The autophagy machinery interacts with EBV capsids during viral envelope release.

Autophagy serves as a defense mechanism against intracellular pathogens, but several microorganisms exploit it for their own benefit. Accordingly, certain herpesviruses include autophagic membranes into their infectious virus particles. In this study, we analyzed the composition of purified virions of the Epstein-Barr virus (EBV), a common oncogenic γ-herpesvirus. In these, we found several components of the autophagy machinery, including membrane-associated LC3B-II, and numerous viral proteins, such as the capsid assembly proteins BVRF2 and BdRF1. Additionally, we showed that BVRF2 and BdRF1 interact with LC3B-II via their common protein domain. Using an EBV mutant, we identified BVRF2 as essential to assemble mature capsids and produce infectious EBV. However, BdRF1 was sufficient for the release of noninfectious viral envelopes as long as autophagy was not compromised. These data suggest that BVRF2 and BdRF1 are not only important for capsid assembly but together with the LC3B conjugation complex of ATG5-ATG12-ATG15L1 are also critical for EBV envelope release.

Design, synthesis and biological evaluation of cajanonic acid A analogues as potent PPAR γ antagonists.

Peroxisome proliferator-activated receptor γ (PPAR γ) antagonists are a key instrument of insulin sensitizers since they have the ability to sensitize insulin and can avoid adverse reactions caused by receptor agonist. In this paper, two series of 28 novel Cajanonic acid A (CAA) derivatives were designed and synthesized. The biological activity showed that a novel CAA derivative 9f was identified as a potential PPAR γ antagonist by medicinal chemistry efforts. The results in vitro displayed that compound 9f could improve the PPAR γ antagonist activity (96.2 % / 50.2 % decrease in PPAR γ transactivation at 10 µM / 1 µM, respectively). It also could improve the glucose consumption activity of insulin-resistant HepG2/3T3-L1 cell line (33.27 % / 72.61 % increase in glucose consumption). And in 3 T3-L1 adipocytes, it showed anti-adipogenesis activity (7.04 % increase in oil red staining). Further, in vivo study suggested that compound 9f could improve the oral glucose tolerance in db/db mice. Taken together, derivative 9f served as a promising candidate for anti-diabetic drug discovery and deserve further study.

Cobalt-promoted synthesis of sulfurated oxindoles via radical annulation of N-arylacrylamides with disulfides.

An efficient radical annulation of N-arylacrylamides with disulfides is developed for the synthesis of sulfurated oxindoles. The reaction occurs in a facile manner using CoBr2 as both an initiator and a promoter for the first time and (NH4)2S2O8 as the oxidant. By controlling the CoBr2/(NH4)2S2O8 ratio, a wide range of sulfurated and brominated/sulfurated oxindoles are selectively prepared in good to excellent yields. The present protocol is simple and highly atom economical, and can tolerate a broad range of substrates.

Iron-Catalyzed Radical Annulation of Unsaturated Carboxylic Acids with Disulfides for the Synthesis of γ-Lactones.

An efficient aerobic iron-catalyzed annulation of unsaturated carboxylic acids with disulfides has been developed. This procedure proceeds using FeCl3 as the catalyst and KI as an iodine source under an air atmosphere, which provides practical access to a wide range of substituted γ-lactone derivatives. The disclosed method is quite simple, highly atom-economic, environmentally friendly, and tolerates a broad substrate scope.

Synthesis and biological evaluation of cajanonic acid A derivatives as potential PPARγ antagonists.

Four series of cajanonic acid A (CAA) derivatives have been designed and synthesized. The newly prepared compounds have been screened for glucose consumption activity in HepG2 cell lines and PPARγ antagonistic activity in HEK293 cell lines. Compound 26g bearing a tetrahydroisoquinolinone scaffold showed the most potent PPARγ antagonistic and hypoglycemic activities. An oral glucose tolerance test (OGTT) was performed and the results further confirmed that 26g was a potent hypoglycemic agent. In addition, the possible binding modes for compound 26g in the PPARγ protein have been investigated in this study.

Hypertonic saline attenuates expression of Notch signaling and proinflammatory mediators in activated microglia in experimentally induced cerebral ischemia and hypoxic BV-2 microglia.

BACKGROUND: Ischemic stroke is a major disease that threatens human health in ageing population. Increasing evidence has shown that neuroinflammatory mediators play crucial roles in the pathophysiology of cerebral ischemia injury. Notch signaling is recognized as the cell fate signaling but recent evidence indicates that it may be involved in the inflammatory response in activated microglia in cerebral ischemia. Previous report in our group demonstrated hypertonic saline (HS) could reduce the release of interleukin-1 beta and tumor necrosis factor-alpha in activated microglia, but the underlying molecular and cellular mechanisms have remained uncertain. This study was aimed to explore whether HS would partake in regulating production of proinflammatory mediators through Notch signaling. RESULTS: HS markedly attenuated the expression of Notch-1, NICD, RBP-JK and Hes-1 in activated microglia both in vivo and in vitro. Remarkably, HS also reduced the expression of iNOS in vivo, while the in vitro levels of inflammatory mediators Phos-NF-κB, iNOS and ROS were reduced by HS as well. CONCLUSION: Our results suggest that HS may suppress of inflammatory mediators following ischemia/hypoxic through the Notch signaling which operates synergistically with NF-κB pathway in activated microglia. Our study has provided the morphological and biochemical evidence that HS can attenuate inflammation reaction and can be neuroprotective in cerebral ischemia, thus supporting the use of hypertonic saline by clinicians in patients with an ischemia stroke.

Hypertonic saline alleviates cerebral edema by inhibiting microglia-derived TNF-α and IL-1ß-induced Na-K-Cl Cotransporter up-regulation.

BACKGROUND: Hypertonic saline (HS) has been successfully used clinically for treatment of various forms of cerebral edema. Up-regulated expression of Na-K-Cl Cotransporter 1 (NKCC1) and inflammatory mediators such as tumor necrosis factor alpha (TNF-α) and interleukin-1 beta (IL-1ß) has been demonstrated to be closely associated with the pathogenesis of cerebral edema resulting from a variety of brain injuries. This study aimed to explore if alleviation of cerebral edema by 10% HS might be effected through down-regulation of inflammatory mediator expression in the microglia, and thus result in decreased NKCC1 expression in astrocytes in the cerebral cortex bordering the ischemic core. METHODS: The Sprague-Dawley (SD) rats that underwent right-sided middle cerebral artery occlusion (MCAO) were used for assessment of NKCC1, TNF-α and IL-1ß expression using Western blotting, double immunofluorescence and real time RT-PCR, and the model also was used for evaluation of brain water content (BWC) and infarct size. SB203580 and SP600125, specific inhibitors of the p38 and JNK signaling pathways, were used to treat primary microglia cultures to determine whether the two signaling pathways were required for the inhibition of HS on microglia expressing and secreting TNF-α and IL-1ß using Western blotting, double immunofluorescence and enzyme-linked immunosorbent assay (ELISA). The effect of TNF-α and IL-1ß on NKCC1 expression in primary astrocyte cultures was determined. In addition, the direct inhibitory effect of HS on NKCC1 expression in primary astrocytes was also investigated by Western blotting, double immunofluorescence and real time RT-PCR. RESULTS: BWC and infarct size decreased significantly after 10% HS treatment. TNF-α and IL-1ß immunoexpression in microglia was noticeably decreased. Concomitantly, NKCC1 expression in astrocytes was down-regulated. TNF-α and IL-1ß released from the primary microglia subjected to hypoxic exposure and treatment with 100 mM HS were decreased. NKCC1 expression in primary astrocytes was concurrently and progressively down-regulated with decreasing concentration of exogenous TNF-α and IL-1ß. Additionally, 100 mM HS directly inhibited NKCC1 up-regulation in astrocytes under hypoxic condition. CONCLUSIONS: The results suggest that 10% HS alleviates cerebral edema through inhibition of the NKCC1 Cotransporter, which is mediated by attenuation of TNF-α and IL-1ß stimulation on NKCC1.

p-Cresol-Enabled Nickel-Catalyzed Intermolecular Redox-Economical Coupling of Allyl Alcohols with Alkynes through oxa-Nickelacycle.

An intermolecular redox-economical coupling reaction of allyl alcohols with alkynes, catalyzed by Ni-Brønsted acid cocatalysis, has been developed. This method allows for the synthesis of a diverse range of γ,δ-unsaturated ketones with yields ranging from 40% to 94%, while maintaining excellent compatibility with various functional groups. The transformation of the resulting product demonstrates the significant practical value of this method. Further mechanistic investigations have revealed that the reaction proceeds through the formation of an oxa-nickelacycle intermediate.

Uncertainty analysis of ecosystem services and implications for environmental management - An experiment in the Heihe River Basin, China.

Integrating the economic values of ecosystem services provided by different land uses into decision analysis is critical to achieving effective environmental management in endorheic basins. However, policymaking often ignores the uncertainty related to the variability of parameters in ecosystem service values. To this end, we identified sensitive parameters in the ecosystem service values under four land uses using the global sensitivity analysis method and quantified the potential monetary outcomes based on the Monte Carlo method. The results indicated that only a few sensitive parameters, such as water yield (Qi) and treatment costs per unit of nitrogen (Cost_N), were the primary sources of uncertainty. Therefore, we suggest that improving the precision of sensitive parameters is essential for reducing uncertainty in the total ecosystem service value. Additionally, the overall monetary outcomes for cropland exhibited negative values and had higher risk and lower benefits than those for forest from the standpoint of ecosystem services. In addition, the nonmarketed service of landscape aesthetic made the monetary outcomes of water bodies higher than those of cropland, yet the value of landscape aesthetic was highly uncertain. Therefore, efforts should be made to improve total monetary outcomes by decreasing the negative values in food provisioning of cropland and the uncertainty in landscape aesthetic for water bodies. The sensitivity analysis and uncertainty analysis provide important guidelines for quantifying and reducing the related uncertainty and provide policy information for environmental management based on a comprehensive consideration of the potential ecosystem service values for various land uses.

Evaluation of the antimicrobial function of Ginkgo biloba exocarp extract against clinical bacteria and its effect on Staphylococcus haemolyticus by disrupting biofilms.

ETHNOPHARMACOLOGICAL RELEVANCE: The fruit of Ginkgo biloba L. (Ginkgo nuts) has been used for a long time as a critical Chinese medicine material to treat cough and asthma, as well as a disinfectant. Similar records were written in the Compendium of Materia Medica (Ben Cao Gang Mu, pinyin in Chinese) and Sheng Nong's herbal classic (Shen Nong Ben Cao Jing, pinyin in Chinese). Recent research has shown that Ginkgo biloba exocarp extract (GBEE) has the functions of unblocking blood vessels and improving brain function, as well as antitumour activity and antibacterial activity. GBEE was shown to inhibit methicillin-resistant Staphylococcus aureus (MRSA) biofilm formation as a traditional Chinese herb in our previous report in this journal. AIM OF THE STUD: yThe antibiotic resistance of clinical bacteria has recently become increasingly serious. Thus, this study aimed to investigate the Ginkgo biloba exocarp extract (GBEE) antibacterial lineage, as well as its effect and mechanism on S. haemolyticus biofilms. This study will provide a new perspective on clinical multidrug resistant (MDR) treatment with ethnopharmacology herbs. METHODS: The microbroth dilution assay was carried out to measure the antibacterial effect of GBEE on 13 types of clinical bacteria. Bacterial growth curves with or without GBEE treatment were drawn at different time points. The potential targets of GBEE against S. haemolyticus were screened by transcriptome sequencing. The effects of GBEE on bacterial biofilm formation and mature biofilm disruption were determined by crystal violet staining and scanning electron microscopy. The metabolic activity of bacteria inside the biofilm was assessed by colony-forming unit (CFU) counting and (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2HY-tetrazolium bromide (MTT) assay. Quantitative polymerase chain reaction (qPCR) was used to measure the gene expression profile of GBEE on S. haemolyticus biofilm-related factors. RESULTS: The results showed that GBEE has bacteriostatic effects on 3 g-positive (G+) and 2 g-negative (G-) bacteria among 13 species of clinical bacteria. The antibacterial effect of GBEE supernatant liquid was stronger than the antibacterial effect of GBEE supernviaould-like liquid. GBEE supernatant liquid inhibited the growth of S. epidermidis, S. haemolyticus, and E. faecium at shallow concentrations with minimum inhibitory concentrations (MICs) of 2 µg/ml, 4 µg/ml and 8 µg/ml, respectively. Genes involved in quorum sensing, two-component systems, folate biosynthesis, and ATP-binding cassette (ABC) transporters were differentially expressed in GBEE-treated groups compared with controls. Crystal violet, scanning electron microscopy (SEM) and MTT assays showed that GBEE suppressed S. haemolyticus biofilm formation in a dose-dependent manner. Moreover, GBEE supernatant liquid downregulated cidA, cidB and atl, which are involved in cell lysis and extracellular DNA (eDNA) release, as well as downregulated the cbp, ebp and fbp participation in encoding cell-surface binding proteins. CONCLUSIONS: GBEE has an excellent antibacterial effect on gram-positive bacteria and also inhibits the growth of gram-negative bacteria, such as A. baumannii (carbapenem-resistant Acinetobacter baumannii) CRABA and S. maltophilia. GBEE inhibits the biofilm formation of S. haemolyticus by altering the regulation and biofilm material-related genes, including the release of eDNA and cell-surface binding proteins.

[Effect of hydroxyethyl starch on intracranial pressure and plasma colloid osmotic pressure in rats with cerebral ischemia/reperfusion injury].

OBJECTIVE: To investigate the regulatory effect of hydroxyethyl starch on colloidal osmotic pressure (COP), and its effect on intracranial pressure (ICP) in rats with cerebral ischemia/reperfusion (I/R) injury. METHODS: Twenty four male Sprague Dawley (SD) rats were randomly divided into sham operation group, model group and the hydroxyethyl starch group, each n =8. Cerebral I/R model was reproduced by middle cerebral artery occlusion (MCAO), followed by reperfusion after ischemia for 2 hours. Rats in hydroxyethyl starch group received hydroxyethyl starch 130/0.4206 ml × kg(-1)× d(-1) ia tail vein at the beginning of reperfusion. ICP and COP were evaluated at 0, 2, 6, 12, 18, 24 hours after the surgery. The rats were sacrificed by decapitation. The water content of the right hemisphere was measured at 24 hours after the surgery, and the ratio of apoptosis of neurons was observed by immunohistochemical method. RESULTS: Two hours after surgery the ICP (mm Hg, 1 mm Hg=0.133 kPa) of model group and hydroxyethyl starch group was significantly increased compared with sham operation group (11.50 ± 1.43, 12.48 ± 0.75 vs. 7.95 ± 0.92, both P <0.05). With prolongation of time, the ICP gradually increased and reached the peak at 24 hours (22.76 ± 0.72, 23.32 ± 0.98 vs. 8.15 ± 1.09, both P <0.05). But there was no significant difference in ICP in the hydroxyethyl starch group compared with that of the model group at all time points. The COP (mm Hg) of hydroxyethyl starch group was significantly higher than the model group and sham operation group at each time point, and peaked at 6 hours after surgery (13.49 ± 0.50 vs. 12.04 ± 0.47, 12.00 ± 0.39, both P <0.01). There was no significant difference in COP between the model group and the sham operation group at all time points. The brain water content, neuronal apoptosis of hydroxyethyl starch group and model group was significantly higher than sham operation group [brain water content: (80.16 ± 0.44)%, (80.59 ± 0.67)% vs. (78.72 ± 0.52)%; neuronal apoptosis:(44.27 ± 7.86)%,(42.82 ± 7.82)%vs. (3.26 ± 0.00)%, P <0.05 or P <0.01], but there was no significant difference between the hydroxyethyl starch group and model group (both P >0.05). CONCLUSION: Intravenous injection of hydroxyethyl starch 130/0.4 can increase the plasma COP, but it can not significantly reduce ICP and brain water content, and it also can not improve the neuronal apoptosis.

Discovery of tetrahydrocarbazoles with potent hypoglycemic and hypolipemic activities.

A series of tetrahydrocarbazole derivatives was designed and synthesized on the basis of the AMP-activated protein kinase activator GY3. All the synthesized compounds were screened in HepG2 cell lines for glucose consumption activity and several of them showed potent glucose decreasing activity. In vivo evaluation of the hypoglycemic and hypolipemic effects indicated that 7a exhibited comparable activity with pioglitazone, but with a weaker body-weight increasing effect. The pharmacokinetic profiles of 7a were also investigated.

Huperzine A protects sepsis associated encephalopathy by promoting the deficient cholinergic nervous function.

Neuroinflammatory deregulation in the brain plays a crucial role in the pathogenesis of sepsis associated encephalopathy (SAE). Given the mounting evidence of anti-inflammatory and neuroprotective effects of the cholinergic nervous system, it is surprising that there is little information about its changes in the brain during sepsis. To elucidate the role of the cholinergic nervous system in SAE, hippocampal choline acetyltransferase, muscarinic acetylcholine receptor-1, acetylcholinesterase and acetylcholine were evaluated in LPS-induced sepsis rats. Expression of pro-inflammatory cytokines, neuronal apoptosis, and animal cognitive performance were also assessed. Furthermore, therapeutic effects of the acetylcholinesterase inhibitor Huperzine A (HupA) on the hippocampal cholinergic nervous function and neuroinflammation were evaluated. A deficiency of the cholinergic nervous function was revealed in SAE, accompanied with over-expressed pro-inflammatory cytokines, increase in neuronal apoptosis and brain cognitive impairment. HupA remarkably promoted the deficient cholinergic nervous function and attenuated the abnormal neuroinflammation in SAE, paralleled with the recovery of brain function. We suggest that the deficiency of the cholinergic nervous function and the abnormal neuroinflammation are synergistically implicated in the pathogenesis of SAE. Thus, HupA is a potential therapeutic candidate for SAE, as it improves the deficient cholinergic nervous function and exerts anti-inflammatory action.

USP18 recruits USP20 to promote innate antiviral response through deubiquitinating STING/MITA.

STING (also known as MITA) mediates the innate antiviral signaling and ubiquitination of STING is key to its function. However, the deubiquitination process of STING is unclear. Here we report that USP18 recruits USP20 to deconjugate K48-linked ubiquitination chains from STING and promotes the stability of STING and the expression of type I IFNs and proinflammatory cytokines after DNA virus infection. USP18 deficiency or knockdown of USP20 resulted in enhanced K48-linked ubiquitination and accelerated degradation of STING, and impaired activation of IRF3 and NF-κB as well as induction of downstream genes after infection with DNA virus HSV-1 or transfection of various DNA ligands. In addition, Usp18-/- mice were more susceptible to HSV-1 infection compared with the wild-type littermates. USP18 did not deubiquitinate STING in vitro but facilitated USP20 to catalyze deubiquitination of STING in a manner independent of the enzymatic activity of USP18. In addition, reconstitution of STING into Usp18-/- MEFs restored HSV-1-induced expression of downstream genes and cellular antiviral responses. Our findings thus uncover previously uncharacterized roles of USP18 and USP20 in mediating virus-triggered signaling and contribute to the understanding of the complicated regulatory system of the innate antiviral responses.

Seasonal fluctuations and temperature dependence in photosynthetic parameters and stomatal conductance at the leaf scale of Populus euphratica Oliv.

A combined model to simulate CO2 and H2O gas exchange at the leaf scale was parameterized using data obtained from in situ leaf-scale observations of diurnal and seasonal changes in CO2 and H2O gas exchange. The Farquhar et al.-type model of photosynthesis was parameterized by using the Bayesian approach and the Ball et al.-type stomatal conductance model was optimized using the linear least-squares procedure. The results show that the seasonal physiological changes in photosynthetic parameters (e.g., V(cmax25), J(max25), R(d25) and g(m25)) in the biochemical model of photosynthesis and m in the stomatal conductance model should be counted in estimating long-term CO2 and H2O gas exchange. Overall, the coupled model successfully reproduced the observed response in net assimilation and transpiration rates.

[A study on the health status and related influencing factors among 3-year-old children in rural area of Anhui province].

OBJECTIVE: To explore the health status and related influencing factors for 3-year-old children in rural area of Anhui province. METHODS: Clustered random sampling method was used to investigate the general situation of the children and their families as well as to measure height, weight and some other indexes of the children. Health status and its influencing factors were then evaluated. RESULTS: A total number of 418 children were investigated and incidence rates of growth retardation, underweight, overweight, emaciation and obesity appeared to be 4.3%, 1.9%, 0.7%, 0.5% and 14.9% respectively. Z value of development index was high among children with the following situations: heavy birth weight, father was tall, being boys, and having habits as washing hands before meals. However, Z value was low among kids with following characteristics: being the youngest among all the siblings, family income was high, mothers were illiterate as well as taking tocolytic agent and exposure to X-rays during pregnancy, delivered at home, major care taker was mother, mixed feeding in the early 4-month-old, age of supplementary food was less than 3 months, being picky on foods and taking snacks frequently etc. CONCLUSION: Health status of the 3-year-old children in rural area of Anhui province was fine in general, however, more attentionn needs to be paid to pregnant women and child health care takers in the rural areas.