Identification and characterization of a Ross River virus variant that grows persistently in macrophages, shows altered disease kinetics in a mouse model, and exhibits resistance to type I interferon

Alphaviruses, such as chikungunya virus, o'nyong-nyong virus, and Ross River virus (RRV), cause outbreaks of human rheumatic disease worldwide. RRV is a positive-sense single-stranded RNA virus endemic to Australia and Papua New Guinea. In this study, we sought to establish an in vitro model of RRV evolution in response to cellular antiviral defense mechanisms. RRV was able to establish persistent infection in activated macrophages, and a small-plaque variant (RRVPERS) was isolated after several weeks of culture. Nucleotide sequence analysis of RRV PERS found several nucleotide differences in the nonstructural protein (nsP) region of the RRV PERS genome. A point mutation was also detected in the E2 gene. Compared to the parent virus (RRV-T48), RRV PERS showed significantly enhanced resistance to beta interferon (IFN-β)-stimulated antiviral activity. RRV PERS infection of RAW 264.7 macrophages induced lower levels of IFN-β expression and production than infection with RRV-T48. RRV PERS was also able to inhibit type I IFN signaling. Mice infected with RRV PERS exhibited significantly enhanced disease severity and mortality compared to mice infected with RRV-T48. These results provide strong evidence that the cellular antiviral response can direct selective pressure for viral sequence evolution that impacts on virus fitness and sensitivity to alpha/beta IFN (IFN-α/β).Facultad de Ciencias Exacta

Carnosine Protects Macrophages against the Toxicity of Aβ1-42 Oligomers by Decreasing Oxidative Stress

Carnosine (β-alanyl-L-histidine) is a naturally occurring endogenous peptide widely distributed in excitable tissues such as the brain. This dipeptide has well-known antioxidant, anti-inflammatory, and anti-aggregation activities, and it may be useful for treatment of neurodegenerative disorders such as Alzheimer’s disease (AD). In this disease, peripheral infiltrating macrophages play a substantial role in the clearance of amyloid beta (Aβ) peptides from the brain. Correspondingly, in patients suffering from AD, defects in the capacity of peripheral macrophages to engulf Aβ have been reported. The effects of carnosine on macrophages and oxidative stress associated with AD are consequently of substantial interest for drug discovery in this field. In the present work, a model of stress induced by Aβ1-42 oligomers was investigated using a combination of methods including trypan blue exclusion, microchip electrophoresis with laser-induced fluorescence, flow cytometry, fluorescence microscopy, and high-throughput quantitative real-time PCR. These assays were used to assess the ability of carnosine to protect macrophage cells, modulate oxidative stress, and profile the expression of genes related to inflammation and pro- and antioxidant systems. We found that pre-treatment of RAW 264.7 macrophages with carnosine counteracted cell death and apoptosis induced by Aβ1-42 oligomers by decreasing oxidative stress as measured by levels of intracellular nitric oxide (NO)/reactive oxygen species (ROS) and production of peroxynitrite. This protective activity of carnosine was not mediated by modulation of the canonical inflammatory pathway but instead can be explained by the well-known antioxidant and free-radical scavenging activities of carnosine, enhanced macrophage phagocytic activity, and the rescue of fractalkine receptor CX3CR1. These new findings obtained with macrophages challenged with Aβ1-42 oligomers, along with the well-known multimodal mechanism of action of carnosine in vitro and in vivo, substantiate the therapeutic potential of this dipeptide in the context of AD pathology

Identification and characterization of a Ross River virus variant that grows persistently in macrophages, shows altered disease kinetics in a mouse model, and exhibits resistance to type I interferon

Alphaviruses, such as chikungunya virus, o'nyong-nyong virus, and Ross River virus (RRV), cause outbreaks of human rheumatic disease worldwide. RRV is a positive-sense single-stranded RNA virus endemic to Australia and Papua New Guinea. In this study, we sought to establish an in vitro model of RRV evolution in response to cellular antiviral defense mechanisms. RRV was able to establish persistent infection in activated macrophages, and a small-plaque variant (RRVPERS) was isolated after several weeks of culture. Nucleotide sequence analysis of RRV PERS found several nucleotide differences in the nonstructural protein (nsP) region of the RRV PERS genome. A point mutation was also detected in the E2 gene. Compared to the parent virus (RRV-T48), RRV PERS showed significantly enhanced resistance to beta interferon (IFN-β)-stimulated antiviral activity. RRV PERS infection of RAW 264.7 macrophages induced lower levels of IFN-β expression and production than infection with RRV-T48. RRV PERS was also able to inhibit type I IFN signaling. Mice infected with RRV PERS exhibited significantly enhanced disease severity and mortality compared to mice infected with RRV-T48. These results provide strong evidence that the cellular antiviral response can direct selective pressure for viral sequence evolution that impacts on virus fitness and sensitivity to alpha/beta IFN (IFN-α/β).Facultad de Ciencias Exacta

Ruxolitinib for Glucocorticoid-Refractory Acute Graft-versus-Host Disease

BACKGROUND: Acute graft-versus-host disease (GVHD) remains a major limitation of allogeneic stem-cell transplantation; not all patients have a response to standard glucocorticoid treatment. In a phase 2 trial, ruxolitinib, a selective Janus kinase (JAK1 and JAK2) inhibitor, showed potential efficacy in patients with glucocorticoid-refractory acute GVHD. METHODS: We conducted a multicenter, randomized, open-label, phase 3 trial comparing the efficacy and safety of oral ruxolitinib (10 mg twice daily) with the investigator's choice of therapy from a list of nine commonly used options (control) in patients 12 years of age or older who had glucocorticoid-refractory acute GVHD after allogeneic stem-cell transplantation. The primary end point was overall response (complete response or partial response) at day 28. The key secondary end point was durable overall response at day 56. RESULTS: A total of 309 patients underwent randomization; 154 patients were assigned to the ruxolitinib group and 155 to the control group. Overall response at day 28 was higher in the ruxolitinib group than in the control group (62% [96 patients] vs. 39% [61]; odds ratio, 2.64; 95% confidence interval [CI], 1.65 to 4.22; P<0.001). Durable overall response at day 56 was higher in the ruxolitinib group than in the control group (40% [61 patients] vs. 22% [34]; odds ratio, 2.38; 95% CI, 1.43 to 3.94; P<0.001). The estimated cumulative incidence of loss of response at 6 months was 10% in the ruxolitinib group and 39% in the control group. The median failure-free survival was considerably longer with ruxolitinib than with control (5.0 months vs. 1.0 month; hazard ratio for relapse or progression of hematologic disease, non-relapse-related death, or addition of new systemic therapy for acute GVHD, 0.46; 95% CI, 0.35 to 0.60). The median overall survival was 11.1 months in the ruxolitinib group and 6.5 months in the control group (hazard ratio for death, 0.83; 95% CI, 0.60 to 1.15). The most common adverse events up to day 28 were thrombocytopenia (in 50 of 152 patients [33%] in the ruxolitinib group and 27 of 150 [18%] in the control group), anemia (in 46 [30%] and 42 [28%], respectively), and cytomegalovirus infection (in 39 [26%] and 31 [21%]). CONCLUSIONS: Ruxolitinib therapy led to significant improvements in efficacy outcomes, with a higher incidence of thrombocytopenia, the most frequent toxic effect, than that observed with control therapy

FFSiOH: a New Force Field for Silica Polymorphs and Their Hydroxylated Surfaces Based on Periodic B3LYP Calculations

A partial charge shell-ion model potential for silica polymorphs and their hydroxylated surfaces(FFSiOH) was parametrized in a self-consistent way using periodic B3LYP results for bulk R-cristobaliteand the (100) and (001) hydroxylated surfaces. The reliability of the new potentials was checked bycomparing structures, vibrational frequencies and relative phase stabilities of dense bulk silica polymorphs,namely R-quartz, R-cristobalite, R-tridymite, and Stishovite with both experimental and B3LYP data.The FFSiOH was also checked for computing structural and vibrational features of representative all-silica microporous materials, namely edingtonite, chabazite, and faujasite. As a last step, FFSiOH wasadopted to predict OH stretching vibrational frequencies and relative thermodynamic stability of themost common fully hydroxylated surfaces of the dense silica polymorphs, the (100) and (001) facesof all-silica edingtonite, the features of the local Si-defect in chabazite and sodalite known as (SiOH)4hydrogarnet and the geometries of H-bonded silanol groups of an amorphous silica surface. In all casesexcellent agreement resulted between FFSiOH and B3LYP periodic data and experimental data, whenavailable. The new FFSiOH force field opens up the molecular simulation of materials in which thesurface hydroxyl groups play a key role, as is the case for amorphous silica surfaces, all-silica zeoliteexternal surfaces, and the internal walls of mesoporous materials

Design, Synthesis, and Evaluation of 5 '-Diphenyl Nucleoside Analogues as Inhibitors of the Plasmodium falciparum dUTPase

Deoxyuridine 5'-triphosphate nucleotidohydrolase (dUTPase) is a potential drug target for malaria. We previously reported some 5'-tritylated deoxyuridine analogues (both cyclic and acyclic) as selective inhibitors of the Plasmodium falciparum dUTPase. Modelling studies indicated that it might be possible to replace the trityl group with a diphenyl moiety, as two of the phenyl groups are buried, whereas the third is exposed to solvent. Herein we report the synthesis and evaluation of some diphenyl analogues that have lower lipophilicity and molecular weight than the trityl lead compound. Co-crystal structures show that the diphenyl inhibitors bind in a similar manner to the corresponding trityl derivatives, with the two phenyl moieties occupying the predicted buried phenyl binding sites. The diphenyl compounds prepared show similar or slightly lower inhibition of PfdUTPase, and similar or weaker inhibition of parasite growth than the trityl compound