Mechanical properties, biosafety, and shearing bonding strength of glass fiber-reinforced PEEK composites used as post-core materials.

OBJECTIVE: To investigate the mechanical properties, biosafety, and shearing bonding strength of glass fibers-reinforced polyether-ether-ketone (PEEK-GF) for post-core materials. METHODS: PEEK-GF composites with different glass fiber contents were prepared by extrusion injection and named PEEK-GF30, PEEK-GF40, and PEEK-GF50. Mechanical properties including flexural modulus, flexural strength, Vickers hardness, and compression strength were tested. The cross-sectional morphology was examined using scanning electron microscopy (SEM). Cytotoxicity was studied in vitro with Cell-counting kit-8 (CCK-8). Cell morphology was observed under a microscope. Cell growth on the composites' surfaces was analyzed with DAPI staining. The shearing bonding strength (SBS) of PEEK-GF50 was assessed after applying different pretreatments. Failure modes were evaluated by microscopy. SEM and contact-angle measurements were performed on the surfaces. Statistical analysis was conducted using one-way ANOVA (P < 0.05). RESULTS: The mechanical properties of PEEK-GF composites improved with increased GF content. The PEEK-GF50 group exhibited flexural modulus (17.4 ± 0.5 GPa) close to that of dentin (18.6 GPa) and showed the highest flexural strength (350.0 ± 2.9 MPa), Vickers hardness (47.6 ± 4.5 HV), and compressive strength (264.0 ± 18.0 MPa). The SEM analysis demonstrated that the PEEK matrix combined well with glass fibers. The CCK-8 results confirmed the biosafety of all groups. DAPI staining indicated that cells were growing well on the composites' surface. The sample that was pretreated with sandblasting and plasma showed the highest SBS (16.0 ± 1.7 MPa). SIGNIFICANCE: The PEEK-GF composites demonstrated excellent mechanical properties, biosafety, and SBS, and have great potential to serve as post-core materials.

MicroRNA-29b modulates the innate immune response by suppressing IFNγs production in orange-spotted grouper (Epinephelus coioides).

Interferon-γ (IFNγ), a type II interferon, is essential to host resistance against various infections. Unlike other vertebrates, fish have two types of IFNγs, IFNγ1 (also named IFNγ-rel) and IFNγ2. MicroRNAs (miRNAs) regulate multiple biological processes by suppressing mRNA translation or inducing mRNA degradation. Among them, miR-29 can directly target IFNγ and affact innate and adaptive immune responses in mice. There are five members of the miR-29 family in orange-spotted grouper (Epinephelus coioides), which share the same miRNA seed region. However, whether miR-29 directly targets E. coioides IFNγs and regulate IFNγ production is still unknown. In the present study, the negative correlation between miR-29b and both IFNγs in immune tissues of healthy E. coioides and grouper spleen cells (GS cells) stimulated with LPS or poly I:C was demonstrated. Moreover, dual-luciferase reporter assays and western blotting were performed to demonstrate that miR-29b suppressed E. coioides IFNγ production. Studies of NO production in GS cells after miR-29b transfection revealed that miR-29b overexpression affected NO production through the downregulation of IFNγ expression. Taken together, our results suggest that miR-29b may directly target E. coioides IFNγs and modulate IFNγ-mediated innate immune responses by suppressing E. coioides IFNγs production.

Two Distinct Interferon-γ in the Orange-Spotted Grouper (Epinephelus coioides): Molecular Cloning, Functional Characterization, and Regulation in Toll-Like Receptor Pathway by Induction of miR-146a.

Interferon gamma (IFNγ) is a Th1 cytokine that is critical for innate and adaptive immunity. Toll-like receptors (TLRs) signaling pathways are critical in early host defense against invading pathogens. miR-146a has been reported to participate in the regulation of host immunity. The known mechanisms of integrations between the IFNγ and TLR signaling pathways are incompletely understood, especially in teleosts. In this study, orange-spotted grouper (Epinephelus coioides) IFNγ1 and IFNγ2, their biological activities, especially their involvements in TLR pathway, were explored. We identified and cloned two IFNγ genes of E. coioides, namely EcIFNγ1 and EcIFNγ2. The produced recombinant E. coioides IFNγ1 (rEcIFNγ1) and IFNγ2 (rEcIFNγ2) proteins showed functions, which are similar to those of other bony fishes, such as enhancing nitric oxide responses and respiratory burst response. rEcIFNγ2 could regulate TLR pathway by enhancing the promoter activity of miR-146a upstream sequence and thus increasing the expression level of miR-146a, which possibly targets TNF receptor-associated factor 6 (TRAF6), a key adapter molecule in TLR signaling pathway. Taken together, these findings unravel a novel regulatory mechanism of anti-inflammatory response by IFNγ2, which could mediate TLR pathway through IFNγ2-miR-146a-TRAF6 negative regulation loop. It is suggested that IFNγ2 may provide a promising therapeutic, which may help to fine tune the immune response.

Intracellular TLR22 acts as an inflammation equalizer via suppression of NF-κB and selective activation of MAPK pathway in fish.

TLR22, a typical member of the fish-specific TLRs, is a crucial sensor in virally triggered innate immune signalling retained from natural selection. To elucidate the role of the TLR22-specific signalling cascade mechanism, we provide evidence that the double-stranded (ds) RNA-sensor TLR22 positively regulates the ERK pathway and negatively regulates the JNK, p38 MAP kinase and NF-κB pathway. Here, we show that TLR22 restrains NF-κB activation and IFN (interferon) ß and AP-1 (activator protein-1) promoter binding (impairing "primary response" genes (TNF and IL-1)), induces "secondary response" genes (IL-12 and IL-6) and mediates the irregular expression of inflammatory genes. Therefore, TLR22 promotes ERK phosphorylation but impairs the JNK and p38 MAP kinases and IκB phosphorylation. Additionally, TLR22 controls the excessive generation of reactive oxygen species (ROS) to avoid damaging the organism. The specific kinetics of TLR22 depends on its distinct cellular localization. We demonstrate that TLR22 is an intracellular receptor localized in the endosome, and the TLR22-TIR domain is the functional structure inducing the signalling cascade post-viral replication in the body. As mentioned above, our data reveal a novel mechanism whereby TLR22-induced positive adjustment and negative regulation evolved independently to avoid harmful and inappropriate inflammatory responses.

Lethal/sublethal responses of Daphnia magna to acute norfloxacin contamination and changes in phytoplankton-zooplankton interactions induced by this antibiotic.

Although the well-known antibiotic norfloxacin (NOR) is recognized as an important environmental pollutant, little is known about its impacts on ecological processes, particularly on species interactions. In this paper, we quantified Daphnia magna (Crustacea, Cladocera) responses in mortality rate at lethal NOR concentrations (0, 25, 50, 100, 200, 300 and 400 mg L-1), and in heartbeat rate, swimming behavior and feeding rate (on the green alga Chlorella pyrenoidosa) at sublethal NOR concentrations (0, 25, 50 and 100 mg L-1) to determine the effects of this antibiotic in plankton systems. In 96-h-long lethal experiment, mortality rates of D. magna increased significantly with increasing NOR concentration and exposure time. In sublethal experiments, heartbeat rate decreased, while time ratio of vertical to horizontal swimming (TVH) and the duration of quiescence increased in D. magna individuals exposed to increasing NOR concentrations after 4 and 12 h of exposure. These collectively led to decreases in both average swimming ability and feeding rate, consistent with the positive relationship between average swimming ability and feeding rate. Overall, results indicate that, by affecting zooplankton heartbeat rate and behavior, NOR decreased feeding efficiency of D. magna even at low doses, therefore, it might seriously compromise ecosystem health and function.