Three point turn: A metallosupramolecular helicate with trigonal‐antiprismatic geometry fits perfectly into the central hydrophobic cavity of a three‐way DNA junction allowing a new mode of DNA ...recognition.
Respiratory syncytial virus (RSV) is a leading cause of infant hospitalization and there remains no pediatric vaccine. RSV live-attenuated vaccines (LAVs) have a history of safe testing in infants; ...however, achieving an effective balance of attenuation and immunogenicity has proven challenging. Here we seek to engineer an RSV LAV with enhanced immunogenicity. Genetic mapping identifies strain line 19 fusion (F) protein residues that correlate with pre-fusion antigen maintenance by ELISA and thermal stability of infectivity in live RSV. We generate a LAV candidate named OE4 which expresses line 19F and is attenuated by codon-deoptimization of non-structural (NS1 and NS2) genes, deletion of the small hydrophobic (SH) gene, codon-deoptimization of the attachment (G) gene and ablation of the secreted form of G. OE4 (RSV-A2-dNS1-dNS2-ΔSH-dG
-Gs
-line19F) exhibits elevated pre-fusion antigen levels, thermal stability, immunogenicity, and efficacy despite heavy attenuation in the upper and lower airways of cotton rats.
Although respiratory syncytial virus (RSV) is the leading cause of lower respiratory tract infections in infants, a safe and effective vaccine is not yet available. Live-attenuated vaccines (LAVs) ...are the most advanced vaccine candidates in RSV-naive infants. However, designing an LAV with appropriate attenuation yet sufficient immunogenicity has proven challenging. In this study, we implemented reverse genetics to address these obstacles with a multifaceted LAV design that combined the codon deoptimization of genes for nonstructural proteins NS1 and NS2 (dNS), deletion of the small hydrophobic protein (ΔSH) gene, and replacement of the wild-type fusion (F) protein gene with a low-fusion RSV subgroup B F consensus sequence of the Buenos Aires clade (BAF). This vaccine candidate, RSV-A2-dNS-ΔSH-BAF (DB1), was attenuated in two models of primary human airway epithelial cells and in the upper and lower airways of cotton rats. DB1 was also highly immunogenic in cotton rats and elicited broadly neutralizing antibodies against a diverse panel of recombinant RSV strains. When vaccinated cotton rats were challenged with wild-type RSV A, DB1 reduced viral titers in the upper and lower airways by 3.8 log10 total PFU and 2.7 log10 PFU/g of tissue, respectively, compared to those in unvaccinated animals (P < 0.0001). DB1 was thus attenuated, highly immunogenic, and protective against RSV challenge in cotton rats. DB1 is the first RSV LAV to incorporate a low-fusion F protein as a strategy to attenuate viral replication and preserve immunogenicity.
RSV is a leading cause of infant hospitalizations and deaths. The development of an effective vaccine for this high-risk population is therefore a public health priority. Although live-attenuated vaccines have been safely administered to RSV-naive infants, strategies to balance vaccine attenuation with immunogenicity have been elusive. In this study, we introduced a novel strategy to attenuate a recombinant RSV vaccine by incorporating a low-fusion, subgroup B F protein in the genetic background of codon-deoptimized nonstructural protein genes and a deleted small hydrophobic protein gene. The resultant vaccine candidate, DB1, was attenuated, highly immunogenic, and protective against RSV challenge in cotton rats.
We previously reported that the Toll-like receptor 4 (TLR4) antagonist Eritoran blocks acute lung injury (ALI) therapeutically in mouse and cotton rat models of influenza. However, secondary (2°) ...bacterial infection following influenza virus infection is associated with excess morbidity and mortality. Wild-type (WT) mice infected with mouse-adapted influenza A/Puerto Rico/8/34 virus (PR8) and, 7 days later, with
serotype 3 (
) exhibited significantly enhanced lung pathology and lethality that was reversed by Eritoran therapy after PR8 infection but before
infection. Cotton rats infected with nonadapted pH1N1 influenza virus and then superinfected with methicillin-resistant
also exhibited increased lung pathology and serum high-mobility-group box 1 (HMGB1) levels, both of which were blunted by Eritoran therapy. In mice, PR8 infection suppressed
-induced CXCL1 and CXCL2 mRNA, reducing neutrophil infiltration and increasing the bacterial burden, all of which were reversed by Eritoran treatment. While beta interferon (IFN-β)-deficient (IFN-β
) mice are highly susceptible to PR8, they exhibited delayed death upon
superinfection, indicating that while IFN-β was protective against influenza, it negatively impacted the host response to
IFN-β-treated WT macrophages selectively suppressed
-induced CXCL1/CXCL2 transcriptionally, as evidenced by reduced recruitment of RNA polymerase II to the CXCL1 promoter. Thus, influenza establishes a "trained" state of immunosuppression toward 2° bacterial infection, in part through the potent induction of IFN-β and its downstream transcriptional regulation of chemokines, an effect reversed by Eritoran.
Enhanced susceptibility to 2° bacterial infections following infection with influenza virus is a global health concern that accounts for many hospitalizations and deaths, particularly during pandemics. The complexity of the impaired host immune response during 2° bacterial infection has been widely studied. Both type I IFN and neutrophil dysfunction through decreased chemokine production have been implicated as mechanisms underlying enhanced susceptibility to 2° bacterial infections. Our findings support the conclusion that selective suppression of CXCL1/CXCL2 represents an IFN-β-mediated "training" of the macrophage transcriptional response to TLR2 agonists and that blocking of TLR4 therapeutically with Eritoran after influenza virus infection reverses this suppression by blunting influenza-induced IFN-β.
Respiratory syncytial virus (RSV) is a significant threat to elderly populations and repeated infections that occur throughout life are poorly protective. To assess the role of prior RSV infections ...as well as elderly immune senescence on vaccine efficacy, we compared immune responses after virus-like particle (VLP) immunization of elderly cotton rats and young cotton rats, both previously RSV infected, in order to mimic the human population. We show that immunization of RSV-experienced young or elderly animals resulted in the same levels of anti-pre-F IgG, anti-G IgG, neutralizing antibody titers, and protection from challenge indicating that the delivery of F and G proteins in a VLP is equally effective in activation of protective responses in both elderly and young populations. Our results suggest that F and G protein-containing VLPs induce anti-RSV memory established in prior RSV infections equally well in both young and elderly animals and thus can be an effective vaccine for the elderly.
The strong dose-dependent association between anthracyclines and cardiomyopathy is further exacerbated by the co-occurrence of cardiovascular risk factors (diabetes and hypertension). The high ...morbidity associated with cardiomyopathy necessitates an understanding of the underlying pathogenesis so that targeted interventions can be developed.
By using a two-stage design, we investigated host susceptibility to anthracycline-related cardiomyopathy by using the ITMAT/Broad CARe cardiovascular single nucleotide polymorphism (SNP) array to profile common SNPs in 2,100 genes considered relevant to de novo cardiovascular disease.
By using a matched case-control design (93 cases, 194 controls), we identified a common SNP, rs2232228, in the hyaluronan synthase 3 (HAS3) gene that exerts a modifying effect on anthracycline dose-dependent cardiomyopathy risk (P = 5.3 × 10(-7)). Among individuals with rs2232228 GG genotype, cardiomyopathy was infrequent and not dose related. However, in individuals exposed to high-dose (> 250 mg/m(2)) anthracyclines, the rs2232228 AA genotype conferred an 8.9-fold (95% CI, 2.1- to 37.5-fold; P = .003) increased cardiomyopathy risk compared with the GG genotype. This gene-environment interaction was successfully replicated in an independent set of 76 patients with anthracycline-related cardiomyopathy. Relative HAS3 mRNA levels measured in healthy hearts tended to be lower among individuals with AA compared with GA genotypes (P = .09).
Hyaluronan (HA) produced by HAS3 is a ubiquitous component of the extracellular matrix and plays an active role in tissue remodeling. In addition, HA is known to reduce reactive oxygen species (ROS) -induced cardiac injury. The high cardiomyopathy risk associated with AA genotype could be due to inadequate remodeling and/or inadequate protection of the heart from ROS-mediated injury on high anthracycline exposure.
Adsorption of organo-sulfur compounds present in liquid fuels on metal–organic framework (MOF) compounds is an efficient alternative to the conventional hydrodesulfurization process. It has been ...demonstrated that the extent of dibenzothiophene (DBT) adsorption at temperatures close to ambient (304
K) is much higher on MOFs systems than on the benchmarked Y-type zeolite. In addition, the DBT adsorption capacity depends strongly on the MOF type as illustrated by the much higher extent of adsorption observed on the copper- (C300) and Al-containing (A100) MOF systems than on the Fe-containing (F300) MOF counterpart. With the aim to investigate the operation in consecutive cycles, the MOFs used in adsorption experiments were regenerated. In addition, the remaining S-containing compounds were identified and quantified by photoelectron spectroscopy (XPS). Examination of S2p core-level spectrum of the adsorbed S-compounds of regenerated MOFs pointed out that a fraction of these molecules become oxidized into S(VI) species.
Many respiratory viruses cause lung damage that may evolve into acute lung injury (ALI), a cytokine storm, acute respiratory distress syndrome, and ultimately, death. Peroxisome proliferator ...activated receptor gamma (PPARγ), a member of the nuclear hormone receptor (NHR) family of transcription factors, regulates transcription by forming heterodimers with another NHR family member, Retinoid X Receptor (RXR). Each component of the heterodimer binds specific ligands that modify transcriptional capacity of the entire heterodimer by recruiting different co-activators/co-repressors. However, the role of PPARγ/RXR ligands in the context of influenza infection is not well understood. PPARγ is associated with macrophage differentiation to an anti-inflammatory M2 state. We show that mice lacking the IL-4Rα receptor, required for M2a macrophage differentiation, are more susceptible to mouse-adapted influenza (A/PR/8/34; "PR8")-induced lethality. Mice lacking
, that encodes COX-2, a key proinflammatory M1 macrophage mediator, are more resistant. Blocking the receptor for COX-2-induced Prostaglandin E
(PGE
) was also protective. Treatment with pioglitazone (PGZ), a PPARγ ligand, increased survival from PR8 infection, decreased M1 macrophage gene expression, and increased PPARγ mRNA in lungs. Conversely, conditional knockout mice expressing PPARγ-deficient macrophages were significantly more sensitive to PR8-induced lethality. These findings were extended in cotton rats: PGZ blunted lung inflammation and M1 cytokine gene expression after challenge with non-adapted human influenza. To study mechanisms by which PPARγ/RXR transcription factors induce canonical M2a genes, WT mouse macrophages were treated with IL-4 in the absence or presence of rosiglitazone (RGZ; PPARγ ligand), LG100754 (LG; RXR ligand), or both. IL-4 dose-dependently induced M2a genes
,
. Treatment of macrophages with IL-4 and RGZ and/or LG differentially affected induction of
and
.
and
gene expression. In PPARγ-deficient macrophages, IL-4 alone failed to induce
and
gene expression; however, concurrent treatment with LG or RGZ + LG enhanced IL-4-induced
and
expression, but to a lower level than in WT macrophages, findings confirmed in the murine alveolar macrophage cell line, MH-S. These findings support a model in which PPARγ/RXR heterodimers control IL-4-induced M2a differentiation, and suggest that PPARγ/RXR agonists should be considered as important tools for clinical intervention against influenza-induced ALI.
MicroRNAs are tiny non-coding small endogenous RNAs that regulate gene expression by translational repression, mRNA cleavage and mRNA inhibition. The aim of this study was to investigate the ...hypermethylation of miR-34b/c and miR-148a in colorectal cancer, and correlate this data to clinicopathological features. We also aimed to evaluate the hypermethylation of miR-34b/c in faeces specimens as a novel non-invasive faecal-DNA-based screening marker.
The 5-aza-2'-deoxycytidine treatment and methylation-specific PCR were carried out to detect the hypermethylation of miR-34b/c and miR-148a.
The miR-34b/c hypermethylation was found in 97.5% (79 out of 82) of primary colorectal tumours, P=0.0110. In 75% (21 out of 28) of faecal specimens we found a hypermethylation of miR-34b/c while only in 16% (2 out of 12) of high-grade dysplasia. In addition, miR-148a was found to be hypermethylated in 65% (51 out of 78) of colorectal tumour tissues with no significant correlation to clinicopathological features. However, a trend with female gender and advanced age was found, P=0.083. We also observed a trend to lower survival rate in patients with miR-148a hypermethylation with 10-year survival probability: 48 vs 65%, P=0.561.
These findings show that aberrant hypermethylation of miR-34b/c could be an ideal class of early screening marker, whereas miR-148a could serve as a disease progression follow-up marker.
A lab-scale Upflow Anaerobic Sludge Blanket (UASB) reactor was used as a model for evaluating synthetic and complex industrial wastewater treatment, using a solar heater to control temperature. Also, ...hydrodynamics was assessed using the Computational Fluid Dynamics (CFD) method. Initially, the UASB reactor was operated with synthetic wastewater at Hydraulic Retention Time (HRT) of 24 h in 20 ± 2 °C and 30 ± 2 °C to measure the biogas bubbles production for CFD study. COD removal efficiencies of 85 ± 3% and 95 ± 3%, respectively, with production of 27 and 39 ml CH
4
/h, correspondingly, were observed. After that, the reactor was fed with complex industrial wastewater. It was evaluated at 24 h in both temperatures. At 30 °C, low COD removal efficiency was observed, being 48 ± 13%, with methane production of 20 ± 3 ml CH
4
/h. The plug flow pattern was observed in the CFD modelling at HRT of 24 h and 20 °C without considering biogas bubbles interaction. Similar hydrodynamic behaviour was observed at HRT of 24 h and 30 °C. Nonetheless, when biogas bubbles were considered in the CFD modelling, hydrodynamics significantly changed, passing from a plug flow to a complete mix flow pattern.
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