Eupholides A-H, abietane diterpenoids from the roots of Euphorbia fischeriana, and their bioactivities.

The roots of Euphorbia fischeriana known as "Langdu" in traditional Chinese medicine have been used for the treatment of tuberculosis in China. Through a bioactive phytochemical investigation of the roots of E. fischeriana, 15 diterpenoids were obtained by various chromatographic techniques. On the basis of wide spectroscopic data, including NMR, UV, IR, HR-ESI-MS, ECD and X-ray crystallography, all of the isolated compounds were elucidated to be ent-abietane diterpenoid analogs, including undescribed eupholides A-H and seven known diterpenoids. In the bioassay for anti-tuberculosis, eupholides F-H moderately inhibited the proliferation of Mycobacterium tuberculosis H37Ra, with the MIC determined to be 50 µM. Furthermore, eupholides G, ent-11α-hydroxyabieta-8(14), 13(15)-dien-16,12α-olide, and jolkinolide F significantly inhibited the lyase activity of human carboxylesterase 2 (HCE 2), with IC50 values of 7.3, 150, and 34.5 nM, respectively.

Comparison of the gut microbiota of short-term and long-term medical workers and non-medical controls: a cross-sectional analysis.

OBJECTIVES: The hospital environment has been implicated in the enrichment and exchange of pathogens and antibiotic resistance, but its potential in shaping the symbiotic microbial community of hospital staff is unclear. This study was designed to evaluate the alteration of the gut microbiome in medical workers compared to non-medical controls. METHODS: A prospective cross-sectional cohort study was conducted in the intensive care unit (ICU) and other departments of a centre in north-eastern China. Faecal samples of 175 healthy medical workers-short-term (1-3 months) workers (n = 80) and long-term (>1 year) workers (n = 95)-and 80 healthy non-medical controls were analysed using 16S rRNA amplicon sequencing. The hospital environmental samples (n = 9) were also analysed. RESULTS: The gut microbiomes of medical workers exhibited marked deviations in diversity and alteration in microbial composition and function. Short-term workers showed significantly higher abundances of taxa such as Lactobacillus, Butyrivibrio, Clostridiaceae, Clostridium, Ruminococcus, Dialister, Bifidobacterium, Odoribacter, and Desulfovibrio and lower abundances of Bacteroides and Blautia than the controls. Long-term workers showed higher abundances of taxa such as Dialister, Veillonella, Clostridiaceae, Clostridium, Bilophila, Desulfovibrio, Pseudomonas, and Akkermansia and lower abundances of Bacteroides and Coprococcus than the controls. The medical workers' department (ICU versus non-ICU) and position (resident doctor versus nursing staff) also impacted their gut microbiome. Compared with the non-ICU workers, workers in the ICU showed a significant increase in the abundances of Dialister, Enterobacteriaceae, Phascolarctobacterium, Pseudomonas, Veillonella, and Streptococcus and a marked depletion of Faecalibacterium, Blautia, and Coprococcus. In contrast with the nursing staff, the resident doctors showed a significant increase in Erysipelotrichaceae and Clostridium and a decrease in Bacteroides, Blautia, and Ruminococcus in the gut microbiome. Moreover, we found that the microbiota of hospital environments potentially correlated with the workers' gut microbiota. CONCLUSIONS: Our findings demonstrated structural changes in the gut microbial community of medical workers.

Synthesis of a pH-Responsive Functional Covalent Organic Framework via Facile and Rapid One-Step Postsynthetic Modification and Its Application in Highly Efficient N1-Methyladenosine Extraction.

A facile and rapid postsynthetic modification strategy for functionalization of covalent organic framework (COF) was developed to synthesize a tailor-made pH-responsive COF called TpPa-1@Au@GSH for highly efficient extraction of N1-methyladenosine (m1A). Glutathione (GSH) was judiciously designed as the functional group for extracting and releasing m1A by pH variations. With the aid of gold nanoparticles (Au NPs) as linkers, GSH was successfully introduced to the robust substrate TpPa-1 in only one step spending only 1 h. Owing to the several-to-one immobilization of GSH on Au NPs and the large surface area of TpPa-1, this functional COF was constructed with abundant m1A binding sites. TpPa-1@Au@GSH showed excellent selectivity for m1A extraction by capturing m1A from a mixture of 14 nucleoside analogues followed by mass spectrometry analysis. It was proved to have ultrafast adsorption ability (only 1 min incubation time), high binding capacity (5 mg g-1, m1A/TpPa-1@Au@GSH), good reusability (at least 5 times), and good storage stability (at least 8 months at room temperature). Great performance was also achieved in extracting m1A from both animal and plant biological samples. The adsorption mechanism was demonstrated to be based on the electrostatic interaction. This work proposed a new approach for m1A extraction, demonstrated the high potential of COFs in biological sample pretreatment, and offered an effective and versatile route for functionalization of COFs.

NCAM regulates temporal specification of neural progenitor cells via profilin2 during corticogenesis.

The development of cerebral cortex requires spatially and temporally orchestrated proliferation, migration, and differentiation of neural progenitor cells (NPCs). The molecular mechanisms underlying cortical development are, however, not fully understood. The neural cell adhesion molecule (NCAM) has been suggested to play a role in corticogenesis. Here we show that NCAM is dynamically expressed in the developing cortex. NCAM expression in NPCs is highest in the neurogenic period and declines during the gliogenic period. In mice bearing an NPC-specific NCAM deletion, proliferation of NPCs is reduced, and production of cortical neurons is delayed, while formation of cortical glia is advanced. Mechanistically, NCAM enhances actin polymerization in NPCs by interacting with actin-associated protein profilin2. NCAM-dependent regulation of NPCs is blocked by mutations in the profilin2 binding site. Thus, NCAM plays an essential role in NPC proliferation and fate decision during cortical development by regulating profilin2-dependent actin polymerization.

A facilely synthesized glutathione-functionalized silver nanoparticle-grafted covalent organic framework for rapid and highly efficient enrichment of N-linked glycopeptides.

The development of facilely synthetic materials for highly efficient enrichment of N-linked glycopeptides is essential in glycoproteome analysis. In this work, by utilizing the self-assembling of glutathione (GSH) on silver nanoparticles (Ag NPs), and the formation and dispersion of Ag NPs on a robust TpPa-1 substrate, a newly functionalized covalent organic framework (COF) called TpPa-1@Ag@GSH was synthesized via a simple two step post-synthetic modification. TpPa-1@Ag@GSH and intermediate products were confirmed and evaluated by nuclear magnetic resonance spectroscopy, Fourier transform infrared spectroscopy, X-ray diffraction, transmission electron microscopy, scanning electron microscopy-energy dispersive spectroscopy, Brunauer-Emmett-Teller and thermogravimetric analyses. Benefiting from the judicious selection of the substrate, the abundance of binding sites, relatively high affinity between GSH and N-linked glycopeptides, and the multivalent interactions between N-linked glycopeptides and unoccupied surfaces of Ag NPs, this porous material showed great performance in N-linked glycopeptide enrichment. By enriching N-linked glycopeptides in tryptic digests of human serum immunoglobulin G (human IgG) followed by mass spectrometry analysis, our method was proved to have good sensitivity (1 fmol), high selectivity (1 : 1500, human IgG to bovine serum albumin), high binding capacity (160 mg g-1, IgG/TpPa-1@Ag@GSH), ultra-fast capture ability (only 1 min incubation time), and good reusability (at least 5 times). It was also successfully applied to the enrichment of N-linked glycopeptides from complex biological samples. Our work improved the enrichment selectivity of COFs, reached the most rapid capture ability among off-column enrichment materials, and provided a very facile and easily popularized post-synthetic modification route for COFs in glycoproteome analysis.

Highly efficient enrichment of N-linked glycopeptides using a hydrophilic covalent-organic framework.

The enrichment of glycopeptides plays an important role in glycoproteomics. In this paper, a covalent-organic framework called TpPa-1, synthesized by the Schiff base reaction of 1,3,5-triformylphloroglucinol and paraphenylenediamine, was first successfully utilized as a hydrophilic porous material for N-linked glycopeptide enrichment. Using this material, interference from non-glycopeptides could be efficiently eliminated, which facilitated the mass spectrometry detection of glycopeptides. By capturing N-linked glycopeptides from tryptic digests of human IgG, our method was proved to have high sensitivity at the femtomole level. And it showed superior selectivity for glycopeptides even when non-glycopeptides were 1000 times more concentrated. Due to the strong covalent bonds, this material possessed good stability and could be repeatedly used for at least 10 times. The ultra-low mass density and abundant binding sites also provided it with high binding capacity (178 mg g-1, IgG/TpPa-1). Moreover, N-linked glycopeptides were easily enriched by this material from only 10 µL human serum, which demonstrated its potential in pretreatment of complex biological samples.

ent-Abietane and Tigliane Diterpenoids from the Roots of Euphorbia fischeriana and Their Inhibitory Effects against Mycobacterium smegmatis.

An investigation on the bioactive chemical constituents of the roots of Euphorbia fischeriana has been conducted, with 21 diterpenoids obtained using various chromatographic techniques. On the basis of spectroscopic data analysis, the new compounds were elucidated as four ent-abietane-type diterpenoids (1-4) and four tigliane-type diterpenoids (13-16). Also obtained were eight known ent-abietane (5-12) and five known tigliane (17-21) diterpenoids. The potential antituberculosis effects of these diterpenoids were evaluated using a Mycobacterium smegmatis model. The most potent compound according to the in vitro bioassay used was 17-hydroxyjolkinolide B (12) (MIC 1.5 µg/mL).

[Inhibiting effects of three components of Astragalus membranaceus on oxidative stress in Chang Liver cells].

The main objective of this research is to investigate the effects of astragaloside IV, calycosin separately glucoside, formononetin on oxidative stress in Chang Liver cells induced by H2O2. In the experiments, Chang Liver cells (a kind of normal human hepatocytes) were used as the research object, bifendate which has a clear hepatoprotective effect was used as the positive control drug, then the oxidative damage model of Chang Liver cells were established by H2O2. Cells were divided into six groups: blank control group, oxidative stress group, astragaloside IV group, calycosin separately glucoside group, formononetin group and positive control group. Then endogenous antioxidant system related indexes were detected by micro plate and colorimetric method; intracellular reactive oxygen species (ROS) were detected by DCFH-DA fluorescent probe; and the expressions of CYP2E1 were evaluated by liver microsomes, mRNA, and protein, respectively with spectrophotometry, Real-time PCR method, and Western blot technique. Results showed that H2O2 decreased antioxidant activity, and increased ROS level and expression of CYP2E1. The above oxidative stress status had been changed with protections of the three components of Astragalus membranaceus (compared with oxidative stress group, P < 0.05, P < 0.01), which taken as a whole had equivalent effects as the drug of positive control group( bifendate). Taken together, three Astragalus membranaceus ingredients all had significant or extremely significant inhibiting effects on oxidative damaged Chang Liver cells which were induced by H2O2, and the oxidative damage of Chang Liver cells had been relieved.

Perfluorooctane sulfonate blocked autophagy flux and induced lysosome membrane permeabilization in HepG2 cells.

Perfluorooctane sulfonate (PFOS) is an emerging persistent organic pollutant widely distributed in the environment, wildlife and human. In this study, as observed under the transmission electron microscope, PFOS increased autophagosome numbers in HepG2 cells, and it was confirmed by elevated LC3-II levels in Western blot analysis. PFOS increased P62 level and chloroquine failed to further increase the expression of LC3-II after PFOS treatment, indicating that the accumulation of autophagosome was due to impaired degradation rather than increased formation. With acridine orange staining, we found PFOS caused lysosomal membrane permeabilization (LMP). In this study, autophasome formation inhibitor 3-methyladenine protected cells against PFOS toxicity, autophagy stimulator rapamycin further decreased cell viability and increased LDH release, cathepsin inhibitor did not influence cell viability of PFOS-treated HepG2 cells significantly. These further supported the notion that autophagic cell death contributed to PFOS-induced hepatotoxicity. In summary, the data of the present study revealed that PFOS induced LMP and consequent blockage of autophagy flux, leading to an excessive accumulation of the autophagosomes and turning autophagy into a destructive process eventually. This finding will provide clues for effective prevention and treatment of PFOS-induced hepatic disease.

[Protective effect of astragaloside IV on oxidative damages of chang liver cell induced by ethanol and H2O2].

OBJECTIVE: To study the protective effect of astragaloside IV on oxidative damages of Chang Liver cells induced by ethanol and H2O2. METHOD: The alcoholic and nonalcoholic oxidative damage models were established on Chang Liver cells with ethanol and H2O2, respectively. The cells viabilities were detected by MTT assay, transaminase activity and antioxidant ability were detected by micro plate and colorimetric method, reactive oxide species (ROS) was detected by DCFH-DA fluorescent probe and cell cycle was detected by flow cytometry. DNA ladder method was used to detect apoptosis. RESULT: Both kinds of oxidative damage could decrease the viability and antioxidant enzyme activity of Chang Liver cells, and increase the transaminase activity and MDA content of extracellular fluid. The protective effects of astragaloside IV against those two kinds of oxidative damages were significant or extremely significant. Meanwhile, ethanol could decline the level of ROS significantly in the damaged cells, while H2O2 could increase it significantly. And the effect of astragaloside IV was to make ROS return to the normal level. Retardation of cell cycle progression of Chang Liver cells in G0/G1 induced by ethanol or H2O2 was relieved, and apoptosis was also inhibited. CONCLUSION: Astragaloside IV had protective effect on oxidative damages of Chang Liver cells induced by ethanol and H2O2.

Cornmeal-induced resistance to ciprofloxacin and erythromycin in enterococci.

OBJECTIVE: To establish a model ecosystem to study the impact of cornmeal on the appearance and persistence of the erythromycin (ERY)- and ciprofloxacin (CIP)-resistant phenotypes in waterborne enterococci. METHODS: After the model ecosystem was established, the system was divided into six dose groups, with the addition of 8, 4, 1, 0.25, 0.05, and 0 g L(-1) sterilized cornmeal. System mud samples were collected at 0, 1, 3, 7, 14, 30, 40, 61, and 130 d, and enterococci present in the mud samples were evaluated for their sensitivities to CIP and ERY. PCR was employed to detect genes such as gyrA and ermB. The gyrA gene was sequenced, and codons 83 and 87 were analyzed for mutations. RESULTS: (1) The addition of 0.05-8 g L(-1) cornmeal had an impact on CIP resistance. The higher the dose of cornmeal added, the larger the impact it generated. Furthermore, the earlier the emergence of CIP-resistant strains, the greater the incidence of drug resistance. The impact of cornmeal on resistance to ERY was less consistent, and the degree of the impact was not in proportion to the dose of cornmeal added. (2) There were no mutations at codons 83 and 87 in the gyrA genes from 102 strains isolated from the model ecosystem. The incidence of ermB-positive strains of ERY-resistant enterococci (28 strains) was 78.6%, and the incidence of ermB-positive strains of ERY-sensitive enterococci (16 strains) was 0%. CONCLUSIONS: (1) Adding different doses of cornmeal can facilitate resistance to CIP and ERY in waterborne enterococci. In this study, the degree of resistance was related to the cornmeal dose. (2) In the model ecosystem, enterococcal CIP resistance was not caused by a gyrA gene mutation; however, in the vast majority of cases, resistance to ERY was related to the ermB resistance gene.

Stage-dependent effect of leptin on development of porcine embryos derived from parthenogenetic activation and transgenic somatic cell nuclear transfer.

Accumulating evidence suggests that leptin may play important roles in preimplantation embryonic development, although this remain controversial, and little is known about whether leptin has a stage-dependent regulatory effect on development of porcine embryos derived by parthenogenetic activation (PA) and somatic cell nuclear transfer (SCNT). The objective of this study was to investigate the effects of addition of leptin to in vitro culture (IVC) medium on development of porcine embryos derived by PA and SCNT. We found that addition of 50 ng/ml human recombinant leptin improved the rate of PA embryos reaching the blastocyst stage and increased the total cell number of blastocysts compared with the control group. The maximal blastocyst rate of SCNT embryos was achieved at 50 ng/ml, and the total cell number of blasocysts was increased significantly at 500 ng/ml leptin concentration. However, the ratio of the inner cell mass (ICM) to total cell number was not affected in any of the groups. Supplementation of leptin (50 ng/ml) from day 3, approximately the 4-8-cell stage, as in the case of the positive control, significantly increased the blatocyst rate of PA embryos compared with the negative control and inhibited cell apoptosis. There were no beneficial effects on embryonic development when 50 ng/ml leptin was added to the culture medium from day 1 to day 3 or from day 4 to day 6. These results indicate that leptin could improve the development and the quality of PA and SCNT embryos; and 50 ng/ml leptin performs its primary stimulatory effect at 4-8-cell stage and that leptin may have no effect on the maternal-zygote transition (MZT) of porcine PA and SCNT embryos.

Inactivation of the mycobacterial rhamnosyltransferase, which is needed for the formation of the arabinogalactan-peptidoglycan linker, leads to irreversible loss of viability.

Temperature-sensitive mutant 2-20/32 of Mycobacterium smegmatis mc(2)155 was isolated and genetically complemented with a Mycobacterium tuberculosis H37Rv DNA fragment that contained a single open reading frame. This open reading frame is designated Rv3265c in the M. tuberculosis H37Rv genome. Rv3265c shows homology to the Escherichia coli gene wbbL, which encodes a dTDP-Rha:alpha-D-GlcNAc-pyrophosphate polyprenol, alpha-3-L-rhamnosyltransferase. In E. coli this enzyme is involved in O-antigen synthesis, but in mycobacteria it is required for the rhamnosyl-containing linker unit responsible for the attachment of the cell wall polymer mycolyl-arabinogalactan to the peptidoglycan. The M. tuberculosis wbbL homologue, encoded by Rv3265c, was shown to be capable of restoring an E. coli K12 strain containing an insertionally inactivated wbbL to O-antigen positive. Likewise, the E. coli wbbL gene allowed 2-20/32 to grow at higher non-permissive temperatures. The rhamnosyltransferase activity of M. tuberculosis WbbL was demonstrated in 2-20/32 as was the loss of this transferase activity in 2-20/32 at elevated temperatures. The wbbL of the temperature-sensitive mutant contained a single-base change that converted what was a proline in mc(2)155 to a serine residue. Exposure of 2-20/32 to higher non-permissive temperatures resulted in bacteria that could not be recovered at the lower permissive temperatures.