Combination of Transcriptomics and Proteomics Reveals Differentially Expressed Genes and Proteins in the Skin of EDAR Gene-Targeted and Wildtype Cashmere Goats.

Cashmere goats play a pivotal role in the animal hair industry and are economically valuable. Cashmere is produced through the periodic growth of secondary hair follicles. To improve their yield of cashmere, the regulatory mechanisms of cashmere follicle growth and development need to be analysed. Therefore, in this study, EDAR gene-targeted cashmere goats were used as an animal model to observe the phenotypic characteristics of abnormal hair growth and development at the top of the head. Transcriptomic and proteomic techniques were used to screen for differentially expressed genes and proteins. In total, 732 differentially expressed genes were identified, including 395 upregulated and 337 downregulated genes. In addition, 140 differentially expressed proteins were identified, including 69 upregulated and 71 downregulated proteins. These results provide a research target for elucidating the mechanism through which EDAR regulates hair follicle growth in cashmere goats. It also enriches the available data on the regulatory network involved in hair follicle growth.

Revealing the Synergistic Mechanism of Multiple Components in Compound Fengshiding Capsule for Rheumatoid Arthritis Therapeutics by Network Pharmacology.

BACKGROUND: Compound Fengshiding capsule (CFC), is a Chinese formulation from herbal origin including Alangium platanifolium, Angelicae dahurica, Cynanchum paniculatum and Glycyrrhiza uralensis. CFC is widely used as clinical therapy against rheumatoid arthritis. However, its exact mechanism of action has not been explored yet. METHODS: In order to explore the synergistic mechanism of CFC, we designed a study adopting network pharmacology scheme to screen the action targets in relation to the CFC components. The study analyses target facts of salicin, paeonol, liquiritin and imperatorin from PubMed database, and explores the potential pharmacological targets of rheumatoid arthritis, cervical neuralgia and sciatica related diseases for their interaction. RESULTS: The results of boosted metabolic pathway showed that the chemical components of CFC interrupted many immune-related pathways, thus participating in immunity regulation of the body and playing a role in the treatment of rheumatism. Collectively, CFC has apoptotic, oxidative stress modulatory and anti-inflammatory effects that accumulatively serve for its clinical application against rheumatoid arthritis. CONCLUSION: Conclusively, our findings from present study reconnoiters and compacts systematic theoretical approach by utilizing the network pharmacology mechanism of four effective components for the treatment of rheumatism indicating sufficient potential drug targets associated with CFC against rheumatism. These interesting findings entreaties for further in vitro and in vivo studies on the mechanism of compound active ingredient against rheumatism.

Liquiritin from Glycyrrhiza uralensis Attenuating Rheumatoid Arthritis via Reducing Inflammation, Suppressing Angiogenesis, and Inhibiting MAPK Signaling Pathway.

Among the various treatments, induction of synoviocyte apoptosis by natural products during a rheumatoid arthritis (RA) pathological condition can be considered to have vast potential. However, it is unclear that liquiritin, a kind of natural flavonoid extracted from the roots of Glycyrrhiza uralensis, induced the apoptosis of the synovial membrane and its molecular mechanism. In this study, interleukin-1ß (IL-1ß)-RA-FLS cells were incubated with different concentrations of liquiritin. An MTT assay, Hoechst 33342 staining, JC-1 staining, and Western blot were used to check the viability, cell apoptosis, mitochondrial membrane potential changes, and the expression of related proteins, respectively. In vivo, a TUNEL assay and HE staining of tissue were used for histopathological evaluation. Our results showed that liquiritin significantly inhibited the proliferation of IL-1ß-induced-RA-FLS, promoted nuclear DNA fragmentation, and changed the mitochondrial membrane potential to accelerate cell apoptosis. Liquiritin downregulated the ratio of Bcl-2/Bax and inhibited the VEGF expression and phosphorylation of JNK and P38. Moreover, liquiritin improved the clinical score of rheumatism, inflammatory infiltration, and angiogenesis and induced apoptosis of the synovial tissue in vivo. Hence, liquiritin ameliorates RA by reducing inflammation, blocking MAPK signaling, and restraining angiogenesis.

Salicin from Alangium chinense Ameliorates Rheumatoid Arthritis by Modulating the Nrf2-HO-1-ROS Pathways.

Rheumatoid arthritis (RA) is a chronic inflammatory disorder linked to oxidative stress of rheumatoid arthritis fibroblast-like synoviocytes (RA-FLSs). The effects and potential mechanism of salicin on inflammation and oxidative stress of RA-FLSs were examined by MTT, ELISA, and Western blot methods. Salicin significantly reduced cell viability (82.03 ± 7.06, P < 0.01), cytokines (47.70 ± 1.48 ng/L for TNF-α, 30.03 ± 3.49 ng/L for IL-6) ( P < 0.01), and matrix metalloproteinases-1/-3 expression ( P < 0.01) in IL-1ß-induced RA-FLSs and inhibited ROS generation and p65 phosphorylation ( P < 0.01) as compared with IL-1ß-induced treatment. Moreover, salicin promoted Nrf2 nuclear translocation (2.15 ± 0.21) and HO-1 expression (1.12 ± 0.05) and reduced ROS production in IL-1ß-induced RA-FLSs ( P < 0.01). Salicin not only reduced the collagen-induced arthritis by reducing the clinical score ( P < 0.01), inflammatory infiltration, and synovial hyperplasia in vivo but also suppressed the oxidative damage indexes (SOD 155.40 ± 6.53 U/mg tissue, MDA 152.80 ± 5.89 nmol/g tissue, GSH 50.98 ± 3.45 nmol/g tissue, and CAT 0.92 ± 0.10 U/g protein) ( P < 0.01) of ankle joint cells. Conclusively, our findings indicate that salicin ameliorates rheumatoid arthritis, which may be associated with oxidative stress and Nrf2-HO-1-ROS pathways in RA-FLSs.

Apoptosis effects of imperatorin on synoviocytes in rheumatoid arthritis through mitochondrial/caspase-mediated pathways.

Rheumatoid arthritis (RA) is a systemic chronic inflammatory disease associated with a potential imbalance between the growth and death of rheumatoid arthritis fibroblast-like synoviocytes (RA-FLSs). Imperatorin (IPT) is a naturally occurring furanocoumarin found in umbelliferous vegetables, citrus fruits, and some herbs. The effects of IPT on the proliferation and apoptosis of RA-FLSs and its potential underlying mechanisms have remained unclear. RA-FLSs obtained from RA patients were induced by interleukin-1ß (IL-1ß) and treated with IPT. Cell viability was determined by MTT assay. Apoptotic cell death was analyzed by Annexin V-FITC/PI double staining and Hoechst 33342 staining. The loss in the mitochondrial membrane potential (ΔΨm) was visualized on the basis of JC-1 staining via fluorescence microscopy, and protein expression changes were assessed by western blot, whereas in vivo studies were conducted in male Wistar rats followed by histopathological assessment via TUNEL assay and HE staining of tissues. The results showed that IPT significantly reduced cell viability, accelerated cell apoptosis and decreased matrix metalloproteinases-1/-3 expression in IL-1ß-induced RA-FLSs. Furthermore, IPT exposure was found to disrupt the ΔΨm compared to the IL-1ß-induced treatment. Moreover, IPT increased the release of mitochondrial cytochrome C, the ratio of Bax/Bcl-2, and the cleavage of caspase-9, caspase-3 and poly (ADP-ribose) polymerase. In vivo studies showed that IPT not only significantly reduced the collagen induced arthritis by reducing synovial hyperplasia, and pannus formation but also enhanced the apoptotic index of ankle joint cells. Conclusively, our findings suggest that IPT inhibits cell proliferation and induces apoptosis in RA-FLSs that may be associated with mitochondrial/caspase-mediated signalling pathways.

Parallel evolution of behaviour during independent host-shifts following maize introduction into Asia and Europe.

Maize was introduced into opposite sides of Eurasia 500 years ago, in Western Europe and in Asia. This caused two host-shifts in the phytophagous genus Ostrinia; O. nubilalis (the European corn borer; ECB) and O. furnacalis (the Asian corn borer; ACB) are now major pests of maize worldwide. They originated independently from Dicot-feeding ancestors, similar to O. scapulalis (the Adzuki bean borer; ABB). Unlike other host-plants, maize is yearly harvested, and harvesting practices impose severe mortality on larvae found above the cut-off line. Positive geotaxis in the ECB has been proposed as a behavioural adaptation to harvesting practices, allowing larvae to move below the cut-off line and thus escape harvest mortality. Here, we test whether the same behavioural adaptation evolved independently in Europe and in Asia. We sampled eight genetically differentiated ECB, ACB and ABB populations in France and China and monitored geotaxis through the entire larval development in artificial stacks mimicking maize stems. We find that all ECB and ACB populations show a similar tendency to move down during the latest larval stages, a behaviour not observed in any European or Asian ABB population. The behaviour is robustly expressed regardless of larval density, development mode or environmental conditions. Our results indicate that maize introduction triggered parallel behavioural adaptations in Europe and Asia, harvest selection presumably being the main driver.

Protective effect of Rabdosia amethystoides (Benth) Hara extract on acute liver injury induced by Concanavalin A in mice through inhibition of TLR4-NF-κB signaling pathway.

Extract of Rabdosia amethystoides (Benth) Hara (ERA), a traditional Chinese medicine has antibacterial, antiviral, anti-tumor, anti-hepatitis and anti-inflammatory properties. However, the hepatoprotective effects and molecular mechanisms of ERA on acute liver injury have not been fully elucidated. This study aims to investigate the anti-inflammatory effect and liver protection of ERA against the acute liver injury induced by Concanavalin A (Con A) and its underlying molecular mechanisms in mice. Mice received ERA (50, 100, 150 mg/kg body weight) by gavage before Con A intravenous administration. We found that ERA pretreatment was able to significantly reduce the elevated serum alanine and aspartate aminotransferase levels and liver necrosis in Con A-induced hepatitis. In addition, ERA treatment significantly decreased the myeloperoxidase, malondialdehyde levels and augmented superoxide dismutase level in the liver tissue, and also suppressed the secretion of proinflammatory cytokines in the serum, compared with Con A group by enzyme linked immunosorbent assay. Furthermore, we observed that ERA pretreatment can significantly decrease the expression level of Toll-like receptor (TLR) 4 mRNA or protein in liver tissues. Further results showed that ERA pretreatment was capable of attenuating the activation of the NF-κB pathway by inhibiting IκBα kinase and p65 phosphorylation in Con A-induced liver injury. Our results demonstrate that ERA pretreatment has hepatoprotective property against Con A-induced liver injury through inhibition of inflammatory mediators in mice. The beneficial effect of ERA may be mediated by the downregulation of TLR4 expression and the inhibition of NF-κB activation.

[Effects of temperature and density on the mortality and reproduction of cotton aphid Aphis gossypii].

A laboratory cotton leaf disc experiment was conducted to study the effects of different temperature (32, 34, 36, 38, and 40 degrees C) and density (5, 25, 50, and 75 individuals per dish) on the mortality and reproduction of Aphis gossypii. With the increase of temperature, density, and culture duration, the cumulative mortality of A. gossypii presented an increasing trend. The parameters estimated by complementary log-log (CLL) model showed that the beta value decreased with the increase of density, indicating that the effects of temperature weakened with increasing density. The gamma value, a parameter for the time effect of temperature, changed with culture duration, indicating that the morality of A. gossypii was co-affected by the temperature and culture duration. The two-way ANOVA analysis of variance showed that temperature and density had significant effects on the fecundity of A. gossypii, and there existed interactive effect. At 32-36 degrees C, the reproduction rate of A. gossypii decreased with the increase of density, but at 40 degrees C, no significant difference was observed in the reproduction rate under different densities, suggesting that the density effect was weakened with increasing temperature, i. e., the contribution of temperature and density to the survival and reproduction of individual varied with the ranges of the temperature and density. This study could provide reference for the monitoring and forecasting of A. gossypii population and for the improvement of pests control.

High-throughput discovery of chloroplast and mitochondrial DNA polymorphisms in Brassicaceae species by ORG-EcoTILLING.

BACKGROUND: Information on polymorphic DNA in organelle genomes is essential for evolutionary and ecological studies. However, it is challenging to perform high-throughput investigations of chloroplast and mitochondrial DNA polymorphisms. In recent years, EcoTILLING stands out as one of the most universal, low-cost, and high-throughput reverse genetic methods, and the identification of natural genetic variants can provide much information about gene function, association mapping and linkage disequilibrium analysis and species evolution. Until now, no report exists on whether this method is applicable to organelle genomes and to what extent it can be used. METHODOLOGY/PRINCIPAL FINDINGS: To address this problem, we adapted the CEL I-based heteroduplex cleavage strategy used in Targeting Induced Local Lesions in Genomes (TILLING) for the discovery of nucleotide polymorphisms in organelle genomes. To assess the applicability and accuracy of this technology, designated ORG-EcoTILLING, at different taxonomic levels, we sampled two sets of taxa representing accessions from the Brassicaceae with three chloroplast genes (accD, matK and rbcL) and one mitochondrial gene (atp6). The method successfully detected nine, six and one mutation sites in the accD, matK and rbcL genes, respectively, in 96 Brassica accessions. These mutations were confirmed by DNA sequencing, with 100% accuracy at both inter- and intraspecific levels. We also detected 44 putative mutations in accD in 91 accessions from 45 species and 29 genera of seven tribes. Compared with DNA sequencing results, the false negative rate was 36%. However, 17 SNPs detected in atp6 were completely identical to the sequencing results. CONCLUSIONS/SIGNIFICANCE: These results suggest that ORG-EcoTILLING is a powerful and cost-effective alternative method for high-throughput genome-wide assessment of inter- and intraspecific chloroplast and mitochondrial DNA polymorphisms. It will play an important role in evolutionary and ecological biology studies, in identification of related genes associated with agronomic importance such as high yield and improved cytoplasmic quality, and for identifying mitochondrial point mutations responsible for diseases in humans and other animals.

Neuroprotective properties of aucubin in diabetic rats and diabetic encephalopathy rats.

In this study, we determined the neuroprotective effect of aucubin on diabetes and diabetic encephalopathy. With the exception of the control group, all rats received intraperitoneal injections of streptozotocin (STZ; 60 mg/kg) to induce type 1 diabetes mellitus (DM). Aucubin (1, 5, 10 mg/kg ip) was used after induction of DM (immediately) and diabetic encephalopathy (65 days after the induction of diabetes). The diabetic encephalopathy treatment groups were divided into short-term and long-term treatment groups. Treatment responses to all parameters were examined (body weight, plasma glucose, Y-maze error rates and proportion of apoptotic cells). In diabetic rats, aucubin controlled blood glucose levels effectively, prevented complications, and improved the quality of life of diabetic rats. In diabetic encephalopathy, aucubin significantly rescued neurons in the hippocampal CA1 subfield and reduced working errors during behavioral testing. The significant neuroprotective effect of aucubin could be seen not only in the short term (15 days) but also in the long term (45 days), which was a highly encouraging finding. These data suggest that aucubin may be a potential neuroprotective agent.

[Effects of high temperature on the mortality and fecundity of two co-existing cotton aphid species Aphis gossypii Glover and Acyrthosiphon gossypii Mordvilko].

Aphis gossypii and Acyrthosiphon gossypii are the coexisting species on cotton plant, with their abundance differed within and among years. To explore whether the abundance difference was related to the different responses of the two aphid species to high temperature, a laboratory experiment was conducted to assess the mortality and reproduction of the two aphid species at high temperatures 32, 34, 36 and 38 degrees C. With the increasing temperature and prolonged exposure period, the cumulative mortality of the two aphid species increased, and Acyrthosiphon gossypii had a higher cumulative mortality than Aphis gossypii. The daily mortality of the aphids could be well simulated by complementary log-log (CLL) model. The median lethal temperature of the two aphid species estimated by CLL model decreased with prolonged exposure period. Under the same exposure period, the median lethal temperature of Aphis gossypii was higher than that of Acyrthosiphon gossypii. Within the range of 32-38 degrees C, the reproduction rate of the two aphid species decreased with increasing temperature, but Aphis gossypii had a significantly higher reproduction rate than Acyrthosiphon gossypii, indicating that at the temperature higher than 32 degrees C, Aphis gossypi had higher tolerance against high temperature than Acyrthosiphon gossypii, and consequently, had more competitive advantage under global warming.

Protective effects of aucubin on H2O2-induced apoptosis in PC12 cells.

The present study investigated the neuroprotective effects of aucubin on hydrogen peroxide (H2O2)-induced apoptosis in PC12 cells. Exposure of PC12 cells to 0.25 mm H2O2 induced a leakage of lactate dehydrogenase and decreased cell viability, as shown by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. In a dose over 0.1 mm, aucubin increased PC12 cellular viability and markedly attenuated H2O2-induced apoptotic cell death. Quantitation of apoptosis by flow cytometry indicated that aucubin inhibited H2O2-induced apoptosis in PC12 cells. Nuclear damage was alleviated by aucubin, as shown by Hoechst staining. In addition, the levels of malondialdehyde were reduced and the activity of superoxide dismutase, catalase and glutathione peroxidase was augmented in these cells. These results indicated that aucubin inhibited H2O2-induced apoptosis in PC12 cells through regulation of the endogenous oxidant-antioxidant balance. Our results suggest that aucubin is a potential protective agent for the treatment of oxidative-stress-induced neurodegenerative disease.

Aucubin modulates Bcl-2 family proteins expression and inhibits caspases cascade in H2O2-induced PC12 cells.

In this study, the effect of aucubin on H(2)O(2)-induced apoptosis was studied by using a rat pheochromocytoma (PC12) cell line. We have analyzed the apoptosis of H(2)O(2)-induced PC12 cells, H(2)O(2)-induced apoptosis appeared to correlate with lower Bcl-2 expression, higher Bax expression and sequential activation of caspase-3 leading to cleavage of poly-ADP-ribose polymerase (PARP). Aucubin not only inhibited lower Bcl-2 expression, high Bax expression, but also modulated caspase-3 activation, PARP cleavage, and eventually protected against H(2)O(2)-induced apoptosis. These results indicated that aucubin can obstruct H(2)O(2)-induced apoptosis by regulating of the expression of Bcl-2 and Bax, as well as suppression of caspases cascade activation.

Quantitative determination of oil content in small quantity of oilseed rape by ultrasound-assisted extraction combined with gas chromatography.

Accurately quantitative determination of oil content in oilseed rape plays an important role in varieties breeding for improving oil content in seeds. However, large quantity of oilseeds were needed in order to obtain accuracy and precision results by using standard Soxhlet extraction method, which may be a handicap in analysis of small, rare and precious samples in plant breeding. In the present work, ultrasound-assisted extraction was evaluated as a simpler and more effective alternative to conventional extraction method for the isolation of oil from small quantity of oilseed rape (<20 mg). The oil of oilseed rape samples was extracted by ultrasound-assisted method, and then the fatty acids and total oil content of the seeds were qualitatively and quantitatively determined by gas chromatography (GC). Extraction efficiency of total oil obtained by ultrasound-assisted extraction through an orthogonal experiment (L(9) (3(4))) were investigated to get the best extraction conditions. Statistical analysis showed that the variable with the largest effect was the ultrasound-assisted extraction time which was followed by the ultrasound-assisted extraction power, and the liquid:solid ratio. A liquid:solid ratio of 1:4 (L:g), an ultrasound-assisted extraction time of 60 min and an ultrasound-assisted extraction power of 500 W were found to be optimal for oil extraction from oilseed rape. By comparing with the conventional method, it was found that the ultrasound-assisted extraction of oil from oilseed rape was about five times faster than the traditional extraction method. By the use of ultrasound-assisted extraction combined with GC analysis, the fatty acids and total oil content in small quantity of seeds (<20 mg) were successfully qualitatively determined and the results are in agreement with that obtained by traditional standard method.

[Molecular mapping of the gene(s) controlling petal-loss trait in Brassica napus].

An F2 population derived from a cross between apetalous line' APT02 'and normal petalled cultivar 'ZS NO.4' was used for molecular marker searching and chromosomal mapping of the gene(s) controlling petal-loss trait in Brassica napus. Twenty pairs of AFLP primers and 170 pairs of SRAP primers were selected and screened from two parents. In further selection through bulked segregant analysis (BSA) approach, one SRAP marker e8m3_4 (600 bp) and one AFLP marker E3247_15 (150 bp) were obtained and found to be linked to the gene(s) controlling petal-loss trait, with the genetic distance of 5 cM and 13.5 cM. A linkage map in Brassica napus was constructed. It consisted of 213 AFLP56 SSR loci and a morphology marker throughout 17 main linkage groups, two triplet and four linkage pairs. Total length of the map covered 2,487.1 cM, and average interval between markers was 10.09 cM. By genetic mapping, the gene(s) controlling petal-loss trait (WHB)was mapped in LG4.

[Study on the mechanism of the interaction of methylene blue and sulfation pachymaran].

In order to understand the mechanism of the interaction of sulfate polysaccharides and small molecules at molecular level, the structure of pachymaran was decorated by Wolfrom, and sulfation pachymaran (SP) was prepared. The maximum binding number (N = 54) and the binding equilibrium constant (K = 1.563 x 10(6)) of the sulfation pachymaran and methylene blue (MB) were obtained by the exoterica model. The influence of the molar ratio of MB/SP, ethanol, hydroxypropyl-beta-cyclodextrin, Triton X-100 and NaCl on the spectra of MB-SP complex was investigated. The mechanism of the color changes of methylene blue and sulfation pachymaran reaction system has been discussed. The authors draw the conclusion that the color changes result mainly from the hydrophobic interaction between methylene blue molecules binding on sulfation pachymaran on the base of the electrostatic interaction of methylene blue and sulfation pachymaran.

[Study on quantitative assay of chondroitin sulfate with a spectrophotometric method of azure A].

A simple, accurate and rapid spectrophotmeric method was proposed for the determination of chondroitin sulfate. The method was based on the absorbances of AA-CS complex at 625 nm being in proportion to the chondroitin sulfate concentrations. The linear range was 0-30 micrograms.mL-1 (R = 0.999), and the recovery range was 97.4%-103.8%. The quantities of chondroitin sulfate in different sample were determined in this way.