The extracellular cyclophilin A-integrin ß2 complex as a therapeutic target of viral pneumonia.

The acute respiratory virus infection can induce uncontrolled inflammatory responses, such as cytokine storm and viral pneumonia, which are the major causes of death in clinical cases. Cyclophilin A (CypA) is mainly distributed in the cytoplasm of resting cells and released into the extracellular space in response to inflammatory stimuli. Extracellular CypA (eCypA) is upregulated and promotes inflammatory response in severe COVID-19 patients. However, how eCypA promotes virus-induced inflammatory response remains elusive. Here, we observe that eCypA is induced by influenza A and B viruses and SARS-CoV-2 in cells, mice, or patients. Anti-CypA mAb reduces pro-inflammatory cytokines production, leukocytes infiltration, and lung injury in virus-infected mice. Mechanistically, eCypA binding to integrin ß2 triggers integrin activation, thereby facilitating leukocyte trafficking and cytokines production via the focal adhesion kinase (FAK)/GTPase and FAK/ERK/P65 pathways, respectively. These functions are suppressed by the anti-CypA mAb that specifically blocks eCypA-integrin ß2 interaction. Overall, our findings reveal that eCypA-integrin ß2 signaling mediates virus-induced inflammatory response, indicating that eCypA is a potential target for antibody therapy against viral pneumonia.

Cyclophilin A facilitates influenza B virus replication by stabilizing viral proteins.

Influenza B circulates annually and causes substantial disease burden in humans. However, little is known about the infection mechanisms of influenza B virus (IBV). Here, we find that the host factor cyclophilin A (CypA) facilitates IBV replication by targeting IBV non-structural protein 1 (BNS1) and nucleoprotein (BNP). CypA promotes OTUD4-mediated K48-linked BNS1 deubiquitination to stabilize BNS1 by upregulating OTUD4 expression. Meanwhile, CypA and the E3 ligase MIB1 competitively interact with BNP to inhibit its proteasomal degradation. Moreover, cyclosporine A treatment or CypA R55A mutation results in an impaired function of CypA in IBV replication. Notably, BNP hijacks CypA into the nucleus to enhance the activity of viral ribonucleoprotein complexes by enhancing the interaction between BNP and IBV polymerase basic protein 1. Taken together, this study unveils the critical role of CypA in facilitating IBV replication, suggesting that CypA is a promising target for anti-IBV drug.

[The I226R protein of African swine fever virus inhibits the cGAS-STING-mediated innate immune response].

This study aimed to explore the mechanism of how African swine fever virus (ASFV) I226R protein inhibits the cGAS-STING signaling pathway. We observed that I226R protein (pI226R) significantly inhibited the cGAS-STING-mediated type Ⅰ interferons and the interferon-stimulated genes production by dual-luciferase reporter assay system and real-time quantitative PCR. The results of co-immunoprecipitation assay and confocal microscopy showed that pI226R interacted with cGAS. Furthermore, pI226R promoted cGAS degradation through autophagy-lysosome pathway. Moreover, we found that pI226R decreased the binding of cGAS to E3 ligase tripartite motif protein 56 (TRIM56), resulting in the weakened monoubiquitination of cGAS, thus inhibiting the activation of cGAS and cGAS-STING signaling. In conclusion, ASFV pI226R suppresses the antiviral innate immune response by antagonizing cGAS, which contributes to an in-depth understanding of the immune escape mechanism of ASFV and provides a theoretical basis for the development of vaccines.

[Eukaryotic expression of GP5 and M protein of porcine reproductive and respiratory syndrome virus and immunogenicity evaluation].

In order to understand the prevalence and evolution of porcine reproductive and respiratory syndrome virus (PRRSV) in China and to develop subunit vaccine against the epidemic lineage, the genetic evolution analysis of PRRSV strains isolated in China from 2001 to 2021 was performed. The representative strains of the dominant epidemic lineage were selected to optimize the membrane protein GP5 and M nucleotide sequences, which were used, with the interferon and the Fc region of immunoglobulin, to construct the eukaryotic expression plasmids pCDNA3.4-IFNα-GP5-Fc and pCDNA3.4-IFNα-M-Fc. Subsequently, the recombinant proteins IFNα-GP5-Fc and IFNα-M-Fc were expressed by HEK293T eukaryotic expression system. The two recombinant proteins were mixed with ISA206VG adjuvant to immunize weaned piglets. The humoral immunity level was evaluated by ELISA and neutralization test, and the cellular immunity level was detected by ELISPOT test. The results showed that the NADC30-like lineage was the main epidemic lineage in China in recent years, and the combination of IFNα-GP5-Fc and IFNα-M-Fc could induce high levels of antibody and cellular immunity in piglets. This study may facilitate the preparation of a safer and more effective new PRRSV subunit vaccine.

SARS-CoV-2 nonstructural protein 6 triggers endoplasmic reticulum stress-induced autophagy to degrade STING1.

ABBREVIATIONS: aa: amino acid; ATF6: activating transcription factor 6; ATG5: autophagy related 5; CCPG1: cell cycle progression 1; CFTR: CF transmembrane conductance regulator; cGAMP: cyclic GMP-AMP; CGAS: cyclic GMP-AMP synthase; CHX: cycloheximide; Co-IP: co-immunoprecipitation; CQ: chloroquine; EIF2A/eIF2α: eukaryotic translation initiation factor 2A; EIF2AK3/PERK: eukaryotic translation initiation factor 2 alpha kinase 3; ER: endoplasmic reticulum; ERN1/IRE1: endoplasmic reticulum to nucleus signaling 1; GFP: green fluorescent protein; HSPA5/GRP78: heat shock protein family A (Hsp70) member 5; HSV-1: herpes simplex virus type 1; IFIT1: interferon induced protein with tetratricopeptide repeats 1; IFNB1/IFN-ß: interferon beta 1; IRF3: interferon regulatory factor 3; ISG15: ISG15 ubiquitin like modifier; MAP1LC3B/LC3B: microtubule associated protein 1 light chain 3 beta; MAP3K7/TAK1: mitogen-activated protein kinase kinase kinase 7; MAVS: mitochondrial antiviral signaling protein; MOI: multiplicity of infection; NFKB/NF-κB: nuclear factor kappa B; NSP6: non-structural protein 6; Δ106-108: deletion of amino acids 106-108 in NSP6 of SARS-CoV-2; Δ105-107: deletion of amino acids 105-107 in NSP6 of SARS-CoV-2; RETREG1/FAM134B: reticulophagy regulator 1; RIGI/DDX58: RNA sensor RIG-I; SQSTM1/p62: sequestosome 1; STING1: stimulator of interferon response cGAMP interactor 1; TBK1: TANK binding kinase 1.

Curative Effect and Mechanisms of Radix Arnebiae Oil on Burn Wound Healing in Rats.

Radix arnebiae oil (RAO) is a clinically useful traditional Chinese medical formula with outstanding curative effects on burns. However, the mechanism of the effect of RAO on wound healing remains unclear. The present study investigates the molecular mechanisms of the potential curative effect of RAO on wound healing. The concentrations of the main constituents, shikonin, imperatorin, and ferulic acid in RAO detected by HPLC were 24.57, 3.15, and 0.13 mg/mL, respectively. A rat burn model was established, and macroscopic and histopathological studies were performed. RAO significantly accelerated wound closure and repair scarring, increased superoxide dismutase activities, and reduced malondialdehyde. RAO also downregulated interleukin (IL)-6, IL-1ß and tumor necrosis factor-α in wound tissues and increased secretion of vascular endothelial growth factor, epidermal growth factor, and transforming growth factor (TGF)-ß1. RAO increased the gene expression of TGF-ß1, type I and III collagen, and increased the protein expression of TGF-ß1 and phosphorylation of PI3K and Akt. In conclusion, RAO likely promotes wound healing via antioxidant and anti-inflammatory activities and increases re-epithelization. Activation of the TGF-ß1/PI3K/Akt pathway may play an important role in the healing efficacy of RAO. These findings suggest that RAO could be a promising alternative local treatment for burn wound healing.

Cyclosporine A Regulates Influenza A Virus-induced Macrophages Polarization and Inflammatory Responses by Targeting Cyclophilin A.

Cyclosporine A (CsA) is an immunosuppressive drug that suppresses T cell responses and is broadly used in transplantation. Its immunosuppressive action is closely linked to its binding of cyclophilin A (CypA), which widely distributed in different cell types. CsA also regulates the functions of innate immune cells, but the mechanism remains elusive. Here, we investigate the role of CsA in regulating macrophages polarization in influenza A virus-infected mice and mouse bone marrow-derived macrophages. CsA downregulates pro-inflammatory cytokines expression and upregulates anti-inflammatory cytokines expression. Mechanically, CsA decreases the polarization of macrophages into pro-inflammatory M1 phenotype and increases the polarization of macrophages into anti-inflammatory M2 phenotype. Further studies show that CsA regulates macrophages polarization-associated IFN-γ/STAT1 and IL-4/STAT6 signaling pathways. Meanwhile, all these roles of CsA are eliminated when CypA is absent, suggesting that CsA regulates macrophages polarization and inflammatory responses depend on its binding to CypA. Collectively, these results reveal a crucial mechanism of CsA in attenuating IAV-induced inflammatory responses by a switch in macrophages polarization.

Wearable multimode sensor with a seamless integrated structure for recognition of different joint motion states with the assistance of a deep learning algorithm.

Accurate motion feature extraction and recognition provide critical information for many scientific problems. Herein, a new paradigm for a wearable seamless multimode sensor with the ability to decouple pressure and strain stimuli and recognize the different joint motion states is reported. This wearable sensor is integrated into a unique seamless structure consisting of two main parts (a resistive component and a capacitive component) to decouple the different stimuli by an independent resistance-capacitance sensing mechanism. The sensor exhibits both high strain sensitivity (GF = 7.62, 0-140% strain) under the resistance mechanism and high linear pressure sensitivity (S = 3.4 kPa-1, 0-14 kPa) under the capacitive mechanism. The sensor can differentiate the motion characteristics of the positions and states of different joints with precise recognition (97.13%) with the assistance of machine learning algorithms. The unique integrated seamless structure is achieved by developing a layer-by-layer casting process that is suitable for large-scale manufacturing. The proposed wearable seamless multimode sensor and the convenient process are expected to contribute significantly to developing essential components in various emerging research fields, including soft robotics, electronic skin, health care, and innovative sports systems applications.

Designs for order-of-addition experiments.

The order-of-addition experiment aims at determining the optimal order of adding components such that the response of interest is optimized. Order of addition has been widely involved in many areas, including bio-chemistry, food science, nutritional science, pharmaceutical science, etc. However, such an important study is rather primitive in statistical literature. In this paper, a thorough study on pair-wise ordering designs for order of addition is provided. The recursive relation between two successive full pair-wise ordering designs is developed. Based on this recursive relation, the full pair-wise ordering design can be obtained without evaluating all the orders of components. The value of the D-efficiency for the full pair-wise ordering model is then derived. It provides a benchmark for choosing the fractional pair-wise ordering designs. To overcome the unaffordability of the full pair-wise ordering design, a new class of minimal-point pair-wise ordering designs is proposed. A job scheduling problem as well as simulation studies are conducted to illustrate the performance of the pair-wise ordering designs for determining the optimal orders. It is shown that the proposed designs are very efficient in determining the optimal order of addition.

Hes1 attenuates type I IFN responses via VEGF-C and WDFY1.

Induction of type I interferons (IFNs) is critical for eliciting competent immune responses, especially antiviral immunity. However, uncontrolled IFN production contributes to pathogenesis of autoimmune and inflammatory diseases. We found that transcription factor Hes1 suppressed production of type I IFNs and expression of IFN-stimulated genes. Functionally, Hes1-deficient mice displayed a heightened IFN signature in vivo, mounted enhanced resistance against encephalomyocarditis virus infection, and showed signs of exacerbated experimental lupus nephritis. Mechanistically, Hes1 did not suppress IFNs via direct transcriptional repression of IFN-encoding genes. Instead, Hes1 attenuated activation of TLR upstream signaling by inhibition of an adaptor molecule, WDFY1. Genome-wide assessment of Hes1 occupancy revealed that suppression of WDFY1 was secondary to direct binding and thus enhancement of expression of VEGF-C by Hes1, making Vegfc a rare example of an Hes1 positively regulated gene. In summary, these results identified Hes1 as a homeostatic negative regulator of type I IFNs for the maintenance of immune balance in the context of antiviral immunity and autoimmune diseases.

PREPARATION AND PROPERTIES OF COMPOUND ARNEBIAE RADIX MICROEMULSION GEL.

BACKGROUND: Compound Arnebiae radix oil has been clinically applied to treat burns and scalds for a long time. However, it is unstable and inconvenient to use. The aim of this study was to prepare a compound Arnebiae radix microemulsion gel for transdermal delivery system and evaluate its characteristics. MATERIALS AND METHODS: Based on the solubility of Shikonin, the active component of Arnebiae radix and the results of phase studies, adequate ratio of each component in microemulsion was determined. The optimized microemulsion gel was prepared using Carbomer 940. The gels were characterized in terms of appearance, preliminary stability test and the content of Shikonin in the compound Arnebiae radix microemulsion gel with HPLC analysis. RESULTS: The optimized conditions for preparing microemulsion were Tween-80, glycerin, isopropyl myristate (IPM) with the ratio of 6:3:2. The optimal microemulsion gel was obtained with Carbomer 940 (1.0%). CONCLUSION: The prepared compound Arnebiae radix microemulsion gel showed good stability over time. It is more convenience in application than the previous used formulations.

Preparation of cationic biodegradable dextran microspheres loaded with BSA and study on the mechanism of protein loading.

AIM: The aim of this study was to investigate the feasibility of cationic biodegradable dextran microspheres loaded with bovine serum albumin (BSA) posterior to gel formation (postloading). METHOD: Positively charged microspheres were prepared by polymerization of hydroxylethyl methacrylate-derivatized dextran (dex-HEMA) and dimethyl aminoethyl methacrylate (DMAEMA) in an aqueous two-phase system and net positive surface charge increased with increasing amounts of DMAEMA. Loading efficiency of dextran microspheres for BSA was analyzed through fluorescence microscopy and measured. The BSA release from the cationic dextran microspheres in vitro was investigated. RESULTS: BSA could penetrate into cationic dextran microspheres, but neutral dextran microspheres could not. Protein-loading efficiency (98.1--100%) by postloading was higher compared with by preloading (60.2--75.9%), when the incubated protein concentration was below 1.5 mg/ml. Even though BSA is incorporated in the hydrogel network based on electrostatic interaction, a controlled release can be achieved by varying the initial network density of the microspheres. CONCLUSION: These findings suggest that it is a feasible method to prepare dextran microspheres with high surface-charge density to efficiently adsorb oppositely charged protein based on electrostatic interactions.

[Preparation of cationic dextran microspheres loaded with tetanus toxoid and study on the mechanism of protein loading].

The aim of this study is to prepare cationic biodegradable dextran microspheres loaded with tetanus toxoid (TT) and to investigate the mechanism of protein loading. Positively charged microspheres were prepared by polymerization of hydroxylethyl methacrylate derivatized dextran (dex-HEMA) and dimethyl aminoethyl methacrylate (DMAEMA) in an aqueous two-phase system. The loading of the microspheres with TT was based on electrostatic attraction. The net positive surface charge increased with increasing amounts of DMAEMA. Confocal images showed fluorescein isothiocyanate labeled bovine serum albumin (FITC-BSA) could penetrate into cationic dextran microspheres but not natural dextran microspheres. TT loading efficiency by post-loading was higher compared with by pre-loading. Even though TT is incorporated in the hydrogel network based on electrostatic interaction, still a controlled release can be achieved by varying the initial network density of the microspheres.

Combustion oxidization synthesis of unique cage-like nanotetrapod ZnO and its optical property.

Cage-like nano-tetrapod ZnO is a novel structure, which was successfully synthesized by combustion oxidation at 850 degrees C. No catalyst or carrier gases were used. Thorough SEM and TEM analyses revealed that the linking legs of the tetrapod ZnO can have or not interface. The formation or not of an interface is discussed and it was attributed to different growth process of the cage-like ZnO nano-tetrapod. Enhanced UV emission peak at the wavelength of 375 nm, featuring high intensity and narrow width, indicates a highly crystalline structure. A green emission, recorded at 502 nm, was related to the defects of the surface of the branching configuration as well as to the ZnO nuclei of the cage-like nano-tetrapod ZnO.