Inhibition of Dual/Mixed Tropic HIV-1 Isolates by CCR5-Inhibitors in Primary Lymphocytes and Macrophages

Background

Dual/mixed-tropic HIV-1 strains are predominant in a significant proportion of patients, though little information is available regarding their replication-capacity and susceptibility against CCR5-antagonists in-vitro. The aim of the study was to analyze the replication-capacity and susceptibility to maraviroc of HIV-1 clinical isolates with different tropism characteristics in primary monocyte-derived-macrophages (MDM), peripheral-blood-mononuclear-cells (PBMC), and CD4⁺T-lymphocytes.

Methods

Twenty-three HIV-1 isolates were phenotipically and genotipically characterized as R5, X4 or dual (discriminated as R5⁺/X4, R5/X4, R5/X4⁺). Phenotypic-tropism was evaluated by multiple-cycles-assay on U87MG-CD4⁺-CCR5⁺−/CXCR4⁺-expressing cells. Genotypic-tropism prediction was obtained using Geno2Pheno-algorithm (false-positive-rate [FPR] = 10%). Replication-capacity and susceptibility to maraviroc were investigated in human-primary MDM, PBMC and CD4⁺T-cells. AMD3100 was used as CXCR4-inhibitor. Infectivity of R5/Dual/X4-viruses in presence/absence of maraviroc was assessed also by total HIV-DNA, quantified by real-time polymerase-chain-reaction.

Results

Among 23 HIV-1 clinical isolates, phenotypic-tropism-assay distinguished 4, 17 and 2 viruses with R5-tropic, dual/mixed-, and X4-tropic characteristics, respectively. Overall, viruses defined as R5⁺/X4-tropic were found with the highest prevalence (10/23, 43.5%). The majority of isolates efficiently replicated in both PBMC and CD4⁺T-cells, regardless of their tropism, while MDM mainly sustained replication of R5- or R5⁺/X4-tropic isolates; strong correlation between viral-replication and genotypic-FPR-values was observed in MDM (rho = 0.710;p-value = 1.4e-4). In all primary cells, maraviroc inhibited viral-replication of isolates not only with pure R5- but also with dual/mixed tropism (mainly R5⁺/X4 and, to a lesser extent R5/X4 and R5/X4⁺). Finally, no main differences by comparing the total HIV-DNA with the p24-production in presence/absence of maraviroc were found.

Conclusions

Maraviroc is effective in-vitro against viruses with dual-characteristics in both MDM and lymphocytes, despite the potential X4-mediated escape. This suggests that the concept of HIV-entry through one of the two coreceptors “separately” may require revision, and that the use of CCR5-antagonists in patients with dual/mixed-tropic viruses may be a therapeutic-option that deserves further investigations in different clinical settings.     

Candida albicans Delays HIV-1 Replication in Macrophages

Macrophages are one of the most important HIV-1 target cells. Unlike CD4⁺ T cells, macrophages are resistant to the cytophatic effect of HIV-1. They are able to produce and harbor the virus for long periods acting as a viral reservoir. Candida albicans (CA) is a commensal fungus that colonizes the portals of HIV-1 entry, such as the vagina and the rectum, and becomes an aggressive pathogen in AIDS patients. In this study, we analyzed the ability of CA to modulate the course of HIV-1 infection in human monocyte-derived macrophages. We found that CA abrogated HIV-1 replication in macrophages when it was evaluated 7 days after virus inoculation. A similar inhibitory effect was observed in monocyte-derived dendritic cells. The analysis of the mechanisms responsible for the inhibition of HIV-1 production in macrophages revealed that CA efficiently sequesters HIV-1 particles avoiding its infectivity. Moreover, by acting on macrophages themselves, CA diminishes their permissibility to HIV-1 infection by reducing the expression of CD4, enhancing the production of the CCR5-interacting chemokines CCL3/MIP-1α, CCL4/MIP-1β, and CCL5/RANTES, and stimulating the production of interferon-α and the restriction factors APOBEC3G, APOBEC3F, and tetherin. Interestingly, abrogation of HIV-1 replication was overcome when the infection of macrophages was evaluated 2-3 weeks after virus inoculation. However, this reactivation of HIV-1 infection could be silenced by CA when added periodically to HIV-1-challenged macrophages. The induction of a silent HIV-1 infection in macrophages at the periphery, where cells are continuously confronted with CA, might help HIV-1 to evade the immune response and to promote resistance to antiretroviral therapy.     

Prognostic factors and predictors of outcome in patients with COVID-19 and related pneumonia: a retrospective cohort study

The aim of the present study was to simultaneously assess several potential predictors of outcome (co-morbidity, previous and in-hospital treatment, radiologic Brixia score) in patients with COVID-19.This retrospective cohort study included 258 consecutive patients with confirmed COVID-19 admitted to a medical ward at Montichiari Hospital, Brescia, Italy from February 28th to April 30rd, 2020. Patients had SARS-CoV-2 related pneumonia with respiratory failure, and were treated with hydroxychloroquine and lopinavir plus ritonavir. In some patients, additional treatment with tocilizumab, dexamethasone and enoxaparin was adopted. Outcomes (death or recovery) were assessed at the end of the discharge period or at the end of the follow-up (August 2020).During hospitalization, 59 patients died, while 6 died after discharge. The following variables were demonstrated to be associated with a worse prognosis: Radiologic Brixia score higher than 8, presence at baseline of hypertension, diabetes, chronic obstructive pulmonary disease, heart disease, cancer, previous treatment with ACE-inhibitors or anti-platelet drugs. Anticoagulant treatment during hospital admission with enoxaparin at a dose higher than 4000 U once daily was associated with a better prognosis.In conclusion, our study demonstrates that some co-morbidities and cardiovascular risk factors may affect prognosis. The radiologic Brixia score may be a useful tool to stratify the risk of death at baseline. Anticoagulant treatment with enoxaparin might be associated to a clinical benefit in terms of survival in patients with COVID-19.     

Evidence for Interrupted Biomagnification of Cadmium in Billfish Food Chain Based on Stable Carbon and Nitrogen Isotopes from Southwestern of Gulf of California

Biological Trace Element Research - We report cadmium (Cd) concentrations in muscle, liver, and blood of striped marlin (Kajikia audax) and blue marlin (Makaira nigricans), caught in the southwest...     

Phenotypic and genetic differentiation between diadromous and landlocked puyen Galaxias maculatus

This study reports the phenotypic and genetic differences between individuals of puyen Galaxias maculatus from two sites in the same river basin in Tierra del Fuego National Park, southern South America. Individuals from the two sampling sites presented morphometric and genetic differences. The morphometric differences indicated that individuals from Laguna Negra (LN) were short and more robust and had large eyes, whereas those from Arroyo Negro (AN) were thin and elongated and had small eyes. Genetic differences showed that AN individuals had a greater genetic structuration and an older demographic history than LN individuals. The results of this study affirmed that the individuals from the two sampling sites belong to different populations with a high degree of isolation. The demographic history could indicate that the individuals of G. maculatus which migrated to northern areas during the last glaciation settled in the Beagle Channel after its formation. The LN population could have originated after the retreat of the glaciers, migrating from AN.     

Thermoregulatory behavior and oxygen consumption of Octopus mimus paralarvae: The effect of age

Octopus mimus is an important cephalopod species in the coastal zone of Peru and Chile that is exposed to temperature variations from time to time due to El Niño/Southern Oscillation (ENSO) episodes when surface temperatures can reach 24 °C, 6 °C above typical temperatures in their habitat. The relationships between temperature and food availability are important factors that determine the recruitment of juveniles into the O. mimus population. The present study was to evaluate the relationship between thermoregulatory behavior and the age of paralarvae (summer population) to determine whether changes in this behavior occur during internal yolk consumption, making larvae more vulnerable to environmental temperature change. Oxygen consumption of paralarvae when 1–4 d old was determined to establish if respiration could be used to monitor the physiological changes that occur during yolk consumption. Horizontal thermal selection (17–30 °C), critical thermal maxima (CTMax), minima (CTMin), and oxygen consumption experiments were conducted with fasting paralarvae 1–4 d old at 20 °C. Preferred temperatures were dependent on the age of O. mimus paralarvae. One day old paralarvae selected a temperature 1.1 °C (23·4 °C) higher than 2 – 4 d old paralarvae (22·3 °C). The CTMax of paralarvae increased with age with values of 31·9±1.1 °C in 1-d-olds and 33·4±0.3 to 4-d-olds. CTMin also changed with age with low values in 2-d-old paralarvae (9.1±1·3 °C) and 11·9±0·9 °C in 4-d-old animals. The temperature tolerance range of paralarvae was age-dependent (TTD=difference between CTMax and CTMin) with higher values in 2 and 3 d old paralarvae (25–26 °C) as compared to 1 d old (23·1 °C) and 4 d old animals (22.7 °C). Oxygen consumption was not affected by the age of paralarvae, suggesting that mechanisms exist that compensate their metabloism until at least 4 d of age. The temperature tolerance range of a planktonic paralarvae of octopus species is presented for the first time. This range was dependent on the age of paralarvae, and so rendered the paralarvae more vunerable to a combination of high temperature and food deprivation during first days of life. Results in the present study provide evidence that O. mimus could be under ecological pressure if a climate change causes increased or decreased temperatures into their distribution range.     

Are Rod Outer Segment ATP-ase and ATP-Synthase Activity Expression of the Same Protein?

Vertebrate retinal rod outer segments (OS) consist of a stack of disks surrounded by the plasma membrane, where phototransduction takes place. Energetic metabolism in rod OS remains obscure. Literature described a so-called Mg²⁺-dependent ATPase activity, while our previous results demonstrated the presence of oxidative phosphorylation (OXPHOS) in OS, sustained by an ATP synthetic activity. Here we propose that the OS ATPase and ATP synthase are the expression of the same protein, i.e., of F₁F_o-ATP synthase. Imaging on bovine retinal sections showed that some OXPHOS proteins are expressed in the OS. Biochemical data on bovine purified rod OS, characterized for purity, show an ATP synthase activity, inhibited by classical F₁F_o-ATP synthase inhibitors. Moreover, OS possess a pH-dependent ATP hydrolysis, inhibited by pH values below 7, suggestive of the functioning of the inhibitor of F1 (IF1) protein. WB confirmed the presence of IF1 in OS, substantiating the expression of F₁F_o ATP synthase in OS. Data suggest that the OS F₁Fo ATP synthase is able to hydrolyze or synthesize ATP, depending on in vitro or in vivo conditions and that the role of IF1 would be pivotal in the prevention of the reversal of ATP synthase in OS, for example during hypoxia, granting photoreceptor survival.