Macrophage NFATc3 prevents foam cell formation and atherosclerosis: evidence and mechanisms.

AIMS: Our previous study demonstrated that Ca2+ influx through the Orai1 store-operated Ca2+ channel in macrophages contributes to foam cell formation and atherosclerosis via the calcineurin-ASK1 pathway, not the classical calcineurin-nuclear factor of activated T-cell (NFAT) pathway. Moreover, up-regulation of NFATc3 in macrophages inhibits foam cell formation, suggesting that macrophage NFATc3 is a negative regulator of atherogenesis. Hence, this study investigated the precise role of macrophage NFATc3 in atherogenesis. METHODS AND RESULTS: Macrophage-specific NFATc3 knockout mice were generated to determine the effect of NFATc3 on atherosclerosis in a mouse model of adeno-associated virus-mutant PCSK9-induced atherosclerosis. NFATc3 expression was decreased in macrophages within human and mouse atherosclerotic lesions. Moreover, NFATc3 levels in peripheral blood mononuclear cells from atherosclerotic patients were negatively associated with plaque instability. Furthermore, macrophage-specific ablation of NFATc3 in mice led to the atherosclerotic plaque formation, whereas macrophage-specific NFATc3 transgenic mice exhibited the opposite phenotype. NFATc3 deficiency in macrophages promoted foam cell formation by potentiating SR-A- and CD36-meditated lipid uptake. NFATc3 directly targeted and transcriptionally up-regulated miR-204 levels. Mature miR-204-5p suppressed SR-A expression via canonical regulation. Unexpectedly, miR-204-3p localized in the nucleus and inhibited CD36 transcription. Restoration of miR-204 abolished the proatherogenic phenotype observed in the macrophage-specific NFATc3 knockout mice, and blockade of miR-204 function reversed the beneficial effects of NFATc3 in macrophages. CONCLUSION: Macrophage NFATc3 up-regulates miR-204 to reduce SR-A and CD36 levels, thereby preventing foam cell formation and atherosclerosis, indicating that the NFATc3/miR-204 axis may be a potential therapeutic target against atherosclerosis.

Protective effect of paeoniflorin on H2O2 induced Schwann cells injury based on network pharmacology and experimental validation.

This study was to investigate the protective effect of paeoniflorin (PF) on hydrogen peroxide-induced injury. Firstly, "SMILES" of PF was searched in Pubchem and further was used for reverse molecular docking in Swiss Target Prediction database to obtain potential targets. Injury-related molecules were obtained from GeenCards database, and the predicted targets of PF for injury treatment were selected by Wayne diagram. For mechanism analysis, the protein-protein interactions were constructed by String, and the KEGG analysis was conducted in Webgestalt. Then, cell viability and cytotoxicity assay were established by CCK8 assay. Also, the experimental cells were allocated to control, model (200 µmol·L-1 H2O2), SB203580 10 µmol·L-1 (200 µmol·L-1 H2O2+ SB203580 10 µmol·L-1), PF 50 µmol·L-1 (200 µmol·L-1 H2O2+ PF 50 µmol·L-1), and PF 100 µmol·L-1 (200 µmol·L-1 H2O2+ PF 100 µmol·L-1) groups. We measured the intracellular ROS, Hoechst 33258 staining, cell apoptosis, the levels of Bcl-xl, Bcl-2, Caspase-3, Cleaved-caspase3, Cleaved-caspase7, TRPA1, TRPV1, and the phosphorylation expression of p38MAPK. There are 96 potential targets that may be associated with PF for injury treatment. Then, we chose the "Inflammatory mediator regulation of TRP channels" pathway for the experimental verification from the first 10 KEGG pathway. In experimental verification, H2O2 decreased the cell viability moderately (P < 0.05), and 100 µmol·L -1 PF increased the cell viability significantly (P < 0.05). Depending on the difference of intracellular ROS fluorescence intensity, PF inhibited H 2O2-induced reactive oxygen species production in Schwann cells. In Hoechst 33258 staining, PF reversed the condensed chromatin and apoptotic nuclei following H2O2 treatment. Moreover, Flow cytometry results showed that PF could substantially inhibit H2O2 induced apoptosis (P < 0.05). Pretreatment with PF obviously reduced the levels of Caspase3, Cleaved-caspase3, Cleaved-caspase7, TRPA1, TRPV1, and the phosphorylation expression of p38MAPK after H 2O2 treatment (P < 0.05), increased the levels of Bcl-2, and Bcl-xl ( P < 0.05). PF inhibited Schwann cell injury and apoptosis induced by hydrogen peroxide, which mechanism was linked to the inhibition of phosphorylation of p38MAPK.

[Differentiation of ectomycorrhizal morphology in Abies faxoniana along an elevation gradient in a subalpine forest of western Sichuan Province, China]. / å·è¥¿äºšé«˜å±±å²·æ±Ÿå†·æ‰å¤–ç”ŸèŒæ ¹å½¢æ€éšæµ·æ‹”æ¢¯åº¦çš„åˆ†åŒ–.

Ectomycorrhizal fungi are an important group of symbiotic fungi beneficial to plant growth and their environmental adaptation. An explicit clarification of the trait of ectomycorrhizal fungi would facilitate our understanding of plant responses to environmental change. We set up sampling plots at five elevations (2850, 3000, 3194, 3413, 3593 m) in the Balong Mountain within the Wolong Nature Reserve of Sichuan Province, and collected cubic soil samples (10 cm×10 cm×10 cm) from those plots by point centered quarter method. Based on examination of the morphological types and diversity of ectomycorrhizal roots of Abies faxoniana in each soil sample, we examined the variations in morphological traits of ectomycorrhizal roots of A. faxoniana along the elevational gra-dient and the effects of soil environmental factors. Results showed that: 1) The major ectomycorrhizae of A. faxoniana in Wolong Nature Reserve was orange or yellow in color, with smooth mantle and no or few extensional hyphae, and cylindric or inflated root tips. This type of ectomycorrhizae occurred with highest rate of colonization (12.4%) in the study area. 2) With the increases of elevation, the morphological diversity of ectomycorrhizae in A. faxoniana declined and the morphological types per cubic soil sample significantly decreased. 3) There were significant differences between the ectomycorrhizae of contact exploration type (CE) and short distance exploration type (SDE) at different elevations, while the colonization rate of CE increased significantly with elevation. 4) Soil factors drove the variations of ectomycorrhizal morphology in A. faxoniana along the elevational gradient. Redundancy analysis (RDA) showed that soil total nitrogen (TN), soil temperature (Ts), soil water content (SWC), pH, soil acid phosphatase (ACP) and soil total phosphorus (TP) had significant effects on ectomycorrhizal morphology in A. faxoniana, among which TN and Ts being the greatest and explaining 5.4% and 4.9% of the total variations. Our results clari-fied the variations in the occurrence of ectomycorrhizal morphology in A. faxoniana along elevational gradient, which provided scientific evidence for further studying the mechanisms underlying the responses to environmental changes in mycorrhizal strategy in coniferous species of subalpine forests.

[Changes of productivity with stand development in broadleaf-Korean pine forest in Changbai Mountain, China]. / 长白山阔叶红松林生产力随林分发育的变化.

Stand age is a key factor affecting carbon stocks and fluxes of forest ecosystem. Quantification of the changes in forest productivity with stand development is critically important for optimizing forest age structure, facilitating maximum utilization of resources, and better realizing the role of forests in regulating the uptake, storage, and emission of CO2. In this study, using space for time substitution approach, we established 12 chronosequence plots in the broadleaf-Korean pine forests of Lushuihe. Using a locally parameterized Biome-BGC model, we simulated the dynamics of net primary productivity (NPP) with stand development and examined the changes with stand development in NPP of broadleaf-Korean pine forests under four developmental scenarios. Results showed that the biomass in broadleaf-Korean pine forests of different age-classes ranked in the order of young stand < mid-age stand < mature stand < over-mature stand, with the average value of (224.35±20.68), (237.23±39.96), (259.16±19.51), and (357.57±84.74) t·hm-2, respectively. Modelled NPP in broadleaf-Korean pine forests of different developmental stages varied in the range of 489.8-588 g C·m-2·a-1, which were consistent with the observed data of MODIS NPP, highlighting the adequacy and accuracy of Biome-BGC model in simulating the carbon flux of broadleaf-Korean pine forests. Simulated NPP displayed a pattern of initial increase and later decrease with stand development, reaching peak in the mid-age stand and being smallest in the over-mature stand. Simulations of NPP in broadleaf-Korean pine forest under four developmental scenarios showed that, for the two scenarios with planted Korean pine forests experiencing either natural development or controlled cutting, NPP was highest in the mature stage; whereas for the two scenarios with initial natural secondary birch forests experiencing either natural development or controlled cutting, NPP was highest in the young stage.

[Characters of ecosystem vulnerability in southwestern China based on vegetation productivity]. / 基于植被生产力的西南地区生态系统脆弱性特征.

Southwestern China is one of the most important areas for global biodiversity conservation. Under the background of global climate change, the vulnerability of this area has showed an increasing trend. According to the IPCC concept of vulnerability, we calculated the spatial distribution of ecosystem's vulnerability in southwestern China based on gross primary productivity (GPP) and then analyzed the spatial variation of different levels of vulnerability. Besides, we analyzed the relationship between environmental factors and ecosystem vulnerability, including precipitation, temperature, altitude, slope and vegetation type. The results showed that ecosystem vulnerability in the southwestern China gradually increased from southeast to northwest, with most area within the region being slight and mild vulnerable area (together occupied 69% of the total). The vulnerability level decreased with the increasing of mean annual precipitation and temperature but increased with increasing elevation and slope. Karst area in southwest and borders between farming and ranging regions in northwest Mountain ecotone of the study area had higher vulnerability, being more easily affected by climate change or other disturbances. The vulnerabilities of needle-leaved forest, grassland, and shrubland were relatively higher than other vegetation types, with the potential to be more easily affected by climate change.

[Spatio-temporal variations and influencing factors of thermal comfort at different elevations.] / æ°”å€™èˆ’é€‚åº¦åœ¨ä¸åŒæµ·æ‹”çš„æ—¶ç©ºå˜åŒ–ç‰¹å¾åŠå ¶å½±å“å› ç´ .

Thermal comfort is one of the ways assessing heat changes in climate change scenarios. Based on the daily observation data of 84 meteorological stations from Guizhou Province, the universal thermal climate index (UTCI) was used to examine the spatio-temporal variations and main influencing factors of thermal comfort at different elevations under the scenarios of climate change. Moreover, the impacts of different climatic factors on UTCI at different elevations were quantitatively analyzed. The results showed that the spatial distribution of annual UTCI in Guizhou Province had a strong consistency with that of annual temperature, which was decreasing with the increased elevation. The number of thermally comfortable days in most parts of the province was between 180 d and 240 d. UTCI increased with the increases of elevation, with the overall trends of UTCI [-0.58-1.38 ℃·(10 a)-1] being higher than the trends of temperature [-0.36-0.45 ℃·(10 a)-1] from 1984 to 2013. In general, the correlation coefficients between UTCI and climatic factors air tempe-rature, wind speed, atmosphere pressure, relative humidity and cloud cover were 0.899, -0.855, 0.818, -0.373, and -0.042, respectively. The correlations between climatic factors and UTCI varied across different elevations. The UTCI was affected lightly by air temperature and heavily by wind speed with increasing elevation.

[Regulatory Role of Nitric Oxide in Development and Hatching of Mouse Blastocysts].

OBJECTIVE: To determine the regulatory role and mechanism of nitric oxide (NO) in the development and hatching of mouse blastocysts. METHODS: The Kunming female mice were superovulated and then mated with mature male mice. On the day 2.5 of their pregnancy, morulae were flushed from their uterine horns with culture media. Morulae were cultured in different concentrations of N-nitro-L arginine methyl ester (L-NAME), sodium nitroprusside (SNP), or the combination of L-NAME and SNP in culture media for 48 hours. The development and hatching of blastocysts were examined on day 4 and day 5 and the total numbers of blastocyst cells and cysteinyl aspartate specific proteinase 3 (caspase 3) were observed under confocal laser scanning microscope. RESULTS: With the increase of the concentration of L-NAME or SNP, the hatching rate of blastocysts and the total number of blastocyst cells were significantly reduced. The addition of 10 nmol/L SNP in culture media with 5 mmol/L L-NAME significantly increased the development of blastocysts and promoted hatching of blastocysts. However, with increase of SNP concentration in culture media with 5 mmol/L L-NAME, the development and hatching rates of blastocysts were significantly decreased. L-NAME had no obvious effect on the expression of active caspase 3 in blastocyst cells. However,when being above 500 nmol/L,SNP significantly increased the expression of caspase 3 in blastocyst cells. CONCLUSIONS: NO plays an important role in development and hatching of mouse blastocysts. Excessively high or low NO can damage the division of blastomeres, resulting in the failure of the blastocyst development and hatching. Also, excessively high NO can lead to the apoptosis of the blastocyst cells.

A carbon nanotube filled polydimethylsiloxane hybrid membrane for enhanced butanol recovery.

The carbon nanotubes (CNTs) filled polydimethylsiloxane (PDMS) hybrid membrane was fabricated to evaluate its potential for butanol recovery from acetone-butanol-ethanol (ABE) fermentation broth. Compared with the homogeneous PDMS membrane, the CNTs filled into the PDMS membrane were beneficial for the improvement of butanol recovery in butanol flux and separation factor. The CNTs acting as sorption-active sites with super hydrophobicity could give an alternative route for mass transport through the inner tubes or along the smooth surface. The maximum total flux and butanol separation factor reached up to 244.3 g/m(2)·h and 32.9, respectively, when the PDMS membrane filled with 10 wt% CNTs was used to separate butanol from the butanol/water solution at 80°C. In addition, the butanol flux and separation factor increased dramatically as temperature increased from 30°C to 80°C in feed solution since the higher temperature produced more free volumes in polymer chains to facilitate butanol permeation. A similar increase was also observed when butanol titer in solution increased from 10 g/L to 25 g/L. Overall, the CNTs/PDMS hybrid membrane with higher butanol flux and selectivity should have good potential for pervaporation separation of butanol from ABE fermentation broth.

Integrated butanol recovery for an advanced biofuel: current state and prospects.

Butanol has recently gained increasing interest due to escalating prices in petroleum fuels and concerns on the energy crisis. However, the butanol production cost with conventional acetone-butanol-ethanol fermentation by Clostridium spp. was higher than that of petrochemical processes due to the low butanol titer, yield, and productivity in bioprocesses. In particular, a low butanol titer usually leads to an extremely high recovery cost. Conventional biobutanol recovery by distillation is an energy-intensive process, which has largely restricted the economic production of biobutanol. This article thus reviews the latest studies on butanol recovery techniques including gas stripping, liquid-liquid extraction, adsorption, and membrane-based techniques, which can be used for in situ recovery of inhibitory products to enhance butanol production. The productivity of the fermentation system is improved efficiently using the in situ recovery technology; however, the recovered butanol titer remains low due to the limitations from each one of these recovery technologies, especially when the feed butanol concentration is lower than 1 % (w/v). Therefore, several innovative multi-stage hybrid processes have been proposed and are discussed in this review. These hybrid processes including two-stage gas stripping and multi-stage pervaporation have high butanol selectivity, considerably higher energy and production efficiency, and should outperform the conventional processes using single separation step or method. The development of these new integrated processes will give a momentum for the sustainable production of industrial biobutanol.

[Spatial distribution of human activities and their influences on landscape patterns in Daqingshan Nature Reserve].

Based on forest inventory data and field survey information, and by using GIS spatial analysis technique and landscape indices, this paper studied the spatial distribution of three categories of human activities (settlement, roads, and other sources of disturbances) and their impacts on landscape patterns in three sub-divided regions, i. e., the west, central and east regions of the Daqingshan Nature Reserve in Inner Mongolia. Results showed that the impacts of human activities were stronger in the east and west regions and weaker in the central region. Among the three subdivided regions, the landscape pattern in the west region was predominantly affected by other sources of disturbances, making the landscape patterns of coniferous forests, broadleaf forests and shrubs tended to be of aggregated distribution; the central region was mainly affected by roads, resulting in reduced landscape patch aggregation of broadleaf forests and shrubs; the east region was mostly affected by settlement, resulting in increased fragmentation of coniferous forests and broadleaf forests and apparent increases in landscape patch aggregation of shrubs and grasslands.

[Effects of forest floor litter and nitrogen addition on soil microbial biomass C and N and microbial activity in a mixed Pinus tabulaeformis and Quercus liaotungensis forest stand in Shanxi Province of China].

From September 2010 to October 2011, a field experiment with randomized block design was conducted in a mixed Pinus tabulaeformis and Quercus liaotungensis forest stand in Lingkong Mountain of Shanxi Province to study the effects of forest floor litter and nitrogen addition on the soil microbial carbon (MBC) and nitrogen (MBN) and microbial activity (MR). The litter treatments included complete litter removal, doubling of leaf litter (L), doubling of woody litter (B), and doubling of mixed leaf and woody litter (LB), and the nitrogen addition rates were 0 (N0), 5 g x m(-2) x yr(-1) (N1), and 10 g x m(-2) x yr(-1) (N2). Except that the treatment of complete litter removal without nitrogen addition decreased the soil organic carbon content significantly, all the other treatments had no significant differences in the effects on soil organic carbon. The soil MBC, MBN, and MR varied in the ranges of 262.42-873.16 mg x kg(-1), 73.55-173.85 mg x kg(-1), and 2.38-3.68 mg x kg(-1) x d(-1), respectively, and the MBC and MBN had significant positive correlations with the MR. Nitrogen addition did not show any effect on the MBC, MBN, and MR, whereas litter treatments affected the MR significantly, with the highest MR in treatment LB, followed by treatments L and B, and the lowest in treatment of complete litter removal. There were no interactive effects between litter and nitrogen addition treatments on any of the variables studied. It was suggested that short-term nitrogen addition and forest floor litter change could have limited effects on soil microbial processes.

[Temporal change in annual air temperature and heat island effect in a coastal city and an inland city at mid-latitude in China during 1956-1998].

Temporal changes in air temperature and urban heat island (UHI) effects during 1956-1998 were compared between a coastal city, Ji' nan, and an inland city, Xi' an, which were similar in latitude, size and development. During 1956-1978, except that the annual mean minimum temperature in Ji' nan increased by 0.37 degrees C x 10 a(-1), the temperature variables in the two cities did not display any apparent trend. During 1979-1998, all temperature variables of the two cities showed an increasing trend. Comparing with that in Ji' nan, the increasing rate of annual mean maximum temperature and annual mean temperature in Xi' an was greater, but that of annual mean minimum temperature was smaller. In the two cities, heat island effect occurred during 1956-1978 but without any apparent trend, whereas during 1979-1998, this effect increased with time, especially in Xi' an where the annual mean minimum temperature and annual mean temperature increased by 0.22 degrees C x 10 a(-1) and 0.32 degrees C x 10 a(-1), respectively. Both the level and the inter-annual variation of the heat island effect were much greater in Ji' nan than in Xi' an, but the increasing rate of this effect was greater in Xi' an than in Ji' nan. Obvious differences were observed in the increasing rate of annual mean maximum air temperature, annual mean air temperature, and annual mean minimum temperature as well as the heat island effect in Ji' nan, whereas negligible differences were found in Xi' an. Among the three temperature variables, annual mean minimum temperature displayed the most obvious increasing trend and was most affected by heat island effect, while annual mean maximum temperature was most variable inter-annually. Geographical location not only affected the magnitude of urban warming, but also affected the mode of urban warming and the strength of heat island effect.

Cloning of Human Glial Neurotrophic Factor and Its High Expression in Insect Cells.

The human glial cell derived neurotrophic factor (GDNF) was expressed in Tn-5B1-4 insect cells using the Bac to Bac baculovirus expression system, accounting for 30% of total cellular soluble proteins. The expressed product was purified by affinity chromatography and the activity assays showed that it could significantly promote the survival of dopaminergic neurons. The study can be the basis for further structural-functional analysis of GDNF.

Helicobacter pylori HspA Heat-shock Protein Gene Cloning, Expression and Immunogenicity.

Helicobacter pylori is the causative agent of gastritis and peptic ulcer in human. The heat-shock protein A (HspA)may stimulate the immunoresponse protecting human body against challenge of H.pylori. The gene encoding the structural A subunit of H.pylori heat-shock protein was amplified from H.pylori chromosomal DNA by PCR, and was inserted in the prokaryotic expression vector pET-22b(+), and then was transformed into the BL-21(DE3)E.coli strain to express the HspA recombinant protein. HspA gene was measured to be 354 base pairs, and the recombinant protein gene encoded polypeptides of 118 amino acid residues, corres-ponding to calculated molecular weight of 15 kD. Western blot analysis of HspA recombinant protein was confirmed that it could be specifically recognized by the serum of H.pylori-infected patients, and could also be re-cognized by the serum of immunized Balb/c mice with HspA itself. This result suggests that HspA may be an effective protein vaccine for prevention and treatment of the infection of H.pylori.

Construction of Biotinylated Annexin V for Detection of Apoptosis.

The biotinylated annexin V was generated by genetic engineering method. The encoding region of annexin V was fused with a gene coding for the carboxyl terminal 87 residues of Escherichia coli biotin carboxyl carrier protein. The fused gene was expressed in E.coli and the annexin V was biotinylated in vivo. The biotinylation efficiency was about 60% as detected by HPLC. The biotinylated annexin V was purified by avidin affinity chromatography and used to detect the apoptosis of the neurons induced by morphine.

Cloning and Expression of Annexin V cDNA in E.coli.

The cDNA encoding the mature annexin V was isolated by using RT-PCR method from total RNAs of fresh human placenta. The result of sequencing indicated that the sequence of isolated annexin V cDNA was the same as the reported nucleotide sequence of annexin V. The annexin V cDNA was cloned into expression plasmid pET24a(+) under T7 promoter and then transformed into E.coli BL21(DE3). SDS-PAGE analysis revealed that the human annexin V was highly expressed and accumulated up to 38% of total bacterial proteins in soluble form after the induction by 1 mmol/L IPTG. The purified human annexin V can significantly prolong activated partial thromboplastin time (APTT).

The Human Recombinant GDNF and Study of Its Biological Activity.

The cDNA encoding the mature human glial cell derived neurotrophic factor(GDNF) was isolated by using RT-PCR method from total RNA extracted of fetal human brain. The expression plasmid pET-GDNF was constructed by inserting GDNF cDNA into plasmid pET-28a(+) containing T7 promoter and transformed into E.coli BL21(DE3). An expression strain BLGDNF was selected. SDS-PAGE analysis revealed that the human GDNF protein was highly expressed and accumulated up to above 30% of the total bacterial proteins in the form of inclusion body after the induction. The antibody to GDNF was prepared by immunization of rat using purified GDNF protein. Purified and refolded GDNF protein could significantly promote the survival of dopaminergic neurons.

Cloning and Expression of the cDNA Encoding Human Tissue Inhibitor of Metalloproteinase-3 and Its Inhibition on Angiogenesis.

We have isolated the cDNA encoding the mature tissue inhibitor of metalloproteinase-3 (TIMP-3) by using RT-PCR method with total RNA extracted from human fresh placenta. The result of sequencing indicated that the mature TIMP-3 polypeptide contained 188 aa residues, including 12 conserved cysteinyl residues in the TIMP family. The expression plasmid pET-TIMP3 was constructed by inserting TIMP-3 cDNA into plasmid pET-24 (a(+)) containing T7 promoter and transformed into E. coli BL21 (DE3). An expression strain BLTIMP3 was selected. SDS-PAGE analysis revealed that the human TIMP-3 protein was highly expressed and accumulated up to above 30% of the total bacterial proteins in the form of inclusion body after induced by 1 mmol/L IPTG for 3--5 h. The partially purified and renatured TIMP-3 can significantly inhibit angiogenesis as shown by CAM assay.