Asymmetric magnesium catalysis for important chiral scaffold synthesis.

Magnesium catalysts are widely used in catalytic asymmetric reactions, and a series of catalytic strategies have been developed in recent years. Herein, in this review, we have tried to summarize asymmetric magnesium catalysis for the synthesis of important chiral scaffolds. Several important optically active motifs that are present in classic chiral ligands or natural products synthesized by Mg(II) catalytic methods are briefly discussed. Moreover, the representative mechanisms for different magnesium catalytic strategies, including in situ generated magnesium catalysts, are also shown in relation to synthetic routes for obtaining these important chiral scaffolds.

Molecular detection and genetic characteristics of porcine circovirus 3 and porcine circovirus 4 in central China.

Porcine circoviruses (PCVs) are a significant cause of concern for swine health, with four genotypes currently recognized. Two of these, PCV3 and PCV4, have been detected in pigs across all age groups, in both healthy and diseased animals. These viruses have been associated with various clinical manifestations, including porcine dermatitis and nephropathy syndrome (PDNS) and respiratory and enteric signs. In this study, we detected PCV3 and PCV4 in central China between January 2022 and February 2023. We tested fecal swabs and tissue samples from growing-finishing and suckling pigs with or without respiratory and systemic manifestations and found the prevalence of PCV3 to be 15.15% (15/99) and that of PCV3/PCV4 coinfection to be 4.04% (4/99). This relatively low prevalence might be attributed to the fact that most of the clinical samples were collected from pigs exhibiting respiratory signs, with only a few samples having been obtained from pigs with diarrhea. In some cases, PCV2 was also detected, and the coinfection rates of PCV2/3, PCV2/4, and PCV2/3/4 were 6.06% (6/99), 5.05% (5/99), and 3.03% (3/99), respectively. The complete genomic sequences of four PCV3 and two PCV4 isolates were determined. All four of the PCV3 isolates were of subtype PCV3b, and the two PCV4 isolates were of subtype PCV4b. Two mutations (A24V and R27K) were found in antibody recognition domains of PCV3, suggesting that they might be associated with immune escape. This study provides valuable insights into the molecular epidemiology and evolution of PCV3 and PCV4 that will be useful in future investigations of genotyping, immunogenicity, and immune evasion strategies.

Molecular and functional characterization of porcine poly C binding protein 1 (PCBP1).

BACKGROUND: Poly C Binding Protein 1 (PCBP1) belongs to the heterogeneous nuclear ribonucleoprotein family. It is a multifunctional protein that participates in several functional circuits and plays a variety of roles in cellular processes. Although PCBP1 has been identified in several mammals, its function in porcine was unclear. RESULTS: In this study, we cloned the gene of porcine PCBP1 and analyzed its evolutionary relationships among different species. We found porcine PCBP1 protein sequence was similar to that of other animals. The subcellular localization of PCBP1 in porcine kidney cells 15 (PK-15) cells was analyzed by immunofluorescence assay (IFA) and revealed that PCBP1 was mainly localized to the nucleus. Reverse transcription-quantitative PCR (RT-qPCR) was used to compare PCBP1 mRNA levels in different tissues of 30-day-old pigs. Results indicated that PCBP1 was expressed in various tissues and was most abundant in the liver. Finally, the effects of PCBP1 on cell cycle and apoptosis were investigated following its overexpression or knockdown in PK-15 cells. The findings demonstrated that PCBP1 knockdown arrested cell cycle in G0/G1 phase, and enhanced cell apoptosis. CONCLUSIONS: Porcine PCBP1 is a highly conserved protein, plays an important role in determining cell fate, and its functions need further study.

Early Application of Sacubitril Valsartan Sodium After Acute Myocardial Infarction and its Influence on Ventricular Remodeling and TGF-ß1/Smad3 Signaling Pathway.

Objective: The objective of this study was to investigate the early application of sacubitril valsartan sodium (LCZ696) following acute myocardial infarction (AMI) and its impact on ventricular remodeling and the TGF-ß1/Smad3 signaling pathway in patients. Methods: The clinical data of 73 patients with AMI admitted to the hospital from June 2021 to September 2022 were retrospectively analyzed, and the patients were grouped according to the treatment methods, including 36 cases in the control group (conventional drug treatment) and 37 cases in the observation group (conventional drug + LCZ696 treatment). The clinical efficacy, cardiac function parameters [left ventricular ejection fraction (LVEF), left ventricular end-diastolic diameter (LVEDD), stroke volume (SV)], cardiac function biochemical indicators [N-terminal pro-B-type natriuretic peptide (NT-proBNP), galectin 3 (Gal-3), amino-terminal peptide of type III procollagen (PIIINP)], ventricular remodeling indicators [left ventricular posterior wall end-diastolic thickness (PWD), posterior wall end-systolic thickness (PWS), ventricular septal end-systolic thickness (IVSS)], ventricular hydrodynamic parameters [left ventricular flow rate in peak ejection (FRPE), flow reversal rate (FRR), flow reversal interval (FRI)], TGF-ß 1/Smad3 signaling pathway-related indicators (TGF-ß1, Smad3), quality of life score (SF-36 Quality of Life Scale) and occurrence of adverse reactions were compared between the two groups. Results: The main findings of the study are as follows: The observation group was significantly better than the control group in many aspects such as overall clinical effectiveness, cardiac function parameters, biochemical indicators, ventricular structure and function, TGF-ß1/Smad3 signaling pathway, and quality of life. Specifically, the observation group showed more significant positive effects in terms of improvement of cardiac function, adjustment of biochemical status, and adjustment of ventricular structure and fluid dynamics parameters. These results provide strong support for the application of new therapeutic approaches in the management of cardiovascular disease. After treatment, the total clinical effective rate in the observation group (89.19%) was significantly higher than that in the control group (69.44%) (P < .05). LVEF and SV in the two groups were significantly increased (P < .05), while LVEDD was significantly decreased (P < .05), and there were statistically significant differences in parameters between the two groups (P < .05). The levels of NT-proBNP, Gal-3 and PIIINP in both groups were significantly reduced (P < .05), and the levels in the observation group were significantly lower than those in the control group (P < .05). The PWD, PWS and IVSS in both groups significantly declined (P < .05), and the indicators in the observation group were significantly lower than those in the control group (P < .05). The FRPE and FRR in the two groups were significantly enhanced (P < .05), while the FRI was significantly reduced (P < .05), and the differences in the above parameters between the two groups were statistically significant (P < .05). The levels of TGF-ß1 and Smad3 in the two groups were significantly declined (P < .05), and the levels in the observation group were significantly lower than those in the control group (P < .05). During the period from before treatment to 6 months of treatment, the quality of life score in the two groups showed a significant downward trend (P < .05), and the score in the observation group after 3 months to 6 months of treatment was significantly lower than that in the control group (P < .05). During treatment, there was no statistical significance in the total incidence rate of adverse reactions between the two groups (P > .05). Conclusion: Early application of LCZ696 after AMI has a significant efficacy, and it can effectively improve the ventricular remodeling, regulate the expression levels of TGF-ß1 and Smad3, inhibit the TGF-ß1/Smad3 signaling pathway, promote the improvements of cardiac function and quality of life, and it has good safety and is worthy of clinical promotion and application. The study's key findings have important clinical implications for understanding and managing acute myocardial infarction (AMI). The observation group showed significant improvements in overall clinical efficacy, cardiac function, biochemical status, ventricular structure and function, etc., providing strong evidence for comprehensive treatment of AMI patients. This treatment method is expected to become an important part of the care and treatment strategy for AMI patients, help reduce cardiovascular risk, improve quality of life, and provide new research directions for future AMI treatment.

Concerted Enantioselective [2+2] Cycloaddition Reaction of Imines Mediated by a Magnesium Catalyst.

Enantioselective [2 + 2] cyclization between an imine and a carbon-carbon double bond is a versatile strategy to build chiral azetidines. However, α-branched allenoates have never been successfully applied in [2 + 2] cyclization reactions with imines, as they always undergo Kwon's [4 + 2] annulation in previous catalytic methods. Herein, a simple in situ generated magnesium catalyst was employed to successfully achieve the enantioselective [2 + 2] cyclization reaction of DPP-imines and α-branched allenoates for the first time. Insightful experiments including KIE experiments, controlled experiments, Hammett plot analysis, and 31P NMR studies of initial intermediates indicate that the current [2 + 2] cyclization of imine most likely involves an asynchronous concerted transition state. Further mechanistic investigations by combining kinetic studies, ESI experiments, 31P NMR studies of coordination complexes, and controlled experiments on reaction rates under different catalyst loading amounts provided the coordination details for this [2 + 2] cyclization reaction between DPP-imines and α-branched allenoates. This new approach was applied to the synthesis of various chiral aza-heterocycles, including the enantioselective synthesis of the key intermediate of a lipid-lowering agent Ezetimibe.

Detection and genetic analysis of porcine circovirus-like virus in pigs with diarrhea between 2016 and 2021 in Henan and Shanxi provinces of China.

Porcine circovirus-like virus (PCLV) is a recently discovered virus that may be associated with diarrhea in pigs. To investigate the epidemic profile and genetic characteristics of this virus, 175 clinical samples (141 intestinal samples, 17 blood samples, and 17 fecal samples) were collected from diseased piglets during outbreaks of diarrhea from 33 pig farms in 19 cities of Henan and Shanxi provinces of China between 2016 and 2021 and were screened by PCR for the presence of PCLV. The results showed that the positive rate for PCLV was 32% (56/175) at the sample level, 60.6% (20/33) at the farm level, and 57.9% (11/19) at the city level, which varied from 5.88% to 44.12% between 2016 and 2021. It was also found that PCLV occurred in coinfections with porcine circovirus type 2 (PCV2), PCV3, PCV4, porcine epidemic diarrhea virus, and porcine reproductive and respiratory syndrome virus, but no nucleic acids were detected for transmissible gastroenteritis virus, porcine deltacoronavirus, or porcine rotavirus in piglets with diarrhea. Notably, PCLV was detected in 13 diarrheal piglets from four different farms that were negative for the other porcine viruses. These findings suggest that PCLV may be associated with porcine diarrhea and that it has been circulating in piglets in Henan and Shanxi provinces of China. In addition, the complete genomes of 13 PCLV strains were sequenced and found to share 35.4%-91.0% nucleotide sequence identity with sequences available in the GenBank database. Phylogenetic analysis based on Rep amino acid sequences revealed that the 13 PCLV strains from this study clustered in group 1 and were closely related to eight Chinese PCLV strains, Bo-Circo-like virus CH, American strains 21 and 22, and Hungarian strains 288_4 and 302_4, but they differed genetically from seven other foreign PCLV strains. The whole genome and rep gene of 13 PCLV strains in this study were 72.2%-82% and 83.8%-89.7% identical, respectively, to those of Bo-Circo-like virus strain CH, indicating that PCLV is a novel virus in pigs that may be involved in cross-species transmission. Evidence of a recombination event was found in the rep region of the 13 PCLV strains sequenced. This study enriches the epidemiological data on PCLV infection in pigs in China and lays a foundation for further study on the pathogenesis of PCLV.

Simultaneous detection and phylogenetic analysis of porcine epidemic diarrhea virus and porcine circovirus 4 in Henan province, China.

Porcine circovirus 4 (PCV4) is a recently discovered circovirus that was first reported in 2019 in several pigs in Hunan province of China and has also been identified in pigs infected with porcine epidemic diarrhea virus (PEDV). To further investigate the coinfection and genetic diversity of these two viruses, 65 clinical samples (including feces and intestinal tissues) were collected from diseased piglets on 19 large-scale pig farms in Henan province of China, and a duplex SYBR Green I-based quantitative real-time polymerase chain reaction (qPCR) assay was developed for detecting PEDV and PCV4 simultaneously. The results showed that the limit of detection was 55.2 copies/µL and 44.1 copies/µL for PEDV and PCV4, respectively. The detection rate for PEDV and PCV4 was 40% (26/65) and 38% (25/65), respectively, and the coinfection rate for the two viruses was 34% (22/65). Subsequently, the full-length spike (S) gene of eight PEDV strains and a portion of the genome containing the capsid (Cap) gene of three PCV4 strains were sequenced and analyzed. Phylogenetic analysis showed that all of the PEDV strains from the present study clustered in the G2a subgroup and were closely related to most of the PEDV reference strains from China from 2011 to 2021, but they differed genetically from a vaccine strain (CV777), a Korean strain (virulent DR1), and two Chinese strains (SD-M and LZC). It is noteworthy that two PEDV strains (HEXX-24 and HNXX-24XIA) were identified in one sample, and the HNXX-24XIA strain had a large deletion at amino acids 31-229 of the S protein. Moreover, a recombination event was observed in strain HEXX-24. Phylogenetic analysis based on the amino acid sequence of the PCV4 Cap protein revealed that PCV4 strains were divided into three genotypes: PCV4a1, PCV4a2, and PCV4b. Three strains in the present study belonged to PCV4a1, and they had a high degree of sequence similarity (>98% identity) to other PCV4 reference strains. This study not only provides technical support for field investigation of PEDV and PCV4 coinfection but also provides data for their prevention and control.

Effect of Hydrophobic Chain Length in Amphiphilic Chitosan Conjugates on Intracellular Drug Delivery and Smart Drug Release of Redox-Responsive Micelle.

Three redox-sensitive nanocarriers were rationally designed based on amphiphilic low molecular weight chitosan-cystamine-octylamine/dodecylamin/cetylamine (LC-Cys-OA, LC-Cys-DA, LC-Cys-CA) conjugates containing disulfide linkage for maximizing therapeutic effect by regulating hydrophobic interaction. The resultant spherical micelles had the characteristics of low CMC, suitable size, excellent biosafety and desired stability. The drug-loaded micelles were fabricated by embedding doxorubicin (Dox) into the hydrophobic cores. The effect of hydrophobic chain lengths of amphiphilic conjugates on encapsulation capacity, redox sensitivity, trigger-release behavior, cellular uptake efficacy, antitumor effect and antimigratory activity of Dox-loaded micelles was systematically investigated. Studies found that Dox-loaded LC-Cys-CA micelle had superior loading capacity and enhanced redox sensitivity compared with the other two micelles. Release assay indicated that the three Dox-loaded micelles maintained sufficiently stability in normal blood circulation but rapidly disintegrated in tumor cells. More importantly, the LC-Cys-CA micelle with a longer hydrophobic chain length exhibited a higher accumulative Dox release percentage than the other two micelles. Additionally, an increase in hydrophobic chain lengths of amphiphilic conjugates improved cellular uptake efficiency, antitumor effect and antimigration activity of Dox-loaded micelles, which could be explained by enhanced loading ability and redox sensitivity. Our research was expected to provide a viable platform for achieving a desired therapeutic efficacy via the alteration of hydrophobic interaction.

Preparation and Antioxidant Activity of New Carboxymethyl Chitosan Derivatives Bearing Quinoline Groups.

A total of 16 novel carboxymethyl chitosan derivatives bearing quinoline groups in four classes were prepared by different synthetic methods. Their chemical structures were confirmed by Fourier-transform infrared spectroscopy (FTIR), nuclear magnetic resonance (NMR), and elemental analysis. The antioxidant experiment results in vitro (including DPPH radical scavenging ability, superoxide anion radical scavenging ability, hydroxyl radical scavenging ability, and ferric reducing antioxidant power) demonstrated that adding quinoline groups to chitosan (CS) and carboxymethyl chitosan (CMCS) enhanced the radical scavenging ability of CS and CMCS. Among them, both N, O-CMCS derivatives and N-TM-O-CMCS derivatives showed DPPH radical scavenging over 70%. In addition, their scavenging of superoxide anion radicals reached more than 90% at the maximum tested concentration of 1.6 mg/mL. Moreover, the cytotoxicity assay was carried out on L929 cells by the MTT method, and the results indicated that all derivatives showed no cytotoxicity (cell viability > 75%) except O-CMCS derivative 1a, which showed low cytotoxicity at 1000 µg/mL (cell viability 50.77 ± 4.67%). In conclusion, the carboxymethyl chitosan derivatives bearing quinoline groups showed remarkable antioxidant ability and weak cytotoxicity, highlighting their potential use in food and medical applications.

Recent Progress in Silicon-Based Materials for Performance-Enhanced Lithium-Ion Batteries.

Silicon (Si) has been considered to be one of the most promising anode materials for high energy density lithium-ion batteries (LIBs) due to its high theoretical capacity, low discharge platform, abundant raw materials and environmental friendliness. However, the large volume changes, unstable solid electrolyte interphase (SEI) formation during cycling and intrinsic low conductivity of Si hinder its practical applications. Various modification strategies have been widely developed to enhance the lithium storage properties of Si-based anodes, including cycling stability and rate capabilities. In this review, recent modification methods to suppress structural collapse and electric conductivity are summarized in terms of structural design, oxide complexing and Si alloys, etc. Moreover, other performance enhancement factors, such as pre-lithiation, surface engineering and binders are briefly discussed. The mechanisms behind the performance enhancement of various Si-based composites characterized by in/ex situ techniques are also reviewed. Finally, we briefly highlight the existing challenges and future development prospects of Si-based anode materials.

Discovery and pharmacodynamic evaluation of the novel butene lactone derivative M355 against influenza A virus in vitro and in vivo.

A new series of butene lactone derivatives were designed according to an influenza neuraminidase target and their antiviral activities against H1N1 infection of Madin-Darby canine kidney cells were evaluated. Among them, a compound that was given the name M355 was identified as the most potent against H1N1 (EC50 = 14.7 µM) with low toxicity (CC50 = 538.13 µM). It also visibly reduced the virus-induced cytopathic effect. Time-of-addition analysis indicated that H1N1 was mostly suppressed by M355 at the late stage of its infectious cycle. M355 inhibited neuraminidase in a dose-dependent fashion to a similar extent as oseltamivir, which was also indicated by a computer modeling experiment. In a mouse model, lung lesions and virus load were reduced and the expression of nucleoprotein was moderated by M355. The enzyme-linked immunosorbent assay and quantitative real-time polymerase chain reaction analyses revealed that the levels of interferon-γ, interferon regulatory factor-3, Toll-like receptor-3, tumor necrosis factor-α, interleukin (IL)-1ß, IL-6, and IL-8 were downregulated in the M355-treated groups, whereas the levels of IL-10 and IL-13 were upregulated. Similarly, IgG was found to be increased in infected mice plasma. These results demonstrate that M355 inhibit the expression of H1N1 in both cellular and animal models. Thus, M355 has the potential to be effective in the treatment of influenza A virus infection.

The role and mechanism of citrus flavonoids in cardiovascular diseases prevention and treatment.

Cardiovascular diseases (CVDs) have been ranked as the leading cause of death in the world, whose global incidence is increasing year by year. Citrus, one of the most popular fruits in the world, is rich in flavonoids. Citrus flavonoids attract special attention due to a variety of biological activities, especially in the prevention and treatment of CVDs. The research progress of citrus flavonoids on CVDs have been constantly updated, but relatively fragmented, which needed to be systematically summarized. Hence, the recent research about citrus flavonoids and CVDs were reviewed, including the types and in vivo processes of citrus flavonoids, epidemiology study and mechanism on prevention and treatment of CVDs by citrus flavonoids. This review would provide a theoretical basis for the citrus flavonoids research and a new idea in the citrus industry development and application.

Preparation of l-Arginine Schiff Bases Modified Chitosan Derivatives and Their Antimicrobial and Antioxidant Properties.

We successfully prepared a series of l-arginine Schiff bases acylated chitosan derivatives, aiming to improve the antioxidant activity and antimicrobial activity of chitosan by introducing a furan ring, pyridine ring, and l-arginine structure. The accuracy of the structures of ten compounds was characterized by FT-IR and 1H NMR. In terms of DPPH radical scavenging activity, except for compound CR3PCA, the scavenging rate of other compounds was higher than chitosan, especially CRCF and CRBF had strong scavenging abilities. At the same time, in the superoxide-radical scavenging activity assay, CRCF, CRBF, CR3PCA, CR2C3PCA, and CR2B3PCA were comparable to positive control at 1.60 mg/mL. Simultaneously, CRFF, CRCF, and CRBF had a certain inhibitory effect on Botrytis cinerea. Furthermore, the inhibitory effect of CRFF, CRCF, and CR3PCA on Staphylococcus aureus was very well, close to the positive control at 1.00 mg/mL. CRCF and CR2B3PCA showed better inhibitory effects on Escherichia coli than other compounds. The MTT assay was used to determine the cytotoxicity of the chitosan derivatives, which proved their safety to fibroblast cells. In summary, the study indicated that some of these compounds have the potential for further development and utilization in the preparation of antioxidants and antimicrobial agents.

Innovative Materials for Energy Storage and Conversion.

The metal chalcogenides (MCs) for sodium-ion batteries (SIBs) have gained increasing attention owing to their low cost and high theoretical capacity. However, the poor electrochemical stability and slow kinetic behaviors hinder its practical application as anodes for SIBs. Hence, various strategies have been used to solve the above problems, such as dimensions reduction, composition formation, doping functionalization, morphology control, coating encapsulation, electrolyte modification, etc. In this work, the recent progress of MCs as electrodes for SIBs has been comprehensively reviewed. Moreover, the summarization of metal chalcogenides contains the synthesis methods, modification strategies and corresponding basic reaction mechanisms of MCs with layered and non-layered structures. Finally, the challenges, potential solutions and future prospects of metal chalcogenides as SIBs anode materials are also proposed.

Conjugated ynones in catalytic enantioselective reactions.

Conjugated ynones are easily accessible feedstock and the existence of an alkyne bond endows ynones with different attractive reactivities, thus making them unique substrates for catalytic asymmetric reactions. Their compatibility under organocatalytic, metal-catalyzed as well as cooperative catalytic conditions has resulted in numerous enantioselective transformations. Importantly, conjugated ynones can act as nucleophiles or electrophiles, and serve as easily accessed synthons for different cyclization pathways. This review summarizes the recent literature examples of the catalytic reactions of conjugated ynones and related compounds such as alkyne conjugated α-ketoesters, and classifies these reaction types alongside mechanistic insights whenever possible. We aim to trigger more intensive research in the future to render the asymmetric transformation of ynones as a common and reliable tool for asymmetric synthesis.

Characterization of a recombinant pseudorabies virus expressing porcine parvovirus VP2 protein and porcine IL-6.

BACKGROUND: Porcine parvovirus (PPV) and pseudorabies virus (PRV) are the important etiological agents of swine infectious diseases, resulting in huge economic losses to the Chinese swine industry. Interleukin-6 (IL-6) has the roles to support host immune response to infections as a pleiotropic cytokine. It is essential to construct a live attenuated vaccine-based recombinant PRV that expresses PPV VP2 protein and porcine IL-6 for prevention and control of PRV and PPV. METHODS: The recombinant plasmid, pGVP2-IL6, was constructed by porcine IL-6 gene substituting for EGFP gene of the PRV transfer plasmid pGVP2-EGFP containing VP2 gene of PPV. Plasmid pGVP2-IL6 was transfected into swine testicle cells pre-infected with the virus rPRV-VP2-EGFP strain through homologous recombination and plaque purification to generate a recombinant virus rPRV-VP2-IL6. The recombinant PRV was further identified by PCR and DNA sequencing, and the expression of the VP2 protein and porcine IL-6 was analyzed by reverse transcription-PCR (RT-PCR) and Western blot. The virus titer was calculated according to Reed and Muench method. The immunogenicity of the recombinant virus was preliminarily evaluated in mice by intramuscular administration twice with the rPRV-VP2-IL6 at 4-week intervals. RESULTS: A recombinant virus rPRV-VP2-IL6 was successfully constructed and confirmed in this study. The properties of rPRV-VP2-IL6 were similar to the parental virus HB98 in terms of growth curve, morphogenesis and virus plaque sizes, and rPRV-VP2-IL6 was proliferated in different cell types. It induced specific antibodies against PPV as well as a strong increase of PPV-specific lymphocyte proliferation responses in mice immunized with rPRV-VP2-IL6, and provided partial protection against the virulent PPV challenge. rPRV-VP2-IL6 also induced a high level of neutralizing antibodies against PRV, and significantly reduced the mortality rate of (1 of 10) following virulent PRV challenge compared with the control (10 of 10). CONCLUSIONS: The recombinant rPRV-VP2-IL6 might be a potential candidate vaccine against PRV and PPV infections in pigs.

Construction and immunogenicity of a gE/gI/TK-deleted PRV based on porcine pseudorabies virus variant.

Pseudorabies (PR) caused by re-emerging pseudorabies virus (PRV) variant has outbroken among PRV vaccine-immunized swine herds on many Chinese pig farms, with severe socioeconomic consequences since late 2011. Here, a gE/gI/TK-deleted recombinant virus (rPRV NY-gE-/gI-/TK-) was constructed based on PRV NY strain from 2012 through homologous DNA recombination and gene-editing technology termed clustered regularly interspaced palindromic repeats (CRISPR)/associated (Cas9) system. The rPRV NY-gE-/gI-/TK- strain showed similar growth kinetics to the parental PRV NY strain in vitro, and was safe for mice. Sixty mice were injected subcutaneously (s.c.) twice with 106.0 TCID50 of rPRV NY-gE-/gI-/TK- and DMEM, respectively, with two-week interval. The levels of PRV gB antibodies and neutralizing antibodies against PRV NY in mice immunized with rPRV NY-gE-/gI-/TK- were higher than those in the DMEM control group. The number of T lymphocyte subclasses CD3+, CD4+ and CD8+ in rPRV NY-gE-/gI-/TK--immunized mice was higher than that in DMEM-injected mice. After challenge with 106.0 TCID50 PRV NY at 42 dpi, all rPRV NY-gE-/gI-/TK--immunized mice survived without exhibiting any pathological lesions in different tissues and intranuclear eosinophilic inclusions of the brain, and the viral genomic copy numbers in various organs of mice were obviously lower than DMEM group. These results showed the rPRV NY-gE-/gI-/TK- could be a promising next-generation vaccine to control now epidemic PR in China.

Cloning, expression profile of the complement component C9 gene and influence of the recombinant C9 protein on peripheral mononuclear leukocytes transcriptome in half-smooth tongue sole (Cynoglossus semilaevis).

The ninth complement component (C9) is a terminal complement component (TCC) that is involved in creating the membrane attack complex (MAC) on the target cell surface. In this study, the CsC9 (C9 of Cynoglossus semilaevis) cDNA sequence was cloned and characterized. The full-length CsC9 cDNA measured 2,150 bp, containing an open reading frame (ORF) of 1,803 bp, a 5'-untranslated region (UTR) of 24 bp and a 3'-UTR of 323 bp. A domain search revealed that the CsC9 protein contains five domains, including two TSP1s, an LDLRA, an EGF, and a MACPF. Quantitative real-time PCR analysis showed that CsC9 at the mRNA level was expressed in all the tested tissues, with the highest expression being observed in the liver. CsC9 expression is significantly upregulated in the tested tissues after challenge with Vibrio anguillarum. To further characterize the role of CsC9, peripheral blood mononuclear cells of C. semilaevis were used for transcriptome analysis after incubation with recombinant CsC9 (rCsC9) protein. A total of 3,775 significant differentially expressed genes (DEGs) were identified between the control and the rCsC9-treated group, including 2,063 upregulated genes and 1,712 downregulated genes. KEGG analyses revealed that the DEGs were enriched in cell adhesion molecules, cytokine-cytokine receptor interactions, T cell receptor signaling pathways, B cell receptor signaling pathways and Toll-like receptor signaling pathways. The results of this study indicate that in addition to participating in MAC formation, CsC9 might play multiple roles in the innate and adaptive immunity of C. semilaevis.

Expression, location and biological effects of four and a half LIM domain protein 2 (FHL2) on granulosa cells in ovine.

Previous studies have shown that four and a half LIM domain protein 2 (FHL2) plays an essential role in the regulation of follicular development in mammals. Although the FHL2 genes of human and mouse have been well characterized, the expression and location of FHL2 in ovary and the biological functions of FHL2 on granulosa cells (GCs) of ovine are still not clear. In this study, full-length complementary DNA (cDNA) of FHL2 from ovine follicular GCs was amplified by real-time PCR (RT-PCR). The expression and location of FHL2 in ovary and GCs of ovine were studied by immunohistochemistry and immunofluorescence, and the biological effects of FHL2 on the cell proliferation, cell apoptosis, cell cycles and expression level of related genes of ovine GCs were also explored by overexpression or knockdown of FHL2. The results indicated that FHL2 was expressed in ovine follicular GCs and the sequence of the FHL2 cDNA was consistent with that predicted in GenBank, which did not cause an amino acid change. According to the results, FHL2 was expressed in ovine ovary and mainly located in the cytoplasm and nucleus of GCs. In addition, overexpression of FHL2 significantly reduced the cell viability, promoted the cell apoptosis and decreased the percentage of G0/G1 and S phase cells. RT-PCR showed that overexpression of FHL2 significantly increased the mRNA expression level of Bax and decreased the expression of Bcl-2 and the Bcl-2/Bax mRNA ratio compared with the control group. Besides, the knockdown of FHL2 gene in ovine GCs significantly improved the cell viability, suppressed the cell apoptosis, decreased the mRNA expression level of Caspase-3 gene, increased the Bcl-2/Bax mRNA ratio and increased the percentage of S and G2/M phase cells. Our results suggest that FHL2 may play an important role in the biological functions of GCs in ovine.

Direct Synthesis of a Dioxabenzobicyclo[3.2.1]octane Core from Salicylaldehydes.

A copper(II) trifluoromethanesulfonate via Prins reaction between salicylaldehydes and allyl alcohols is realized under mild conditions. A series of compounds containing dioxabenzobicyclo[3.2.1]octane motifs are obtained in moderate to good yields. The current method can efficiently construct the valuable structural motif from easily accessed starting materials, providing a smooth method to build benzobicyclo skeletons.