CD36 Participates in PrP106–126-Induced Activation of Microglia

Microglial activation is a characteristic feature of the pathogenesis of prion diseases. The molecular mechanisms that underlie prion-induced microglial activation are not very well understood. In the present study, we investigated the role of the class B scavenger receptor CD36 in microglial activation induced by neurotoxic prion protein (PrP) fragment 106–126 (PrP106–126). We first examined the time course of CD36 mRNA expression upon exposure to PrP106–126 in BV2 microglia. We then analyzed different parameters of microglial activation in PrP106–126-treated cells in the presence or not of anti-CD36 monoclonal antibody (mAb). The cells were first incubated for 1 h with CD36 monoclonal antibody to block the CD36 receptor, and were then treated with neurotoxic prion peptides PrP106–126. The results showed that PrP106–126 treatment led to a rapid yet transitory increase in the mRNA expression of CD36, upregulated mRNA and protein levels of proinflammatory cytokines (IL-1β, IL-6 and TNF-α), increased iNOS expression and nitric oxide (NO) production, stimulated the activation of NF-κB and caspase-1, and elevated Fyn activity. The blockade of CD36 had no effect on PrP106–126-stimulated NF-κB activation and TNF-α protein release, abrogated the PrP106–126-induced iNOS stimulation, downregulated IL-1β and IL-6 expression at both mRNA and protein levels as well as TNF-α mRNA expression, decreased NO production and Fyn phosphorylation, reduced caspase-1 cleavage induced by moderate PrP106–126 –treatment, but had no effect on caspase-1 activation after treatment with a high concentration of PrP106–126. Together, these results suggest that CD36 is involved in PrP106–126-induced microglial activation and that the participation of CD36 in the interaction between PrP106–126 and microglia may be mediated by Src tyrosine kinases. Our findings provide new insights into the mechanisms underlying the activation of microglia by neurotoxic prion peptides and open perspectives for new therapeutic strategies for prion diseases by modulation of CD36 signaling

Numerical Study on Aerodynamic Noise Reduction of Pantograph

A hybrid method incorporating the simulations of noise sources with delayed detached eddy simulation (DDES) and calculations of far-field noise with the Ffowcs Williams–Hawkings (FW-H) equation is used to study the suppression technique for the aerodynamic noise of a Faiveley CX-PG pantograph. Considering that China’s Fuxing bullet trains operate at 350 km/h, the inflow velocity of 350 km/h is applied in this paper. The noise radiated from the panhead area, middle area, and bottom area at an inflow velocity of 350 km/h is distinguished. The noise intensities at the standard observer show that the noise radiated from the panhead area is the strongest, and the sound pressure level spectrum value is larger than the other two in the range above 500 Hz. The influence of applying the wavy rods and modifying the contact strip shape on the aerodynamic noise is discussed in detail. By comparing the acoustic source distribution and the far-field noise intensity, it is found that applying the wavy rods can effectively reduce the panhead noise, especially around the peak frequency. Modifying the shape of the contact strip to a hexagon can suppress the vortex shedding, leading to a lower surface pressure level. Combining the strip modification and wavy rods, the total noise intensity can be diminished by about 3.0 dB

Are vegetated areas of mangroves attractive to juvenile and small fish? The case of Dongzhaigang Bay, Hainan Island, China

Well-developed aerial roots of mangroves make it difficult to study how fish utilize the mangrove forest as a habitat. In the present study, we compared the differences in fish assemblages in three major types of habitats of mangrove estuary (vegetated area, treeless mudflat, and creek) of a mangrove bay in Hainan Island, China, at different seasons during two consecutive years. Three types of gears, centipede net, gill net and cast net, were used in the different habitats of mangrove estuary and sampling efficiencies among gears were evaluated. Centipede nets were used in all the three types of habitats and cast nets and gill nets in treeless mudflats and creeks. Fish assemblages were dependent on gears used. Centipede net could efficiently catch fish occurring both inside and outside of vegetated areas efficiently. A total of 115 fish species in 51 families were collected. In terms of numbers of species per family, Gobiidae was the most diverse (17 species), followed by Mugilidae (5 species). Almost all of the fish were juvenile or small fish and few predators were recorded, implying low predation pressure in the bay. ANOVA analysis showed that significant seasonal and spatial variation existed in species richness, abundance, and biomass, which were less in the vegetated areas than those of treeless mudflats and creeks. The attraction of vegetated areas to fish was less than that of creeks and mudflats. Many species were specific to a particular habitat type, 4 species occurring exclusively in the creeks, 45 species occurring exclusively in the treeless mudflats, and 5 species occurring exclusively in the vegetated areas. The results indicated that mangrove estuaries were potentially attractive habitats for juvenile and small fish, but this attraction was accomplished by a connection of vegetated areas, treeless mudflats and creeks, not only by vegetated areas. (C) 2009 Elsevier Ltd. All rights reserved

Overcoming the Undesirable CRISPR-Cas9 Expression in Gene Correction

The CRISPR-Cas9 system is attractive for gene therapy, as it allows for permanent genetic correction. However, as a new technology, Cas9 gene editing in clinical applications faces major challenges, such as safe delivery and gene targeting efficiency. Cas9 is a foreign protein to recipient cells; thus, its expression may prompt the immune system to eliminate gene-edited cells. To overcome these challenges, we have engineered a novel delivery system based on the helper-dependent adenoviral (HD-Ad) vector, which is capable of delivering genes to airway basal stem cells in vivo. Using this system, we demonstrate the successful co-delivery of both CRISPR-Cas9/single-guide RNA and the LacZ reporter or CFTR gene as donor DNA to cultured cells. HD-Ad vector genome integrity is compromised following donor DNA integration, and because the CRISPR-Cas9/single-guide RNA and donor DNA are carried on the same vector, CRISPR-Cas9 expression is concurrently eliminated. Thus, we show the feasibility of site-specific gene targeting with limited Cas9 expression. In addition, we achieved stable CFTR expression and functional correction in cultured cells following successful gene integration. Keywords: gene targeting, cystic fibrosis, lung disease, gene therapy, CRISP

DNAJA3 Interacts with PEDV S1 Protein and Inhibits Virus Replication by Affecting Virus Adsorption to Host Cells

Porcine epidemic diarrhea virus (PEDV) infection causes huge economic losses to the pig industry worldwide. DNAJA3, a member of the Hsp40 family proteins, is known to play an important role in the replication of several viruses. However, it remains unknown if it interacts with PEDV. We found that DNAJA3 interacted with PEDV S1, initially with yeast two-hybrid screening and later with Co-IP, GST pull-down, and confocal imaging. Further experiments showed the functional relationship between DNAJA3 and PEDV in the infected IPEC-J2 cells. DNAJA3 overexpression significantly inhibited PEDV replication while its knockdown had the opposite effect, suggesting that it is a negative regulator of PEDV replication. In addition, DNAJA3 expression could be downregulated by PEDV infection possibly as the viral strategy to evade the suppressive role of DNAJA3. By gene silencing and overexpression, we were able to show that DNAJA3 inhibited PEDV adsorption to IPEC-J2 cells but did not affect virus invasion. In conclusion, our study provides clear evidence that DNAJA3 mediates PEDV adsorption to host cells and plays an antiviral role in IPEC-J2 cells

Porcine Epidemic Diarrhea Virus Antagonizes Host IFN-λ-Mediated Responses by Tilting Transcription Factor STAT1 toward Acetylation over Phosphorylation To Block Its Activation

ABSTRACT Porcine epidemic diarrhea virus (PEDV) is the main etiologic agent causing acute swine epidemic diarrhea, leading to severe economic losses to the pig industry. PEDV has evolved to deploy complicated antagonistic strategies to escape from host antiviral innate immunity. Our previous study demonstrated that PEDV downregulates histone deacetylase 1 (HDAC1) expression by binding viral nucleocapsid (N) protein to the transcription factor Sp1, inducing enhanced protein acetylation. We hypothesized that PEDV inhibition of HDAC1 expression would enhance acetylation of the molecules critical in innate immune signaling. Signal transducer and activator of transcription 1 (STAT1) is a crucial transcription factor regulating expression of interferon (IFN)-stimulated genes (ISGs) and anti-PEDV immune responses, as shown by overexpression, chemical inhibition, and gene knockdown in IPEC-J2 cells. We further show that PEDV infection and its N protein overexpression, although they upregulated STAT1 transcription level, could significantly block poly(I·C) and IFN-λ3-induced STAT1 phosphorylation and nuclear localization. Western blotting revealed that PEDV and its N protein promote STAT1 acetylation via downregulation of HDAC1. Enhanced STAT1 acetylation due to HDAC1 inhibition by PEDV or MS-275 (an HDAC1 inhibitor) impaired STAT1 phosphorylation, indicating that STAT1 acetylation negatively regulated its activation. These results, together with our recent report on PEDV N-mediated inhibition of Sp1, clearly indicate that PEDV manipulates the Sp1-HDAC1-STAT1 signaling axis to inhibit transcription of OAS1 and ISG15 in favor of its replication. This novel immune evasion mechanism is realized by suppression of STAT1 activation through preferential modulation of STAT1 acetylation over phosphorylation as a result of HDAC1 expression inhibition. IMPORTANCE PEDV has developed sophisticated evasion mechanisms to escape host IFN signaling via its structural and nonstructural proteins. STAT1 is one of the key transcription factors in regulating expression of ISGs. We found that PEDV and its N protein inhibit STAT1 phosphorylation and nuclear localization via inducing STAT1 acetylation as a result of HDAC1 downregulation, which, in turn, dampens the host IFN signaling activation. Our study demonstrates a novel mechanism that PEDV evades host antiviral innate immunity through manipulating the reciprocal relationship of STAT1 acetylation and phosphorylation. This provides new insights into the pathogenetic mechanisms of PEDV and even other coronaviruses

Beyond canonical PROTAC: biological targeted protein degradation (bioTPD)

Abstract Targeted protein degradation (TPD) is an emerging therapeutic strategy with the potential to modulate disease-associated proteins that have previously been considered undruggable, by employing the host destruction machinery. The exploration and discovery of cellular degradation pathways, including but not limited to proteasomes and lysosome pathways as well as their degraders, is an area of active research. Since the concept of proteolysis-targeting chimeras (PROTACs) was introduced in 2001, the paradigm of TPD has been greatly expanded and moved from academia to industry for clinical translation, with small-molecule TPD being particularly represented. As an indispensable part of TPD, biological TPD (bioTPD) technologies including peptide-, fusion protein-, antibody-, nucleic acid-based bioTPD and others have also emerged and undergone significant advancement in recent years, demonstrating unique and promising activities beyond those of conventional small-molecule TPD. In this review, we provide an overview of recent advances in bioTPD technologies, summarize their compositional features and potential applications, and briefly discuss their drawbacks. Moreover, we present some strategies to improve the delivery efficacy of bioTPD, addressing their challenges in further clinical development