The global succinylation of SARS-CoV-2-infected host cells reveals drug targets.

SARS-CoV-2, the causative agent of the COVID-19 pandemic, undergoes continuous evolution, highlighting an urgent need for development of novel antiviral therapies. Here we show a quantitative mass spectrometry-based succinylproteomics analysis of SARS-CoV-2 infection in Caco-2 cells, revealing dramatic reshape of succinylation on host and viral proteins. SARS-CoV-2 infection promotes succinylation of several key enzymes in the TCA, leading to inhibition of cellular metabolic pathways. We demonstrated that host protein succinylation is regulated by viral nonstructural protein (NSP14) through interaction with sirtuin 5 (SIRT5); overexpressed SIRT5 can effectively inhibit virus replication. We found succinylation inhibitors possess significant antiviral effects. We also found that SARS-CoV-2 nucleocapsid and membrane proteins underwent succinylation modification, which was conserved in SARS-CoV-2 and its variants. Collectively, our results uncover a regulatory mechanism of host protein posttranslational modification and cellular pathways mediated by SARS-CoV-2, which may become antiviral drug targets against COVID-19.

Identification of a Novel Interferon Lambda Splice Variant in Chickens.

Type III interferons (IFNLs) have critical roles in the host's innate immune system, also serving as the first line against pathogenic infections of mucosal surfaces. In mammals, several IFNLs have been reported; however, only limited data on the repertoire of IFNLs in avian species is available. Previous studies showed only one member in chicken (chIFNL3). Herein, we identified a novel chicken IFNL for the first time, termed chIFNL3a, which contains 354 bp, and encodes 118 amino acids. The predicted protein is 57.1% amino acid identity with chIFNL. Genetic, evolutionary, and sequence analyses indicated that the new open reading frame (ORF) groups with type III chicken IFNs represent a novel splice variant. Compared to IFNs from different species, the new ORF is clustered within the type III IFNs group. Further study showed that chIFNL3a could activate a panel of IFN-regulated genes and function mediated by the IFNL receptor, and chIFNL3a markedly inhibited the replication of Newcastle disease virus (NDV) and influenza virus in vitro. These data collectively shed light on the repertoire of IFNs in avian species and provide useful information that further elucidate the interaction of the chIFNLs and viral infection of poultry. IMPORTANCE Interferons (IFNs) are critical soluble factors in the immune system, and are composed of 3 types (I, II, and III) that utilize different receptor complexes (IFN-αR1/IFN-αR2, IFN-γR1/IFN-γR2, and IFN-λR1/IL-10R2, respectively). Herein, we identified IFNL from the genomic sequences of chicken and termed it chIFNL3a, located on chromosome 7 of chicken. Phylogenetically clustered with all known types of chicken IFNs, the finding of this IFN is considered a type III IFN. To further evaluate the biological properties of chIFNL3a, the target protein was prepared by the baculovirus expression system (BES), which could markedly inhibit the replication of NDV and influenza viruses. In this study, we uncovered a new interferon lambda splice variant of chicken, termed chIFNL3a, which could inhibit viral replication in cells. Importantly, these novel findings may extend to other viruses, offering a new direction for therapeutic interventions.

Constructing a Ready-to-Use mRNA Vaccine Delivery System for the Prevention of Influenza A virus, Utilizing FDA-Approved Raw Materials.

Messenger ribonucleic acid (mRNA) vaccines, serving as a rapid and easily scalable emergency preventive measure, have played a pivotal role in preventing infectious diseases. The effectiveness of mRNA vaccines heavily relies on the delivery carrier, but the current market options are predominantly lipid nanoparticles. Their intricate preparation process and high transportation costs pose challenges for widespread use in remote areas. In this study, we harnessed FDA-approved polymer PLGA and lipid components widely employed in clinical experiments to craft a ready-to-use mRNA vaccine delivery system known as lipid-polymer hybrid nanoparticles (LPP). Following formulation optimization, the PDCD nanoparticles emerged as the most effective, showcasing exceptional mRNA delivery capabilities both in vitro and in vivo. Loading PDCD nanoparticles with mRNA encoding the H1N1 influenza virus HA antigen-fused M2e peptide enabled the successful induction of M2e-specific antibodies and T cell immune responses in immunized mice. After three rounds of vaccine immunization, the mice demonstrated weight recovery to normal levels and maintained a survival rate exceeding 80% following an encounter with the H1N1 influenza virus. The innovative mRNA delivery system that we designed demonstrates outstanding effectiveness in preventing infectious diseases, with the potential to play an even more significant role in future clinical applications.

Integrating Magnetic-Bead-Based Sample Extraction and Molecular Barcoding for the One-Step Pooled RT-qPCR Assay of Viral Pathogens without Retesting.

Pooling multiple samples prior to real-time reverse-transcription polymerase chain reaction (RT-PCR) analysis has been proposed as a strategy to minimize expenses and boost test throughput during the COVID-19 pandemic. Nevertheless, the traditional pooling approach cannot be effectively deployed in high-prevalence settings due to the need for secondary tests in the case of a positive pool. In this study, we present a pooling test platform with high adaptability and simplicity that allows sample-specific detection of multiple-tagged samples in a single run without the need for retesting. This was accomplished by labeling distinct samples with predefined ID-Primers and identifying tagged pooled samples using one-step RT-PCR followed by melting curve analysis with rationally designed universal fluorescence- and quencher-tagged oligo probes. Using magnetic beads (MBs), nucleic acid targets from different individuals can be tagged and extracted concurrently and then pooled before RT, eliminating the need for extra RNA extraction and separate RT and enzyme digestion steps in the recently developed barcoding strategies. Pools of six samples (positive and negative) were successfully identified by melting temperature values under two fluorescent channels, with a detection sensitivity of 5 copies/µL. We validated the reproducibility of this assay by running it on 40 clinical samples with a hypothetical infection rate of 15%. In addition, to aid the scenario of large-scale pooling tests, we constructed a melting curve autoreadout system (MCARS) for statistical analysis of melting curve plots to eliminate error-prone manual result readout. Our results suggest that this strategy could be a simple and adaptable tool for alleviating existing bottlenecks in diagnostic pooling testing.

Efficient large-scale screening of viral pathogens by fragment length identification of pooled nucleic acid samples (FLIPNAS).

The necessity for the large-scale screening of viral pathogens has been amply demonstrated during the COVID-19 pandemic. During this time, SARS-CoV-2 nucleic acid pooled testing, such as Dorfman-based group testing, was widely adopted in response to the sudden increased demand for detection. However, the current approach still necessitates the individual retesting of positive pools. Here, we established an efficient method termed the fragment-length identification of pooled nucleic acid samples (FLIPNAS), where all subsamples (n = 8) can be uniquely labelled and tested in a single-time detection among pools of samples. We used a novel and simple design of unique primers (UPs) to generate amplicons of unique lengths after reverse transcription and polymerase chain reaction to reach this aim. As a result, the unique lengths of the amplicons can be recognized and traced back to the corresponding UPs and specific samples. Our results demonstrated that FLIPNAS could recognize one to eight positive subsamples in a single test without retesting positive pools. The system also showed sufficient sensitivity for the mass monitoring of SARS-CoV-2 and no cross-reactivity against three common respiratory diseases. Moreover, the FLIPNAS results of 40 samples with a positive ratio of 7.8% were in 100% agreement with their individual detection results using the gold standard. Collectively, this study shows that the efficiency of nucleic acid pooling detection can be further improved by FLIPNAS, which can speed up testing and mitigate the urgent demand for resources.

Study on Related Factors of the Treatment Zone After Wearing Paragon CRT and Euclid Orthokeratology Lenses.

OBJECTIVE: To explore the influence factors of the treatment zone diameter (TZD) and its relationship with axial length growth (ALG) after wearing Paragon CRT and Euclid orthokeratology lenses. METHODS: The right eye data of myopic patients wearing Paragon CRT and Euclid orthokeratology in the ophthalmology department of The First Affiliated Hospital of Soochow University were retrospectively reviewed from April 2019 to October 2022. The TZD and ALG were compared between the Paragon CRT and Euclid groups. The correlation factors of TZD after wearing lens for 1 month and the relationship between the overlapping treatment zone-to-pupil area ratio and the ALG after wearing lens for 1 year were analyzed between the two groups. RESULTS: There were 160 patients (160 eyes) in the Paragon CRT group and 155 patients (155 eyes) in the Euclid group. After wearing lens for 1 month, the TZD in the Paragon CRT group (3.72±0.37 mm) was larger than that in the Euclid group (3.26±0.37 mm) ( P <0.001). The stepwise multivariate linear regression analysis showed that the eccentricity at the flattest meridians (Em) and the central corneal thickness were correlated with the TZD in both groups ( P <0.05). After wearing lens for 1 year, the ALG in the Paragon CRT group (0.32±0.20 mm) was larger than that in the Euclid group (0.25±0.20 mm) ( P =0.001). The stepwise multivariate linear regression analysis showed that the initial wearing age and the overlapping treatment zone area-to-pupil area ratio were correlated with the ALG in both groups ( P <0.05). CONCLUSION: For both the Paragon CRT and Euclid orthokeratology, the wearers with thicker central corneal thickness and smaller Em usually had a smaller TZD. In both groups, the overlapping treatment zone area-to-pupil area ratio was correlated with the ALG.

Unique Barcoded Primer-Assisted Sample-Specific Pooled Testing (Uni-Pool) for Large-Scale Screening of Viral Pathogens.

Pooled testing has been widely adopted recently to facilitate large-scale community testing during the COVID-19 pandemic. This strategy allows to collect and screen multiple specimen samples in a single test, thus immensely saving the assay time and consumable expenses. Nevertheless, when the outcome of a pooled testing is positive, it necessitates repetitive retesting steps for each sample which can pose a serious challenge during a rising infection wave of increasing prevalence. In this work, we develop a unique barcoded primer-assisted sample-specific pooled testing strategy (Uni-Pool) where the key genetic sequences of the viral pathogen in a crude sample are extracted and amplified with concurrent tagging of sample-specific identifiers. This new process improves the existing pooled testing by eliminating the need for retesting and allowing the test results-positive or negative-for all samples in the pool to be revealed by multiplex melting curve analysis right after real-time polymerase chain reaction. It significantly reduces the total assay time for large-scale screening without compromising the specificity and detection sensitivity caused by the sample dilution of pooling. Our method was able to successfully differentiate five samples, positive and negative, in one pool with negligible cross-reactivity among the positive and negative samples. A pooling of 40 simulated samples containing severe acute respiratory syndrome coronavirus-2 pseudovirus of different loads (min: 10 copies/µL; max: 103 copies/µL) spiked into artificial saliva was demonstrated in eight randomized pools. The outcome of five samples in one pool with a hypothetical infection prevalence of 15% in 40 samples was successfully tested and validated by a typical Dorman-based pooling.

Two-Dimensional Cobalt-Doped Ti3C2 MXene Nanozyme-Mediated Homogeneous Electrochemical Strategy for Pesticides Assay Based on In Situ Generation of Electroactive Substances.

Common homogeneous electrochemical (HEC) sensors usually suffer from the drawbacks of high background signal, low signal-to-noise ratio, and even false positive results due to the preaddition of electroactive substances. Thus, it is necessary to develop novel HEC sensors based on in situ generation of electroactive substances to overcome these shortcomings, which, however, is underexplored. In this work, two-dimensional (2D) nanozymes, i.e., cobalt-doped 2D Ti3C2 MXene nanosheets (CMNSs), with excellent peroxidase-like properties were utilized to develop HEC sensors based on the in situ generation of electroactive substances for organophosphate pesticides (OPs) detection. The 2D CMNSs were synthesized via a template-directed wet chemical approach and displayed outstanding features of hydrophilia and water dispersibility, which could catalyze the oxidation of o-phenylenediamine (OPD) to generate significantly increased reduction current. Interestingly, the 2D CMNSs with peroxidase-like properties exhibited a unique response to thiol compounds and were thus employed as highly efficient catalysts to develop HEC sensors for OPs based on the hydrolysis of acetylthiocholine (ATCh) to form thiocholine catalyzed by acetylcholinesterase (AChE) and the inhibition of AChE activity by OPs. The recovery for OPs analysis of pakchoi extract solutions ranged from 97.4% to 103.3%. The as-proposed HEC sensor based on in situ generation of electroactive substances will provide a new way for the development of high-performance electrochemical sensors and demonstrate potential applicability for the determination of pesticide residues in real samples.

Inhibition of Autophagy Suppresses SARS-CoV-2 Replication and Ameliorates Pneumonia in hACE2 Transgenic Mice and Xenografted Human Lung Tissues.

Autophagy is thought to be involved in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. However, how SARS-CoV-2 interferes with the autophagic pathway and whether autophagy contributes to virus infection in vivo is unclear. In this study, we identified SARS-CoV-2-triggered autophagy in animal models, including the long-tailed or crab-eating macaque (Macaca fascicularis), human angiotensin-converting enzyme 2 (hACE2) transgenic mice, and xenografted human lung tissues. In Vero E6 and Huh-7 cells, SARS-CoV-2 induces autophagosome formation, accompanied by consistent autophagic events, including inhibition of the Akt-mTOR pathway and activation of the ULK-1-Atg13 and VPS34-VPS15-Beclin1 complexes, but it blocks autophagosome-lysosome fusion. Modulation of autophagic elements, including the VPS34 complex and Atg14, but not Atg5, inhibits SARS-CoV-2 replication. Moreover, this study represents the first to demonstrate that the mouse bearing xenografted human lung tissue is a suitable model for SARS-CoV-2 infection and that autophagy inhibition suppresses SARS-CoV-2 replication and ameliorates virus-associated pneumonia in human lung tissues. We also observed a critical role of autophagy in SARS-CoV-2 infection in an hACE2 transgenic mouse model. This study, therefore, gives insights into the mechanisms by which SARS-CoV-2 manipulates autophagosome formation, and we suggest that autophagy-inhibiting agents might be useful as therapeutic agents against SARS-CoV-2 infection. IMPORTANCE Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) caused a global pandemic with limited therapeutics. Insights into the virus-host interactions contribute substantially to the development of anti-SARS-CoV-2 therapeutics. The novelty of this study is the use of a new animal model: mice xenografted with human lung tissues. Using a combination of in vitro and in vivo studies, we have obtained experimental evidence that induction of autophagy contributes to SARS-CoV-2 infection and improves our understanding of potential therapeutic targets for SARS-CoV-2.

Immunogenicity and protective efficacy of a DNA vaccine inducing optimal expression of the SARS-CoV-2 S gene in hACE2 mice.

The wide spread of coronavirus disease 2019 (COVID-19) has significantly threatened public health. Human herd immunity induced by vaccination is essential to fight the epidemic. Therefore, highly immunogenic and safe vaccines are necessary to control SARS-CoV-2, whose S protein is the antigenic determinant responsible for eliciting antibodies that prevent viral entry and fusion. In this study, we developed a SARS-CoV-2 DNA vaccine expressing the S protein, named pVAX-S-OP, which was optimized according to the human-origin codon preference and using polyinosinic-polycytidylic acid as an adjuvant. pVAX-S-OP induced specific antibodies and neutralizing antibodies in BALB/c and hACE2 transgenic mice. Furthermore, we observed 1.43-fold higher antibody titers in mice receiving pVAX-S-OP plus adjuvant than in those receiving pVAX-S-OP alone. Interferon gamma production in the pVAX-S-OP-immunized group was 1.58 times (CD3+CD4+IFN-gamma+) and 2.29 times (CD3+CD8+IFN-gamma+) lower than that in the pVAX-S-OP plus adjuvant group but higher than that in the control group. The pVAX-S-OP vaccine was also observed to stimulate a Th1-type immune response. When, hACE2 transgenic mice were challenged with SARS-CoV-2, qPCR detection of N and E genes showed that the viral RNA loads in pVAX-S-OP-immunized mice lung tissues were 104 times and 106 times lower than those of the PBS control group, which shows that the vaccine could reduce the amount of live virus in the lungs of hACE2 mice. In addition, pathological sections showed less lung damage in the pVAX-S-OP-immunized group. Taken together, our results demonstrated that pVAX-S-OP has significant immunogenicity, which provides support for developing SARS-CoV-2 DNA candidate vaccines.

Experiment and numerical investigation of inhalable particles and indoor environment with ventilation system.

After the outbreak of COVID-19, the indoor environment has become particularly important in closed spaces, being a common concern in environmental science and public health, and of great significance for the building environment. To improve the indoor air quality and control the spread of viruses, the analysis of inhalable particles in indoor environments is critical. In this research, we study standards focused on inhalable particles and indoor environmental quality, as well as analyzing the movement and diffusion of indoor particles. Based on our analysis, we conduct an experimental study to determine the distribution of indoor inhalable particles of different sizes before and after diffusion under the conditions of underfloor air distribution. Furthermore, the mathematical modeling method is adopted to simulate the indoor flow field, particle trajectories, and pollutant dispersion process. The k-ε two-equation model is applied as the turbulence model in the numerical simulation, while the Lagrangian discrete phase model is adopted to trace the motion of particles and analyze the distribution characteristics of indoor particles. The results demonstrate that fine particles (i.e., those with size less than 0.5 µm) have a significant impact on the indoor particle concentration, while coarse particles (i.e., with size above 2.5 µm) have a greater influence on the total mass concentration of indoor particles. Small-sized particles can easily follow the airflow and diffuse to upper parts of the room. Overall, the effects of indoor particles on indoor air quality, including the potential threat of aerosol transmission of respiratory infectious diseases, are non-negligible. Application of the presented research can contribute to improving the health-related aspects of the building environment.

Inhibitors of endosomal acidification suppress SARS-CoV-2 replication and relieve viral pneumonia in hACE2 transgenic mice.

BACKGROUND: Coronavirus disease 2019 (COVID-19) is caused by SARS-CoV-2 and broke out as a global pandemic in late 2019. The acidic pH environment of endosomes is believed to be essential for SARS-CoV-2 to be able to enter cells and begin replication. However, the clinical use of endosomal acidification inhibitors, typically chloroquine, has been controversial with this respect. METHODS: In this study, RT-qPCR method was used to detect the SARS-CoV-2N gene to evaluate viral replication. The CCK-8 assay was also used to evaluate the cytotoxic effect of SARS-CoV-2. In situ hybridization was used to examine the distribution of the SARS-CoV-2 gene in lung tissues. Hematoxylin and eosin staining was also used to evaluate virus-associated pathological changes in lung tissues. RESULTS: In this study, analysis showed that endosomal acidification inhibitors, including chloroquine, bafilomycin A1 and NH4CL, significantly reduced the viral yields of SARS-CoV-2 in Vero E6, Huh-7 and 293T-ACE2 cells. Chloroquine and bafilomycin A1 also improved the viability and proliferation of Vero E6 cells after SARS-CoV-2 infection. Moreover, in the hACE2 transgenic mice model of SARS-CoV-2 infection, chloroquine and bafilomycin A1 reduced viral replication in lung tissues and alleviated viral pneumonia with reduced inflammatory exudation and infiltration in peribronchiolar and perivascular tissues, as well as improved structures of alveolar septum and pulmonary alveoli. CONCLUSIONS: Our research investigated the antiviral effects of endosomal acidification inhibitors against SARS-CoV-2 in several infection models and provides an experimental basis for further mechanistic studies and drug development.

Retrospective surveillance of porcine circovirus 4 in pigs in Inner Mongolia, China, from 2016 to 2018.

A novel circovirus designated "porcine circovirus type 4" (PCV4) was recently reported in pigs with severe clinical disease in Hunan Province, China. Relatively little is known about the molecular epidemiology of this recently discovered virus. In order to assess the prevalence of PCV4 infection in pigs and to analyze its genomic characteristics, 1683 clinical samples were collected in Inner Mongolia, China, from 2016 to 2018. The overall infection rate of PCV4 was 1.6% (27/1683) at the sample level and 21.6% (11/51) at the farm level, with rates ranging from 3.2% (1/31) to 20.0% (6/30) on different PCV4-positive pig farms. In addition, the PCV4 infection rates at both the sample and farm level increased from 2016 to 2018. This also showed that PCV4 was present in pigs in 2016 in China and therefore did not arrive later than this date. Additionally, our findings showed that PCV4 infections had no association with PCV2 or PCV3 infections. We sequenced the complete genomes of three PCV4 strains and found that the PCV4 strains had a high degree of genetic stability but shared less than 80% sequence identity with other circoviruses. We identified six amino acid mutations in the Rep protein and seven in the Cap protein. Phylogenetic analysis based on Cap and Rep sequences confirmed that the PCV4 strains grouped in an independent branch. Our findings provide important information about the prevalence and genetic characteristics of PCV4 strains.

Characterization of porcine reproductive and respiratory syndrome virus (ORF5 RFLP 1-7-4 viruses) in northern China.

The porcine reproductive and respiratory syndrome virus (PRRSV) is the causative agent of porcine reproductive and respiratory syndrome (PRRS). Disease outbreaks caused by NADC30-like PRRSV strains were a bit prevalent in China in recent years. In the present study, two newly emerged PRRSV strains, which were designated as PRRSV-ZDXYL-China-2018-1 and PRRSV-ZDXYL-China-2018-2 strains were found from piglets' lung tissues in Northern China. The virus belongs to lineage 1 of the PRRSV genotype 2 and is closely related to US strains that possess the open reading frame (ORF5) restriction fragment length polymorphism (RFLP) 1-7-4. The two strains were identified from infected weaning piglet herds in Zhaodong City, Heilongjiang province of China. The complete genome of the PRRSV-ZDXYL-China-2018-1 and PRRSV-ZDXYL-China-2018-2 strains were 15093 nt and 15110 nt, and shared 96.7%-97.0% and 97.1%-97.4% similarities with the US identified, ISU10 and NADC34 strains respectively. Then the PRRSV-ZDXYL-China-2018-1 strain was successfully isolated from the clinical sample. Our results demonstrate, that the emergence of ORF5 RFLP 1-7-4-like PRRSVs in China, could pose a significant challenge to PRRSV epidemic prevention.

Inhibition of TRPA1 Promotes Cardiac Repair in Mice After Myocardial Infarction.

Recent studies have shown that TRPA1, a nonselective cation channel with high permeability to calcium, is expressed in many tissues of the cardiovascular system and is involved in the pathogenesis of many cardiovascular diseases. However, the role of TRPA1 in cardiac repair after myocardial infarction (MI) has not been clearly defined. The aim of this study was to confirm whether inhibition of TRPA1 could attenuate MI-induced cardiac ischemia injury. The C57BL/6 mice were subjected to ligation of the left anterior descending coronary artery and treated with TRPA1-specific inhibitor HC-030031 (HC) for 4 weeks. Echocardiography was performed to assess cardiac function. The results showed that HC significantly attenuated MI-induced cardiac dysfunction 4 weeks after MI. Similarly, HC reduced cardiac fibrosis and cell apoptosis after MI and significantly increased angiogenesis in the border zone of the infarct. In vitro, we found that HC promoted the proliferation and migration of human umbilical vein endothelial cells (HUVECs). Importantly, HC treatment decreased phosphatase and tensin homolog expression and augmented the expression of phosphorylated Akt in the myocardium post MI and HUVECs. However, treatment of HUVECs with a PI3K inhibitor, LY294002, before HC administration almost completely abolished HC-induced migration in HUVECs. In conclusion, we demonstrate that the inhibition of TRPA1 promotes angiogenesis after MI, thereby alleviating myocardial ischemia injury via mechanisms involving inhibition of phosphatase and tensin homolog expression and subsequent activation of the PI3K/Akt signaling.

Functional Annotations of Single-Nucleotide Polymorphism (SNP)-Based and Gene-Based Genome-Wide Association Studies Show Genes Affecting Keratitis Susceptibility.

BACKGROUND Keratitis is a complex condition in humans and is the second most common cause of legal blindness worldwide. MATERIAL AND METHODS To reveal the genomic loci underlying keratitis, we performed functional annotations of SNP-based and gene-based genome-wide association studies of keratitis in the UK Biobank (UKB) cohort with 337 199 subjects of European ancestry. RESULTS The publicly available SNP-based association results showed a total of 34 SNPs, from 14 distinct loci, associated with keratitis in the UKB. Gene-based association analysis identified 2 significant genes: IQCF3 (p=2.0×10⁻6) and SOD3 (p=2.0×10⁻6). Thirty-two candidate genes were then prioritized using information from multiple sources. The overlap of IQCF3 in these 2 analyses resulted in a total of 33 hub genes. Functional annotation of hub genes was performed and transcriptional factors of IQCF3 and SOD3 were predicted. CONCLUSIONS A total of 34 SNPs from 14 distinct loci were identified as being associated with keratitis, and 32 candidate genes were then prioritized. In addition, IQCF3 and SOD3 were identified by their p values through gene-based tests on the basis of individual SNP-based tests. The functional relationship between these suspect genes and keratitis suggest that IQCF3 and SOD3 are candidate genes underlying keratitis.

miR-221-3p inhibits oxidized low-density lipoprotein induced oxidative stress and apoptosis via targeting a disintegrin and metalloprotease-22.

Oxidized low-density lipoprotein (ox-LDL)-induced oxidative stress and apoptosis are considered as a critical contributor to atherosclerosis. MicroRNAs (miRNAs) have been reported versatile functions in all biological processes via directly suppressing target messenger RNA at a posttranscriptional level. Although miRNA-221 has been implied to be involved in the regulation of atherosclerosis, the underlying mechanism remains unclear. Here, we showed that ox-LDL treatment remarkably suppressed the expression of miR-221-3p in a concentration-dependent and time-dependent manner. Transfection of miR-221-3p mimic significantly reduced the foam cell formation and expression of lipid biomarkers, while transfection of the miR-221-3p inhibitor showed completely opposite effects. Moreover, miR-221-3p was also found to inhibit the process of cell apoptosis in macrophages. A disintegrin and metalloprotease-22 (ADAM22) is predicted as a direct target of miR-221-3p, and silencing AMAM22 resulted in a reduced foam cell formation and cell apoptosis. Furthermore, silencing AMAM22 restored the stimulatory effect of the miR-221-3p inhibitor in ox-LDL-induced foam cell formation and apoptosis. These findings suggest that miR-221-3p inhibits ox-LDL and apoptosis via directly targeting ADAM22.

Photoresponsive Self-Assembled DNA Nanomaterials: Design, Working Principles, and Applications.

Photoresponsive DNA nanomaterials represent a new class of remarkable functional materials. By adjusting the irradiation wavelength, light intensity, and exposure time, various photocontrolled DNA-based systems can be reversibly or irreversibly regulated in respect of their size, shape, conformation, movement, and dissociation/association. This Review introduces the most updated progress in the development of photoresponsive DNA-based system and emphasizes their advantages over other stimuli-responsive systems. Their design and mechanisms to trigger the photoresponses are shown and discussed. The potential application of these photon-responsive DNA nanomaterials in biology, biomedicine, materials science, nanophotonic and nanoelectronic are also covered and described. The challenges faced and further directions of the development of photocontrolled DNA-based systems are also highlighted.

An epidemiological investigation of porcine circovirus 3 infection in cattle in Shandong province, China.

BACKGROUND: Porcine circovirus type 3 (PCV3) is a single-stranded, closed circular DNA virus, which causes porcine dermatitis and nephropathy syndrome (PDNS), multisystemic inflammation, and reproductive failure. The present study aimed to investigate the seroprevalence of PCV3 in cattle (Bos taurus) in Shandong province, China, and examine its genome diversity. RESULTS: PCR amplification and sequencing showed that 74 of 213 bovine samples (34.7%) were positive for PCV3. Among them, the capsid gene (n = 12) and the complete genome (n = 4) were sequenced. These sequences had high identities to the reference capsid gene (98.0-100%) and the complete genome (97.5-99.8%). The PCV3 strains were classified into two different genotypes (PCV3a and PCV3b), according to phylogenetic analysis based on the complete genome and capsid gene sequences. Specifically, the bovine-origin strains in this study were grouped into PCV3a, showing a close relationship with PCV3-US/SD2016 (American strain; GenBank: KX966193.1). Notably, a comparison of the inferred amino acid sequences revealed a mutation from D124 to Y124. CONCLUSION: This was the first seroprevalence and genetic investigation of PCV3 in cattle in Shandong province, China. The results could provide insights into the epidemiology and pathogenesis of this important virus.

The Impact of Circulating Mitochondrial DNA on Cardiomyocyte Apoptosis and Myocardial Injury After TLR4 Activation in Experimental Autoimmune Myocarditis.

BACKGROUND/AIMS: Mitochondrial DNA (mtDNA), acting as a newly found 'danger-associated molecular patterns' (DAMPs), is released into circulation upon tissue injury and performs as a considerable activator of inflammation and immune response. However, the role of circulating mtDNA in experimental autoimmune myocarditis (EAM) as well as Toll like receptor4 (TLR4) mediated cardiac inflammation and injury remains unknown. METHODS: A model of EAM was established in BALB/c mice by immunization with porcine cardiac myosin. Lipopolysaccharide (LPS) was used to stimulate TLR4 activation in EAM mice and H9C2 cells. RESULTS: LPS stimulation significantly aggravated cardiac inflammation and tissue injury in EAM, as demonstrated by increased myocardium inflammatory cell infiltration, and up-regulated inflammatory cytokines and troponin I(TnI) level in serum. Circulating mtDNA level was increased in EAM and TLR4 activation led to a greater elevation, which may be related to Reactive oxygen species (ROS) stress involved mtDNA damage characterized by reduced mtDNA copy number in myocardium tissue. In addition, the expression of Toll like receptor9 (TLR9), a ligand of mtDNA, was significantly up-regulated in the myocardium of EAM and EAM LPS group; meanwhile, TLR9 inhibition by ODN 2088 caused an inhibited apoptosis in LPS treated H9C2 cells. Moreover, in EAM and EAM LPS group, simultaneously giving ODN 2088 treatment significantly ameliorated cardiac inflammation and tissue injury compared with untreated group. CONCLUSION: Increased circulating mtDNA combined with upregulated TLR9 expression may corporately play a role in EAM as well as TLR4 activation mediated cardiac inflammation and injury.