ILRUN Downregulates ACE2 Expression and Blocks Infection of Human Cells by SARS-CoV-2.

The human protein-coding gene ILRUN (inflammation and lipid regulator with UBA-like and NBR1-like domains; previously C6orf106) was identified as a proviral factor for Hendra virus infection and was recently characterized to function as an inhibitor of type I interferon expression. Here, we have utilized transcriptome sequencing (RNA-seq) to define cellular pathways regulated by ILRUN in the context of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection of Caco-2 cells. We find that inhibition of ILRUN expression by RNA interference alters transcription profiles of numerous cellular pathways, including upregulation of the SARS-CoV-2 entry receptor ACE2 and several other members of the renin-angiotensin aldosterone system. In addition, transcripts of the SARS-CoV-2 coreceptors TMPRSS2 and CTSL were also upregulated. Inhibition of ILRUN also resulted in increased SARS-CoV-2 replication, while overexpression of ILRUN had the opposite effect, identifying ILRUN as a novel antiviral factor for SARS-CoV-2 replication. This represents, to our knowledge, the first report of ILRUN as a regulator of the renin-angiotensin-aldosterone system (RAAS). IMPORTANCE There is no doubt that the current rapid global spread of COVID-19 has had significant and far-reaching impacts on our health and economy and will continue to do so. Research in emerging infectious diseases, such as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is growing rapidly, with new breakthroughs in the understanding of host-virus interactions to assist with the development of innovative and exciting therapeutic strategies. Here, we present the first evidence that modulation of the human protein-coding gene ILRUN functions as an antiviral factor for SARS-CoV-2 infection, likely through its newly identified role in regulating the expression of SARS-CoV-2 entry receptors ACE2, TMPRSS2, and CTSL. These data improve our understanding of biological pathways that regulate host factors critical to SARS-CoV-2 infection, contributing to the development of antiviral strategies to deal with the current SARS-CoV-2 pandemic.

Comparing Gene Silencing and Physiochemical Properties in siRNA Bound Cationic Star-Polymer Complexes.

The translation of siRNA into clinical therapies has been significantly delayed by issues surrounding the delivery of naked siRNA to target cells. Here we investigate siRNA delivery by cationic acrylic polymers developed by Reversible Addition-Fragmentation chain Transfer (RAFT) mediated free radical polymerization. We investigated cell uptake and gene silencing of a series of siRNA-star polymer complexes both in the presence and absence of a protein "corona". Using a multidisciplinary approach including quantitative nanoscale mechanical-atomic force microscopy, dynamic light scattering and nanoparticle tracking analysis we have characterized the nanoscale morphology, stiffness, and surface charge of the complexes with and without the protein corona. This is one of the first examples of a comprehensive physiochemical analysis of siRNA-polymer complexes being performed alongside in vitro biological assays, allowing us to describe a set of desirable physical features of cationic polymer complexes that promote gene silencing. Multifaceted studies such as this will improve our understanding of structure-function relationships in nanotherapeutics, facilitating the rational design of polymer-mediated siRNA delivery systems for novel treatment strategies.

[The Effects of Skin Thickness on Optical Transmission Characteristics in Fruits Tissues].

Fruit quality inspection techniques play a very important role in the production and consumption of fruits. In the field of quality non-destructive inspection and grading for fruits, the light-based techniques using optical properties of fruit products were widely used as one of the most practical and the most successful techniques. Quantitative understanding of light interaction with fruits is critical to designing better optical systems for inspection of food quality. In this paper, a fruit model consisted of two layer tissues was developed using Monte Carlo simulations to explore the light transport process and properties in the pome fruits, such as apples and mandarins, which were used as the thin-skinned and thick-skinned fruits respectively. The simulation results obtained are based on the assumption that the light source is a Gaussian beam at the wavelength 808 nm. This paper reports that the effects of skin thickness on light transmission characteristics in fruit tissues, including diffuse reflectance, transmittance, absorptivity, penetration depth etc. The inspection efficiency of flesh tissues was also demonstrated. The results indicated that the transmittance and the penetration depth decreases with the fruit skin increasing. As for the absorbed energy density, the fruit skin tissues have the wider distribution at the radial distance than the fruit flesh tissues. The absorbed energy density always tended to decrease with the inside depth of the fruit tissues increasing, especially decreased more apparently at the radial direction. The diffuse reflectance at the radial distance from 0.2 to 1.2 cm decreased with the decreasing of fruit skin, however it showed the inverse relationship in the radial distance range from 1.2 to 4.0 cm, the diffuse reflectance decreases with the increasing fruit skin. This paper proposed that the interaction between light and fruits skin in transmission or reflective approach, should be considered for developing optical techniques of non-destructive fruit quality inspection. And it provided a theoretical basis for designing more efficient optical detection device, including how to confirm the light source power, size and position of the detector, etc. It has very important significance for fruit quality inspection by optical techniques.

[Mass spectrum characterization of five valepotriates by electrospray ionization tandem mass spectrometry].

OBJECTIVE: To discuss mass spectrum characterization of five valepotriates including 'monoene' type (didrovaltrate), 'diene' type (valtrate, acevaltrate) and 'four-olefinic' type (baldrinal and homobaldrinal) by electrospray ionization tandem mass spectrometry (ESI-MS(n)). METHOD: This study was carried out on the basis of electrospray ionization tandem mass spectrometric method and analysis of multistage fragments. RESULT: The fragmentation patterns and structural assignment of 'monoene' type, 'diene' type and 'four-olefinic' type valepotriates in ESI-MSn under positive mode were summarized. CONCLUSION: The compounds have a strong pyrolysis rules and it can provide reference date for valepotriates in rapid structural identification, quantitative analysis and pharmacokinetic study.

Glycerol monooleate-based nanocarriers for siRNA delivery in vitro.

We present studies of the delivery of short interfering ribonucleic acid (siRNA) into a green fluorescent protein (GFP) expressing cell line, using lipid nanocarriers in cubic lyotropic liquid crystal form. These carriers are based on glycerol monooleate (GMO) and employ the use of varying concentrations of cationic siRNA binding lipids. The essential physicochemical parameters of the cationic lipid/GMO/siRNA complexes such as particle size, ζ otential, siRNA uptake stability, lyotropic mesophase behavior, cytotoxicity,and gene silencing efficiency were systematically assessed. We find that the lipid nanocarriers were effectively taken up by mammalian cells and that their siRNA payload was able to induce gene silencing in vitro. More importantly, it was found that the nonlamellar structure of some of the lipid nanocarrier formulations were more effective at gene silencing than their lamellar structured counterparts. The development of cationic lipid functionalized nonlamellar GMO-based nanostructured nanoparticles may lead to improved siRNA delivery vehicles.

Effects of Different Factors on the Performance of Recycled Aggregate Permeable Pavement Concrete.

Urban construction has produced a large amount of construction waste which has caused huge environmental problems. The sponge city is the development direction of urban construction, and permeable pavement concrete is an important material for sponge city construction. To see the law influencing different factors on the performance of recycled aggregate permeable pavement concrete, different water binder ratios, recycled aggregate particle gradations, ordinary aggregate substitution rates, and fly ash and admixture contents are designed to prepare permeable concrete. The compressive strength, permeability coefficient, frost resistance, and pore structure of permeable concrete are tested. The results show that when the replacement rate of recycled aggregate is 50%, the 28-d strength of concrete with a 0.25 water binder ratio can reach 28.9 MPa, and the permeability coefficient is 13.26 mm/s. The addition of fly ash will reduce the compressive strength, and the permeability coefficient increases first and then decreases with the increase of the fly ash content. When the mass fraction of fly ash instead of cement is 12%, the 28-d strength is 94.8% of that of the cement group, and the permeability coefficient can reach 14.03 mm/s. A water-reducing agent can obviously improve the workability of permeable concrete; the best content of the water-reducing agent is 0.2% of the cement mass. A reasonable amount of fly ash and water-reducing agent can optimize the number of harmless holes and less harmful holes in the concrete to improve the frost resistance and strength after the freeze-thaw, and the frost resistance is F150. This study provides a theoretical basis and technical guarantee for the resource utilization of recycled aggregate in permeable pavement concrete.

Polyanionic Macromolecular Prodrugs of Ribavirin: Antiviral Agents with a Broad Spectrum of Activity.

Macromolecular prodrugs are developed as multiarmed agents against diverse viral pathogens. Lead candidates inhibit infectivity and replication of HIV, Ebola, influenza, measles, RSV, etc-thus being broad-spectrum antiviral agents.

Analysis of Light Transport Features in Stone Fruits Using Monte Carlo Simulation.

The propagation of light in stone fruit tissue was modeled using the Monte Carlo (MC) method. Peaches were used as the representative model of stone fruits. The effects of the fruit core and the skin on light transport features in the peaches were assessed. It is suggested that the skin, flesh and core should be separately considered with different parameters to accurately simulate light propagation in intact stone fruit. The detection efficiency was evaluated by the percentage of effective photons and the detection sensitivity of the flesh tissue. The fruit skin decreases the detection efficiency, especially in the region close to the incident point. The choices of the source-detector distance, detection angle and source intensity were discussed. Accurate MC simulations may result in better insight into light propagation in stone fruit and aid in achieving the optimal fruit quality inspection without extensive experimental measurements.

Synthesis and Antimicrobial Activity of the Hybrid Molecules between Sulfonamides and Active Antimicrobial Pleuromutilin Derivative.

A series of novel hybrid molecules between sulfonamides and active antimicrobial 14-o-(3-carboxy-phenylsulfide)-mutilin were synthesized, and their in vitro antibacterial activities were evaluated by the broth microdilution. Results indicated that these compounds displayed potent antimicrobial activities in vitro against various drug-susceptible and drug-resistant Gram-positive bacteria such as Staphylococci and streptococci, including methicillin-resistant Staphylococcus aureus, and mycoplasma. In particular, sulfapyridine analog (6c) exhibited more potent inhibitory activity against Gram-positive bacteria and mycoplasma, including Staphylococcus aureus (MIC = 0.016-0.063 µg/mL), methicillin-resistant Staphylococcus aureus (MIC = 0.016 µg/mL), Streptococcus pneumoniae (MIC = 0.032-0.063 µg/mL), Mycoplasma gallisepticum (MIC = 0.004 µg/mL), with respect to other synthesized compounds and reference drugs sulfonamide (MIC = 8-128 µg/mL) and valnemulin (MIC = 0.004-0.5 µg/mL). Furthermore, comparison between MIC values of pleuromutilin-sulfonamide hybrids 6a-f with pleuromutilin parent compound 3 revealed that these modifications at 14 position side chain of the pleuromutilin with benzene sulfonamide could greatly improve the antibacterial activity especially against Gram-positives.

The anxiolytic effects of valtrate in rats involves changes of corticosterone levels.

Valtrate is a principle compound isolated from Valeriana jatamansi Jones, which is a Traditional Chinese Medicine used to treat various mood disorders. The aim of the present study was to investigate the anxiolytic effects of valtrate in rats. The animals were orally administered valtrate (5, 10, and 20 g/kg daily) for 10 days and exposed to open field test (OFT) and elevated plus-maze (EPM). Then the corticosterone levels in the rat serum were measured by enzyme-linked immunosorbent assay (ELISA). The valtrate (10 mg/kg, p.o.) exhibited the anxiolytic effect in rats by increasing the time and entry percentage into the open arms in the EPM and the number of central entries in the OFT. Valtrate (10 mg/kg, p.o.) significantly reduced the corticosterone level in the rat serum. Taken together, these results suggest that the valtrate has anxiolytic activity in behavioral models that might be mediated via the function of hypothalamus-pituitary-adrenal axis.

Inhibition of influenza virus in vivo by siRNA delivered using ABA triblock copolymer synthesized by reversible addition-fragmentation chain-transfer polymerization.

AIM: Influenza virus remains a major threat, with outbreaks continuing to occur. Few treatment options are available and drug resistance can emerge rapidly. New drugs that can quickly be adapted to virus mutations are needed. Several highly effective siRNAs targeting influenza that inhibit virus replication are known; however, effective delivery of these siRNAs remains a challenge. The aim of this study was to demonstrate the safety and efficacy of ABA triblock copolymer-delivered siRNA to inhibit influenza virus replication in vivo. MATERIALS & METHODS: We report on the delivery of a siRNA targeting the influenza virus in chicken embryos using an ABA triblock copolymer prepared by reversible addition-fragmentation chain-transfer polymerization, containing a central cationic block and two outer hydrophilic polyethylene glycol blocks. RESULTS: A significant reduction of virus titer was observed with the polymer/anti-influenza siRNA complexes, whereas the control with polymer/control siRNA complexes showed no effect. CONCLUSION: These data suggest that a reversible addition-fragmentation chain transfer-based siRNA delivery platform may be suitable for combating infectious diseases in vivo.

Synthesis and evaluation of degradable polyurea block copolymers as siRNA delivery agents.

Chain extension by diisocyanate condensation provides a versatile and convenient means for preparing block copolymers. We have utilized this chemistry to prepare reducible multiblock polycations for siRNA delivery. This approach, an alternative to oxidative coupling, was suitable for preparing multiblock polycations with defined molecular weight and architecture. The polymer, PEG-b-multi-(polyhexylurea-co-oligo-L-lysine)-b-PEG, was capable of electrostatically condensing siRNA to form nano-sized polyplexes across a broad compositional range. We demonstrated that the polyplexes enter the cells via endocytosis and interact with the endosome membrane leading to destabilization and hence endosome escape. Another feature of these polymers is their multiple intra-chain disulfide linkages. This enables weakening of the polyplex via chain scission within the cytosol's reductive environment. In addition to the controlled preparation of the polymer, the polyplexes were capable of delivering siRNA in vitro to silence greater than 50% green fluorescent protein expression with negligible toxicity.

The effect of RAFT-derived cationic block copolymer structure on gene silencing efficiency.

In this work a series of ABA tri-block copolymers was prepared from oligo(ethylene glycol) methyl ether methacrylate (OEGMA(475)) and N,N-dimethylaminoethyl methacrylate (DMAEMA) to investigate the effect of polymer composition on cell viability, siRNA uptake, serum stability and gene silencing. Reversible Addition-Fragmentation Chain Transfer (RAFT) polymerization was used as the method of polymer synthesis as this technique allows the preparation of well-defined block copolymers with low polydispersity. Eight block copolymers were prepared by systematically varying the central cationic block (DMAEMA) length from 38 to 192 monomer units and the outer hydrophilic block (OEGMA(475)) from 7 to 69 units. The polymers were characterized using size exclusion chromatography and (1)H NMR. Chinese Hamster Ovary-GFP and Human Embryonic Kidney 293 cells were used to assay cell viability while the efficiency of block copolymers to complex with siRNA was evaluated by agarose gel electrophoresis. The ability of the polymer-siRNA complexes to enter into cells and to silence the targeted reporter gene enhanced green fluorescent protein (EGFP) was measured by using a CHO-GFP silencing assay. The length of the central cationic block appears to be the key structural parameter that has a significant effect on cell viability and gene silencing efficiency with block lengths of 110-120 monomer units being the optimum. The ABA block copolymer architecture is also critical with the outer hydrophilic blocks contributing to serum stability and overall efficiency of the polymer as a delivery system.

Biodistribution of polymer hydrogel capsules for the delivery of therapeutics.

A key phase in the development of intelligently designed nanoparticle delivery vehicles for new therapeutic agents is to gain an understanding of their interaction with tissues and cells. We report a series of in vitro and in vivo experiments aimed at tracking a potential delivery vehicle for therapeutic agents, including vaccine peptides and drugs derived from poly(methacrylic acid) hydrogel capsules in certain organs and cell types. For the in vitro studies, two immortal liver-derived cell lines (Huh7 and Hepa1-6) and primary cultures of mouse hepatocytes were incubated with Alexa 647 labelled fluorescent capsules to track their internalization and intracellular distribution by confocal microscopy. Capsules, 500nm in diameter, were taken up into the cells in a time-dependent manner in all three cell lines. Capsules were observed in plasma membrane-derived vesicles within the cells. After 24h a significant proportion of the capsules was observed in lysosomes. To understand the behaviour of the capsules in vivo, Alexa 488 labelled fluorescent capsules were intravenously injected into Sprague-Dawley rats and after 24h the fate of the capsules in a number of organs was determined by flow cytometry and confocal microscopy. By flow cytometry, the majority of the capsules were detected in the spleen whilst similar numbers were found in the lung and liver. By confocal microscopy, the majority of the capsules were found in the liver and spleen with significantly less capsules in the lung, heart and kidney. Colocalization of capsules with cell-type specific markers indicated that in lung, heart and kidney, the majority of the capsules were located in endothelial cells. In the spleen ~50% of the capsules were found in CD163-positive cells, whereas in the liver, almost all capsules were located in CD163-positive cells, indicating uptake by Kupffer cells. Electron microscopy confirmed the presence of capsules within Kupffer cells.

Vibrio fluvialis peritonitis in a patient receiving continuous ambulatory peritoneal dialysis.

We describe a case of peritonitis due to Vibrio fluvialis in a patient receiving continuous ambulatory peritoneal dialysis; we believe the case to be associated with the consumption of poorly prepared seafood. This was shown to be an important but rare cause of recurrent infection in our patient.