), and COX-2 (91.88%) in LPS-stimulated RAW 264.7 cells were significantly inhibited by 75 μg/mL CPS1 (P < 0.0001), while CPW2 showed lower inhibitory effects than CPS1. Compared with CPW2, CPS1 showed stronger scavenging abilities for hydroxyl (EC50 = 520.46 μg/mL), ABTS (EC50 = 533.99 μg/mL), and superoxide (EC50 = 1512.06 μg/mL) radicals. CPS1 with four-helix conformation exhibited more outstanding bioactivities than CPW2 without entangled chains

Fig 1 -

Effects of liquid-solid ratio (A), extraction time (B), and extraction temperature (C) on the yields of total sugars from C. decumbens.</p

The maximum absorption wavelengths of Congo red-polysaccharide (CPS1 or CPW2) complex at various concentrations of NaOH.

The maximum absorption wavelengths of Congo red-polysaccharide (CPS1 or CPW2) complex at various concentrations of NaOH.</p

Structural characterization of a novel polysaccharide from Sargassum thunbergii and its antioxidant and anti-inflammation effects.

A novel polysaccharide STSP-I was isolated and purified from Sargassum thunbergii. Its structure and bioactivity were studied using gas chromatography (GC), fourier transform infrared spectroscopy (FTIR), periodate oxidation-smith degradation, partial acid hydrolysis, methylation-GC-MS, nuclear magnetic resonance (NMR), transmission electron microscopy (TEM), radicals scavenging assays and anti-inflammatory assays. STSP-I was consisted of fucose and galactose with a molar ratio of 1.2:1, and its mass was 373 kDa. The main structural components of STSP-I were →4)-α-D-Galp-(1→ and →3)-β-L-Fucp-(1→, STSP-I was a non-branched polysaccharide, and TEM further revealed the existence of entangled chains and linear forms. Compared with Vitamin C (Vc), STSP-I showed a higher scavenging effect of superoxide radical (EC50 = 0.22 mg/mL) and an equivalent scavenging effect of hydroxyl radical (EC50 = 0.88 mg/mL). STSP-I also exhibited good inhibitory effects of TNF-α, IL-6 and COX-2 mRNA expressions in LPS-stimulated RAW 264.7 mouse macrophage cells, and the inhibitory effects were more than 91% at the concentrations of 75 and 150 μg/ml. The results indicate that the polysaccharide STSP-I from S. thunbergii with the linear structure may serve as potential antioxidant and anti-inflammatory agents