Ubiquitin-specific proteinase 1 stabilizes PRRSV nonstructural protein Nsp1ß to promote viral replication by regulating K48 ubiquitination.

The porcine reproductive and respiratory syndrome virus (PRRSV) can lead to severe reproductive problems in sows, pneumonia in weaned piglets, and increased mortality, significantly negatively impacting the economy. Post-translational changes are essential for the host-dependent replication and long-term infection of PRRSV. Uncertainty surrounds the function of the ubiquitin network in PRRSV infection. Here, we screened 10 deubiquitinating enzyme inhibitors and found that the ubiquitin-specific proteinase 1 (USP1) inhibitor ML323 significantly inhibited PRRSV replication in vitro. Importantly, we found that USP1 interacts with nonstructural protein 1ß (Nsp1ß) and deubiquitinates its K48 to increase protein stability, thereby improving PRRSV replication and viral titer. Among them, lysine at position 45 is essential for Nsp1ß protein stability. In addition, deficiency of USP1 significantly reduced viral replication. Moreover, ML323 loses antagonism to PRRSV rSD16-K45R. This study reveals the mechanism by which PRRSV recruits the host factor USP1 to promote viral replication, providing a new target for PRRSV defense.IMPORTANCEDeubiquitinating enzymes are critical factors in regulating host innate immunity. The porcine reproductive and respiratory syndrome virus (PRRSV) nonstructural protein 1ß (Nsp1ß) is essential for producing viral subgenomic mRNA and controlling the host immune system. The host inhibits PRRSV proliferation by ubiquitinating Nsp1ß, and conversely, PRRSV recruits the host protein ubiquitin-specific proteinase 1 (USP1) to remove this restriction. Our results demonstrate the binding of USP1 to Nsp1ß, revealing a balance of antagonism between PRRSV and the host. Our research identifies a brand-new PRRSV escape mechanism from the immune response.

Rapid and Complete Digestion of Refractory Geological Samples Using Ultrafine Powder for Accurate Analyses of Trace Elements.

Complete sample digestion is a prerequisite for acquiring high-quality analytical results for geological samples. Closed-vessel acid digestion (bomb) has typically been used for the total digestion of refractory geological samples. However, the long digestion time (4-5 days) and insoluble fluoride complexes still pose challenges for digesting refractory geological samples using this approach. In this study, an efficient and simplified digestion technique combining ultrafine powders from planetary ball milling with bomb digestion was developed for trace element analysis of refractory geological samples: peridotite and granitoid. The method shows two significant improvements compared with previous approaches. (1) By performing dry planetary ultrafine milling, the initial 200 mesh peridotite (<74 µm) could be reduced to 800 mesh (<20 µm) in 6 min at a ball-to-powder mass ratio of approximately 15 using 3 mm tungsten carbide milling balls. (2) Complete peridotite and granitoid dissolution were achieved in approximately 2 h, 60 times faster than what is achievable using previous methods (2 h vs 120 h). Moreover, ultrafine powders effectively suppressed insoluble fluoride formation during bomb digestion. A suite of peridotite and granitoid reference materials were measured to evaluate the stability of this method. This efficient, simple, and reliable sample digestion method could benefit geological, food, environmental, and other fields requiring solid sample decomposition via wet acid, fusion, combustion, or dry ashing.

The African swine fever virus I10L protein inhibits the NF-κB signaling pathway by targeting IKKß.

Proinflammatory factors play important roles in the pathogenesis of African swine fever virus (ASFV), which is the causative agent of African swine fever (ASF), a highly contagious and severe hemorrhagic disease. Efforts in the prevention and treatment of ASF have been severely hindered by knowledge gaps in viral proteins responsible for modulating host antiviral responses. In this study, we identified the I10L protein (pI10L) of ASFV as a potential inhibitor of the TNF-α- and IL-1ß-triggered NF-κB signaling pathway, the most canonical and important part of host inflammatory responses. The ectopically expressed pI10L remarkably suppressed the activation of NF-κB signaling in HEK293T and PK-15 cells. The ASFV mutant lacking the I10L gene (ASFVΔI10L) induced higher levels of proinflammatory cytokines production in primary porcine alveolar macrophages (PAMs) compared with its parental ASFV HLJ/2018 strain (ASFVWT). Mechanistic studies suggest that pI10L inhibits IKKß phosphorylation by reducing the K63-linked ubiquitination of NEMO, which is necessary for the activation of IKKß. Morever, pI10L interacts with the kinase domain of IKKß through its N-terminus, and consequently blocks the association of IKKß with its substrates IκBα and p65, leading to reduced phosphorylation. In addition, the nuclear translocation efficiency of p65 was also altered by pI10L. Further biochemical evidence supported that the amino acids 1-102 on pI10L were essential for the pI10L-mediated suppression of the NF-κB signaling pathway. The present study clarifies the immunosuppressive activity of pI10L, and provides novel insights into the understanding of ASFV pathobiology and the development of vaccines against ASF. IMPORTANCE African swine fever (ASF), caused by the African swine fever virus (ASFV), is now widespread in many countries and severely affects the commercial rearing of swine. To date, few safe and effective vaccines or antiviral strategies have been marketed due to large gaps in knowledge regarding ASFV pathobiology and immune evasion mechanisms. In this study, we deciphered the important role of the ASFV-encoded I10L protein in the TNF-α-/IL-1ß-triggered NF-κB signaling pathway. This study provides novel insights into the pathogenesis of ASFV and thus contributes to the development of vaccines against ASF.

Outcomes with allogeneic stem cell transplant using cryopreserved versus fresh hematopoietic progenitor cell products.

BACKGROUND: Allogeneic hematopoietic stem cell transplant (alloHSCT) is a mainstay of treatment for hematologic malignancies such as acute leukemias and aggressive lymphomas. Historically, fresh hematopoietic progenitor cell (HPC) products have been preferred to cryopreserved products (cryo-HPC) due to concerns of loss of stem cell viability and number with the cryopreservation procedure. OBJECTIVE: We aimed to analyze the outcomes of patients who received cryo-HPCs during the COVID-19 pandemic and compare this against historical cohorts that received fresh HPC. STUDY DESIGN: A retrospective chart review was conducted on all adult patients who received a peripheral blood alloHSCT in British Columbia, Canada between June 2017 and November 2021. Baseline characteristics, Kaplan-Meier (KM) overall survival (OS), engraftment, and incidences of acute and chronic graft versus host disease were compared between patients who received cryo-HPCs and fresh HPCs. Univariable analysis followed by multivariable analysis was performed using a backward stepwise selection procedure to generate predictors of OS, cumulative incidence of relapse (CIR), nonrelapse mortality (NRM), and primary and secondary graft failure. RESULTS: Three hundred eighty-three patients were included in the analysis, with cryo-HPC representing 40%. Median viability was higher in the fresh-HPC group at 99.2% (IQR 98.3-99.5) versus cryo-HPCs at 97.0% (96.0, 98.6) (P < 0.01). The 12-month actuarial survivals were 77% in the fresh HPC and 75% in the cryo-HPC groups (P = 0.21). There were no differences between cryo-HPCs and fresh HPCs on univariable analysis of OS, CIR, or NRM. There was a shorter median time to platelet engraftment in patients receiving fresh HPC at 17 days (IQR 16, 20) versus cryo-HPC at 21 days (IQR 18, 29), P < 0.001. There was a shorter median time to neutrophil engraftment in the fresh HPC group at 17 days (IQR 14, 20) versus 20 days (17, 23), P < 0.001. Cryo-HPC accounted for 5 out of 6 cases of primary graft failure (P = 0.04), and 3 out of five cases of secondary graft failure (P = 0.39). There were no significant differences in acute GVHD between the fresh HPC and cryo-HPC groups (P = 0.34). The incidence of moderate or severe chronic GVHD was 32% in the fresh-HPC group and 17% in the cryo-HPC group (P < 0.001). In multivariable analysis, cryopreservation did not emerge as an independent predictor of OS, CIR, NRM, primary GF or secondary GF. However, viability <90% on arrival at our center was a significant predictor of OS (HR 5.3, 2.3-12.3, P < 0.01), primary graft failure (OR 36.3, 5.4-210.2, P < 0.01), and secondary graft failure (OR 18.4, 1.7-121.1, P < 0.01). CONCLUSIONS: Patients who received cryo-HPCs had similar OS and relapse rates to those who received fresh-HPCs but typically took 2-3 days longer to achieve engraftment of platelets or neutrophils and were associated increased primary graft failure. However, after accounting for multiple variables, cryopreservation was no longer a significant predictor of survival or engraftment while viability <90% emerged as an important predictor of OS, primary graft failure, and secondary graft failure. If confirmed, this suggests that viability on arrival at the infusion center may be a good quality control indicator used to identify HPC products that may warrant recollection if the risk of graft failure is sufficiently increased.

Outcomes of a Double-Cup Construct to Treat Paprosky 3A and 3B Acetabular Defects at a Mean of 39 Months.

BACKGROUND: Modular reconstruction systems based on porous tantalum (PT) prosthetic components have been increasingly used for the treatment of complex acetabular bone defects in revision total hip arthroplasty. We report a novel technique that applies a revision cup as a "super-augment" to form a "double-cup" construct for Paprosky type III defects. METHODS: A retrospective review was conducted on revision total hip arthroplasty cases, comparing those treated with double-cup constructs (DC group, n = 48) to those treated with PT shells and augments (PT group, n = 48). All procedures were performed at the same institute between 2017 and 2022. Clinical outcome evaluation utilized the Harris Hip Score, Oxford Hip Score, and the 36-Item Short Form Survey. Preoperative and postoperative radiographic assessments measured hip center of rotation (COR) position and leg length discrepancy. Additionally, postoperative complications and implant survivorship were monitored during the follow-up period. RESULTS: The clinical outcomes improved substantially in both groups, which showed no significant difference in the Harris Hip Score (P = .786), the Oxford Hip Score (P = .570), and the 36-Item Short Form Survey (P = .691). Compared to the PT group, the reconstruction COR was significantly closer to the anatomic COR (vertical distance: 2.630 versus 7.355 mm, P = .0034; horizontal distance: 1.881 versus -6.413 mm, P < .0001) in Paprosky 3B type defects. Additionally, postoperative leg length discrepancy was less in the DC group (-8.252 versus -1.821 mm, P = .0008). Dislocation was the main complication in the DC group, and only 1 patient received re-revision due to repeated dislocation. The cumulative survival rate of the DC group (100%; 95% confidence interval 100) was better than the PT group (83.4%; 95% confidence interval 70.5 to 98.6) when re-revisions for aseptic loosening were the endpoint (P = .046). CONCLUSIONS: The DC is a reliable revision technique for the reconstruction of Paprosky type III bone defects. Although dislocation remains challenging, the biomechanically superior restoration achieved by this technique lowers the risk of aseptic loosening.

Stress modulates Ahi1-dependent nuclear localization of ten-eleven translocation protein 2.

Major depression disorder is one of the most common psychiatric diseases. Recent evidence supports that environmental stress affects gene expression and promotes the pathological process of depression through epigenetic mechanisms. Three ten-eleven translocation (Tet) enzymes are epigenetic regulators of gene expression that promote 5-hydroxymethylcytosine (5hmC) modification of genes. Here, we show that the loss of Tet2 can induce depression-like phenotypes in mice. Paradoxically, using the paradigms of chronic stress, such as chronic mild stress and chronic social defeat stress, we found that depressive behaviors were associated with increased Tet2 expression but decreased global 5hmC level in hippocampus. We examined the genome-wide 5hmC profile in the hippocampus of Tet2 knockout mice and identified 651 dynamically hydroxymethylated regions, some of which overlapped with known depression-associated loci. We further showed that chronic stress could induce the abnormal nuclear translocation of Tet2 protein from cytosol. Through Tet2 immunoprecipitation and mass spectrum analyses, we identified a cellular trafficking protein, Abelson helper integration site-1 (Ahi1), which could interact with Tet2 protein. Ahi1 knockout or knockdown caused the accumulation of Tet2 in cytosol. The reduction of Ahi1 protein under chronic stress explained the abnormal Ahi1-dependent nuclear translocation of Tet2. These findings together provide the evidence for a critical role of modulating Tet2 nuclear translocation in regulating stress response.

RhoA suppresses pseudorabies virus replication in vitro.

The porcine pseudorabies virus (PRV) is one of the most devastating pathogens and brings great economic losses to the swine industry worldwide. Viruses are intracellular parasites that have evolved numerous strategies to subvert and utilize different host processes for their life cycle. Among the different systems of the host cell, the cytoskeleton is one of the most important which not only facilitate viral invasion and spread into neighboring cells, but also help viruses to evade the host immune system. RhoA is a key regulator of cytoskeleton system that may participate in virus infection. In this study, we characterized the function of RhoA in the PRV replication by chemical drugs treatment, gene knockdown and gene over-expression strategy. Inhibition of RhoA by specific inhibitor and gene knockdown promoted PRV proliferation. On the contrary, overexpression of RhoA or activation of RhoA by chemical drug inhibited PRV infection. Besides, our data demonstrated that PRV infection induced the disruption of actin stress fiber, which was consistent with previous report. In turn, the actin specific inhibitor cytochalasin D markedly disrupted the normal fibrous structure of intracellular actin cytoskeleton and decreased the PRV replication, suggesting that actin cytoskeleton polymerization contributed to PRV replication in vitro. In summary, our data displayed that RhoA was a host restriction factor that inhibited PRV replication, which may deepen our understanding the pathogenesis of PRV and provide further insight into the prevention of PRV infection and the development of anti-viral drugs.

COVID-19 information exposure and vaccine hesitancy: The influence of trust in government and vaccine confidence.

As the novel coronavirus (COVID-19) continues to spread, vaccine hesitancy increasingly threats public health worldwide. Health information from traditional, online and social media may influence vaccine hesitancy. The purpose of this study was to explore the influence of exposure to COVID-19 information from various media on vaccine hesitancy, as well as the mediating roles of public trust in government and vaccine confidence. With a sample of 438 online participants (mean age = 30.69 years) responding to an anonymous questionnaire, the study tested a mediation model using bias-corrected bootstrap. The results indicated that exposure to COVID-19 information from online news media and traditional media can reduce vaccine hesitancy indirectly. Whereas a positive and indirect relationship between COVID-19 information exposure on social media and vaccine hesitancy was revealed. Trust in government and vaccine confidence were found to be salient mediators between exposure to COVID-19 information from various media and vaccine hesitancy. Findings from this study offer implications for strategies to address vaccine hesitancy.

Ahi1 regulates serotonin production by the GR/ERß/TPH2 pathway involving sexual differences in depressive behaviors.

BACKGROUND: Depression is one of the most common psychiatric diseases. The monoamine transmitter theory suggests that neurotransmitters are involved in the mechanism of depression; however, the regulation on serotonin production is still unclear. We previously showed that Ahi1 knockout (KO) mice exhibited depression-like behavior accompanied by a significant decrease in brain serotonin. METHODS: In the present study, western blot, gene knockdown, immunofluorescence, dual-luciferase reporter assay, and rescue assay were used to detect changes in the Ahi1/GR/ERß/TPH2 pathway in the brains of male stressed mice and male Ahi1 KO mice to explain the pathogenesis of depression-like behaviors. In addition, E2 levels in the blood and brain of male and female mice were measured to investigate the effect on the ERß/TPH2 pathway and to reveal the mechanisms for the phenomenon of gender differences in depression-like behaviors. RESULTS: We found that the serotonin-producing pathway-the ERß/TPH2 pathway was inhibited in male stressed mice and male Ahi1 KO mice. We further demonstrated that glucocorticoid receptor (GR) as a transcription factor bound to the promoter of ERß that contains glucocorticoid response elements and inhibited the transcription of ERß. Our recent study had indicated that Ahi1 regulates the nuclear translocation of GR upon stress, thus proposing the Ahi1/GR/ERß/TPH2 pathway for serotonin production. Interestingly, female Ahi1 KO mice did not exhibit depressive behaviors, indicating sexual differences in depressive behaviors compared with male mice. Furthermore, we found that serum 17ß-estradiol (E2) level was not changed in male and female mice; however, brain E2 level significantly decreased in male but not female Ahi1 KO mice. Further, ERß agonist LY-500307 increased TPH2 expression and 5-HT production. Therefore, both Ahi1 and E2 regulate the ERß/TPH2 pathway and involve sexual differences in brain serotonin production and depressive behaviors. CONCLUSIONS: In conclusion, although it is unclear how Ahi1 controls E2 secretion in the brain, our findings demonstrate that Ahi1 regulates serotonin production by the GR/ERß/TPH2 pathway in the brain and possibly involves the regulation on sex differences in depressive behaviors. Video Abstract.

Heterometallic Polyoxotitanium Clusters as Bifunctional Electrocatalysts for Overall Water Splitting.

Incorporating heterometal into titanium-oxygen clusters (TOCs) is an effective way to improve its catalytic activity. Herein, we synthesize three novel heterometallic TOCs with the formula of [Ti6Cu2O7(Dmg)2(OAc)4(iPrO)6][H2Ti6Cu2O7(Dmg)2(OAc)4(iPrO)8] ({Ti6Cu2}), [Ti8Cu2O9(Dmg)2(OAc)2(iPrO)12] ({Ti8Cu2}), and [Ti10Co2O6(Dmg)2(Pdc)4(iPrO)18Cl3] ({Ti10Co2}, DmgH2 = dimethylglyoxime; PdcH2 = pyridine-2,3-dicarboxylic acid) using dimethylglyoxime and different carboxylates as the synergistic ligands. By depositing the clusters {Ti6Cu2} and {Ti10Co2} on carbon cloth as electrodes, we investigated the electrocatalytic performance of TOCs for full water splitting for the first time. To reach a 10 mA cm-2 current density in an alkaline solution, the {Ti10Co2}@CC electrode needs an overpotential as low as 120 and 400 mV for the hydrogen evolution reaction (HER) and the oxygen evolution reaction (OER), respectively. In addition, full water-splitting equipment with {Ti10Co2}@CC as a cathode and an anode need only 1.67 V to deliver a current density of 10 mA cm-2. Our work confirmed the potential of noble metal-free TOCs as bifunctional cluster-based electrocatalysts for water splitting, and their activities can be tuned by doping with different metal ions.

HRP-conjugated-nanobody-based cELISA for rapid and sensitive clinical detection of ASFV antibodies.

African swine fever (ASF), which is caused by the ASF virus (ASFV), is a highly contagious hemorrhagic disease that causes high mortality to domestic porcine and wild boars and brings huge economic losses to world swine industry. Due to the lack of an effective vaccine, the control of ASF must depend on early, efficient, and cost-effective detection and strict control and elimination strategies. Traditional serological testing methods are generally associated with high testing costs, complex operations, and high technical requirements. As a promising alternative diagnostic tool to traditional antibodies, nanobodies (Nb) have the advantages of simpler and faster generation, good stability and solubility, and high affinity and specificity, although the system is dependent on the immunization of Bactrian camels to obtain the specific VHH library of the target protein. The application of Nbs in the detection of ASFV antibodies has not yet been reported yet. Using a phage display technology, one Nb against the ASFV p54 protein that exhibited high specificity and affinity, Nb8, was successfully screened. A HEK293T cell line stably expressing Nb8-horseradish peroxidase (HRP) fusion protein was established using the lentiviral expression system. Following the optimization of the reaction conditions, the Nb8-HRP fusion protein was successfully used to establish a competitive enzyme-linked immunosorbent assay (cELISA) to detect ASFV-specific antibodies in pig serum, for the first time. There was no cross-reaction with healthy pig serum, porcine pseudorabies virus (PRV), porcine reproductive and respiratory syndrome virus (PRRSV), classical swine fever virus (CSFV), porcine epidemic diarrhea virus (PEDV), and classical swine fever virus (CSFV) positive sera. The optimal cut-off value for the cELISA by ROC analysis was 52.5%. A total of 209 serum samples were tested using the developed cELISA and a commercial ELISA kit. The results showed that the relative specificity of the cELISA was 98.97%, and the relative sensitivity of the cELISA was 93.3%, with the percent agreement between the two ELISA methods being 98.56%. In conclusion, a specific, sensitive, and repeatable cELISA was successfully developed based on the Nb8 as a probe, providing a promising method for the detection of anti-ASFV antibodies in clinical pig serum. KEY POINTS: • We successfully screened a specific, high affinity nanobody against ASFV p54 protein. • We establish a method for continuous and stable expression of Nb-HRP fusion protein using a lentiviral packaging system. • We establish a nanobody cELISA detection method that can monitor an ASF infection.

Splenic-targeting biomimetic nanovaccine for elevating protective immunity against virus infection.

BACKGROUND: The prevalence of viral infectious diseases has become a serious threat to public safety, economic and social development. Vaccines have been served as the most effective platform to prevent virus transmission via the activation of host immune responses, while the low immunogenicity or safety, the high cost of production, storage, transport limit their effective clinical application. Therefore, there is a need to develop a promising strategy to improve the immunogenicity and safety of vaccines. METHODS: We developed a splenic-targeting biomimetic nanovaccine (NV) that can boost protective humoral and cellular immunity against african swine fever virus (ASFV) infection. The universal PLGA nanoparticles (CMR-PLGA/p54 NPs) coated with mannose and CpG (TLR9 agonist) co-modified red blood cell (RBC) membrane were prepared, which comprised a viral antigen (p54) and can be served as a versatile nanovaccine for elevating protective immunity. RESULTS: CMR-PLGA/p54 NVs could be effectively uptaken by BMDC and promoted BMDC maturation in vitro. After subcutaneous immunization, antigen could be effectively delivered to the splenic dendritic cells (DCs) due to the splenic homing ability of RBC and DC targeting capacity of mannose, which promoted antigen presentation and DCs maturation, and further elicited higher levels of cytokines secretion and specific IgG titers, CD4+ and CD8+ T cells activation and B maturation. Moreover, NVs demonstrated notable safety during the immunization period. CONCLUSIONS: This study demonstrates the high potential of CMR-PLGA NPs as vaccine delivery carriers to promote humoral and cellular immune responses, and it provides a promising strategy to develop safe and effective vaccines against viral infectious diseases.

Octopaminergic neurons have multiple targets in Drosophila larval mushroom body calyx and can modulate behavioral odor discrimination.

Discrimination of sensory signals is essential for an organism to form and retrieve memories of relevance in a given behavioral context. Sensory representations are modified dynamically by changes in behavioral state, facilitating context-dependent selection of behavior, through signals carried by noradrenergic input in mammals, or octopamine (OA) in insects. To understand the circuit mechanisms of this signaling, we characterized the function of two OA neurons, sVUM1 neurons, that originate in the subesophageal zone (SEZ) and target the input region of the memory center, the mushroom body (MB) calyx, in larval Drosophila We found that sVUM1 neurons target multiple neurons, including olfactory projection neurons (PNs), the inhibitory neuron APL, and a pair of extrinsic output neurons, but relatively few mushroom body intrinsic neurons, Kenyon cells. PN terminals carried the OA receptor Oamb, a Drosophila α1-adrenergic receptor ortholog. Using an odor discrimination learning paradigm, we showed that optogenetic activation of OA neurons compromised discrimination of similar odors but not learning ability. Our results suggest that sVUM1 neurons modify odor representations via multiple extrinsic inputs at the sensory input area to the MB olfactory learning circuit.

[Up-regulated expression of miR-576 inhibits ALK4 expression, regulates JAK/STAT signaling pathway and promotes proliferation and migration of prostatic cancer cells].

OBJECTIVE: To explore the mechanism of the action of the miR-576/ALK4 axis on the progression of prostate cancer (PCa). METHODS: PCa cells were transfected with miR-576 mimics/inhibitor, the proliferation and migration distance of the cells were detected by MTT and scratch wound healing assay, respectively. The targeted regulation effect of miR-576 on ALK4 was verified by dual-luciferase reporter assay. The effects of miR-576 on the mRNA and protein expressions and phosphorylation levels of the ALK4 and JAK/STAT signaling pathway factors JAK2 and STAT3 were determined by qPCR and Western blot, respectively. The C4-2 cells were co-treated with sh-ALK4 and Ruxolitinib for measurement of the proliferation and migration of the PCa cells. RESULTS: Bioinformatics analysis and binding site prediction showed that miR-576 was up-regulated in the PCa cells, and dual-luciferase reporter assay revealed its targeted regulation effect on ALK4 and its impact on the phosphorylation levels of JAK2 and STAT3. Overexpressed miR-576 promoted while knocked-down miR-576 inhibited the proliferation and migration of the PCa cells. sh-ALK4 increased the proliferation and migration of the cells, while Ruxolitinib suppressed the promoting effect of sh-ALK4. CONCLUSION: The expression of miR-576 is up-regulated in PCa, inhibits the expression of ALK4, regulates the activity of the JAK and STAT signaling pathways, and promotes the proliferation and migration of PCa cells.

Interferon-Induced Transmembrane Protein 3 Is a Virus-Associated Protein Which Suppresses Porcine Reproductive and Respiratory Syndrome Virus Replication by Blocking Viral Membrane Fusion.

Porcine reproductive and respiratory syndrome virus (PRRSV) infection eliminates production of type I interferons (IFNs) in host cells, which triggers an antiviral immune response through the induction of downstream IFN-stimulated genes (ISGs), thus escaping the fate of host-mediated clearance. The IFN-induced transmembrane 3 (IFITM3) has recently been identified as an ISG and plays a pivotal role against enveloped RNA viruses by restricting cell entry. However, the role of IFITM3 in PRRSV replication is unknown. The present study demonstrated that overexpression of IFITM3 suppresses PRRSV replication, while silencing of endogenous IFITM3 prominently promoted PRRSV replication. Additionally, it was shown that IFITM3 undergoes S-palmitoylation and ubiquitination modification, and both posttranslational modifications contribute to the anti-PRRSV activity of IFITM3. Further study showed that PRRSV particles are transported into endosomes and then into lysosomes during the early stages of infection, and confocal microscopy results revealed that PRRSV particles are transported to IFITM3-positive cellular vesicles. By using a single virus particle fluorescent labeling technique, we confirmed that IFITM3 can restrict PRRSV membrane fusion by inducing accumulation of cholesterol in cellular vesicles. Additionally, we found that both endogenous and exogenous IFITM3 are incorporated into newly producing PRRS virions and diminish viral intrinsic infectivity. By using cell coculture systems, we found that IFITM3 effectively restricted PRRSV intercellular transmission, which may have been caused by disrupted membrane fusion and reduced viral infectivity. In conclusion, our results demonstrate, for the first time, that swine IFITM3 interferes with the life cycle of PRRSV, and possibly other enveloped arteritis viruses, at multiple steps.IMPORTANCE Porcine reproductive and respiratory syndrome (PRRS), which is caused by PRRS virus (PRRSV), is of great economic significance to the swine industry. Due to the complicated immune escape mechanisms of PRRSV, there are no effective vaccines or therapeutic drugs currently available against PRRS. Identification of cellular factors and underlying mechanisms that establish an effective antiviral state against PRRSV can provide unique strategies for developing antiviral vaccines or drugs. As an interferon (IFN)-stimulated gene, the role of IFN-induced transmembrane 3 (IFITM3) in PRRSV infection has not been reported as of yet. In the present study, it was shown that IFITM3 can exert a potent anti-PRRSV effect, and PRRS virions are trafficked to IFITM3-containing cell vesicles, where viral membrane fusion is impaired by cholesterol accumulation that is induced by IFITM3. Additionally, both endogenous and exogenous IFITM3 are incorporated into newly assembled progeny virions, and this decreased their intrinsic infectivity.

MiR-410-3p activates the NF-κB pathway by targeting ZCCHC10 to promote migration, invasion and EMT of colorectal cancer.

BACKGROUND: Colorectal cancer (CRC) is a prevalent malignancy of the digestive tract. miR-410-3p is involved in oncogenesis and development of CRC, but the specific regulation mechanism is still not known clearly. METHODS: The expression of miR-410-3p and zinc finger CCHC-type containing 10 (ZCCHC10) in CRC cells was detected by qRT-PCR and western blot method, respectively. The dual-luciferase reporter gene detection was applied for determination of interaction between miR-410-3p and ZCCHC10. The wound healing assay and transwell assay were carried out to measure cell migration and invasive ability, respectively. RESULTS: The miR-410-3p expression levels were markedly increased, but ZCCHC10 levels were reduced in CRC cells and tissues. Dual-luciferase reporter gene detection indicated that miR-410-3p targeted ZCCHC10 directly. Functionally knockdown of ZCCHC10 or overexpression of miR-410-3p activated nuclear factor-κB (NF-κB) signaling pathway, promoted epithelial-mesenchymal transition (EMT) process, as well as cell migration and invasion of CRC cells. After adding NF-κB inhibitor BAY 11-708 to inhibit NF-κB pathway, the promoting effects of si-ZCCHC10 on cell migration, invasion and EMT of HT29 and SW480 cells were suppressed. Meanwhile, overexpression of ZCCHC10 inhibited the effects of miR-410-3p on cell migration, invasion and EMT of HT29 and SW480. CONCLUSION: miR-410-3p-mediated ZCCHC10 suppression regulates NF-κB activation, thereby promoting EMT process, cell migration and invasion of CRC cells. This study provides a new insight into the specific mechanism by which miR-410-3p mediates CRC progression.

Self-reported pain in breast cancer patients receiving adjuvant radiotherapy.

PURPOSE: Breast cancer patients receiving radiotherapy (RT) commonly report pain, contributing to physical and emotional distress, and potentially resulting in poor quality of life. This study prospectively identified trends and risk factors in patient-reported pain associated with breast irradiation using the Edmonton Symptom Assessment Scale (ESAS) and a study-specific Skin Symptom Assessment (SSA). METHODS: Before RT and once per week during RT, patients completed the ESAS and SSA. Upon RT completion, patients were contacted via telephone to complete both assessments weekly for 6 weeks, and a final assessment was conducted 1-3 months post-RT. Only data from patients who had completed both assessments before, at least once during, and at least once after RT were included in our analysis. RESULTS: A total of 426 patients provided data for the analysis. Overall acute pain increased significantly at week 1-2 (p < 0.0001), week 5 (p = 0.0011), and at 1-3 months (p < 0.0001) post-RT compared with baseline, and acute breast pain increased significantly at week 1 (p < 0.001) and week 2 (p = 0.0002) post-RT compared with baseline. Previous chemotherapy (adjuvant or neoadjuvant) in mastectomy patients was associated with increased overall pain compared with mastectomy patients without previous chemotherapy (p = 0.017). Younger patients (40-49 or 50-59 years of age) reported more overall pain (p = 0.0001, p = 0.038) and breast pain (p = 0.0003, p = 0.0038) compared with patients ≥ 60 years of age. CONCLUSIONS: Patient-reported pain associated with breast irradiation peaked 1 week after RT completion. Our findings provide support for closer monitoring of acute pain associated with breast RT in younger patients.

Efficacy of a live attenuated highly pathogenic PRRSV vaccine against a NADC30-like strain challenge: implications for ADE of PRRSV.

BACKGROUND: Porcine reproductive and respiratory syndrome virus (PRRSV) infection can cause severe reproductive failure in sows and respiratory distress in pigs of all ages, leading to major economic losses. To date, there are still no effective strategies to prevent and control PRRSV. Antibody-dependent enhancement (ADE), a phenomenon in which preexisting non-neutralizing antibodies or sub-neutralizing antibodies facilitate virus entry and replication, may be a significant obstacle in the development of effective vaccines for many viruses, including PRRSV. However, the contribution of ADE to PRRSV infection remains controversial, especially in vivo. Whether attenuated PRRSV vaccines prevent or worsen subsequent disease in pigs infected by novel PRRSV strains requires more research. In the present study, in vivo experiments were conducted to evaluate ADE under different immune statuses, which were produced by waiting different lengths of time after vaccination with a commercially available attenuated highly pathogenic PRRSV (HP-PRRSV) vaccine (JXA1-R) before challenging the pigs with a novel heterologous NADC30-like strain. RESULTS: Piglets that were vaccinated before being challenged with PRRSV exhibited lower mortality rates, lower body temperatures, higher bodyweight gain, and lower viremia. These results demonstrate that vaccination with JXA1-R alleviated the clinical signs of PRRSV infection in all vaccinated groups. CONCLUSIONS: The obtained data indicate that the attenuated vaccine test here provided partial protection against the NADC30-like strain HNhx. No signs of enhanced PRRSV infection were observed under the applied experimental conditions. Our results provide some insight into the molecular mechanisms underlying vaccine-induced protection or enhancement in PRRSV.

Association between serum 25-hydroxy vitamin D level and metabolic associated fatty liver disease (MAFLD)-a population-based study.

Metabolic associated fatty liver disease (MAFLD) is a new concept proposed in 2020. This study aimed to explore the relationship between serum 25-hydroxy vitamin D (25(OH)D) level and MAFLD based on a population survey dataset (the third National Health and Nutrition Examination Surveys of the United States). Multivariate logistic regression was used to estimate the odds ratio (OR) of serum 25(OH)D level for MAFLD. A total of 12,878 participants were included in this analysis. Among them, 4,027 (31.27%) cases were diagnosed with MAFLD and 8,851 (66.40%) were without MAFLD (non-MAFLD). Patients with vitamin D sufficiency and insufficiency totaled 6,983 (54.22%) and 5,895 (45.78%), respectively. The incidence of MAFLD and the grade of hepatic steatosis were both significantly higher in vitamin D insufficiency group. Multivariate analysis showed that vitamin D insufficiency was an independent risk factor for MAFLD after adjusted for other confounders (OR: 1.130, 95%CI: 1.035 to 1.234). In MAFLD population, the average serum 25(OH)D level decreased with the numbers of metabolic risks in MAFLD cases. Serum 25(OH)D level was not associated with the severity of fibrosis or steatosis in MAFLD group. In Conclusion, lower serum 25(OH)D level is associated with higher prevalence of MAFLD in general population. No relationship was found between serum 25(OH)D level and the severity of hepatic steatosis or fibrosis in MAFLD.

Patient-Reported Pain in Patients with Breast Cancer Who Receive Radiotherapy.

BACKGROUND: Patients who receive radiation treatment (RT) for breast cancer often report pain, which contributes negatively to quality of life (QoL). AIMS: To identify demographic, treatment, and disease characteristics associated with pain and changes in pain before and after RT using the Edmonton Symptom Assessment Scale (ESAS). DESIGN: Retrospective study. SETTINGS: Odette Cancer Centre. PARTICIPANTS: Patients diagnosed with nonmetastatic breast cancer from January 2011-June 2017 with at least one ESAS completed pre-RT and one completed post-RT. METHODS: Data on systemic treatment, radiation, patient demographics, and disease stage were extracted. To identify factors associated with pain before and after RT and changes in pain, univariate and multivariate general linear regression analysis were conducted. p < .05 was considered statistically significant. RESULTS: This study included 1,222 female patients with a mean age of 59 years. ESAS was completed an average of 28 days before RT (baseline) and 142 days after RT, respectively. In multivariable analysis, higher baseline pain scores were associated with having recently completed adjuvant chemotherapy (p = .002) and eventual receipt of locoregional (p = .026) or chest wall (p = .003) radiation. Adjuvant chemotherapy (p = .002) and chest wall radiation (p = .03), were associated with a significant reduction in pain score after radiotherapy, while locoregional RT was associated with a higher pain score after RT (p < .001). CONCLUSIONS: Patients with locoregional RT had higher baseline pain that remained elevated after RT completion and should be screened for pain and provided with pain management and support when necessary.