Search

The SARS-CoV-2 life cycle is strictly dependent on the environmental redox state that influences both virus entry and replication. A reducing environment impairs the binding of the spike protein (S) to the angiotensin-converting enzyme 2 receptor (ACE2), while a highly oxidizing environment is thought to favor S interaction with ACE2. Moreover, SARS-CoV-2 interferes with redox homeostasis in infected cells to promote the oxidative folding of its own proteins. Here we demonstrate that synthetic low molecular weight (LMW) monothiol and dithiol compounds induce a redox switch in the S protein receptor binding domain (RBD) toward a more reduced state. Reactive cysteine residue profiling revealed that all the disulfides present in RBD are targets of the thiol compounds. The reduction of disulfides in RBD decreases the binding to ACE2 in a cell-free system as demonstrated by enzyme-linked immunosorbent and surface plasmon resonance (SPR) assays. Moreover, LMW thiols interfere with protein oxidative folding and the production of newly synthesized polypeptides in HEK293 cells expressing the S1 and RBD domain, respectively. Based on these results, we hypothesize that these thiol compounds impair both the binding of S protein to its cellular receptor during the early stage of viral infection, as well as viral protein folding/maturation and thus the formation of new viral mature particles. Indeed, all the tested molecules, although at different concentrations, efficiently inhibit both SARS-CoV-2 entry and replication in Vero E6 cells. LMW thiols may represent innovative anti-SARS-CoV-2 therapeutics acting directly on viral targets and indirectly by inhibiting cellular functions mandatory for viral replication. [ABSTRACT FROM AUTHOR]

[Display omitted] The mechanisms by which myeloid-derived suppressor cells (MDSCs) mediate inhibition prominently include the production of reactive nitrogen species, in particular those generated by inducible nitric oxide synthase (iNOS), and reactive oxygen species. LP-BM5 murine retroviral infection results in a profound immunodeficiency, known as murine AIDS, as well as in increased numbers and activity of monocytic-type MDSCs (M-MDSCs) that suppress both T and B cell responses. While M-MDSCs suppress T cells ex vivo in a fully iNOS/NO-dependent manner, M-MDSC suppression of B cell responses is only partially due to iNOS/NO. This study preliminarily explored the role of two redox-modulating compounds in inhibiting the M-MDSC suppressive activity in LP-BM5 infection. The tested molecules were: I-152 consisting in a conjugate of N-acetyl-cysteine (NAC) and S-acetyl-cysteamine (SMEA) and C4-GSH that is the n-butanoyl glutathione (GSH) derivative. The results show that both molecules, tested in a concentration range between 3 and 20 mM, blocked the M-MDSC suppression of activated B and T cells ex vivo and restored their proliferative capacity in vivo. Ex vivo I-152 blockade of M-MDSC suppressiveness was more significant for T cell (about 70%) while M-MDSC blockade by C4-GSH was preferential for B cell responsiveness (about 60%), which was also confirmed by in vivo investigation. Beyond insights into redox-dependent suppressive effector mechanism(s) of M-MDSCs in LP-BM5 infection, these findings may ultimately be important to identify new immunotherapeutics against infectious diseases. [ABSTRACT FROM AUTHOR]

Reduced glutathione (GSH) is the most prevalent non-protein thiol in animal cells. Its de novo and salvage synthesis serves to maintain a reduced cellular environment, which is important for several cellular functions. Altered intracellular GSH levels are observed in a wide range of pathologies, including several viral infections, as well as in aging, all of which are also characterized by an unbalanced Th1/Th2 immune response. A central role in influencing the immune response has been ascribed to GSH. Specifically, GSH depletion in antigen-presenting cells (APCs) correlates with altered antigen processing and reduced secretion of Th1 cytokines. Conversely, an increase in intracellular GSH content stimulates IL-12 and/or IL-27, which in turn induces differentiation of naive CD4+ T cells to Th1 cells. In addition, GSH has been shown to inhibit the replication/survival of several pathogens, i.e. viruses and bacteria. Hence, molecules able to increase GSH levels have been proposed as new tools to more effectively hinder different pathogens by acting as both immunomodulators and antimicrobials. Herein, the new role of GSH and its derivatives as immunotherapeutics will be discussed. [ABSTRACT FROM AUTHOR]

Macrophages are an important defense against in vivo herpes simplex virus (HSV) infection by early cytokine secretion; however, they can be infected by HSV-1 and they may be compromised in their ability to produce cytokines. In this paper, we studied the expression of two Th1 cytokines, interleukin (IL)-12 and IL-27, upon HSV-1 infection of human macrophages, and how it is regulated by treatment with two antiviral drugs exerting their anti-HSV-1 activity through different mechanisms of action. We found that infection does not alter intra-macrophage thiol content, while it induces mRNA expression of IL-12 p35 and IL-12 p40 as well as of IL-27 p28 and IL-27 EBI3, as revealed by RT-PCR. The increased expression of mRNA is accompanied by increased production of IL-12 p40 and IL-27 p28 protein, as detected in the culture supernatants by ELISA. The two antiviral drugs tested were acyclovir (ACV), commonly used to treat herpes virus infections, and an N-butanoyl glutathione (GSH) derivative, GSH-C4. While ACV inhibits viral DNA polymerase, GSH-C4 inhibits virus replication by interfering with protein folding and maturation of viral particles. Indeed, GSH-C4, altering the intracellular redox state, may modulate the Th1/Th2 balance favoring Th1-type response. Our data show that both drugs inhibit HSV-1 replication in macrophages, without significantly affecting cytokine mRNA levels. Nonetheless, lower levels of IL-12 p40 and IL-27 p28 proteins were found in the supernatants of macrophages treated with either GSH-C4 or ACV, likely as an indirect consequence of inhibited HSV-1 replication. [ABSTRACT FROM AUTHOR]

Background: The aim of this study was to investigate the molecular mechanisms involved in the production of Th1 cytokines, namely IL-12 and IL-27, when the intra-macrophage redox state was altered by different chemical entities such as GSH-C4, which is reduced glutathione carrying an aliphatic chain, or I-152, a pro-drug of N-acetyl-cysteine (NAC) and beta-mercaptoethylamine. We had already demonstrated that GSH-C4 and I-152 could shift the immune response towards Th1 in Ovalbumin-immunized mice as well as enhance Th1 response in HIV-1 Tat-immunized mice. Methodology/Principal Findings: By a new high performance liquid chromatography method, we found that 20 mM GSH-C4 provided a number of thiol species in the form of GSH, while 20 mM I-152 decreased GSH and increased the thiols in the form of NAC and I-152. Under these experimental conditions, GSH-C4 and I-152 enhanced and suppressed respectively the mRNA expression levels of IL-12 p40 induced by LPS/IFN-γ as assessed by Real-Time PCR. The protein production of IL-12 p40 was increased by GSH-C4 and decreased by I-152 as determined by Enzyme-linked immunosorbent assay. Western immunoblot and electrophoretic mobility shift assays revealed that Nuclear Factor -kB (NF-kB) activation was inhibited by I-152 and prolonged by GSH-C4. Twenty mM I-152 stimulated IL-27 p28 gene expression and sustained Signal Transducer and Activator of Transcription (STAT)-mediated interferon regulator factor 1 (IRF-1) de novo synthesis. By contrast, 20 mM GSH-C4 did not exert any effect on IL-27 p28 gene expression. Conclusions and Significance: an increase in the intra-macrophage redox state by GSH-C4 and I-152 enhances Th1 cytokine production although the chemical structure and the intra-cellular metabolism influence differently signalling pathways involved in IL-27 or IL-12 production. GSH-C4 and I-152 may be used as Th1 immunomodulators in some pathologies and in ageing where GSH depletion may contribute to the Th1/Th2 imbalance, and in new immunization strategies. [ABSTRACT FROM AUTHOR]

Abstract: We have previously demonstrated that in Ova-immunized mice the increase in intra-macrophage thiol pool induced by pro-GSH molecules modulates the Th1/Th2 balance in favour of a Th1-type immune response. We show now that the same molecules can support a Th1-type over Th2-type immunity against Tat, which is an early HIV-1 regulatory protein and a Th1 polarizing immunomodulator that is increasingly considered in new anti-HIV vaccination strategies. Our results indicate that Tat-immunized mice pre-treated with the C4 (n-butanoyl) derivative of reduced glutathione (GSH-C4) or a pro-drug of N-acetylcysteine (NAC) and beta-mercaptoethylamine (MEA) (I-152), have decreased levels of anti-Tat IgG1 as well as increased levels of anti-Tat IgG2a and IgG2b isotypes suggesting a Th1-type response. Moreover, Th1-(IFN-γ and IL-2) Ag-specific cellular responses were detected by ELISPOT assay in splenocytes of the same animals as well as an increase of IL-12 levels in the plasma. These findings suggest that the Th1 immune response to HIV-1 Tat could be further polarized by these molecules. These results together with those previously reported suggest that pro-GSH molecules could be used to modulate the immune response towards different antigens and may be further exploited for inducing specific Th1 immune responses against other HIV antigens as well as other intracellular pathogens in new Tat-based vaccination protocols. [Copyright &y& Elsevier]

Abstract: In the present work, the capacity of new pro-GSH molecules to increase the intra-macrophage thiol content in vitro and in vivo as well as to shift the immune response to Th1 in ovalbumin (Ova)-sensitized mice were examined. The molecules were the N-butanoyl GSH derivative, GSH-C4, and a pro-drug of N-acetylcysteine (NAC) and beta-mercaptoethylamine (MEA), I-152. In vitro, 2h-incubation with both molecules was found to increase intra-macrophage thiol content; in vivo, Ova-sensitized mice pre-treated by intraperitoneal administration of the pro-GSH molecules showed an increase in plasma anti-Ova IgG2a and IgG2b, characterizing Th1 immune response, and a decrease in IgG1, typical of the Th2 response. Such findings were connected to a shift to a Th1 response also involving splenocyte IFN-γ production as revealed by ELISPOT assay and higher levels of IL-12 in circulation. Although immune responses are in vivo mediated both by dendritic cells and macrophages, the data reported in this paper corroborate the suggestion that the pro-GSH molecules, increasing the intra-cellular thiol pool, modulate the Th1/Th2 balance favouring Th1-type responses and may be employed as Th1-directing adjuvants in new vaccination protocols and as immunomodulators in those diseases where Th1 response patterns are compromised in favour of Th2. [ABSTRACT FROM AUTHOR]

PNAPR2 is a peptide nucleic acid (PNA) complementary to a sequence of the viral protease-encoding gene, effective in blocking HIV release, when used at high doses. Erythrocytes (RBC) were used to target PNAPR2 to the macrophage compartment. The antiviral activity was assessed in human HIV-infected macrophages both as inhibition of p24 production and reduction of HIV DNA content. PNAPR2, either added to the medium at a concentration of 100 μM or loaded into RBC at about 40 μM, inhibited p24 production ~80% compared with infected samples and reduced HIV DNA content by 83% and 90%, respectively. The results show that (1) a stronger anti-HIV effect is achievable with higher doses of PNAPR2, both when given free and encapsulated into RBC; (2) the antiviral effect obtained by free PNAPR2 at a concentration of 100 μM is achievable by encapsulating it into RBC at a concentration of 40 μM, suggesting that RBC can be used as a delivery system to increase the antisense effect of PNAPR2. [ABSTRACT FROM AUTHOR]

Abstract: Reduced glutathione (GSH) is the most prevalent non-protein thiol in animal cells. Its de novo and salvage synthesis serves to maintain a reduced cellular environment. GSH is the most powerful intracellular antioxidant and plays a role in the detoxification of a variety of electrophilic compounds and peroxides via catalysis by glutathione-S-transferases (GST) and glutathione peroxidases (GPx). As a consequence, the ratio of reduced and oxidized glutathione (GSH:GSSG) serves as a representative marker of the antioxidative capacity of the cell. A deficiency in GSH puts the cell at risk for oxidative damage. An imbalance in GSH is observed in a wide range of pathologies, such as cancer, neurodegenerative diseases, cystic fibrosis (CF), several viral infections including HIV-1, as well as in aging. Several reports have provided evidence for the use of GSH and molecules able to replenish intracellular GSH levels in antiviral therapy. This non-conventional role of GSH and its analogs as antiviral drugs is discussed in this chapter. [Copyright &y& Elsevier]

The results of the combined use of Fludarabine, an anticancer agent that may be able to target latently infected cells, and conventional antiretroviral therapy (AZT+DDI) in a murine model of AIDS, i.e., LP-BM5 infection, are reported. Eighty percent of infected mice, treated with four cycles of alternate administration of Fludarabine and AZT+DDI, showed undetectable levels of proviral DNA in lymph nodes. After 8 weeks of treatment interruption, the infected/treated animals, although still alive at a time when all untreated animals had succumbed to the infection, showed disease progression and reappearance of proviral DNA in lymph nodes. The retrospective analysis of proviral DNA content in spleen and bone marrow at the end of the fourth cycle of treatment revealed a low but detectable amount of BM5d proviral DNA. We thus concluded that the spleen and bone marrow may be less sensitive to lympholitic drugs and therefore act as viral reservoirs in LP-BM5 infection. This study suggests that optimized protocols of alternate administration of cytolytic and antiretroviral drugs may represent a useful strategy to eradicate retroviral infections. [Copyright &y& Elsevier]

Host-directed therapy using drugs that target cellular pathways required for virus lifecycle or its clearance might represent an effective approach for treating infectious diseases. Changes in redox homeostasis, including intracellular glutathione (GSH) depletion, are one of the key events that favor virus replication and contribute to the pathogenesis of virus-induced disease. Redox homeostasis has an important role in maintaining an appropriate Th1/Th2 balance, which is necessary to mount an effective immune response against viral infection and to avoid excessive inflammatory responses. It is known that excessive production of reactive oxygen species (ROS) induced by viral infection activates nuclear factor (NF)-kB, which orchestrates the expression of viral and host genes involved in the viral replication and inflammatory response. Moreover, redox-regulated protein disulfide isomerase (PDI) chaperones have an essential role in catalyzing formation of disulfide bonds in viral proteins. This review aims at describing the role of GSH in modulating redox sensitive pathways, in particular that mediated by NF-kB, and PDI activity. The second part of the review discusses the effectiveness of GSH-boosting molecules as broad-spectrum antivirals acting in a multifaceted way that includes the modulation of immune and inflammatory responses. [ABSTRACT FROM AUTHOR]

Demonstrates that in murine AIDS, the adenosine triphosphate-dependent and ubiquitin-dependent proteolytic system is strongly affected at least in the lymph nodes of infected mice. Features of retrovirus-induced murine AIDS including lymphoproliferation, chronic B-cell activation resulting in hypergammaglobulinemia and profound immunodeficiency; Coincidence of the accumulation of ubiquitin conjugates and induction of the conjugating system with an increase in the proteolytic activity.

Focuses on the inhibitory effect of a combined chemotherapy on the retrovirus-induced model of AIDS in mice. Levels of the drugs AZT and LP-BM5 administered in experimental mice; Reduction of lymphadenopathy, splenomegaly and hypergammaglobulinemia; Increase in the stimulation index of T cells in mice receiving multi-drug treatments.

Excessive production of immunoglobulins (Ig) causes endoplasmic reticulum (ER) stress and triggers the unfolded protein response (UPR). Hypergammaglobulinemia and lymphadenopathy are hallmarks of murine AIDS that develops in mice infected with the LP-BM5 murine leukemia retrovirus complex. In these mice, Th2 polarization and aberrant humoral response have been previously correlated to altered intracellular redox homeostasis. Our goal was to understand the role of the cell's redox state in Ig secretion and plasma cell (PC) maturation. To this aim, LP-BM5-infected mice were treated with I-152, an N -acetyl-cysteine and cysteamine supplier. Intraperitoneal I-152 administration (30 μmol/mouse three times a week for 9 weeks) decreased plasma IgG and increased IgG/Syndecan 1 ratio in the lymph nodes where IgG were in part accumulated within the ER. PC containing cytoplasmic inclusions filled with IgG were present in all animals, with fewer mature PC in those treated with I-152. Infection induced up-regulation of signaling molecules involved in the UPR, i.e. CHAC1, BiP, sXBP-1 and PDI, that were generally unaffected by I-152 treatment except for PDI and sXBP-1, which have a key role in protein folding and PC maturation, respectively. Our data suggest that one of the mechanisms through which I-152 can limit hypergammaglobulinemia in LP-BM5-infected mice is by influencing IgG folding/assembly as well as secretion and affecting PC maturation. • I-152, a pro-glutathione molecule, inhibits IgG production in an in vivo model. • I-152 affects the function and maturation of PC. • I-152 interferes with IgG folding/assembly. • I-152 mainly influences the UPR signaling branch IRE1α-XBP1. • The redox state has a role in the maturation of PC. [ABSTRACT FROM AUTHOR]

• C4-GSH and I-152 have bacteriostatic and bactericidal activity against M. avium in vitro. • I-152 shows immunomodulatory activity by inducing macrophage cytokines. • Redox homeostasis is an important target to design new antimycobacterial drugs. • Studies are required to establish the exact mechanism of action of C4-GSH and I-152. Despite early treatment with antimycobacterial combination therapy, drug resistance continues to emerge. Maintenance of redox homeostasis is essential for Mycobacterium avium (M. avium) survival and growth. The aim of the present study was to investigate the antimycobacterial activity of two pro-glutathione (pro-GSH) drugs that are able to induce redox stress in M. avium and to modulate cytokine production by macrophages. Hence, we investigated two molecules shown to possess antiviral and immunomodulatory properties: C4-GSH, an N -butanoyl GSH derivative; and I-152, a prodrug of N -acetyl-cysteine (NAC) and β-mercaptoethylamine (MEA). Both molecules showed activity against replicating M. avium , both in the cell-free model and inside macrophages. Moreover, they were even more effective in reducing the viability of bacteria that had been kept in water for 7 days, proving to be active both against replicating and non-replicating bacteria. By regulating the macrophage redox state, I-152 modulated cytokine production. In particular, higher levels of interferon-gamma (IFN-γ), interleukin 1 beta (IL-1β), IL-18 and IL-12, which are known to be crucial for the control of intracellular pathogens, were found after I-152 treatment. Our results show that C4-GSH and I-152, by inducing perturbation of redox equilibrium, exert bacteriostatic and bactericidal activity against M. avium. Moreover, I-152 can boost the host response by inducing the production of cytokines that serve as key regulators of the Th1 response. [ABSTRACT FROM AUTHOR]

Reduction in oxygen levels is a key feature in the physiology of the bone marrow (BM) niche where hematopoiesis occurs. The BM niche is a highly vascularized tissue and endothelial cells (ECs) support and regulate blood cell formation from hematopoietic stem cells (HSCs). While in vivo studies are limited, ECs when cultured in vitro at low O2 (<5%), fail to support functional HSC maintenance due to oxidative environment. Therefore, changes in EC redox status induced by antioxidant molecules may lead to alterations in the cellular response to hypoxia likely favoring HSC self‐renewal. To evaluate the impact of redox regulation, HUVEC, exposed for 1, 6, and 24 h to 3% O2 were treated with N‐(N‐acetyl‐l‐cysteinyl)‐S‐acetylcysteamine (I‐152). Metabolomic analyses revealed that I‐152 increased glutathione levels and influenced the metabolic profiles interconnected with the glutathione system and the redox couples NAD(P)+/NAD(P)H. mRNA analysis showed a lowered gene expression of HIF‐1α and VEGF following I‐152 treatment whereas TRX1 and 2 were stimulated. Accordingly, the proteomic study revealed the redox‐dependent upregulation of thioredoxin and peroxiredoxins that, together with the glutathione system, are the main regulators of intracellular ROS. Indeed, a time‐dependent ROS production under hypoxia and a quenching effect of the molecule were evidenced. At the secretome level, the molecule downregulated IL‐6, MCP‐1, and PDGF‐bb. These results suggest that redox modulation by I‐152 reduces oxidative stress and ROS level in hypoxic ECs and may be a strategy to fine‐tune the environment of an in vitro BM niche able to support functional HSC maintenance. [ABSTRACT FROM AUTHOR]

Sinusoidal endothelial cells are the predominant vascular surface of the bone marrow and constitute the functional hematopoietic niche where hematopoietic stem and progenitor cells receive cues for self-renewal, survival, and differentiation. In the bone marrow hematopoietic niche, the oxygen tension is usually very low, and this condition affects stem and progenitor cell proliferation and differentiation and other important functions of this region. Here, we have investigated in vitro the response of endothelial cells to a marked decrease in O2 partial pressure to understand how the basal gene expression of some relevant biological factors (i.e., chemokines and interleukins) that are fundamental for the intercellular communication could change in anoxic conditions. Interestingly, mRNA levels of CXCL3, CXCL5, and IL-34 genes are upregulated after anoxia exposure but become downmodulated by sirtuin 6 (SIRT6) overexpression. Indeed, the expression levels of some other genes (such as Leukemia Inhibitory Factor (LIF)) that were not significantly affected by 8 h anoxia exposure become upregulated in the presence of SIRT6. Therefore, SIRT6 mediates also the endothelial cellular response through the modulation of selected genes in an extreme hypoxic condition. [ABSTRACT FROM AUTHOR]

Drug delivery is a growing field of interdisciplinary activities that combine the use of new materials with the biochemical properties of selected drugs, with the aim of improving their therapeutic action and reducing their toxicity. In few cases, proper medical devices have been also realized to implement new drug delivery modalities. In this article, we have summarized available information and our experience on the use of autologous Red Blood Cells as carriers for drugs to be released within the vascular system. This is not a comprehensive review, but it focusses on the mechanisms that are available to distribute drugs in circulation by carrier red blood cells and provide illustrative examples on how this is currently obtained. We have not included a summary of clinical data collected in recent years using this technology but simply provided proper references for the interested readers. Finally, a special attention is devoted to the possibility of entrapping, into autologous red blood cells, recombinant drug-binding proteins. This new strategy is opening the way at a new modality to influence the vascular distribution of drugs by realizing a dynamic circulating container (the engineered red cell) capable of reversible binding and transportation of one or more drugs of interest selected on the bases of the red cell entrapped target proteins. This new modality is not yet fully developed and explored but will certainly provide a technical solution to the problem of stabilizing drug concentration in circulation improving drug efficacy and reducing drug toxicity. © 2011 IUBMB IUBMB Life, 2011 [ABSTRACT FROM AUTHOR]

Abstract: Peptide nucleic acids (PNAs) provide a powerful tool to study the mechanism of transcription and translation, an innovative strategy to regulate target gene expression. They have been successfully used in antisense technology, for their ability to specifically bind to messenger RNA (mRNA) targets and to inhibit translation of the target genes. However, unlike most of the DNA and RNA oligonucleotides, PNAs are poorly penetrated through the cell membrane, partially due to their uncharged property. To enhance the efficiency in PNA delivery, many strategies have been explored. We here compare the efficacy of three different delivery strategies for antisense PNA: 1) conjugation to hydrophobic peptides, 2) adsorption onto polymeric microspheres and 3) encapsulation in autologous erythrocytes. To this purpose, we designed and prepared PNA sequences able to inhibit the expression of macrophage enzymes involved in inflammatory process, i.e. nitric oxide synthase (iNOS) and cyclooxygenase 2 (COX-2) and tested their antisense activity in a murine macrophage cellular model. Both delivery through polymeric microspheres and encapsulation into erythrocytes allowed the antisense activity of unmodified PNAs at nanomolar concentration. [Copyright &y& Elsevier]

Oxidized LDLs (oxLDLs) and oxysterols play a key role in endothelial dysfunction and the development of atherosclerosis. The loss of vascular endothelium function negatively impacts vasomotion, cell growth, adhesiveness and barrier functions. While for some of these disturbances, a reasonable explanation can be provided from a mechanistic standpoint, for many others, the molecular mediators that are involved are unknown. Caveolae, specific plasma membrane domains, have recently emerged as targets and mediators of oxLDL-induced endothelial dysfunction. Caveolae and their associated protein caveolin-1 (Cav-1) are involved in oxLDLs/LDLs transcytosis, mainly through the scavenger receptor class B type 1 (SR-B1 or SCARB1). In contrast, oxLDLs endocytosis is mediated by the lectin-like oxidized LDL receptor 1 (LOX-1), whose activity depends on an intact caveolae system. In addition, LOX-1 regulates the expression of Cav-1 and vice versa. On the other hand, oxLDLs may affect cholesterol plasma membrane content/distribution thus influencing caveolae architecture, Cav-1 localization and the associated signalling. Overall, the evidence indicate that caveolae have both active and passive roles in oxLDL-induced endothelial cell dysfunction. First, as mediators of lipid uptake and transfer in the subendothelial space and, later, as targets of changes in composition/dynamics of plasma membrane lipids resulting from increased levels of circulating oxLDLs. Gaining a better understanding of how oxLDLs interact with endothelial cells and modulate caveolae-mediated signalling pathways, leading to endothelial dysfunction, is crucial to find new targets for intervention to tackle atherosclerosis and the related clinical entities. LINKED ARTICLES: This article is part of a themed issue on Oxysterols, Lifelong Health and Therapeutics. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v178.16/issuetoc. [ABSTRACT FROM AUTHOR]

The causative agent of murine AIDS (MAIDS) in C57BL/6 mice, is a defective murine leukemia virus (BM5d) that requires the replication-competent helper virus (BM5e). Since this animal model of immunodeficiency, which shows many similarities to human AIDS, is also used to test the efficacy and toxicity of antiretroviral drugs, a method that allows the quantitative detection of both viruses would be very useful also if hampered potentially by endogenous viral sequences usually present in mice. While BM5d alone could induce the disease, the effect of BM5e on the immune system of diseased mice is unclear. A specific and reliable one-step RT-PCR method was developed for the co-amplification, with the same efficiency, of BM5d or BM5e with ß-actin used as an internal standard. The standard curves produced with cloned cDNA sequences (ß-actin and BM5d or BM5e) assure that all samples are analyzed during the exponential phase of the reaction. Using this new assay which provided a dynamic range of at least four-log-unit, the ratio of initial absolute amounts of the virus and ß-actin RNA was determined, obtaining quantitative information on virus-specific cellular-transcript in the lymph nodes and spleen during the natural history of the disease and during therapeutic regimens. [Copyright &y& Elsevier]

Examines the influence of peptide nucleic acids (PNA) in the inhibition of anti-inducible nitric oxide synthase. Delivery of PNA into macrophages; Pleiotropic activities of nitric oxide; Mechanisms of PNA-DNA hydrogen-bonded double helix formation.

Thiol molecules have been recently re-considered as drug candidates in viral infections because of their ability to induce redox changes which interfere with virus life cycle and modulate the host immune response. Little is known about the molecular mechanisms of their immunomodulatory properties. Here we show that I-152, a thiol molecule metabolized to release N -acetyl- l -cysteine and cysteamine and acting as a pro-glutathione agent, causes early up-regulation of immunoproteasome subunits in the lymph nodes of murine leukemia virus infected mice. This evidence suggests that the immunoproteasome may be modulated by thiol-based compounds with important implications in understanding redox-controlled immunoregulation. [ABSTRACT FROM AUTHOR]

Biostimulants improve yield, quality, and stress acclimation in crops. In this work, we tested the possibility of using phenolics-rich extracts from spelt (Triticum dicoccum L.) husks to attenuate the effects of salt stress (100–200 mM NaCl) in maize. Two methanolic extracts were prepared from the soluble-conjugated (SC), and the insoluble-bound (IB) phenolic acid fractions of the spelt husk, and their effects were investigated on several stress-associated biochemical parameters, such as proline, lipid peroxidation, H 2 O 2 , GSH levels, and ion content. Results show that SC and IB fractions of husk extracts behaved very differently, no doubt due to their greatly divergent chemical composition, as revealed by both GC-MS and HPLC analyses. The efficacy of treatments in mitigating salt stress was also dose- and timing-dependent. IB, even at the lower concentration tested, was able to recover the performance of stressed plants in terms of growth, photosynthetic pigments content, and levels of salt stress markers. Recovery of shoot growth to control levels and reduction of stress-induced proline accumulation occurred regardless of whether plants were pre-treated or post-treated with IB, whereas only pre-treatment with the higher dose of IB was effective in mitigating oxidative stress. Although in some cases SC and even methanol alone exerted some positive effects, they could also be deleterious whereas IB never was. Overall, results indicate that a polyphenol-containing extract obtained from spelt by-products can behave as biostimulant in maize plants and can mitigate their response to salt stress, by acting on different biochemical targets. Image 1 • Two polyphenol-enriched extracts were prepared from spelt by-products. • Chemical composition of the two extracts was assessed by HPLC-DAD and GC-MS analysis. • Maize plants subjected to salt stress were treated with polyphenol-enriched extracts. • The efficacy of spelt husk extracts as biostimulant on maize plants was evaluated. [ABSTRACT FROM AUTHOR]

During aging, glutathione (GSH) content declines and the immune system undergoes a deficiency in the induction of Th1 response. Reduced secretion of Th1 cytokines, which is associated with GSH depletion, could weaken the host defenses against viral infections. We first evaluated the concentration of GSH and cysteine in organs of old mice; then, the effect of the administration of the N‐butanoyl GSH derivative (GSH‐C4) on the response of aged mice infected with influenza A PR8/H1N1 virus was studied through the determination of GSH concentration in organs, lung viral titer, IgA and IgG1/IgG2a production, and Th1/Th2 cytokine profile. Old mice had lower GSH than young mice in organs. Also the gene expression of endoplasmic reticulum (ER) stress markers involved in GSH metabolism and folding of proteins, that is, Nrf2 and PDI, was reduced. Following infection, GSH content remained low and neither infection nor GSH‐C4 treatment affected Nrf2 expression. In contrast, PDI expression was upregulated during infection and appeared counterbalanced by GSH‐C4. Moreover, the treatment with GSH‐C4 increased GSH content in organs, reduced viral replication and induced a predominant Th1 response. In conclusion, GSH‐C4 treatment could be used in the elderly to contrast influenza virus infection by inducing immune response, in particular the Th1 profile. [ABSTRACT FROM AUTHOR]

Ataxia telangiectasia (A-T) is a rare incurable neurodegenerative disease caused by biallelic mutations in the gene for ataxia-telangiectasia mutated ( ATM). The lack of a functional ATM kinase leads to a pleiotropic phenotype, and oxidative stress is considered to have a crucial role in the complex physiopathology. Recently, steroids have been shown to reduce the neurological symptoms of the disease, although the molecular mechanism of this effect is largely unknown. In the present study, we have demonstrated that dexamethasone treatment of A-T lymphoblastoid cells increases the content of two of the most abundant antioxidants [glutathione ( GSH) and NADPH] by up to 30%. Dexamethasone promoted the nuclear accumulation of the transcription factor nuclear factor (erythroid-derived 2)-like 2 to drive expression of antioxidant pathways involved in GSH synthesis and NADPH production. The latter effect was via glucose 6-phosphate dehydrogenase activation, as confirmed by increased enzyme activity and enhancement of the pentose phosphate pathway rate. This evidence indicates that glucocorticoids are able to potentiate antioxidant defenses to counteract oxidative stress in ataxia telangiectasia, and also reveals an unexpected role for dexamethasone in redox homeostasis and cellular antioxidant activity. [ABSTRACT FROM AUTHOR]

Injection of the LP-BM5 murine leukemia virus into mice causes murine AIDS, a disease characterized by many dysfunctions of immunocompetent cells. To establish whether the disease is characterized by glutathione imbalance, reduced glutathione (GSH) and cysteine were quantified in different organs. A marked redox imbalance, consisting of GSH and/or cysteine depletion, was found in the lymphoid organs, such as the spleen and lymph nodes. Moreover, a significant decrease in cysteine and GSH levels in the pancreas and brain, respectively, was measured at 5 weeks postinfection. The Th2 immune response was predominant at all times investigated, as revealed by the expression of Th1/Th2 cytokines. Furthermore, investigation of the activation status of peritoneal macrophages showed that the expression of genetic markers of alternative activation, namely, Fizz1, Ym1, and Arginase1, was induced. Conversely, expression of inducible nitric oxide synthase, a marker of classical activation of macrophages, was detected only when Th1 cytokines were expressed at high levels. In vitro studies revealed that during the very early phases of infection, GSH depletion and the downregulation of interleukin-12 (IL-12) p40 mRNA were correlated with the dose of LP-BM5 used to infect the macrophages. Treatment of LP-BM5-infected mice with N-(N-acetyl-L-cysteinyl)-S-acetylcysteamine (I-152), an N-acetyl-cysteine supplier, restored GSH/cysteine levels in the organs, reduced the expression of alternatively activated macrophage markers, and increased the level of gamma interferon production, while it decreased the levels of Th2 cytokines, such as IL-4 and IL-5. Our findings thus establish a link between GSH deficiency and Th1/Th2 disequilibrium in LP-BM5 infection and indicate that I-152 can be used to restore the GSH level and a balanced Th1/Th2 response in infected mice. [ABSTRACT FROM AUTHOR]

Despite the great strides in healthcare during the last century, some challenges still remained unanswered. The development of multi-drug resistant bacteria, the alarming growth of fungal infections, the emerging/re-emerging of viral diseases are yet a worldwide threat. Since the discovery of natural antimicrobial peptides able to broadly hit several pathogens, peptide-based therapeutics have been under the lenses of the researchers. This review aims to focus on synthetic peptides and elucidate their multifaceted mechanisms of action as antiviral, antibacterial and antifungal agents. Antimicrobial peptides generally affect highly preserved structures, e.g., the phospholipid membrane via pore formation or other constitutive targets like peptidoglycans in Gram-negative and Gram-positive bacteria, and glucan in the fungal cell wall. Additionally, some peptides are particularly active on biofilm destabilizing the microbial communities. They can also act intracellularly, e.g., on protein biosynthesis or DNA replication. Their intracellular properties are extended upon viral infection since peptides can influence several steps along the virus life cycle starting from viral receptor-cell interaction to the budding. Besides their mode of action, improvements in manufacturing to increase their half-life and performances are also taken into consideration together with advantages and impairments in the clinical usage. Thus far, the progress of new synthetic peptide-based approaches is making them a promising tool to counteract emerging infections. [ABSTRACT FROM AUTHOR]

5,6,11,12,17,18,23,24-octahydrocyclododeca[1,2-b:4,5-b′:7,8-b″:10,11- b′″]tetrai ndole (CTet), an indole-3-carbinol (I3C) metabolite endowed with anticancer properties, is poorly soluble in the solvents most frequently used in biological tests. This study indicates that the use of γ-cyclodextrin (γ-CD) avoids this problem. Formulated with γ-CD CTet is a potent inhibitor of DNA synthesis in both estrogen receptor positive (MCF-7) and estrogen receptor negative (MDA-MB-231) human breast cell lines (IC50 = 1.20 ± 0.04 μM and 1.0 ± 0.1 μM, respectively). [ABSTRACT FROM AUTHOR]

The accumulation of chromium by germinating kiwifruit pollen appears to be significantly affected by Cr species, Cr concentration and calcium availability. Cr(III) accumulation always occurred in a linear manner while Cr(VI) uptake followed a logarithmic model. In the absence of exogenous calcium, Cr(III) accumulation was much higher than that of Cr(VI). It was observed that, as the Cr(III) concentration increased, there was a significant decrease in the endogenous calcium content of pollen, ultimately leading to complete calcium depletion after 90min of incubation at 150μM Cr(III). This loss of calcium could be responsible for the strong inhibition of tube emergence and growth following exposure of pollen to Cr(III). Indeed, when exogenous calcium was added to the kiwifruit pollen culture medium, significant growth recovery and reduced Cr(III) uptake occurred; the opposite was true in Cr(VI)-treatments. A significant rise in lipid peroxide production occurs in the presence of both Cr species; the effect was more pronounced following Cr(VI) exposure. Finally, glutathione pool dynamics appears to be differentially affected by chromium species and concentrations. In conclusion, results of the present study have provided important information regarding the different activity profiles of Cr(III) and Cr(VI) in relation to kiwifruit pollen performance, and have also demonstrated differences in some biochemical responses of pollen to metal stress. [Copyright &y& Elsevier]

Viruses use cell machinery to replicate their genome and produce viral proteins. For this reason, several intracellular factors, including the redox state, might directly or indirectly affect the progression and outcome of viral infection. In physiological conditions, the redox balance between oxidant and antioxidant species is maintained by enzymatic and non-enzymatic systems, and it finely regulates several cell functions. Different viruses break this equilibrium and induce an oxidative stress that in turn facilitates specific steps of the virus lifecycle and activates an inflammatory response. In this context, many studies highlighted the importance of redox-sensitive pathways as novel cell-based targets for therapies aimed at blocking both viral replication and virus-induced inflammation. In the review, we discuss the most recent findings in this field. In particular, we describe the effects of natural or synthetic redox-modulating molecules in inhibiting DNA or RNA virus replication as well as inflammatory pathways. The importance of the antioxidant transcription factor Nrf2 is also discussed. Most of the data reported here are on influenza virus infection. We believe that this approach could be usefully applied to fight other acute respiratory viral infections characterized by a strong inflammatory response, like COVID-19. [ABSTRACT FROM AUTHOR]