African swine fever virus pB318L, a trans-geranylgeranyl-diphosphate synthase, negatively regulates cGAS-STING and IFNAR-JAK-STAT signaling pathways.

African swine fever (ASF) is an acute, hemorrhagic, and severe infectious disease caused by the ASF virus (ASFV). ASFV has evolved multiple strategies to escape host antiviral immune responses. Here, we reported that ASFV pB318L, a trans-geranylgeranyl-diphosphate synthase, reduced the expression of type I interferon (IFN-I) and IFN-stimulated genes (ISGs). Mechanically, pB318L not only interacted with STING to reduce the translocation of STING from the endoplasmic reticulum to the Golgi apparatus but also interacted with IFN receptors to reduce the interaction of IFNAR1/TYK2 and IFNAR2/JAK1. Of note, ASFV with interruption of B318L gene (ASFV-intB318L) infected PAMs produces more IFN-I and ISGs than that in PAMs infected with its parental ASFV HLJ/18 at the late stage of infection. Consistently, the pathogenicity of ASFV-intB318L is attenuated in piglets compared with its parental virus. Taken together, our data reveal that B318L gene may partially affect ASFV pathogenicity by reducing the production of IFN-I and ISGs. This study provides a clue to design antiviral agents or live attenuated vaccines to prevent and control ASF.

The Importance of Sintering-Induced Grain Boundaries in Copper Catalysis to Improve Carbon-Carbon Coupling.

Syngas conversion serves as a gas-to-liquid technology to produce liquid fuels and valuable chemicals from coal, natural gas, or biomass. During syngas conversion, sintering is known to deactivate the catalyst owing to the loss of active surface area. However, the growth of nanoparticles might induce the formation of new active sites such as grain boundaries (GBs) which perform differently from the original nanoparticles. Herein, we reported a unique Cu-based catalyst, Cu nanoparticles with in situ generated GBs confined in zeolite Y (denoted as activated Cu/Y), which exhibited a high selectivity for C5+ hydrocarbons (65.3 C%) during syngas conversion. Such high selectivity for long-chain products distinguished activated Cu/Y from typical copper-based catalysts which mainly catalyze methanol synthesis. This unique performance was attributed to the GBs, while the zeolite assisted the stabilization through spatial confinement. Specifically, the GBs enabled H-assisted dissociation of CO and subsequent hydrogenation into CHx*. CHx* species not only serve as the initiator but also directly polymerize on Cu GBs, known as the carbide mechanism. Meanwhile, the synergy of GBs and their vicinal low-index facets led to the CO insertion where non-dissociative adsorbed CO on low-index facets migrated to GBs and inserted into the metal-alkyl bond for the chain growth.

African swine fever virus pH240R enhances viral replication via inhibition of the type I IFN signaling pathway.

African swine fever (ASF) is an acute, hemorrhagic, and severe infectious disease caused by ASF virus (ASFV) infection. At present, there are still no safe and effective drugs and vaccines to prevent ASF. Mining the important proteins encoded by ASFV that affect the virulence and replication of ASFV is the key to developing effective vaccines and drugs. In this study, ASFV pH240R, a capsid protein of ASFV, was found to inhibit the type I interferon (IFN) signaling pathway. Mechanistically, pH240R interacted with IFNAR1 and IFNAR2 to disrupt the interaction of IFNAR1-TYK2 and IFNAR2-JAK1. Additionally, pH240R inhibited the phosphorylation of IFNAR1, TYK2, and JAK1 induced by IFN-α, resulting in the suppression of the nuclear import of STAT1 and STAT2 and the expression of IFN-stimulated genes (ISGs). Consistent with these results, H240R-deficient ASFV (ASFV-∆H240R) infection induced more ISGs in porcine alveolar macrophages compared with its parental ASFV HLJ/18. We also found that pH240R enhanced viral replication via inhibition of ISGs expression. Taken together, our results clarify that pH240R enhances ASFV replication by inhibiting the JAK-STAT signaling pathway, which highlights the possibility of pH240R as a potential drug target.IMPORTANCEThe innate immune response is the host's first line of defense against pathogen infection, which has been reported to affect the replication and virulence of African swine fever virus (ASFV) isolates. Identification of ASFV-encoded proteins that affect the virulence and replication of ASFV is the key step in developing more effective vaccines and drugs. In this study, we found that pH240R interacted with IFNAR1 and IFNAR2 by disrupting the interaction of IFNAR1-TYK2 and IFNAR2-JAK1, resulting in the suppression of the expression of interferon (IFN)-stimulated genes (ISGs). Consistent with these results, H240R-deficient ASFV (ASFV-∆H240R) infection induces more ISGs' expression compared with its parental ASFV HLJ/18. We also found that pH240R enhanced viral replication via inhibition of ISGs' expression. Taken together, our findings showed that pH240R enhances ASFV replication by inhibiting the IFN-JAK-STAT axis, which highlights the possibility of pH240R as a potential drug target.

Tanshinone IIA modulates cancer cell morphology and movement via Rho GTPases-mediated actin cytoskeleton remodeling.

Actin filaments form unique structures with robust actin bundles and cytoskeletal networks affixed to the extracellular matrix and interact with neighboring cells, which are crucial structures for cancer cells to acquire a motile phenotype. This study aims to investigate a novel antitumor mechanism by which Tanshinone IIA (Tan IIA) modulates the morphology and migration of liver cancer cells via actin cytoskeleton regulation. 97H and Huh7 exhibited numerous tentacle-like protrusions that interacted with neighboring cells. Following treatment with Tan IIA, 97H and Huh7 showed a complete absence of cytoplasmic protrusion and adherens junctions, thereby effectively impeding their migration capability. The fluorescence staining of F-actin and microtubules indicated that these tentacle-like protrusions and cell-cell networks were actin-based structures that led to morphological changes after Tan IIA treatment by retracting and reorganizing beneath the membrane. Tan IIA can reverse the actin depolymerization and cell morphology alterations induced by latrunculin A. Tan IIA down-regulated actin and Rho GTPases expression significantly, as opposed to inducing Rho signaling activation. Preventing the activity of proteasomes and lysosomes had no discernible impact on the modifications in cellular structure and protein expression induced by Tan IIA. However, as demonstrated by the puromycin labeling technique, the newly synthesized proteins were significantly inhibited by Tan IIA. In conclusion, Tan IIA can induce dramatic actin cytoskeleton remodeling by inhibiting the protein synthesis of actin and Rho GTPases, resulting in the suppression of tumor growth and migration. Targeting the actin cytoskeleton of Tan IIA is a promising strategy for HCC treatment.

Improving cardiovascular risk prediction through machine learning modelling of irregularly repeated electronic health records.

Aims: Existing electronic health records (EHRs) often consist of abundant but irregular longitudinal measurements of risk factors. In this study, we aim to leverage such data to improve the risk prediction of atherosclerotic cardiovascular disease (ASCVD) by applying machine learning (ML) algorithms, which can allow automatic screening of the population. Methods and results: A total of 215 744 Chinese adults aged between 40 and 79 without a history of cardiovascular disease were included (6081 cases) from an EHR-based longitudinal cohort study. To allow interpretability of the model, the predictors of demographic characteristics, medication treatment, and repeatedly measured records of lipids, glycaemia, obesity, blood pressure, and renal function were used. The primary outcome was ASCVD, defined as non-fatal acute myocardial infarction, coronary heart disease death, or fatal and non-fatal stroke. The eXtreme Gradient boosting (XGBoost) algorithm and Least Absolute Shrinkage and Selection Operator (LASSO) regression models were derived to predict the 5-year ASCVD risk. In the validation set, compared with the refitted Chinese guideline-recommended Cox model (i.e. the China-PAR), the XGBoost model had a significantly higher C-statistic of 0.792, (the differences in the C-statistics: 0.011, 0.006-0.017, P < 0.001), with similar results reported for LASSO regression (the differences in the C-statistics: 0.008, 0.005-0.011, P < 0.001). The XGBoost model demonstrated the best calibration performance (men: Dx = 0.598, P = 0.75; women: Dx = 1.867, P = 0.08). Moreover, the risk distribution of the ML algorithms differed from that of the conventional model. The net reclassification improvement rates of XGBoost and LASSO over the Cox model were 3.9% (1.4-6.4%) and 2.8% (0.7-4.9%), respectively. Conclusion: Machine learning algorithms with irregular, repeated real-world data could improve cardiovascular risk prediction. They demonstrated significantly better performance for reclassification to identify the high-risk population correctly.

The attenuated African swine fever vaccine HLJ/18-7GD provides protection against emerging prevalent genotype II variants in China.

Genetic changes have occurred in the genomes of prevalent African swine fever viruses (ASFVs) in the field in China, which may change their antigenic properties and result in immune escape. There is usually poor cross-protection between heterogonous isolates, and, therefore, it is important to test the cross-protection of the live attenuated ASFV vaccines against current prevalent heterogonous isolates. In this study, we evaluated the protective efficacy of the ASFV vaccine candidate HLJ/18-7GD against emerging isolates. HLJ/18-7GD provided protection against a highly virulent variant and a lower lethal isolate, both derived from genotype II Georgia07-like ASFV and isolated in 2020. HLJ/18-7GD vaccination prevented pigs from developing ASF-specific clinical signs and death, decreased viral shedding via the oral and rectal routes, and suppressed viral replication after challenges. However, HLJ/18-7GD vaccination did not provide solid cross-protection against genotype I NH/P68-like ASFV challenge in pigs. HLJ/18-7GD vaccination thus shows great promise as an alternative strategy for preventing and controlling genotype II ASFVs, but vaccines providing cross-protection against different ASFV genotypes may be needed in China.

[Association Between Dietary Knowledge and Food Preferences in Residents Aged 18 and Over in China Based on Latent Class Analysis].

Objective To investigate the food preferences and explore the potential association between dietary knowledge and food preferences in residents aged 18 and over in China,so as to provide a basis for promoting healthy diets.Methods The latent class analysis was carried out with the 2015 cross-sectional data of China health and nutrition survey to categorize the food preferences among 8 783 residents aged 18 and over.Multinomial Logistic regression was adopted to assess the association between and dietary knowledge and food preferences.Results The food preferences of the residents aged 18 and over in China were classified into preference for less vegetable(3.28%),lack of preference(11.20%),diverse preferences(4.19%),and preference for healthy diets(81.33%).The proportion of the adults with dietary knowledge was 36.87%(3 238/8 783).The dietary knowledge varied in the adults with different food preferences(all P<0.001).After adjusting for gender,age,urban and rural distribution,education background,and annual household income,for each point increase in the dietary knowledge score,there was an estimated reduction of 22% in the probability of preferring less vegetables(OR=0.78,95%CI=0.76-0.80, P<0.001),13% in the probability of lacking preference(OR=0.87,95%CI=0.86-0.89, P<0.001),and 3% in the probability of having diverse preferences(OR=0.97,95%CI=0.94-1.00, P=0.030).Compared with those lacking dietary knowledge,the individuals with dietary knowledge had a 77% less probability of preferring less vegetables(OR=0.23,95%CI=0.16-0.32, P<0.001),a 55% less probability of lacking preference(OR=0.45,95%CI=0.39-0.53, P<0.001),and a 23% less probability of having diverse preferences(OR=0.77,95%CI=0.61-0.96, P=0.023).Conclusions The residents aged 18 and over in China presented four food preferences,including preference for less vegetables,lack of preference,diverse preferences,and preference for healthy diets,the last of which had the highest proportion.The individuals with lower levels of dietary knowledge have higher probability of preferring unhealthy food.

[Comparison on Performance of Quantitative Ultrasound and Dual-Energy X-ray Absorptiometry in Evaluating Bone Health of Adults Aged 18-40 Years].

Objective To compare the consistency of quantitative ultrasound(QUS)and dual-energy X-ray absorptiometry(DXA)in measuring bone mineral density(BMD)of adults aged 18-40 years in Guangzhou and evaluate the diagnostic value of QUS for identifying low bone mass.Methods DXA was employed to measure the BMD and QUS to measure the speed of sound(SOS)in 731 participants.The Bland-Altman analysis was performed to evaluate the consistency of Z scores between SOS and BMD.With the BMD Z ≤-2.00 as the diagnostic criterion for low bone mass,the receiver operating characteristics curve of QUS was established,and the area under the curve(AUC)and the sensitivity,specificity,and correct diagnostic index for the optimal cut-off of SOS Z score were calculated.Results The results of Bland-Altman analysis showed that the mean differences in the Z scores of SOS and BMD in males and females were 1.27(-0.94 to 3.47)and 0.93(-1.33 to 3.18),respectively.The AUC of SOS Z score in the diagnosis of low bone mass in males and females was 0.734(95%CI=0.380-0.788)and 0.679(95%CI=0.625-0.732),respectively.In males,the optimal cut-off of SOS Z score for low bone mass was -0.35,with the sensitivity,specificity,and correct diagnostic index of 64.1%,68.6%,and 0.327,respectively.In females,the optimal cut-off value of SOS Z scores for low bone mass was -1.14,with the sensitivity,specificity,and correct index of 73.9%,54.8%,and 0.285,respectively.Conclusion QUS and DXA show poor consistency in the diagnosis of BMD in the adults aged 18-40 years in Guangzhou,while QUS demonstrates an acceptable value in identifying low bone mass.

Development of a new effective African swine fever virus vaccine candidate by deletion of the H240R and MGF505-7R genes results in protective immunity against the Eurasia strain.

IMPORTANCE: African swine fever (ASF) caused by ASF virus (ASFV) is a highly contagious and acute hemorrhagic viral disease in domestic pigs. Until now, no effective commercial vaccine and antiviral drugs are available for ASF control. Here, we generated a new live-attenuated vaccine candidate (ASFV-ΔH240R-Δ7R) by deleting H240R and MGF505-7R genes from the highly pathogenic ASFV HLJ/18 genome. Piglets immunized with ASFV-ΔH240R-Δ7R were safe without any ASF-related signs and produced specific antibodies against p30. Challenged with a virulent ASFV HLJ/18, the piglets immunized with high-dose group (105 HAD50) exhibited 100% protection without clinical symptoms, showing that low levels of virus replication with no observed pathogenicity by postmortem and histological analysis. Overall, our results provided a new strategy by designing live-attenuated vaccine candidate, resulting in protection against ASFV infection.

African Swine Fever Virus H240R Protein Inhibits the Production of Type I Interferon through Disrupting the Oligomerization of STING.

African swine fever (ASF) is a highly contagious and acute hemorrhagic viral disease in domestic pigs and wild boars. Domestic pigs infected with virulent African swine fever virus (ASFV) isolates have a high mortality, approaching 100%. Identification of ASFV genes related to virulence/pathogenicity and deletion of them are considered to be key steps in the development of live attenuated vaccines, because the ability of ASFV to escape host innate immune responses is related to viral pathogenicity. However, the relationship between the host antiviral innate immune responses and the pathogenic genes of ASFV has not been fully understood. In this study, the ASFV H240R protein (pH240R), a capsid protein of ASFV, was found to inhibit type I interferon (IFN) production. Mechanistically, pH240R interacted with the N-terminal transmembrane domain of stimulator of interferon genes (STING) and inhibited its oligomerization and translocation from the endoplasmic reticulum to the Golgi apparatus. Additionally, pH240R inhibited the phosphorylation of interferon regulatory factor 3 (IRF3) and TANK binding kinase 1 (TBK1), leading to reduced production of type I IFN. Consistent with these results, infection with H240R-deficient ASFV (ASFV-ΔH240R) induced more type I IFN than infection with its parental strain, ASFV HLJ/18. We also found that pH240R may enhance viral replication via inhibition of type I IFN production and the antiviral effect of interferon alpha (IFN-α). Taken together, our findings provide a new explanation for the reduction of ASFV's replication ability by knockout of the H240R gene and a clue for the development of live attenuated ASFV vaccines. IMPORTANCE African swine fever (ASF), caused by African swine fever virus (ASFV), is a highly contagious and acute hemorrhagic viral disease with a high mortality, approaching 100% in domestic pigs. However, the relationship between viral pathogenicity and immune evasion of ASFV is not fully understood, which limits the development of safe and effective ASF vaccines, specifically, live attenuated vaccines. In this study, we found that pH240R, as a potent antagonist, inhibited type I IFN production by targeting STING and inhibiting its oligomerization and translocation from the endoplasmic reticulum to the Golgi apparatus. Furthermore, we also found that deletion of the H240R gene reduced viral pathogenicity by enhancing type I IFN production, which decreases ASFV replication. Taken together, our findings provide a clue for the development of an ASFV live attenuated vaccine via deleting the H240R gene.

Harnessing heterogeneity in space with statistically guided meta-learning.

Spatial data are ubiquitous, massively collected, and widely used to support critical decision-making in many societal domains, including public health (e.g., COVID-19 pandemic control), agricultural crop monitoring, transportation, etc. While recent advances in machine learning and deep learning offer new promising ways to mine such rich datasets (e.g., satellite imagery, COVID statistics), spatial heterogeneity-an intrinsic characteristic embedded in spatial data-poses a major challenge as data distributions or generative processes often vary across space at different scales, with their spatial extents unknown. Recent studies (e.g., SVANN, spatial ensemble) targeting this difficult problem either require a known space-partitioning as the input, or can only support very limited number of partitions or classes (e.g., two) due to the decrease in training data size and the complexity of analysis. To address these limitations, we propose a model-agnostic framework to automatically transform a deep learning model into a spatial-heterogeneity-aware architecture, where the learning of arbitrary space partitionings is guided by a learning-engaged generalization of multivariate scan statistic and parameters are shared based on spatial relationships. Moreover, we propose a spatial moderator to generalize learned space partitionings to new test regions. Finally, we extend the framework by integrating meta-learning-based training strategies into both spatial transformation and moderation to enhance knowledge sharing and adaptation among different processes. Experiment results on real-world datasets show that the framework can effectively capture flexibly shaped heterogeneous footprints and substantially improve prediction performances.

Comprehensive mapping of antigenic linear B-cell epitopes on K205R protein of African swine fever virus with monoclonal antibodies.

African swine fever virus causes an acute, highly contagious swine disease with high mortality, leading to enormous losses in the pig industry. The K205R, a nonstructural protein of African swine fever virus, is abundantly expressed in the cytoplasm of infected cells at the early stage of infection and induces a strong immune response. However, to date, the antigenic epitopes of this immunodeterminant have not been characterized. In the present study, the K205R protein was expressed in a mammalian cell line and purified using Ni-affinity chromatography. Furthermore, three monoclonal antibodies (mAbs; 5D6, 7A8, and 7H10) against K205R were generated. Indirect immunofluorescence assay and western blot results showed that all three mAbs recognized native and denatured K205R in African swine fever virus (ASFV)-infected cells. To identify the epitopes of the mAbs, a series of overlapping short peptides were designed and expressed as fusion proteins with maltose-binding protein. Subsequently, the peptide fusion proteins were probed with monoclonal antibodies using western blot and enzyme-linked immunosorbent assay. The three target epitopes were fine-mapped; the core sequences of recognized by the mAbs 5D6, 7A8, and 7H10 were identified as 157FLTPEIQAILDE168, 154REKFLTP160, and 136PTNAMFFTRSEWA148, respectively. Probing with sera from ASFV-infected pigs in a dot blot assay demonstrated that epitope 7H10 was the immunodominant epitope of K205R. Sequence alignment showed that all epitopes were conserved across ASFV strains and genotypes. To our knowledge, this is the first study to characterize the epitopes of the antigenic K205R protein of ASFV. These findings may serve as a basis for the development of serological diagnostic methods and subunit vaccines.

African Swine Fever Virus HLJ/18 CD2v Suppresses Type I IFN Production and IFN-Stimulated Genes Expression through Negatively Regulating cGMP-AMP Synthase-STING and IFN Signaling Pathways.

African swine fever is a fatal infectious disease caused by African swine fever virus (ASFV). The high mortality caused by this infectious disease is a significant challenge to the swine industry worldwide. ASFV virulence is related to its ability to antagonize IFN response, yet the mechanism of antagonism is not understood. Recently, a less virulent recombinant virus has emerged that has a EP402R gene deletion within the parental ASFV HLJ/18 (ASFV-ΔEP402R) strain. EP402R gene encodes CD2v. Hence we hypothesized that ASFV uses CD2v protein to evade type I IFN-mediated innate immune response. We found that ASFV-ΔEP402R infection induced higher type I IFN response and increased the expression of IFN-stimulated genes in porcine alveolar macrophages when compared with parental ASFV HLJ/18. Consistent with these results, CD2v overexpression inhibited type I IFN production and IFN-stimulated gene expression. Mechanistically, CD2v, by interacting with the transmembrane domain of stimulator of IFN genes (STING), prevented the transport of STING to the Golgi apparatus, and thereby inhibited the cGMP-AMP synthase-STING signaling pathway. Furthermore, ASFV CD2v disrupted IFNAR1-TYK2 and IFNAR2-JAK1 interactions, and thereby inhibited JAK-STAT activation by IFN-α. In vivo, specific pathogen-free pigs infected with the mutant ASFV-ΔEP402R strain survived better than animals infected with the parental ASFV HLJ/18 strain. Consistent with this finding, IFN-ß protein levels in the peripheral blood of ASFV-ΔEP402R-challenged pigs were significantly higher than in the blood of ASFV HLJ/18-challenged pigs. Taken together, our findings suggest a molecular mechanism in which CD2v inhibits cGMP-AMP synthase-STING and IFN signaling pathways to evade the innate immune response rendering ASFV infection fatal in pigs.

Experimental and Updated Kinetic Modeling Study of Neopentane Low Temperature Oxidation.

Neopentane is an ideal fuel model to study low-temperature oxidation chemistry. The significant discrepancies between experimental data and simulations using the existing neopentane models indicate that an updated study of neopentane oxidation is needed. In this work, neopentane oxidation experiments are carried out using two jet-stirred reactors (JSRs) at 1 atm, at a residence time of 3 s, and at three different equivalence ratios of 0.5, 0.9, and 1.62. Two different analytical methods (synchrotron vacuum ultraviolet photoionization mass spectrometry and gas chromatography) were used to investigate the species distributions. Numerous oxidation intermediates were detected and quantified, including acetone, 3,3-dimethyloxetane, methacrolein, isobutene, 2-methylpropanal, isobutyric acid, and peroxides, which are valuable for validating the kinetic model describing neopentane oxidation. In the model development, the pressure dependencies of the rate constants for the reaction classes Q̇OOH + O2 and Q̇OOH decompositions are considered. This addition improves the prediction of the low-temperature oxidation reactivity of neopentane. Another focus of model development is to improve the prediction of carboxylic acids formed during the low-temperature oxidation of neopentane. The detection and identification of isobutyric acid indicates the existence of the Korcek mechanism during neopentane oxidation. Regarding the formation of acetic acid, the reaction channels are considered to be initiated from the reactions of ȮH radical addition to acetaldehyde/acetone. This updated kinetic model is validated extensively against the experimental data in this work and various experimental data available in the literature, including ignition delay times (IDTs) from both shock tubes (STs) and rapid compression machines (RCMs) and JSR speciation data at high temperatures.

Deletion of African Swine Fever Virus (ASFV) H240R Gene Attenuates the Virulence of ASFV by Enhancing NLRP3-Mediated Inflammatory Responses.

African swine fever (ASF) is a highly contagious infectious disease of domestic pigs and wild boars caused by African swine fever virus (ASFV), with a mortality rate of up to 100%. In order to replicate efficiently in macrophages and monocytes, ASFV has evolved multiple strategies to evade host antiviral responses. However, the underlying molecular mechanisms by which ASFV-encoded proteins execute immune evasion are not fully understood. In this study, we found that ASFV pH240R strongly inhibits transcription, maturation, and secretion of interleukin-1ß (IL-1ß). Importantly, pH240R not only targeted NF-κB signaling but also impaired NLRP3 inflammasome activation. In this mechanism, pH240R interacted with NF-kappa-B essential modulator (NEMO), a component of inhibitor of kappa B kinase (IKK) complex and subsequently reduced phosphorylation of IκBα and p65. In addition, pH240R bonded to NLRP3 to inhibit NLRP3 inflammasome activation, resulting in reduced IL-1ß production. As expected, infection with H240R-deficient ASFV (ASFV-ΔH240R) induced more inflammatory cytokine expression both in vitro and in vivo than its parental ASFV HLJ/18 strain. Consistently, H240R deficiency reduced the viral pathogenicity in pigs compared with its parental strain. These findings reveal that the H240R gene is an essential virulence factor, and deletion of the H240R gene affects the pathogenicity of ASFV HLJ/18 by enhancing antiviral inflammatory responses, which provides insights for ASFV immune evasion mechanisms and development of attenuated live vaccines and drugs for prevention and control of ASF. IMPORTANCE African swine fever (ASF), caused by African swine fever virus (ASFV), is a highly contagious and acute hemorrhagic viral disease of domestic pigs, with a high mortality approaching 100%. ASFV has spread rapidly worldwide and caused huge economic losses and ecological consequences. However, the pathogenesis and immune evasion mechanisms of ASFV are not fully understood, which limits the development of safe and effective ASF attenuated live vaccines. Therefore, investigations are urgently needed to identify virulence factors that are responsible for escaping the host antiviral innate immune responses and provide a new target for development of ASFV live-attenuated vaccine. In this study, we determined that the H240R gene is an essential virulence factor, and its depletion affects the pathogenicity of ASFV by enhancing NLRP3-mediated inflammatory responses, which provides theoretical support for the development of an ASFV attenuated live vaccine.

Platelet Desialylation Is a Novel Mechanism and Therapeutic Target in Daboia siamensis and Agkistrodon halys Envenomation-Induced Thrombocytopenia.

Venom-induced thrombocytopenia (VIT) is one of the most important hemotoxic effects of a snakebite, which is often associated with venom-induced consumptive coagulopathy (VICC). Refractory thrombocytopenia without significant coagulation abnormalities has also been reported after envenomation by some viperid snakes; however, the mechanisms are not well understood and therapeutic strategies are lacking. Here, we found that patients injured by Daboia siamensis or Agkistrodon halys snakes, who were resistant to standard antivenom treatment, had developed coagulopathy-independent thrombocytopenia. Venoms from these viperid snakes, rather than from the elapid snake (Bungarus multicinctus), induced platelet surface expression of neuraminidase-1 (NEU-1), and significantly increased the desialylation of the glycoproteins on human platelets. The desialylated platelets caused by viperid snake venoms were further internalized by macrophages, which resulted in reduced platelet numbers in peripheral blood. Importantly, neuraminidase inhibitor significantly decreased viper venom-induced platelet desialylation, therefore inhibiting platelet phagocytosis by macrophages, and alleviating venom-induced thrombocytopenia. Collectively, these findings support an important role for desialylated platelet clearance in the progression of viper envenomation-induced, coagulopathy-independent thrombocytopenia. Our study demonstrates that the neuraminidase inhibitor may be a potential therapy or adjuvant therapy to treat snakebite-induced thrombocytopenia.

Host Responses to Live-Attenuated ASFV (HLJ/18-7GD).

African swine fever (ASF) is a highly contagious and fatal disease caused by the African swine fever virus. Recently, the multigene family and CD2v gene-deleted ASF vaccine candidate HLJ/18-7GD was found to be safe and effective in laboratory and clinical trials. However, the immune-protective mechanisms underlying the effects of HLJ/18-7GD remain unclear. We assessed samples from pigs immunized with a single dose of 106 TCID50 HLJ/18-7GD. We found that pigs immunized with HLJ/18-7GD showed high levels of specific antibodies. T lymphocyte subsets (helper T cells (Th); cytotoxic T lymphocytes (CTL); double-positive T cells (DP-T cells)) were temporarily increased in peripheral blood mononuclear cells (PBMCs) after HLJ/18-7GD immunization. Once the HLJ/18-7GD-immunized pigs had been challenged with virulent HLJ/18, the percentage of Th, CTL, and DP-T cells increased significantly. PBMCs extracted from the pigs induced higher levels of CD8+ T cells after infection with the HLJ/18 strain in vitro. The levels of GM-CSF, IFN-γ, and TNF-α were upregulated at 7 days post-inoculation; this finding was contrary to the results obtained after HLJ/18 or HLJ/18ΔCD2v infection. The immune protection from HLJ/18-7GD resulted from many synergies, which could provide a theoretical basis for HLJ/18-7GD as a safe and effective ASF vaccine.

A modified LC-MS/MS method for the detection of whole blood tacrolimus and its clinical value in Chinese kidney transplant patients.

Background: For patients who treated with tacrolimus after kidney transplant, therapeutic drug monitoring is essential to improve their prognosis. However, previous detection methods have limitations, such as the overestimation and unacceptable bias in the immunoassays. Precision medicine has been challenged. The liquid chromatography-tandem mass spectrometry (LC-MS/MS) method is recognized as the gold standard due to its accuracy and specificity, but lack of throughput and complex process limits its clinical application. Therefore, an accurate, simple and high throughput method for tacrolimus monitoring is needed for clinical practice. Methods: A modified LC-MS/MS method was introduced and validated. Whole blood samples were prepared by a one-step protein precipitation method. Chromatographic separation was achieved using a Phenomenex Kinetex 2.6 µm XB-C18 2.1 × 50 mm column with a total run time of 3.5 min to avoid matrix effect. An electrospray ionization source (ESI) was used in positive ion multiple reaction monitoring (MRM) mode for mass spectrometric detection. In order to protect the mass spectrometer, only part of the sample after LC separation was allowed to enter the mass spectrum, through a two HPLC systems coupled one mass spectrometry design. In this way, the instrument throughput is also improved and realizing the detection of 2 samples within 3.5 min and carried out a shorter analyzing time for each sample of 1.75 min. Additionally, we calculated tacrolimus-intrapatient variant (Tac-IPV) based on this modified method and assessed the prognostic value of Tac-IPV in Chinese kidney transplant patients. Results: The LC-MS/MS was modified by streamlining the procedure and increasing the throughput. The method proved to be accurate and reproducible with all performance parameters suitably meeting the clinical requirements over a calibration ranged from 0.37 to 42.90 ng/mL. Parameters such as linearity, limit of quantification (LoQ) and dilution integrity were validated with a clinical reportable range from 0.37 to 343.20 ng/mL, which was particularly useful for high drug concentrations patients (rare but very serious). Both cross-contamination and matrix effects were negligible. Clinical data of 83 patients showed that Tac-IPV was associated with poor kidney transplant outcome in Chinese (Hazard Ratio (HR) = 3.96, 4.75; 95% Cl: 1.10-14.21, 1.23-18.36; P < 0.05). Conclusions: This modified LC-MS/MS method possessed high throughput and simple sample preparation, allowing it to meet daily clinical needs. At the same time, Tac-IPV based on this modified LC-MS/MS had excellent prognostic value in kidney transplantation. These advantages have great significance for the individualized treatment of Chinese kidney transplant patients and broad application of Tac-IPV.

Development of an ELISA Method to Differentiate Animals Infected with Wild-Type African Swine Fever Viruses and Attenuated HLJ/18-7GD Vaccine Candidate.

African swine fever (ASF) is a highly contagious hemorrhagic disease of pigs, posing a significant threat to the world pig industry. Several researchers are investigating the possibilities for developing a safe and efficient vaccine against ASF. In this regard, significant progress has been made and some gene-deleted ASFVs are reported as potential live attenuated vaccines. A seven-gene-deleted live attenuated vaccine candidate HLJ/18-7GD (among which CD2v is included) has been developed in our laboratory and reported to be safe and protective, and it is expected to be commercialized in the near future. There is an urgent need for developing a diagnostic method that can clearly discriminate between wild-type-ASFV-infected and vaccinated animals (DIVA). In the present study, a dual indirect ELISA based on p54 and CD2v proteins was successfully established to specifically distinguish serum antibodies from pigs infected with wild-type ASFV or possessing vaccine immunization. To evaluate the performance of the assay, a total of 433 serum samples from four groups of pigs experimentally infected with the wild-type HLJ/18 ASFV, immunized with the HLJ/18-7GD vaccine candidate, infected with the new lower virulent variant, and specific-pathogen-free pigs were used. Our results showed that the positive rate of immunized serum was 96.54% (p54) and 2.83% (CD2v), and the positive rate of the infection by wild-type virus was 100% (p54) and 97.8% (CD2v). Similarly, the positive rate to infection by the new low-virulent ASFV variant in China was 100% (p54) and 0% (CD2v), indicating the technique was also able to distinguish antibodies from wild-type and the new low-virulent ASFV variant in China. Moreover, no cross-reaction was observed in immune sera from other swine pathogens, such as CSFV, PEDV, PRRSV, HP-PRRSV, PCV2, and PrV. Overall, the developed dual indirect ELISA exhibited high diagnostic sensitivity, specificity, and repeatability and will provide a new approach to differentiate serum antibodies between wild virulent and CD2v-unexpressed ASFV infection, which will play a great role in serological diagnosis and epidemiological monitoring of ASF in the future.

Long Noncoding RNA RP11-732M18.3 Promotes Glioma Angiogenesis by Upregulating VEGFA.

Gliomas are the most aggressive and common type of malignant brain tumor, with limited treatment options and a dismal prognosis. Angiogenesis, a hallmarks of cancer, is one of two critical events in the progression of gliomas. Accumulating evidence has demonstrated that in glioma dysregulated molecules like long noncoding RNAs (lncRNAs), are closely linked to tumorigenesis and prognosis. However, the effects of and mechanisms of action of lncRNAs during tumor angiogenesis are poorly understood. The effect of lncRNA RP11-732M18.3 on angiogenesis was elucidated through an intracranial orthotopic glioma model, immunohistochemistry, and an in vitro angiogenesis assay. Co-culture experiments and cell migration assays were performed to investigate the function of lncRNA RP11-732M18.3 in vitro. lncRNA RP11-732M18.3 increased CD31+ microvessel density, and overexpression of lncRNA RP11-732M18.3 resulted in poor mouse survival. lncRNA RP11-732M18.3 promoted endothelial cell migration and tube formation. Nomogram and Kaplan-Meier survival analyses indicated that higher VEGFA is correlated with a poor prognosis. Mechanistically, lncRNA RP11-732M18.3 promotes angiogenesis by increasing the nuclear level of EP300 and facilitating the transcription and secretion of VEGFA. Our study contributes to the latest understanding of glioma angiogenesis and prognosis. lncRNA RP11-732M18.3 may be a potential treatment target in glioma.