Potent antiviral activity of Agrimonia pilosa, Galla rhois, and their components against SARS-CoV-2.

Agrimonia pilosa (AP), Galla rhois (RG), and their mixture (APRG64) strongly inhibited SARS-CoV-2 by interfering with multiple steps of the viral life cycle including viral entry and replication. Furthermore, among 12 components identified in APRG64, three displayed strong antiviral activity, ursolic acid (1), quercetin (7), and 1,2,3,4,6-penta-O-galloyl-ß-d-glucose (12). Molecular docking analysis showed these components to bind potently to the spike receptor-binding-domain (RBD) of the SARS-CoV-2 and its variant B.1.1.7. Taken together, these findings indicate APRG64 as a potent drug candidate to treat SARS-CoV-2 and its variants.

Nanoformulated Single-Stranded RNA-Based Adjuvant with a Coordinative Amphiphile as an Effective Stabilizer: Inducing Humoral Immune Response by Activation of Antigen-Presenting Cells.

As agonists of TLR7/8, single-stranded RNAs (ssRNAs) are safe and promising adjuvants that do not cause off-target effects or innate immune overactivation. However, low stability prevents them from mounting sufficient immune responses. This study evaluates the adjuvant effects of ssRNA derived from the cricket paralysis virus intergenic region internal ribosome entry site, formulated as nanoparticles with a coordinative amphiphile, containing a zinc/dipicolylamine complex moiety as a coordinative phosphate binder, as a stabilizer for RNA-based adjuvants. The nanoformulated ssRNA adjuvant was resistant to enzymatic degradation in vitro and in vivo, and that with a coordinative amphiphile bearing an oleyl group (CA-O) was approximately 100 nm, promoted effective recognition, and improved activation of antigen-presenting cells, leading to better induction of neutralizing antibodies following single immunization. Hence, CA-O may increase the efficacy of ssRNA-based adjuvants, proving useful to meet the urgent need for vaccines during pathogen outbreaks.

Dual inhibition of EGFR and MET by Echinatin retards cell growth and induces apoptosis of lung cancer cells sensitive or resistant to gefitinib.

Patients with non-small-cell lung cancer (NSCLC) containing epidermal growth factor receptor (EGFR) amplification or sensitive mutations initially respond to tyrosine kinase inhibitor gefitinib; however, the treatment is less effective over time. Gefitinib resistance mechanisms include MET gene amplification. A therapeutic strategy targeting MET as well as EGFR can overcome resistance to gefitinib. In the present study we identified Echinatin (Ecn), a characteristic chalcone in licorice, which inhibited both EGFR and MET and strongly altered NSCLC cell growth. The antitumor efficacy of Ecn against gefitinib-sensitive or -resistant NSCLC cells with EGFR mutations and MET amplification was confirmed by suppressing cell proliferation and anchorage-independent colony growth. During the targeting of EGFR and MET, Ecn significantly blocked the kinase activity, which was validated with competitive ATP binding. Inhibition of EGFR and MET by Ecn decreases the phosphorylation of downstream target proteins ERBB3, AKT and ERK compared with total protein expression or control. Ecn induced the G2/M cell cycle arrest, and apoptosis via the intrinsic pathway of caspase-dependent activation. Ecn induced ROS production and GRP78, CHOP, DR5 and DR4 expression as well as depolarized the mitochondria membrane potential. Therefore, our results suggest that Ecn is a promising therapeutic agent in NSCLC therapy.

Endogenous n-3 Polyunsaturated Fatty Acids Are Beneficial to Dampen CD8+ T Cell-Mediated Inflammatory Response upon the Viral Infection in Mice.

Omega-3 (n-3) polyunsaturated fatty acids (PUFAs) have been known to exert anti-inflammatory effects on various disease states. However, its effect on CD8+ T cell-mediated immunopathology upon viral infection has not been well elucidated yet. In this study, we investigated the possible implication of n-3 PUFAs in CD8+ T cell responses against an acute viral infection. Infection of FAT-1 transgenic mice that are capable of synthesizing n-3 PUFAs from n-6 PUFAs with lymphocytic choriomeningitis virus (LCMV) resulted in significant reduction of anti-viral CD8+ T cell responses. Interestingly, expansion of adoptively transferred wild-type (WT) LCMV-specific T cell receptor (TCR) transgenic CD8+ (P14) T cells into FAT-1 mice was significantly decreased. Also, activation of anti-viral CD4+ helper T cells was reduced in FAT-1 mice. Importantly, P14 cells carrying the fat-1 gene that were adoptively transferred into WT mice exhibited a substantially decreased ability to proliferate and produce cytokines against LCMV infection. Together, n-3 PUFAs attenuated anti-viral CD8+ T cell responses against an acute viral infection and thus could be used to alleviate immunopathology mediated by the viral infection.

A hot-water extract of Sanguisorba officinalis ameliorates endotoxin-induced septic shock by inhibiting inflammasome activation.

The inflammasome is a multiprotein signaling complex that mediates inflammatory innate immune responses through caspase 1 activation and subsequent IL-1ß secretion. However, because its aberrant activation often leads to inflammatory diseases, targeting the inflammasome holds promise for the treatment of inflammation-related diseases. In this study, it was found that a hot-water extract of Sanguisorba officinalis (HSO) suppresses inflammasome activation triggered by adenosine 5'-triphosphate, nigericin, microbial pathogens, and double stranded DNA in bone marrow-derived macrophages. HSO was found to significantly suppress IL-1ß production in a dose-dependent manner; this effect correlated well with small amounts of caspase 1 and little ASC pyroptosome formation in HSO-treated cells. The anti-inflammatory activity of HSO was further confirmed in a mouse model of endotoxin-induced septic shock. Oral administration of HSO reduced IL-1ß titers in the serum and peritoneal cavity, increasing the survival rate. Taken together, our results suggest that HSO is an inhibits inflammasome activation through nucleotide-binding domain and leucine-rich repeat pyrin domain 3, nucleotide-binding domain and leucine-rich repeat caspase recruitment domain 4 and absent in melanoma 2 pathways, and may be useful for treatment of inflammasome-mediated diseases.

A hot water extract of Aralia cordata activates bone marrow-derived macrophages via a myeloid differentiation protein 88-dependent pathway and protects mice from bacterial infection.

In traditional Asian medicine, Aralia cordata (AC) is a known as a pain reliever and anti-inflammatory drug. Although several of its biological activities have been reported, the immunomodulatory effects of a hot water extract of AC (HAC) have not yet been described. The aim of this study was to investigate whether HAC modulates the activation of macrophages, which play important roles in innate immune responses against microbial pathogens, and if so, to determine the molecular mechanisms by which HAC mediates this process. It was found that HAC activates bone marrow-derived macrophages (BMDM) and increases amounts of nitric oxide and proinflammatory cytokines in a dose-dependent manner. In addition, HAC was found to induce phosphorylation of NF-κB and mitogen-activated protein kinases (MAPKs), including c-Jun N-terminal kinases, extracellular signal-regulated kinases and p38. Interestingly, these effects were absent in BMDM prepared from myeloid differentiation protein 88-knockout mice. Polysaccharides from HAC exerted stronger immunostimulatory effects than HAC itself. Furthermore, orally administered HAC clearly enhanced clearance of the intracellular pathogen Listeria monocytogenes by boosting innate immune responses. These results demonstrate that HAC exerts immunostimulatory effects through the TLR/MyD88 and NF-κB/MAPK signal transduction pathways.

Eco-friendly approach towards green synthesis of zinc oxide nanocrystals and its potential applications.

In the present study, we investigated a novel green route for synthesis of zinc oxide (ZnO) nanocrystals using Prunus × yedoensis Matsumura leaf extract as a reducing agent without using any surfactant or external energy. Standard characterization studies were carried out to confirm the obtained product using UV-Vis spectra, SEM-EDS, FTIR, TEM, and XRD. In addition, the synthesized ZnO nanocrystals were coated onto fabric and leather samples to study their bacteriostatic effect against odor-causing bacteria Brevibacterium linens and Staphylococcus epidermidis. Zinc oxide nanocrystal-coated fabric and leather showed good activity against both bacteria.

Synthesis and characterization of nanosilver with antibacterial properties using Pinus densiflora young cone extract.

This study describes an eco-friendly, rapid method for green synthesis of silver nanoparticles (Ag NPs) from an aqueous solution of silver nitrate using Pinus densiflora for. multicaulis Uyeki young cone extract in a single-pot process. Color changes, ultraviolet-visible spectra (444.5 nm), X-ray diffraction peaks (2θ=39.68, 46.92, 68.12, and 79.10), and Fourier transform infrared spectroscopy (FT-IR) confirmed the presence of Ag NPs and phytochemicals. Transmission electron microscopy showed that the nanoparticles were mostly oval in shape, with a few triangular-shaped particles. Average particle size was 30-80 nm. Phytochemicals present in the young pine cone extract were likely responsible for the reduction of Ag(+) ions. The synthesized Ag NPs (40 µg) had a 7 mm larger zone of inhibition against the skin pathogen Brevibacterium linens than commercial Ag NPs, Propionibacterium acnes (14 mm), Bacillus cereus (9 mm) and Staphylococcus epidermidis (10mm).

Differential Modulation of Lipopolysaccharide-Induced Inflammatory Cytokine Production by and Antioxidant Activity of Fomentariol in RAW264.7 Cells.

Medicinal mushrooms have been used worldwide to treat cancer and modulate the immune system. Over the last several years, there has been increasing interest in isolating bioactive compounds from medicinal mushrooms and evaluating their health beneficial effects. Fomes fomentarius is used in traditional oriental medicine and is known to possess antioxidant, anti-inflammatory, antidiabetic, and antitumor effects. In the present study, we isolated fomentariol from Fomes fomentarius and investigated its anti-inflammatory effect in murine macrophages (RAW264.7 cells) stimulated with lipopolysaccharides. Fomentariol inhibited the production of nitric oxide and intracellular reactive oxygen species triggered by lipopolysaccharides. Interestingly, fomentariol differentially regulated cytokine production triggered by lipopolysaccharides. Fomentariol effectively suppressed the production of interleukin-1ß and interleukin-6 but not tumor necrosis factor-α. The inhibitory effect of fomentariol against nitric oxide, interleukin-1ß, and interleukin-6 production was possibly mediated by downregulation of the extracellular signal-regulated kinase signaling pathway. Taken together, our results suggest that fomentariol differentially modulated inflammatory responses triggered by lipopolysaccharides in macrophages and is one of the bioactive compounds that mediate the physiological effects of Fomes fomentarius.

Antimicrobial fabrication of cotton fabric and leather using green-synthesized nanosilver.

This study aims to investigate the green synthesis of silver nanoparticles (AgNPs) by Erigeron annuus (L.) pers flower extract as reducing and capping agent, and evaluation of their antibacterial activities for the first time. The obtained product was confirmed by UV-Vis spectrum, high resolution-transmission electron microscopy, energy-dispersive X-ray spectroscopy, Fourier transform infrared spectroscopy, and X-ray diffraction studies. The optimum AgNPs production was achieved at pH 7, metal silver (Ag(+) ion) concentration of 2.0mM, flower extract concentration 4%, and time 335 min. In addition, the antibacterial activity of cotton fabrics and tanned leather loaded with AgNPs, commercial AgNPs, flower extract, Ag(+) ion and blend of flower extract with AgNPs were evaluated against Gram-positive odor causing bacteria Brevibacterium linens and Staphylococcus epidermidis. The results showed maximum zone of inhibition (ZOI) by the cotton fabrics embedded with blend of flower extract and AgNPs against B. linens. The structure and morphology of cotton fabric and leather samples embedded with AgNPs, Ag(+) ion and blend of flower extract with AgNPs were examined under field emission scanning electron microscope.

Green synthesis of silver and gold nanoparticles using Zingiber officinale root extract and antibacterial activity of silver nanoparticles against food pathogens.

In the present study, we synthesized silver and gold nanoparticles with a particle size of 10-20 nm, using Zingiber officinale root extract as a reducing and capping agent. Chloroauric acid (HAuCl4) and silver nitrate (AgNO3) were mixed with Z. officinale root extract for the production of silver (AgNPs) and gold nanoparticles (AuNPs). The surface plasmon absorbance spectra of AgNPs and AuNPs were observed at 436-531 nm, respectively. Optimum nanoparticle production was achieved at pH 8 and 9, 1 mM metal ion, a reaction temperature 50 °C and reaction time of 150-180 min for AgNPs and AuNPs, respectively. An energy-dispersive X-ray spectroscopy (SEM-EDS) study provides proof for the purity of AgNPs and AuNPs. Transmission electron microscopy images show the diameter of well-dispersed AgNPs (10-20 nm) and AuNPs (5-20 nm). The nanocrystalline phase of Ag and Au with FCC crystal structures have been confirmed by X-ray diffraction analysis. Fourier transform infrared spectroscopy analysis shows the respective peaks for the potential biomolecules in the ginger rhizome extract, which are responsible for the reduction in metal ions and synthesized AgNPs and AuNPs. In addition, the synthesized AgNPs showed a moderate antibacterial activity against bacterial food pathogens.

Effects of n-3 PUFA on the CD4⁺ type 2 helper T-cell-mediated immune responses in Fat-1 mice.

SCOPE: It has been suggested that n-3 PUFA can be used as a preventive or therapeutic strategy to control allergic asthma. But little is known about the exact mechanisms by which n-3 PUFA modulates it. Here, the effects of elevated n-3 PUFA on ovalbumin (OVA) induced airway inflammation were investigated using Fat-1 transgenic mice that can convert n-6 PUFA to n-3 PUFA endogenously. METHODS AND RESULTS: First, we tested whether Fat-1 expression modulates CD4⁺ T-cell activation, proliferation, and differentiation in vitro and found that the Fat-1 expression attenuated all of these CD4⁺ T-cell responses by suppression of T-cell receptor mediated signaling and cytokine-mediated phosphorylation of STATs. When the Fat-1 mice were sensitized and challenged with the OVA, they showed a significant decrease in the recruitment of inflammatory cells into airway, the production of Th2 cytokines, eotaxin, and mucin in the lung, and the concentration of OVA-specific IgE in the serum. Furthermore, the differentiation of CD4⁺ T cells into Th2 was also decreased in the spleen of Fat-1 mice. CONCLUSION: Our results showed that an elevated level of n-3 PUFA was effective in preventing allergic airway inflammation by modulating the activation and differentiation of CD4⁺ T cells in Fat-1 mice.

Overexpression of the small GTPase Arl4D suppresses adipogenesis.

Arl4D is a developmentally-regulated member of the ADP-ribosylation factor/ARF-like protein (ARF/Arl) family of Ras-related GTPases. Although Arl4 protein is reported to be expressed in adipose tissue, the function of Arl4D is unknown. To investigate the potential role of Arl4D in adipogenesis, we examined Arl4D expression during adipocyte differentiation and the effects of Arl4D overexpression on adipogenesis. Arl4D protein increased early in adipogenesis, with the highest expression at 4 h after adipogenesis initiation, followed by a decrease thereafter. Overexpression of Arl4D in 3T3-L1 cells potently inhibited their ability to differentiate and accumulate lipid, and reduced the expression of adipogenic genes. Furthermore, treatment with valproic acid, an Arl4D inducer, suppressed adipogenesis. These results suggest that rapid reduction of Arl4D is required for adipogenesis to proceed.

Synoviolin promotes IRE1 ubiquitination and degradation in synovial fibroblasts from mice with collagen-induced arthritis.

The E3 ubiquitin ligase synoviolin (SYVN1) functions as an anti-apoptotic factor that is responsible for the outgrowth of synovial cells during the development of rheumatoid arthritis. The molecular mechanisms underlying SYVN1 regulation of cell death are largely unknown. Here, we report that elevated SYVN1 expression correlates with decreased levels of the protein inositol-requiring enzyme 1 (IRE1)-a pro-apoptotic factor in the endoplasmic reticulum (ER)-stress-induced apoptosis pathway-in synovial fibroblasts from mice with collagen-induced arthritis (CIA). SYVN1 interacts with and catalyses IRE1 ubiquitination and consequently promotes IRE1 degradation. Suppression of SYVN1 expression in synovial fibroblasts from CIA mice restores IRE1 protein expression and reverses the resistance of ER-stress-induced apoptosis of CIA synovial fibroblasts. These results show that SYVN1 causes the overgrowth of synovial cells by degrading IRE1, and therefore antagonizes ER-stress-induced cell death.

RLE-1, an E3 ubiquitin ligase, regulates C. elegans aging by catalyzing DAF-16 polyubiquitination.

The forkhead transcription factor, DAF-16, a downstream target of the insulin/IGF-I signaling pathway in C. elegans, is indispensable both for lifespan regulation and stress resistance. The molecular mechanisms involved in regulating DAF-16 transcriptional activation remain undefined. Here, we have identified an E3 ubiquitin ligase, RLE-1 (regulation of longevity by E3), which regulates aging in C. elegans. Disruption of RLE-1 expression in C. elegans increases lifespan; this extension of lifespan is due to elevated DAF-16 protein but not to changes of daf-16 mRNA levels. We have also found that RLE-1 catalyzes DAF-16 ubiquitination, leading to degradation by the proteasome. Elimination of RLE-1 expression in C. elegans causes increased transcriptional activation and sustained nuclear localization of DAF-16. Overexpression of DAF-16 in rle-1 mutants increases worm lifespan, while disruption of DAF-16 expression in rle-1 mutants reverses their longevity. Thus, RLE-1 is an E3 ubiquitin ligase of DAF-16 that regulates C. elegans aging.