mRNA Splicing of UL44 and Secretion of Alphaherpesvirinae Glycoprotein C (gC) Is Conserved among the Mardiviruses.

Marek's disease (MD), caused by gallid alphaherpesvirus 2 (GaAHV2) or Marek's disease herpesvirus (MDV), is a devastating disease in chickens characterized by the development of lymphomas throughout the body. Vaccine strains used against MD include gallid alphaherpesvirus 3 (GaAHV3), a non-oncogenic chicken alphaherpesvirus homologous to MDV, and homologous meleagrid alphaherpesvirus 1 (MeAHV1) or turkey herpesvirus (HVT). Previous work has shown most of the MDV gC produced during in vitro passage is secreted into the media of infected cells although the predicted protein contains a transmembrane domain. We formerly identified two alternatively spliced gC mRNAs that are secreted during MDV replication in vitro, termed gC104 and gC145 based on the size of the intron removed for each UL44 (gC) transcript. Since gC is conserved within the Alphaherpesvirinae subfamily, we hypothesized GaAHV3 (strain 301B/1) and HVT also secrete gC due to mRNA splicing. To address this, we collected media from 301B/1- and HVT-infected cell cultures and used Western blot analyses and determined that both 301B/1 and HVT produced secreted gC. Next, we extracted RNAs from 301B/1- and HVT-infected cell cultures and chicken feather follicle epithelial (FFE) skin cells. RT-PCR analyses confirmed one splicing variant for 301B/1 gC (gC104) and two variants for HVT gC (gC104 and gC145). Interestingly, the splicing between all three viruses was remarkably conserved. Further analysis of predicted and validated mRNA splicing donor, branch point (BP), and acceptor sites suggested single nucleotide polymorphisms (SNPs) within the 301B/1 UL44 transcript sequence resulted in no gC145 being produced. However, modification of the 301B/1 gC145 donor, BP, and acceptor sites to the MDV UL44 sequences did not result in gC145 mRNA splice variant, suggesting mRNA splicing is more complex than originally hypothesized. In all, our results show that mRNA splicing of avian herpesviruses is conserved and this information may be important in developing the next generation of MD vaccines or therapies to block transmission.

Monoterpenoids from the root bark of Acanthopanax gracilistylus and their inhibitory effects on neutrophil elastase, 5-lipoxygenase, andcyclooxygenase-2 in vitro.

Twenty-four monoterpenoids, including three previously undescribed compounds (1-3), were isolated from the root bark of Acanthopanax gracilistylus W. W. Smith (Acanthopanacis Cortex). Their structures were unambiguously established based on spectroscopic analysis (HR-ESIMS, IR, 1D, and 2D NMR), and the absolute configurations of 1-3 were elucidated by comparing their experimental and calculated electronic circular dichroism spectra. In addition, the structure of 8 was confirmed by single-crystal X-ray diffraction. The inhibitory activities of 1-24 against neutrophil elastase, 5-lipoxygenase, and cyclooxygenase-2 (COX-2) were studied in vitro for the first time, and the results showed that compound 24 possessed a significant inhibitory effect on COX-2 with an IC50 value of 1.53 ± 0.10 µΜ. This research first reported the presence of monoterpenoids in Acanthopanacis Cortex, including one monoterpenoid 2 with an unusual 4/5 bicyclic lactone system, and compounds 4 and 5 have never been reported in nature.

Transcription level differences in Taxus wallichiana var. mairei elicited by Ce3+, Ce4+ and methyl jasmonate.

Taxol is a precious and effective anticancer drug. Cerium and methyl jasmonate (MJ) have been shown to increase the yield of taxol in taxus cells. However, the mechanisms of cerium-mediated and MJ-mediated taxol biosynthesis remain unknown. RNA-Seq was applied to study the overall regulation mechanism of cerium and MJ on taxol biosynthesis and analyze the differences among T. mairei cells elicited by Ce3+, Ce4+ and MJ on transcriptional level . Using sequence homology, 179 unigenes were identified as taxol synthesis genes. Under the condition of 100 µM MJ, taxol synthesis genes were up-regulated. Notably, taxol synthesis genes were down-regulated expression at 1 mM Ce3+ and 1 mM Ce4+. Differential expression genes involved in some related functions were analyzed, such as MAPK signaling pathway and plant-pathogen interaction. Sequence alignment and phylogenetic analysis of nine differentially expressed WRKYs in our data were carried out.

Lysosomal dysfunction is associated with NLRP3 inflammasome activation in chronic unpredictable mild stress-induced depressive mice.

NLRP3 inflammasome pathway-mediated inflammatory response is closely associated with depression. Increasing attention has been recently paid to the links between autophagy and depression, however, the relationship between autophagy and NLRP3 inflammasome in depressive behavior remain poorly understood. In the present study, the potential roles of autophagy-lysosome pathway in NLRP3 inflammasome regulation were investigated both in vivo (chronic unpredictable mild stress (CUMS)-induced depressive mouse model) and in vitro (LPS-induced cellular model) model. It demonstrated that CUMS induces depressive-like behaviors in mice, accompanied by increased expression of NLRP3 inflammasome and inflammatory responses. Meanwhile, it promoted the autophagosome marker LC3 and autophagic adapter protein p62 accumulation, accompanied by the decrease of lysosomal cathepsins B and D expression in the prefrontal cortex of mice. Notably, a significant colocalization of NLRP3 and LC3 in CUMS mice by immunofluorescence co-staining were observed. For the in vitro study, disrupting the lysosomal function with Baf A1 significantly increased the LPS-induced NLRP3 inflammasome accumulation and pro-inflammatory factors (IL-1ß and IL-18) production in BV2 cells. Collectively, our results suggested that the autophagic process is related to NLRP3 inflammasome activation, and dysfunctional lysosome in autophagy-lysosomal pathway may retard NLRP3 inflammasome degradation, facilitating the production of pro-inflammatory factors, thereby contributing to depressive behavior in CUMS mice.

Effect of Thermal Treatment on the Internal Structure, Physicochemical Properties and Storage Stability of Whole Grain Highland Barley Flour.

In this study, to improve the processing performance of whole grain highland barley flour (whole grain HB flour), they were prepared by sand-roasting, far-infrared baking, steam explosion, and extrusion, and the effects of on functional properties and storage characteristics were measured. The results indicated that sand-roasting, far-infrared baking, and steam explosion all caused cracks and honeycomb structures in the outer layer and endosperm of the highland barley. The XRD analysis results indicated that highland barley starch treated by far-infrared baking exhibited typical A-type crystal structure, while sand-roasting, steam explosion, and extrusion presented the typical V-type. The results of DSC analysis revealed that the onset temperature (To), peak temperature (Tp), gelatinization enthalpy (ΔH), peak viscosity (PV), trough viscosity (TV), and final viscosity (FV) decreased significantly, while the swelling power, water-holding capacity and oil-holding capacity increased significantly. During the storage period, the moisture content and lipase activity of the whole grain HB flour after thermal treatment remained at a low level; the fatty acid value, peroxide value, and malondialdehyde value increased; finally, the cooked whole grain HB flour was unstable during storage. The functional properties of whole grain HB flour can be improved by steam explosion, and will then have better storage stability.

Evaluation of Nutritional Components, Phenolic Composition, and Antioxidant Capacity of Highland Barley with Different Grain Colors on the Qinghai Tibet Plateau.

The nutritional composition, polyphenol and anthocyanin composition, and antioxidant capacity of 52 colored highland barley were evaluated. The results showed that the protein content of highland barley in the black group was the highest, the total starch and fat contents in the blue group were the highest, the amylose content in the purple group was quite high, the fiber content in the yellow group was quite high, and the ß-glucan content of the dark highland barley (purple, blue and black) was quite high. The polyphenol content and its antioxidant capacity in the black group were the highest, while the anthocyanin content and its antioxidant capacity in the purple highland barley were the highest. Ten types of monomeric phenolic substances were the main contributors to DPPH, ABTS, and FRAP antioxidant capacity. All varieties could be divided into four categories according to nutrition or function. The grain color could not be used as an absolute index to evaluate the quality of highland barley, and the important influence of variety on the quality of highland barley also needed to be considered. In actual production, suitable raw materials must be selected according to the processing purpose and variety characteristics of highland barley.

Coinfection in the host can result in functional complementation between live vaccines and virulent virus.

One of the greatest achievements of the last century is the development of vaccines against viral diseases. Vaccines are essential for battling infectious diseases and many different formulations are available, including live attenuated vaccines. However, the use of live attenuated vaccines has the potential for adverse effects, including reversion of pathogenicity, recombination, and functional complementation in the host. Marek's disease is a serious disease in poultry controlled by live attenuated vaccines that has resulted in increased virulence over the decades. Recombination between circulating field viruses or vaccines is a proposed mechanism for the increase in virulence, however, complementation between vaccines and field strains has not been demonstrated in chickens. Here, we describe functional complementation of vaccines with virulent virus to functionally complement transmission and spread in the host. Using the natural virus-host model of Marek's disease in chickens, our results show dual infection of target cells in chickens with vaccine and virulent virus providing the opportunity for recombination or complementation to transpire. Interestingly, our controlled results showed no evidence of recombination between vaccine and virulent virus, but functional complementation occurred in two independent experiments providing proof for complementation during natural infection in vaccinated individuals. These results suggest complementation as a potential mechanism for vaccine-mediated viral evolution and the potential for complementation should be taken into consideration when developing novel vaccines.

Screening 5-lipoxygenase inhibitors from selected traditional Chinese medicines and isolation of the active compounds from Polygoni Cuspidati Rhizoma by an on-line bioactivity evaluation system.

To identify natural products as new prototypes for 5-lipoxygenase (5-LOX), 12 traditional Chinese medicines (TCMs) were selected for screening their 5-LOX inhibition activities. The results showed that the methanol extracts of all selected TCMs (n = 12) possessed inhibitory activities against 5-LOX at 200 µg/mL, of which six extracts of the TCMs showed significant inhibitory effects with IC50 values in the range from 33.2 ± 1.4 µg/mL to 153.5 ± 1.7 µg/mL, and the extract of Polygoni Cuspidati Rhizoma (RPC) was the most active sample. An on-line ultra-performance liquid chromatography-photodiode array-MSn -5-LOX-fluorescence detector (UPLC-PDA-MSn -5-LOX-FLD) method was applied to further identify the potential 5-LOX inhibitory constituents in RPC extracts, which resulted in the identification of seven components with 5-LOX-binding activities. Finally, four compounds (polydatin, resveratrol, emodin-8-O-glucoside, and emodin) were successfully purified from RPC extracts. The 5-LOX inhibition action was assayed in vitro, and the results showed that these compounds possessed potent inhibitory effects against 5-LOX with IC50 values of 15.3 ± 2.1, 4.5 ± 1.2, 23.8 ± 0.4, and 11.8 ± 1.5 µg/mL, respectively. This was the first study to reveal the 5-LOX inhibitory constituents of RPC, and the present investigation might provide a valuable approach for the rapid discovery of natural inhibitors from TCMs.

The Conserved Herpesviridae Protein Kinase (CHPK) of Gallid alphaherpesvirus 3 (GaHV3) Is Required for Horizontal Spread and Natural Infection in Chickens.

We have formerly identified the conserved herpesvirus protein kinase (CHPK) as essential for horizontal transmission of Marek's disease virus (MDV). Thus far, it has been confirmed that the mutation of the invariant lysine (K) of CHPKs abrogates kinase activity and that CHPK activity is required for MDV horizontal transmission. Since CHPK is conserved among all members of the Herpesviridae, we hypothesized that CHPK, and specifically its kinase activity, is important for the horizontal transmission of other herpesviruses. To test this hypothesis, we utilized our experimental and natural infection model in chickens with MD vaccine strain 301B/1 of Gallid alphaherpesvirus 3 (GaHV3). First, we mutated the invariant lysine (K) 157 of 301B/1 CHPK to alanine (A) and determined whether it was required for horizontal transmission. To confirm the requirement of 301B/1 CHPK activity for transmission, a rescued virus was generated in which the A157 was changed back to a K (A157K). Despite both the CHPK mutant (K157A) and rescuant (A157K) viruses having replication defects in vivo, only the CHPK mutant (K157A) was unable to spread to contact chickens, while both wild-type and rescuant (A157K) viruses transmitted efficiently, confirming the importance of CHPK activity for horizontal spread. The data confirm that CHPK is required for GaHV3 transmission and suggest that the requirement of avian CHPKs for natural infection is conserved.

Links Between Gut Dysbiosis and Neurotransmitter Disturbance in Chronic Restraint Stress-Induced Depressive Behaviours: the Role of Inflammation.

Accumulating evidence has shown that inflammation, the gut microbiota, and neurotransmitters are closely associated with the pathophysiology of depression. However, the links between the gut microbiota and neurotransmitter metabolism remain poorly understood. The present study aimed to investigate the neuroinflammatory reactions in chronic restraint stress (CRS)-induced depression and to delineate the potential links between the gut microbiota and neurotransmitter metabolism. C57BL/6 mice were subjected to chronic restraint stress for 5 weeks, followed by behavioural tests (the sucrose preference test, forced swim test, open field test, and elevated plus maze) and analysis. The results showed that CRS significantly increased interleukin-1 beta (IL-1ß), interleukin-2 (IL-2), interleukin-6 (IL-6), and tumour necrosis factor α (TNFα) levels and decreased brain-derived neurotrophic factor (BDNF) expression, accompanied by the activation of IkappaB-alpha-phosphorylation-nuclear factor kappa-B (IκBα-p-NF-κB) signalling in the mouse hippocampus. In addition, the neurotransmitter metabolomics results showed that CRS resulted in decreased levels of plasma 5-hydroxytryptamine (5-HT), dopamine (DA), and noradrenaline (NE) and their corresponding metabolites, and gut microbiota faecal metabolites with the 16S rRNA gene sequencing indicated that CRS caused marked microbiota dysbiosis in mice, with a significant increase in Helicobacter, Lactobacillus, and Oscillibacter and a decrease in Parabacteroides, Ruminococcus, and Prevotella. Notably, CRS-induced depressive behaviours and the disturbance of neurotransmitter metabolism and microbiota dysbiosis can be substantially restored by dexamethasone (DXMS) administration. Furthermore, a Pearson heatmap focusing on correlations between the microbiota, behaviours, and neurotransmitters showed that Helicobacter, Lactobacillus, and Oscillibacter were positively correlated with depressive behaviours but were negatively correlated with neurotransmitter metabolism, and Parabacteroides and Ruminococcus were negatively correlated with depressive behaviours but were positively correlated with neurotransmitter metabolism. Taken together, the results suggest that inflammation is involved in microbiota dysbiosis and the disturbance of neurotransmitter metabolism in CRS-induced depressive changes, and the delineation of the potential links between the microbiota and neurotransmitter metabolism will provide novel strategies for depression treatment.

Diet, food availability, and climatic factors drive ranging behavior in white-headed langurs in the limestone forests of Guangxi, southwest China.

Changes in abiotic and biotic factors can affect the efficiency of biological systems in animals, forcing them to adjust their behaviors in response to daily and seasonal variations. From September 2016 to August 2017, we collected ranging behavior data on four groups of white-headed langurs ( Trachypithecus leucocephalus) in the Guangxi Chongzuo White-Headed Langur National Nature Reserve, Guangxi, southwest China. We simultaneously analyzed how multiple ecological factors affect langur ranging behavior, which should facilitate our understanding of the potential mechanisms underlying their adaptation to limestone habitats. Results showed that langur ranging behavior was significantly affected by diet composition, food availability, and climatic factors. Specifically, moving time and daily path length increased with the increase in dietary diversity. Furthermore, moving time and daily path length were positively associated with the availability of fruit and relative humidity of the forest, and moderately associated with temperature and relative humidity of bare rock. Our study demonstrated that langurs maintain stable moving and feeding times and exhibit a short daily travel distance, likely adopting an energy-conserving behavioral strategy in response to food shortages and high temperatures in the fragmented karst forest. These results highlight the importance of food availability and temperature in shaping the ranging behavior of these karst-dwelling primates.

The requirement of glycoprotein C (gC) for interindividual spread is a conserved function of gC for avian herpesviruses.

We have formerly shown that glycoprotein C (gC) of Gallid alphaherpesvirus 2, better known as Marek's disease (MD) alphaherpesvirus (MDV), is required for interindividual spread in chickens. Since gC is conserved within the Alphaherpesvirinae subfamily, we hypothesized gC was important for interindividual spread of other alphaherpesviruses. To test this hypothesis, we first generated a fluorescent protein tagged clone of Gallid alphaherpesvirus 3 MD vaccine strain 301B/1 to track virus replication in cell culture and chickens using fluorescent microscopy. Following validation of this system, we removed the open reading frame of 301B/1 gC from the genome and determined whether it was required for interindividual spread using experimental and natural infection studies. Interindividual spread of MD vaccine 301B/1 was abrogated by removal of 301B/1 gC. Rescuent virus in which 301B/1 gC was inserted back into the genome efficiently spread among chickens. To further study the conserved function of gC, we replaced 301B/1 gC with MDV gC and this virus also efficiently spread in chickens. These data suggest the essential function of alphaherpesvirus gC proteins is conserved and can be exploited during the generation of future vaccines against MD that affects the poultry industry worldwide.

Cyperane-Type and Related (Nor)Sesquiterpenoids from the Root Bark of Acanthopanax gracilistylus and Their Inhibitory Effects on Nitric Oxide Production.

An enantiomeric pair of rare cyperane-type sesquiterpenoids, (+)- and (-)-gracilistones C (1a, 1b), together with a novel norsesquiterpenoid, gracilistone D (2), bearing a bicyclic lactone system were isolated from the root bark of Acanthopanax gracilistylus using LC-MS-IT-TOF analyses. The structures and absolute configurations of 1a, 1b, and 2 were elucidated by 1D and 2D NMR spectroscopy, X-ray diffraction, and ECD spectroscopic methods. Intermediate 1b suggests a possible biosynthesis process involving compound 2. The bioassay results showed that compounds 1a, 1b, and 2 exhibited significant inhibitory effects against lipopolysaccharide-induced nitric oxide production in RAW 264.7 cells, with IC50 values of 7.7 ± 0.6, 6.8 ± 1.5, and 2.6 ± 0.4 µM, respectively. Additional docking analyses provided some perspective of this activity in human inducible nitric oxide synthase.

LAMP real-time turbidity detection for fowl adenovirus.

BACKGROUND: Fowl adenovirus (FAdV) is an infectious agent that can cause jaundice, severe anaemia, dyspnoea and reproductive disorders in fowls. Since 2015, FAdV disease has been rapidly spreading among broiler chickens in China, where it has caused significant economic losses. In this study, a loop-mediated isothermal amplification (LAMP) real-time turbidity method with strong specificity to FAdV was established. RESULTS: The established assay was specific to FAdV-4, and the lower limit of detection was 75 copies/µL of extracted DNA. The positive detection rate for the suspected tissue samples was 33.3% (14/42), which was consistent with that of the real-time PCR. CONCLUSION: The proposed LAMP method can quickly and accurately detect prevalent FAdV via real-time turbidity assay, thereby providing a diagnostic platform for the prevention and control of the FAdV disease.

[Preparation and quality standard of standard decoction of Scrophulariae Radix pieces].

The standard decoction of Chinese herbal decoction pieces is a standard reference substance to measure whether different dosage forms of Chinese medicine are basically consistent with those of clinical decoction,and provides new ideas and methods for effectively solving the problems of uneven quality in Chinese medicine dispensing granules. In this study,a systematic method for evaluating the quality of Scrophulariae Radix decoction was established from the perspective of " standard decoction",providing reference for the quality control of the Scrophulariae Radix dispensing granules. 15 batches of Scrophulariae Radix decoction pieces from different origins were collected,and 15 batches of standard decoctions were prepared according to the standardized process with water as solvent.Harpagide and harpagoside were used as quantitative detection indicators to determine the content,calculate the transfer rates and determine the extraction rate. The high performance liquid chromatography( HPLC) was used to establish a standard decoction fingerprint analysis method. The results showed that the transfer rates of harpagide and harpagoside in 15 batches of Scrophulariae Radix pieces standard decoction were( 70. 84±13. 39) % and( 48. 56±6. 40) % respectively; the extraction rate was( 57. 47±5. 89) %. Nine peaks were identified in the HPLC fingerprint,and the similarity was higher than 0. 97 between the fingerprints of 15 batches of standard decoction and the control fingerprint. In this study,the preparation process of standard decoction of Scrophulariae Radix pieces conformed to the traditional decoction preparation method. The sources of the samples were representative,and the established fingerprint method was stable and feasible,which can provide reference for the preparation and quality control of Scrophulariae Radix dispensing granules.

NO in the dPAG modulates panic-like responses and ASIC1a expression in the prefrontal cortex and hippocampus in mice.

Panic disorder (PD) is a multifactorial neuropsychiatric disorder. Our previous study has demonstrated that the nitric oxide (NO) pathway and the acid-sensing ion channel 1a (ASIC1a) level in the dorsal midbrain periaqueductal gray (dPAG) are involved in the modulation of panic-like responses. In addition, the prefrontal cortex (PFC) and the hippocampus also play a role in panic-like responses. However, no studies have investigated the protein level of ASIC1a in the PFC and hippocampus in a mouse model of panic-like disorders after alteration of the NO pathway in the dPAG. We investigated the production of a panic attack with intra-dPAG injections of SNAP, an NO donor, and 7-NI, an nNOS inhibitor. Moreover, we measured ASIC1a protein levels in the PFC and hippocampus. The rat exposure test (RET) is frequently used as an animal model of panic. In our study, C57BL/6 mice received an intra-dPAG injection of SNAP or 7-NI before RET; neurobehavioral tests were then conducted, followed by mechanistic evaluation through western blot analysis in the PFC and hippocampus. An intra-dPAG infusion of SNAP significantly increased the panic-like effect, whereas treatment with 7-NI decreased fear behavior. Mice treated with SNAP/7-NI showed significantly increased/decreased ASIC1a expression in the PFC, and a decreasing/increasing trend in the hippocampus. The present study suggests that the NO pathway in the dPAG plays a key role in panic-like responses in mice confronted by a rat, further, NO intra-dPAG injection also modulates the ASIC1a expression levels in the PFC and hippocampus.

LC-MS guided isolation of gracilistones A and B, a pair of diastereomeric sesquiterpenoids with an unusual tetrahydrofuran-fused tricyclic skeleton from Acanthopanax gracilistylus and their potential anti-inflammatory activities.

Gracilistones A (1) and B (2), two new eudesmane-type sesquiterpenoids with an unusual tetrahydrofuran-fused 6/6/5 tricyclic ring system, were obtained from Acanthopanax gracilistylus under the guidance of LC-MS investigation. Their structures and absolute configurations were assigned by extensive spectroscopic analyses and quantum calculation methods. Compounds 1 and 2 showed potent inhibitory activity against LPS-induced nitric oxide production in RAW 264.7 macrophages, compared with the positive control L-NMMA. In addition, compounds 1 and 2 were also evaluated for their antioxidant (DPPH• and ABTS•+) and xanthine oxidase (XO) inhibitory activities, and they exhibited weak inhibitory effects at 100 µM.

Regulatory role of microRNA-320a in the proliferation, migration, invasion, and apoptosis of trophoblasts and endothelial cells by targeting estrogen-related receptor γ.

Recently, microRNAs (miRNAs) have been demonstrated to participate in many physiological and biological processes, especially by acting as circulating biomarkers or modulators in cell differentiation. Therefore, the aim of the current study was to clarify whether microRNA-320a (miR-320a) regulates the proliferation, migration, invasion, and apoptosis of trophoblasts and endothelial cells. In this study, miR-320a mimics and inhibitors were transfected into HTR.8/SVneo cells and human umbilical vein endothelial cells (HUVECs) using liposomes. Subsequently, the expression of miR-320a and estrogen-related receptor γ (ERRγ) mRNA was detected by a reverse transcription quantitative polymerase chain reaction, whereas the protein expression of ERRγ, vascular endothelial growth factor (VEGF), angiogenin 1 (Ang-1), human 3beta-hydroxysteroid dehydrogenase type 1 (HSD3B1), and human chorionic gonadotropin (HCG) was detected by western blot analysis. Furthermore, the proliferation, invasion/migration, and apoptosis of cells were analyzed by the cell counting kit-8 assay, transwell assay, and flow cytometry, respectively. The results showed that overexpression of miR-320a decreased the optical density (OD) values and the proliferation rate of HTR.8/SVneo cells and HUVECs, while inhibiting the expression of VEGF, Ang-1, HSD3B1, and HCG in these cells. Furthermore, miR-320a reduced the ability of cell invasion and migration, while increasing the rate of cell apoptosis. After cotransfecting the cells with miR-320a and ERRγ small (or short) interfering RNA (siRNA), the decreased ERRγ expression led to inhibited proliferation, migration, and invasion, but increased apoptosis of HTR.8/SVneo cells and HUVECs. Our results further revealed that miR-320a induced the apoptosis of trophoblasts and endothelial cells while inhibiting their proliferation, migration, and invasion by decreasing the expression of ERRγ and by indirectly suppressing the expression of VEGF, Ang-1, HSD3B1, and HCG.

Mechanism of nitric oxide and acid-sensing ion channel 1a modulation of panic-like behaviour in the dorsal periaqueductal grey of the mouse.

Predators induce defensive responses and fear behaviours in prey. The rat exposure test (RET) is frequently used as an animal model of panic. Nitric oxide (NO) which has been reported to be activated by the NMDA receptor, in turn mediates calcium/calmodulin-dependent protein kinase II (CaMKII) signalling pathways in defensive responses. ACCN2, the orthologous human gene of acid-sensing ion channel 1a (ASIC1a), is also associated with panic disorder; however, few studies have focused on the role of ASIC1a in the modulation of panic and calcium/CaMKII signalling by NO. In the present study, NG-nitro-L-arginine-methyl-ester (L-NAME; non-selective NOS inhibitor), S-nitroso-N-acetyl-D,L-penicillamine (SNAP; NO donor), and psalmotoxin (PcTx-1; selective ASIC1a blocker) were administered to the dorsal periaqueductal grey (dPAG) before the predator stimulus, and the roles of NO in the expression of ASIC1a, phosphorylation of CaMKIIα (p-CaMKIIα) and expression of calmodulin (CaM) were investigated. The effects of ASIC1a, p-CaMKIIα and CaM regulation were also examined. Our results showed that intra-dPAG infusion of L-NAME weakened panic-like behaviour and decreased ASIC1a, p-CaMKIIα and CaM expression levels, whereas intra-dPAG infusion of SNAP enhanced panic-like behaviour and increased ASIC1a, p-CaMKIIα and CaM levels. Intra-dPAG infusion of PcTx-1 also weakened panic-like behaviour and decreased p-CaMKIIα expression level. Taken together, these results indicate that NO and ASIC1a are involved in the modulation of RET-induced panic-like behaviour in the dPAG. NO regulates the calcium/CaMKII signalling pathways, and ASIC1a participates in this regulation.

Overexpression of long non-coding RNA CRNDE facilitates epithelial-mesenchymal transition and correlates with poor prognosis in intrahepatic cholangiocarcinoma.

The clinical significance and essential role of long non-coding RNA colorectal neoplasia differentially expressed (lncRNA CRNDE) have been well illuminated in various cancers. However, the function of CRNDE in intrahepatic cholangiocarcinoma (IHCC) has not been reported at present. The aim of the present study was to investigate the role of CRNDE in IHCC. Firstly, the relative expression of CRNDE was observed to be upregulated in IHCC cell lines and tissues. And high CRNDE expression was statistically associated with IHCC differentiation grade, lymph node metastasis, tumor-nodes-metastasis (TNM) stage and size. Survival analysis identified that high CRNDE expression is a predictor of worse overall survival (OS) and progression-free survival (PFS) in patients with IHCC. Moreover, high CRNDE expression was identified as an independent risk factor of IHCC poor OS and PFS. Further studies of in vitro assays suggested that CRNDE silencing could suppress the proliferation of HuCCT1 cells following CCK-8 and colony formation assays, while CRNDE ectopic expression in HCCC9810 cells promoted proliferation. Moreover, the migration and invasion of HuCCT1 cells were greatly repressed with CRNDE deficiency following Transwell and Matrigel assays. Accordingly, the motility of HCCC9810 cells was notably accelerated with CRNDE overexpression. Mechanistically, CRNDE was revealed to facilitate the epithelial-mesenchymal transition (EMT) of IHCC cells. In conclusion, these observations indicated that CRNDE could promote the clinical progression and metastasis of IHCC by facilitating EMT. CRNDE may be a novel prognostic marker and therapeutic target in IHCC.