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Background: CD4 T-cell counts are still widely used to assess treatment eligibility and follow-up of HIV-infected patients. The World Health Organization (WHO) prequalification of in vitro diagnostics requested a manufacturer independent laboratory evaluation of the analytical performance at the Institute of Tropical Medicine (ITM) Antwerp, Belgium, of the Muse Auto CD4/CD4% system (Millipore), a new small capillary-flow cytometer dedicated to count absolute CD4-T cells and percentages in venous blood samples from HIV-infected patients. Methods: Two hundred and fifty (250) patients were recruited from the HIV outpatient clinic at ITM. Accuracy and precision of CD4 T cell counting on fresh EDTA anticoagulated venous blood samples were assessed in the laboratory on a Muse Auto CD4/CD4% system. Extensive precision analyses were performed both on fresh blood and on normal and low stabilized whole blood controls. Accuracy ((bias) was assessed by comparing results from Muse CD4/CD4% to the reference (single-platform FACSCalibur). Clinical misclassification was measured at 500, 350, 200 and 100 cells/μL thresholds. Results: Intra-assay precision was < 5%, and inter-assay was < 9%. CD4 T cell counts measured on Muse Auto CD4/CD4% System and on the reference instrument resulted in regression slopes of 0.97 for absolute counts and 1.03 for CD4 T cell percentages and a correlation coefficient of 0.99 for both. The average absolute bias as compared to the reference was negligible (4 cells/μL or 0.5%). The absolute average bias on CD4 T cell percentages was < 1%. Clinical misclassification at different CD4 T cell thresholds was small resulting in sensitivities and specificities equal or >90% at all thresholds except at 100 cells/μL (sensitivity = 87%). All samples could be analyzed as there was no repetitive rejection errors recorded. Conclusions: The Muse Auto CD4/CD4% System performed very well on fresh venous blood samples and met all WHO acceptance criteria for analytical performance of CD4 technologies. [ABSTRACT FROM AUTHOR]

The classical form of α1-antitrypsin deficiency (ATD) is characterized by intracellular accumulation of the misfolded variant α1-antitrypsin Z (ATZ) and severe liver disease in some of the affected individuals. In this study, we investigated the possibility of discovering novel therapeutic agents that would reduce ATZ accumulation by interrogating a C. elegans model of ATD with high-content genome-wide RNAi screening and computational systems pharmacology strategies. The RNAi screening was utilized to identify genes that modify the intracellular accumulation of ATZ and a novel computational pipeline was developed to make high confidence predictions on repurposable drugs. This approach identified glibenclamide (GLB), a sulfonylurea drug that has been used broadly in clinical medicine as an oral hypoglycemic agent. Here we show that GLB promotes autophagic degradation of misfolded ATZ in mammalian cell line models of ATD. Furthermore, an analog of GLB reduces hepatic ATZ accumulation and hepatic fibrosis in a mouse model in vivo without affecting blood glucose or insulin levels. These results provide support for a drug discovery strategy using simple organisms as human disease models combined with genetic and computational screening methods. They also show that GLB and/or at least one of its analogs can be immediately tested to arrest the progression of human ATD liver disease. [ABSTRACT FROM AUTHOR]

Jellyfish are a prominent component of the plankton community. They frequently form conspicuous blooms which may interfere with different human enterprises. Among the aspects that remain understudied are jellyfish associations with microorganisms having potentially important implications for organic matter cycling. To the best of our knowledge, this study is the first to investigate the bacterial community associated with live moon jellyfish (Aurelia solida, Scyohozoa) in the Adriatic Sea. Using 16S rRNA clone libraries and culture-based methods, we have analyzed the bacterial community composition of different body parts: the exumbrella surface, oral arms, and gastric cavity, and investigated possible differences in medusa-associated bacterial community structure at the time of the jellyfish population peak, and during the senescent phase at the end of bloom. Microbiota associated with moon jellyfish was different from ambient seawater bacterial assemblage and varied between different body parts. Betaproteobacteria (Burkholderia, Cupriavidus and Achromobacter) dominated community in the gastral cavity of medusa, while Alphaproteobacteria (Phaeobacter, Ruegeria) and Gammaproteobacteria (Stenotrophomonas, Alteromonas, Pseudoalteromonas and Vibrio) prevailed on ‘outer’ body parts. Bacterial community structure changed during senescent phase, at the end of the jellyfish bloom, showing an increased abundance of Gammaproteobacteria, exclusively Vibrio. The results of cultured bacterial isolates showed the dominance of Gammaproeteobacteria, especially Vibrio and Pseudoalteromonas in all body parts. Our results suggest that jellyfish associated bacterial community might have an important role for the host, and that anthropogenic pollution in the Gulf of Trieste might affect their community structure. [ABSTRACT FROM AUTHOR]

Amyotrophic lateral sclerosis (ALS) is a lethal motor neuron disease with no cure. Currently there are only two ALS drugs approved by the FDA, both with a limited therapeutic effect. In the search for drug candidates for ALS, we studied the effect of known stem cell mobilizing agents (treatment) and antimetabolite 5-fluorouracil (5-FU) (anti-treatment) in SOD1G93A model of ALS. Surprisingly, we found that anti-cancer drug 5-FU increases lifespan, delays the disease onset and improves motor performance in ALS mice. Although we were not able to demonstrate the mechanistic basis of the beneficial 5-FU action in ALS mice, our findings suggest that 5-FU or similar drugs are possible drug candidates for the treatment of motor neuron diseases through drug repurposing. [ABSTRACT FROM AUTHOR]

Objective: Prg4, also known as Lubricin, acts as a joint/boundary lubricant. Prg4 has been used to prevent surgically induced osteoarthritis (OA) in mice. Surgically induced OA serves as a good model for post-traumatic OA but is not ideal for recapitulating age-related OA. Reduced expression of the TGF-β type II receptor (TGFβR2) is associated with age-related OA in clinical samples, so we previously characterized a mouse model that exhibits OA due to expression of a mutated dominant-negative form of TGFβR2 (DNIIR). Prg4 expression was significantly reduced in DNIIR mice. Furthermore, we showed that Prg4 was a transcriptional target of TGF-ß via activation of Smad3, the main signal transducing protein for TGF-ß. The objective of the present study was to determine whether maintenance of Prg4, a down-stream transcriptional target of TGF-ß, prevents OA associated with attenuated TGF-ß signaling in mice. Design: Wild-type, DNIIR, and bitransgenic mice that express both DNIIR and Prg4, were compared. Mice were assessed with a foot misplacement behavioral test, μCT, histology, and Western blot. Results: Compared to DNIIR mice, bitransgenic DNIIR+Prg4 mice missed 1.3 (0.4, 2.1) fewer steps while walking (mean difference (95% confidence interval)), exhibited a cartilage fibrillation score that was 1.8 (0.4, 3.1) points lower, exhibited cartilage that was 28.2 (0.5, 55.9) μm thicker, and exhibited an OARSI score that was 6.8 (-0.9, 14.5) points lower. However, maintenance of Prg4 expression did not restore levels of phosphorylated Smad3 in DNIIR mice, indicating Prg4 does not simply stimulate TGF-ß signaling. Conclusions: Our results indicate that maintenance of Prg4 expression prevents OA progression associated with reduced TGF-β signaling in mice. Since there was no evidence that Prg4 acts by stimulating the TGF-ß signaling cascade, we propose that Prg4, a transcriptional target of TGF-ß, attenuates OA progression through its joint lubrication function. [ABSTRACT FROM AUTHOR]

Antimicrobial peptides (AMP), part of the innate immune system, are well studied for their ability to kill pathogenic microorganisms. However, many also possess important immunomodulatory effects, and this area has potential for the development of novel therapies to supplement traditional methods such as the use of antibiotics. Here, we characterise the microbicidal and immunomodulatory potential of the proline-rich bovine AMP, Bactenecin 5 (Bac5). We demonstrate broad antimicrobial activity, including against some mycobacterial species, which are important pathogens of fish, cattle and humans. Bac5 is able to activate macrophage-like THP-1 cells and can synergistically trigger the upregulation of tnf-α when co-stimulated with M. marinum. Furthermore, Bac5 sensitises A549 epithelial cells to stimulation with TNF-α. For the first time, we characterise the activity of Bac5 in vivo, and show it to be a potent chemokine for macrophages in the zebrafish (Danio rerio) embryo model of infection. Bac5 also supports the early recruitment of neutrophils in the presence of M. marinum. In the absence of host adaptive immunity, exogenous injected Bac5 is able to slow, although not prevent, infection of zebrafish with M. marinum. [ABSTRACT FROM AUTHOR]

Seminal plasma is a key biological fluid that modulates sperm function in the reproduction process. However, its role in sperm biotechnologies is scarce in poultry. The aims of the present study were to study the amino acids profile and total proteins of seminal plasma in 12 Spanish chicken breeds and to investigate the role of seminal plasma on cryoresistance of rooster sperm. To investigate the role of seminal plasma on cryoresistance, diluted pooled semen samples were cryopreserved in the presence and absence of seminal plasma. Glutamic acid was the most abundant free amino acid in seminal plasma, followed by alanine, serine, valine, and glycine. There was an influence of breed (P<0.05) on the percentage of viable sperm after freezing-thawing of samples with seminal plasma. Cluster analysis revealed that White Prat, Black Castellana, Blue Andaluza, Quail Castellana, and Red-Barred Vasca returned the best freezing-thawing response (good freezers). There was a positive correlation between seminal plasma concentrations of valine, isoleucine lysine, leucine and post thaw viability. The evaluation of fertilization capacity of frozen-thawed semen from the breeds White Prat (‘good freezer’) and Black-Red Andaluza (‘bad freezer’) showed that good freezer had higher fertility (20/68, 29.4%) compared to bad freezer breed (14/76, 18.4%), even if the difference was not significant (P = 0.08). The TUNEL assay revealed that freezing/thawing procedures in presence of seminal plasma provoked higher DNA fragmentation in most of the breeds, with a positive correlation between seminal alanine, valine, isoleucine, methionine, leucine, tyrosine, phenylalanine concentrations and DNA integrity. DNA fragmentation was lower in absence of seminal plasma and the breed effect on sperm viability was highly reduced. It is concluded that specific seminal plasma amino acids were associated with post-thaw percentage of viable sperm and DNA integrity. The removal of seminal plasma decreases the variability of the results and DNA fragmentation damages. [ABSTRACT FROM AUTHOR]

Although the study of pathogen sensing by host defense systems continues to uncover a role for inflammasome components specific to particular pathogens, gaps remain in our knowledge. After internalization, Francisella escapes from the phagosome in mononuclear cells and is thought to be detected by intracellular pathogen-response-receptors pyrin and Aim2 in human and murine models, respectively. However, it remains controversial as to the role of pyrin in detecting Francisella. Our current work aims to study the contribution of inflammasome sensor, Pyrin in regulating microparticulate caspase-1/GSDM-D activation by Francisella. Our findings suggest that NLRP3 is central to the activation/release of active caspase-1/GSDM-D encapsulated in microparticles (MP) by Francisella. We also provide evidence that this regulation is independent of pyrin, implicated in sensing cytosolic Francisella in NLRP3-/- conditions where endogenous Pyrin is present. Absence of NLRP3 completely abrogated Francisella mediated MP caspase-1/GSDM-D activation and release both before and after internalization of the pathogen. However, deletion of pyrin not only enhanced both LPS and Francisella mediated MP active caspase-1/GSDM-D release, but pyrin overexpression resulted in a reduction of inflammasome activation and release; suggesting an inhibitory role of pyrin in LPS and Francisella mediated MP responses. This NLRP3 dependence and inhibitory effect of pyrin correlated with cytokine release as well. These observations also correlated with MPs ability to induce cell death; as LPS and Francisella-induced MPs from pyrin-deficient cells were more potent than wild-type monocytes whereas, NLRP3-/- MPs failed to induce cell death. Taken together, we report that NLPR3 not only mediates Francisella induced cytokine responses, but is also critical for cytokine-independent microparticle-induced inflammasome activation and endothelial cell injury independent of pyrin. [ABSTRACT FROM AUTHOR]

Repairing damaged articular cartilage is particularly challenging because of the limited ability of cartilage to perform self-repair. Intra-articular injections of N-acetylglucosamine (GlcNAc) comprise a method of repairing full-thickness articular cartilage defects in the rabbit knee joint model. To date, the effects of administration of GlcNAc and hyaluronic acid (HA) have been investigated only in the context of osteoarthritis treatment. Therefore, we evaluated the therapeutic effects of using cell-free porous poly lactic-co-glycolic acid (PLGA) graft implants and intra-articular injections of GlcNAc or HA in a rabbit model of osteochondral regeneration to investigate whether they have the potential for inducing osteochondral regeneration when used alone or simultaneously. Twenty-four rabbits were randomized into one of four groups: the scaffold-only group (PLGA), the scaffold with intra-articular injections of GlcNAc (PLGA+G) group, twice per week for four weeks; the scaffold with intra-articular injections of HA group (PLGA+HA) group, once per week for three weeks; and the scaffold with intra-articular injections of GlcNAc and HA (PLGA+G+HA) group, once per week for three weeks. Knees were evaluated at 4 and 12 weeks after surgery. At the end of testing, only the PLGA+G+HA group exhibited significant bone reconstruction, chondrocyte clustering, and good interactions with adjacent surfaces at 4 weeks. Additionally, the PLGA+G+HA group demonstrated essentially original hyaline cartilage structures that appeared to have sound chondrocyte orientation, considerable glycosaminoglycan levels, and reconstruction of the bone structure at 12 weeks. Moreover, the PLGA+G+HA group showed organized osteochondral integration and significantly higher bone volume per tissue volume and trabecular thickness. However, there were no significant differences between the PLGA+G and PLGA+HA groups except for gap formation on subchondral bone in the PLGA+G group. This study demonstrated that PLGA implantation combined with intra-articular injections of GlcNAc and HA allowed for cartilage and bone regeneration and significantly promoted osteochondral regeneration in rabbits without supplementation of exogenous growth factors. And the combination of this two supplements with PLGA scaffold could also prolong injection interval and better performance than either of them alone for the reconstruction of osteochondral tissue in the knee joints of rabbits. [ABSTRACT FROM AUTHOR]

Traditional pyramidotomy models have a high mortality rate from breathing difficulties and show early recovery from the induced motor deficits. This study establishes a novel pyramidotomy technique in Sprague Dawley rats that generates persistent motor deficits and has a reduced mortality rate. We used viral neural tracing to identify the course and relative distribution of forelimb and hindlimb motor fibers (n = 9). On basis of the neural tracing data, the medullary pyramid was targeted dorsally from the cerebellar cortex for photothrombotic infarct lesioning (n = 18). The photothrombotic technique selectively destroyed the corticospinal fibers in the medullary pyramid with relative preservation of neighboring grey-matter tissue. MicroPET imaging using 2-deoxy-2-[18F]-fluoro-D-glucose (FDG-microPET) showed a decrease in regional cerebral glucose metabolism (rCGM) in the bilateral pyramid and ipsilateral sensory cortex (p < 0.001, FDR q < 0.05). In addition, the trapezoid bodies and superior olivary nuclei showed a decrease in rCGM, compatible with damage caused during the introduction of the optical fiber. Connected structures such as the inferior colliculi and auditory cortices also showed decreases in rCGM in both hemispheres (p < 0.001, FDR q < 0.05). There was a significant and persistent decrease in motor and sensory function in the contralateral limb following pyramidotomy, as demonstrated by performance in the single pellet reaching task and the foot-fault test. There was no operative mortality or loss of respiratory function in this study. These results indicate that photothrombotic pyramidotomy with a dorsal transcortical approach is a safe and reliable technique for generating a pyramidotomy model with persistent motor deficits. [ABSTRACT FROM AUTHOR]

Background: The mortality from cryptococcal meningitis remains high, despite the availability of antiretroviral therapy (ART) and amphotericin-based fungal regimens. The role of neutrophils in cryptococcosis is controversial. Our objective was to examine the association between blood neutrophil counts and outcomes in terms of mortality, the incidence of bacterial infections (including Mycobacterium tuberculosis) and hospitalization among HIV-infected patients presenting with cryptococcal meningitis. Methods: We used data from participants from the Cryptococcal Optimal ART Timing (COAT) trial (2010–2012; Uganda and South Africa) and the Adjunctive Sertraline for Treatment of Cryptococcal Meningitis (ASTRO-CM) trial (2013–2017; Uganda). We estimated 30-day mortality risk with Cox proportional hazards models by baseline neutrophil counts (a) on a continuous scale and (b) with indicators for both relatively high (> 3,500 cells/mm3) and low (≤ 1,000 cells/mm3) counts. Follow-up neutrophil counts from the COAT trial were used to examine the time-dependent association of neutrophil counts with 12-month mortality and rehospitalization. Results: 801 participants had an absolute neutrophil value at meningitis diagnosis. The median baseline absolute neutrophil count was 2100 cells/mm3 (IQR, 1400 to 3300 cells/mm3). Baseline neutrophil count was positively associated with 30-day mortality (adjusted hazard ratio = 1.09, 95%CI, 1.04–1.13, per 1000 cells/mm3 increase; p<0.001). Baseline absolute neutrophil counts ≤ 1000 cells/mm3 did not have increased risk of 30-day mortality compared to those with baseline neutrophils of 1001–3500 cells/mm3; however, baseline >3500 cells/mm3 had significantly increased risk, with an adjusted hazard ratio of 1.85(95%CI, 1.40–2.44; p<0.001). Among the COAT participants with follow-up neutrophil data, there was a strong association between time-updated neutrophil count and 12-month mortality (adjusted hazard ratio = 1.16, 95% CI 1.09–1.24; p<0.001. Conclusion: Higher blood neutrophil counts in HIV-infected patients with cryptococcal meningitis were associated with mortality. Neutrophils role requires further investigation as to whether this may be a mediator directly contributing to mortality or merely a marker of underlying pathologies that increase mortality risk. [ABSTRACT FROM AUTHOR]

Candidatus Neoehrlichia mikurensis (Ca. N. mikurensis; family Anaplasmataceae) is an emerging tick-borne pathogen that causes a systemic inflammatory syndrome with thrombotic complications. We report here the first identification of Ca. N. mikurensis in organ samples from small mammals captured in southwest South Korea. Nested PCR of groEL and 16S rRNA genes was used to confirm the identity of the bacteria present, and successfully amplified fragments were sequenced. All captured animals were identified as striped field mice (Apodemus agrarius), approximately 28.6% (4/14) and 21.4% (3/14) of which were found to be PCR-positive for Ca. N. mikurensis and Anaplasma phagocytophilum, respectively. The detection of Ca. N. mikurensis in these animals represents the first evidence of this pathogen in South Korea. Carriage of this bacterium by rodents highlights the need for more detailed investigation of their role in its transmission to humans. [ABSTRACT FROM AUTHOR]

The present study was aimed to develop a liposomal formulation of thymoquinone (Lip-TQ) to treat Candida albicans infection in diabetic mice. Streptozotocin (STZ) was injected to induce hyperglycemia and on day 3 post STZ administration, mice were intravenously infected with C. albicans. Various doses (2, 5 and 10 mg/kg) of Free or Lip-TQ were administered in C. albicans infected diabetic mice. The effect of Lip-TQ was also determined on the organ indices, liver and kidney function parameters. Lip-TQ at a dose of 10 mg/kg significantly reduced the level of the blood glucose and alleviated the systemic C. albicans infection in diabetic mice. C. albicans infected diabetic mice treated with Lip-TQ at a dose of 10 mg/kg showed the survival rate of 70% as compared to that of 20% in the group treated with free TQ. The treatment with Lip-TQ resulted in the recovery of the organ indices, liver inflammation, kidney functioning and pancreas regeneration in diabetic mice. Moreover, TQ formulations also showed the direct therapeutic effect against candidiasis in the untreated or metformin-treated diabetic mice. Therefore, the findings of the present study support the use of Lip-TQ in the treatment of candidiasis in the diabetic patients. [ABSTRACT FROM AUTHOR]

Nociceptive sensitization involves an increase in responsiveness of pain sensing neurons to sensory stimuli, typically through the lowering of their nociceptive threshold. Nociceptive sensitization is common following tissue damage, inflammation, and disease and serves to protect the affected area while it heals. Organisms can become sensitized to a range of noxious and innocuous stimuli, including thermal stimuli. The basic mechanisms underlying sensitization to warm or painfully hot stimuli have begun to be elucidated, however, sensitization to cold is not well understood. Here, we develop a Drosophila assay to study cold sensitization after UV-induced epidermal damage in larvae. Larvae respond to acute cold stimuli with a set of unique behaviors that include a contraction of the head and tail (CT) or a raising of the head and tail into a U-Shape (US). Under baseline, non-injured conditions larvae primarily produce a CT response to an acute cold (10°C) stimulus, however, we show that cold-evoked responses shift following tissue damage: CT responses decrease, US responses increase and some larvae exhibit a lateral body roll (BR) that is typically only observed in response to high temperature and noxious mechanical stimuli. At the cellular level, class III neurons are required for the decrease in CT, chordotonal neurons are required for the increase in US, and chordotonal and class IV neurons are required for the appearance of BR responses after UV. At the molecular level, we found that the transient receptor potential (TRP) channel brivido-1 (brv1) is required for these behavioral shifts. Our Drosophila model will allow us to precisely identify the genes and circuits involved in cold nociceptive sensitization. [ABSTRACT FROM AUTHOR]

Histological evidence points to the presence of dopamine (DA) in the cephalic sensory organs of multiple gastropod molluscs, suggesting a possible sensory role for the neurotransmitter. We investigated the sensory function of DA in the nudipleuran Pleurobranchaea californica, in which the central neural correlates of sensation and foraging behavior have been well characterized. Tyrosine hydroxylase-like immunoreactivity (THli), a signature of the dopamine synthetic pathway, was similar to that found in two other opisthobranchs and two pulmonates previously studied: 1) relatively few (<100) THli neuronal somata were observed in the central ganglia, with those observed found in locations similar to those documented in the other snails but varying in number, and 2) the vast majority of THli somata were located in the peripheral nervous system, were associated with ciliated, putative primary sensory cells, and were highly concentrated in chemotactile sensory organs, giving rise to afferent axons projecting to the central nervous system. We extended these findings by observing that applying a selective D2/D3 receptor antagonist to the chemo- and mechanosensory oral veil-tentacle complex of behaving animals significantly delayed feeding behavior in response to an appetitive stimulus. A D1 blocker had no effect. Recordings of the two major cephalic sensory nerves, the tentacle and large oral veil nerves, in a deganglionated head preparation revealed a decrease of stimulus-evoked activity in the former nerve following application of the same D2/D3 antagonist. Broadly, our results implicate DA in sensation and engender speculation regarding the foraging-based decisions the neurotransmitter may serve in the nervous system of Pleurobranchaea and, by extension, other gastropods. [ABSTRACT FROM AUTHOR]

Cognitive decline is a common occurrence of the natural aging process in animals and studying age-related changes in gene expression in the brain might shed light on disrupted molecular pathways that play a role in this decline. The fruit fly is a useful neurobiological model for studying aging due to its short generational time and relatively small brain size. We investigated age-dependent changes in the Drosophila melanogaster whole-brain transcriptome by comparing 5-, 20-, 30- and 40-day-old flies of both sexes. We used RNA-Sequencing of dissected brain samples followed by differential expression, temporal clustering, co-expression network and gene ontology enrichment analyses. We found an overall decline in expression of genes from the mitochondrial oxidative phosphorylation pathway that occurred as part of aging. We also detected, in females, a pattern of continuously declining expression for many neuronal function genes, which was unexpectedly reversed later in life. This group of genes was highly enriched in memory-impairing genes previously identified through an RNAi screen. We also identified deficits in short-term olfactory memory performance in older flies of both sexes, some of which matched the timing of certain changes in the brain transcriptome. Our study provides the first transcriptome profile of aging brains from fruit flies of both sexes, and it will serve as an important resource for those who study aging and cognitive decline in this model. [ABSTRACT FROM AUTHOR]

Broadly neutralizing antibodies (bNAbs) are rarely elicited by current human immunodeficiency virus type 1 (HIV-1) vaccine designs, but the presence of bNAbs in naturally infected individuals may be associated with high plasma viral loads, suggesting that the magnitude, duration, and diversity of viral exposure may contribute to the development of bNAbs. Here, we report the isolation and characterization of a panel of human monoclonal antibodies (mAbs) from two subjects who developed broadly neutralizing autologous antibody responses during HIV-1 infection. In both subjects, we identified collections of mAbs that exhibited specificity only to a few autologous envelopes (Envs), with some mAbs exhibiting specificity only to a subset of Envs within the quasispecies of a particular sample at one time point. Neutralizing antibodies (NAbs) isolated from these subjects mapped mostly to epitopes in the Env V3 loop region and the CD4 binding site. None of the individual neutralizing mAbs recovered exhibited the cumulative breadth of neutralization present in the serum of the subjects. Surprisingly, however, the activity of polyclonal mixtures comprising individual mAbs that each possessed limited neutralizing activity, could achieve increased breadth of neutralizing activity against autologous isolates. While a single broadly neutralizing antibody targeting one epitope can mediate neutralization breadth, the findings presented here suggest that a cooperative polyclonal process mediated by diverse antibodies with more limited breadth targeting multiple epitopes also can achieve neutralization breadth against HIV-1. [ABSTRACT FROM AUTHOR]

A study has been made of the effect of chronic exercise on myocardial electrophysiological heterogeneity and stability, as well as of the role of cholinergic neurons in these changes. Determinations in hearts from untrained and trained rabbits on a treadmill were performed. The hearts were isolated and perfused. A pacing electrode and a recording multielectrode were located in the left ventricle. The parameters determined during induced VF, before and after atropine (1μM), were: fibrillatory cycle length (VV), ventricular functional refractory period (FRPVF), normalized energy (NE) of the fibrillatory signal and its coefficient of variation (CV), and electrical ventricular activation complexity, as an approach to myocardial heterogeneity and stability. The VV interval was longer in the trained group than in the control group both prior to atropine (78±10 vs. 68±10 ms) and after atropine (76±8 vs. 67±10 ms). Likewise, FRPVF was longer in the trained group than in the control group both prior to and after atropine (53±8 vs. 42±7 ms and 50±6 vs. 40±6 ms, respectively), and atropine did not modify FRPVF. The CV of FRPVF was lower in the trained group than in the control group prior to atropine (12.5±1.5% vs. 15.1±3.8%) and, decreased after atropine (15.1±3.8% vs. 12.2±2.4%) in the control group. The trained group showed higher NE values before (0.40±0.04 vs. 0.36±0.05) and after atropine (0.37±0.04 vs. 0.34±0.06; p = 0.08). Training decreased the CV of NE both before (23.3±2% vs. 25.2±4%; p = 0.08) and after parasympathetic blockade (22.6±1% vs. 26.1±5%). Cholinergic blockade did not modify these parameters within the control and trained groups. Activation complexity was lower in the trained than in the control animals before atropine (34±8 vs. 41±5), and increased after atropine in the control group (41±5 vs. 48±9, respectively). Thus, training decreases the intrinsic heterogeneity of the myocardium, increases electrophysiological stability, and prevents some modifications due to muscarinic block. [ABSTRACT FROM AUTHOR]

Rationale: HSP90B1, also known as gp96, is a chaperone for multiple Toll-like receptors (TLRs) and is necessary for TLR-mediated inflammatory responses in murine myeloid cells. The molecule is also expressed in T-cells though its specific role is unknown. We hypothesized that human HSP90B1 regulates monocyte and T-cell responses to Mycobacterium tuberculosis (Mtb) and bacilli Calmette-Guerin (BCG) and that its variants are associated with susceptibility to TB disease. Methods: We screened 17 haplotype-tagging SNPs in the HSP90B1 gene region for association with BCG-induced T-cell cytokine responses using both an ex-vivo whole blood assay (N = 295) and an intracellular cytokine staining assay (N = 180) on samples collected 10 weeks after birth. Using a case-control study design, we evaluated the same SNPs for association with TB disease in a South African pediatric cohort (N = 217 cases, 604 controls). A subset of these SNPs was evaluated for association with HSP90B1 expression in human monocytes, monocyte-derived dendritic cells, and T-cells using RT-PCR. Lastly, we used CRISPR/Cas9 gene editing to knock down HSP90B1 expression in a human monocyte cell line (U937). Knockdown and control cell lines were tested for TLR surface expression and control of Mtb replication. Results: We identified three SNPs, rs10507172, rs10507173 and rs1920413, that were associated with BCG-induced IL-2 secretion (p = 0.017 for rs10507172 and p = 0.03 for rs10507173 and rs1920413, Mann-Whitney, dominant model). SNPs rs10507172 and rs10507173 were associated with TB disease in an unadjusted analysis (p = 0.036 and 0.025, respectively, dominant model) that strengthened with sensitivity analysis of the definite TB cases, which included only those patients with microbiologically confirmed Mtb (p = 0.007 and 0.012, respectively). Knockdowns of HSP90B1 in monocyte cell lines with CRISPR did not alter TLR2 surface expression nor influence Mtb replication relative to controls. Conclusion: Among infants, an HSP90B1 gene-region variant is associated with BCG-induced IL-2 production and may be associated with protection from TB disease. HSP90B1 knockdown in human monocyte-like cell lines did not influence TLR2 surface localization nor Mtb replication. Together, these data suggest that HSP90B1 regulates T-cell, but not monocyte, responses to mycobacteria in humans. [ABSTRACT FROM AUTHOR]

The guinea pig has been used as a model to study various human infectious diseases because of its similarity to humans regarding symptoms and immune response, but little is known about the humoral immune response. To better understand the mechanism underlying the generation of the antibody repertoire in guinea pigs, we performed deep sequencing of full-length immunoglobulin variable chains from naïve B and plasma cells. We gathered and analyzed nearly 16,000 full-length VH, Vκ and Vλ genes and analyzed V and J gene segment usage profiles and mutation statuses by annotating recently reported genome data of guinea pig immunoglobulin genes. We found that approximately 70% of heavy, 73% of kappa and 81% of lambda functional germline V gene segments are integrated into the actual V(D)J recombination events. We also found preferential use of a particular V gene segment and accumulated mutation in CDRs 1 and 2 in antigen-specific plasma cells. Our study represents the first attempt to characterize sequence diversity in the expressed guinea pig antibody repertoire and provides significant insight into antibody repertoire generation and Ig-based immunity of guinea pigs. [ABSTRACT FROM AUTHOR]

Cystic fibrosis (CF) is the most common life-shortening genetic disease and is caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene. Several current therapies aim at improving availability and/or function of the mutant CFTR proteins. The combination therapeutic lumacaftor/ivacaftor (Orkambi, luma/iva) partially corrects folding and potentiates CFTR function impaired by the F508del mutation. Despite the potential for clinical benefit, a substantial number of patients discontinue treatment due to intolerable adverse effects. The aim of the present study is to identify differences between individuals who continued treatment and those who discontinued due to adverse respiratory effects to potentially inform treatment decisions. Clinical data from the year prior to treatment initiation were analyzed from 82 patients homozygous for the F508del mutation treated at the Colorado Adult CF Program. Blood samples were collected from 30 of these subjects before initiation of treatment to examine expression of circulating leukocyte surface antigens and cytokines. Clinical and demographic characteristics were analyzed along with inflammatory markers to determine biomarkers of drug discontinuation. The use of oral prednisone and/or nasal budesonide in the year prior to luma/iva initiation was more prevalent in CF subjects who did not tolerate luma/iva (82% vs. 43%). Increased age, but not gender or initial lung function, was associated with higher probability of discontinuing treatment due to side effects overall. Worse lung function (lower ppFEV1, ppFEF25-75 ≤ 60%) was associated with higher incidence of discontinuing treatment due to pulmonary adverse effects. In a nested cohort of patients, increased surface levels of CXCR2 on CD14+CD16- monocytes were associated with discontinuation. Overall, the patients who tolerated luma/iva were distinguishable from those who did not tolerate the drug based on clinical and cellular markers obtained prior to treatment initiation. [ABSTRACT FROM AUTHOR]

Species are shifting their ranges due to climate change, many moving to cooler and higher locations. However, with elevation increase comes oxygen decline, potentially limiting a species’ ability to track its environment depending on what mechanisms it has available to compensate for hypoxic stress. Pikas (Family Ochotonidae), cold-specialist small mammal species, are already undergoing elevational range shifts. We collected RNA samples from one population of Ochotona roylei in the western Himalaya at three sites– 3,600, 4,000, and 5,000 meters–and found no evidence of significant population genetic structure nor positive selection among sites. However, out of over 10,000 expressed transcripts, 26 were significantly upregulated at the 5,000 m site and were significantly enriched for pathways consistent with physiological compensation for limited oxygen. These results suggest that differences in gene expression may play a key role in enabling hypoxia tolerance on this local scale, indicating elevational flexibility that may facilitate successful range shifts in response to climate change. [ABSTRACT FROM AUTHOR]

Indole-3-acetic acid is a ubiquitous small molecule found in all domains of life. It is the predominant and most active auxin in seed plants, where it coordinates a variety of complex growth and development processes. The potential origin of auxin signaling in algae remains a matter of some controversy. In order to clarify the evolutionary context of algal auxin signaling, we undertook a genomic survey to assess whether auxin acts as a signaling molecule in the emerging model chlorophyte Chlorella sorokiniana UTEX 1230. C. sorokiniana produces the auxin indole-3-acetic acid (IAA), which was present in both the cell pellet and in the supernatant at a concentration of ~ 1 nM, and its genome encodes orthologs of genes related to auxin synthesis, transport, and signaling in higher plants. Candidate orthologs for the canonical AUX/IAA signaling pathway were not found; however, auxin-binding protein 1 (ABP1), an alternate auxin receptor, is present and highly conserved at essential auxin binding and zinc coordinating residues. Additionally, candidate orthologs for PIN proteins, responsible for intercellular, vectorial auxin transport in higher plants, were not found, but PILs (PIN-Like) proteins, a recently discovered family that mediates intracellular auxin transport, were identified. The distribution of auxin related gene in this unicellular chlorophyte demonstrates that a core suite of auxin signaling components was present early in the evolution of plants. Understanding the simplified auxin signaling pathways in chlorophytes will aid in understanding phytohormone signaling and crosstalk in seed plants, and in understanding the diversification and integration of developmental signals during the evolution of multicellular plants. [ABSTRACT FROM AUTHOR]

Cornus kousa the Korean dogwood has been traditionally used in East Asia as therapeutic traditional medicine however biological activities of Cornus kousa have not been investigated previously. The aim of the present study was to evaluate anti-obesity activities coupled with anti-angiogenic activities of anthocyanins rich fraction of ethanolic leaf extract of Cornus kousa (ELECk) in HUVECs and 3T3- L1 cells. Dried plants leaves were extracted with 70% ethanol and anthocyanin fraction (AnT Fr) was obtained by eluting the ethanolic extract through non-polar macroporous resin and further purification by HPLC. Antiangiogenic activities were determined by antiproliferative effect of AnT Fr on HUVECs. In the presence of various concentrations of AnT Fr, 3T3-L1 preadipocytes were induced to differentiate. Lipid accumulation in differentiated adipocytes were quantified by Oil-Red O staining. AnT Fr significantly suppressed angiogenesis by inhibiting proliferation and tube formation of HUVECs via downregulating VEGRF 2, PI3K, β-catenin, NF-kB, and Akt1 in a dose dependent manner. AnT Fr inhibited lipid accumulation by down-regulating adipogenesis and lipogenesis promoting signaling proteins, PPARγ, CCAAT, C/EBPα, aP2, FAS, and LPL, however enhanced AMPK activation to p-AMPK in 3T3 cells quantified and expressed by western blotting. AnT Fr inhibit lipid accumulation by regulating adipogenesis and lipogenesis related genes and signaling proteins. The anti-obesity activities exerted by Cornus kousa are associated with antiangiogenic activities of anthocyanins rich fraction of Cornus kousa. Hence the presence of bioactive anthocyanins, Cornus kosa, is a good candidate for nutraceutical and pharmaceutical formulation for treating or controlling obesity. [ABSTRACT FROM AUTHOR]

Increased proinflammatory interleukin-6 (IL-6) levels are associated with acquired long QT-syndrome (LQTS) in patients with systemic inflammation, leading to higher risks for life-threatening polymorphic ventricular tachycardia such as Torsades de Pointes. However, the functional and molecular mechanisms of this association are not known. In most cases of acquired LQTS, the target ion channel is the human ether-á-go-go-related gene (hERG) encoding the rapid component of the delayed rectifier K current, IKr, which plays a critical role in cardiac repolarization. Here, we tested the hypothesis that IL-6 may cause QT prolongation by suppressing IKr. Electrophysiological and biochemical assays were used to assess the impact of IL-6 on the functional expression of IKr in HEK293 cells and adult guinea-pig ventricular myocytes (AGPVM). In HEK293 cells, IL-6 alone or in combination with the soluble IL-6 receptor (IL-6R), produced a significant depression of IKr peak and tail current densities. Block of IL-6R or Janus kinase (JAK) reversed the inhibitory effects of IL-6 on IKr. In AGPVM, IL-6 prolonged action potential duration (APD) which was further prolonged in the presence of IL-6R. Similar to heterologous cells, IL-6 reduced endogenous guinea pig ERG channel mRNA and protein expression. The data are first to demonstrate that IL-6 inhibition of IKr and the resulting prolongation of APD is mediated via IL-6R and JAK pathway activation and forms the basis for the observed clinical QT interval prolongation. These novel findings may guide the development of targeted anti-arrhythmic therapeutic interventions in patients with LQTS and inflammatory disorders. [ABSTRACT FROM AUTHOR]

Objectives: We hypothesized that systemic administration of high-mobility group box 1 fragment attenuates the progression of myocardial fibrosis and cardiac dysfunction in a hamster model of dilated cardiomyopathy by recruiting bone marrow mesenchymal stem cells thus causing enhancement of a self-regeneration system. Methods: Twenty-week-old J2N-k hamsters, which are δ-sarcoglycan-deficient, were treated with systemic injection of high-mobility group box 1 fragment (HMGB1, n = 15) or phosphate buffered saline (control, n = 11). Echocardiography for left ventricular function, cardiac histology, and molecular biology were analyzed. The life-prolonging effect was assessed separately using the HMGB1 and control groups, in addition to a monthly HMGB1 group which received monthly systemic injections of high-mobility group box 1 fragment, 3 times (HMGB1, n = 11, control, n = 9, monthly HMGB1, n = 9). Results: The HMGB1 group showed improved left ventricular ejection fraction, reduced myocardial fibrosis, and increased capillary density. The number of platelet-derived growth factor receptor-alpha and CD106 positive mesenchymal stem cells detected in the myocardium was significantly increased, and intra-myocardial expression of tumor necrosis factor α stimulating gene 6, hepatic growth factor, and vascular endothelial growth factor were significantly upregulated after high-mobility group box 1 fragment administration. Improved survival was observed in the monthly HMGB1 group compared with the control group. Conclusions: Systemic high-mobility group box 1 fragment administration attenuates the progression of left ventricular remodeling in a hamster model of dilated cardiomyopathy by enhanced homing of bone marrow mesenchymal stem cells into damaged myocardium, suggesting that high-mobility group box 1 fragment could be a new treatment for dilated cardiomyopathy. [ABSTRACT FROM AUTHOR]

Traumatic insemination (TI) is an extraordinary style of mating behavior wherein the female integument is pierced by the male extragenital structure to transfer the spermatozoa into the female’s body through wounding. Flower bugs of the genus Orius belong to the family Anthocoridae (Heteroptera), which is referred to as the “TI family”. Males possess sharp shaped extragenitalia, and females receive the extragenitalia using the copulatory tubes, which are specialized extragenital structures in Orius species. Since TI is not well studied in insects possessing the copulatory tube, we examined the genital structures and copulatory processes of three species, Orius strigicollis, O. sauteri, and O. minutus. Scanning electron microscopic observations revealed the positions of male extragenital structures during copulation. A needle-like flagellum was deeply inserted into the female intersegment between the abdominal VII and VIII segments, while the curved part of a sickle-like cone forced the intersegment to expand. No scars were detected around the copulation region after copulation. The copulatory tube adhered to the interior of segment VII, and the interior integument around the copulatory tube remained intact after copulation. On the basis of these results, TI does not occur in these Orius species. A pair of seminal conceptacles, which exists in typical TI insects, was found at the base of the oviducts in O. strigicollis. The distal end of the copulatory tube connected to a closed bag with a double-membrane, termed the sperm pouch. The sperm pouch was filled with filamentous structures after copulation and structures with equivalent forms were observed in adult male testis. These structures, considered to be spermatozoa, persisted in the pouch for at least two weeks after copulation, suggesting that the pouch is a long-term spermatozoa storage organ. [ABSTRACT FROM AUTHOR]

In a prospective study of twenty sexually transmitted infection (STI)-free women, we examined the impact of an intrauterine contraceptive device (IUCD) insertion on cervico-vaginal cytokine levels. Nine women chose the levonorgestrel-containing IUCD and eight chose a copper IUCD. A cervico-vaginal swab was collected for cytokine analysis pre-insertion and four weeks post-insertion. Significant increases were noted in levels of IL-1α (median 483.4 versus 316.6 pg/mL, p = 0.046), IL-1β (median 605.7 versus 147.3 pg/mL, p = 0.018), IL-6 (median 570.1 versus 157.3 pg/mL, p = 0.046), TNFα (median 1.19 versus 0.6 pg/mL, p = 0.029) and the chemokine MCP-1 (median 340.2 versus 135.2 pg/mL, p = 0.003). No significant changes were noted in the levels of GM-CSF, IL-8, MIG, MIP-3α, RANTES, IL-10, IL-17, IP-10, MIP-1β. Whether this increase in pro-inflammatory cytokine levels decreases epithelial barrier integrity and enhances susceptibility to STIs, including HIV, merits further study. [ABSTRACT FROM AUTHOR]

Feline sporotrichosis due to Sporothrix brasiliensis is frequently severe and often correlated to zoonotic transmission. Feline Immunodeficiency Virus (FIV) and Feline Leukemia Virus (FeLV) cause immunodeficiency in cats; no association has been identified with critical cases of sporotrichosis. Moreover, the cytokine profile in Sporothrix-infected cats and a potential impact of retrovirus co-infections on their immunity is unknown. This study assessed immunological parameters in cats with sporotrichosis with and without FIV or FeLV co-infection. FeLV infection was detected by antigen ELISA and by provirus PCR. FIV infection was investigated through ELISA and Western blot. Cytokine transcription (IFN-γ, IL-4, IL-5, IL-6, IL-10, IL-12, TNF-α) was quantified using RT-qPCR and lymphocyte subpopulations (CD4, CD8, CD5 and CD21) were assessed by flow cytometry. Thirty cats with sporotrichosis were recruited to the study, including three FIV-positive and five FeLV-positive (progressive infection) cats. One cat with regressive FeLV infection was excluded from statistics. In comparison to retrovirus-negative cats, FIV-positive cats and FeLV-positive cats had higher IL-10 levels, FeLV-positive cats had lower IL-4 levels and FIV-positive cats had lower IL-12 levels and a lower CD4+/CD8+ ratio. Remarkably, all cats with poor general condition were FeLV (progressive infection) or FIV-positive, but the retrovirus status was not associated with the sporotrichosis treatment length or outcome. The immunological changes and the more severe clinical presentation observed in cats with retrovirus co-infections encourage future prospective studies that address the impact of these changes on prognostic determinants of feline sporotrichosis and the development of new therapy strategies that control disease spread. [ABSTRACT FROM AUTHOR]

Most patients with chronic lower back pain (CLBP) exhibit degenerative disc disease. Disc specimens obtained during initial therapeutic discectomies are often infected/colonized with Propionibacterium acnes, a Gram-positive commensal of the human skin. Although pain associated with infection is typically ascribed to the body’s inflammatory response, the Gram-positive bacterium Staphylococcus aureus was recently observed to directly activate nociceptors by secreting pore-forming α-hemolysins that disrupt neuronal cell membranes. The hemolytic activity of P. acnes in cultured disc specimens obtained during routine therapeutic discectomies was assessed through incubation on sheep-blood agar. The β-hemolysis pattern displayed by P. acnes on sheep-blood agar was variable and phylogroup-dependent. Their molecular phylogroups were correlated with their hemolytic patterns. Our findings raise the possibility that pore-forming proteins contribute to the pathogenesis and/or symptomology of chronic P. acnes disc infections and CLBP, at least in a subset of cases. [ABSTRACT FROM AUTHOR]

The natural polysaccharides extracted from the pollen of Pinus massoniana (TPPPS) have been shown to be a promising immune adjuvant against several viral chicken diseases. However, the exact mechanism through which TPPPS enhances the host immune response in chicken remains poorly understood. In the current study, chicken peripheral blood lymphocytes were treated with varying concentrations of TPPPS and pro-inflammatory cytokines such as IFN-γ, iIL-2 and IL-6 were measured to determine the optimal dose of the polysaccharide. A comparative analysis was subsequently performed between the proteome of lymphocytes subjected to the best treatment conditions and that of untreated cells. Protein identification and quantitation revealed a panel of three up-regulated and seven down-regulated candidates in TPPPS-treated chicken peripheral blood lymphocytes. Further annotation and functional analysis suggested that a number of those protein candidates were involved in the regulation of host innate immune response, inflammation and other immune-related pathways. We believe that our results could serve as a stepping stone for further research on the immune-enhancing properties of TPPPS and other polysaccharide-based immune adjuvants. [ABSTRACT FROM AUTHOR]

The nucleus accumbens (NAc) contains multiple subpopulations of medium spiny neurons (MSNs). One subpopulation expresses D1-type dopamine receptors, another expresses D2-type receptors, and a third expresses both. The relative roles in NAc of D1 neurons versus D2 neurons in appetitive motivation were assessed here. Specifically, we asked whether D1-Cre mice would instrumentally seek optogenetic self-stimulation specifically targeted at D1 MSNs in NAc, and similarly if D2-Cre mice would self-stimulate D2 neurons in NAc. Mice were implanted with Cre-targeted channelrhodopsin (ChR2) virus and optic fibers in NAc. Subsequently, mice could earn brief NAc laser illuminations by actively touching a metal spout in one task, or by going to a particular location in a separate task. Results indicated that D1 neuronal excitation in NAc supported intense self-stimulation in both tasks. D1-Cre mice earned hundreds to thousands of spout-touches per half-hour session, and also sought out locations that delivered NAc laser to excite D1 MSNs. By comparison, D2 ChR2 mice showed lower but still positive levels of self-stimulation in the spout-touch task, earning dozens to hundreds of NAc laser illuminations. However, in the location task, D2 mice failed to show positive self-stimulation. If anything, a few D2 individuals gradually avoided the laser location. Brain-wide measures indicated that D1 and D2 stimulations in NAc recruited heavily overlapping patterns of Fos activation in distant limbic structures. These results confirm that excitation of D1 MSNs in NAc supports strong incentive motivation to self-stimulate. They also suggest that excitation of D2 neurons in NAc supports self-stimulation under some conditions, but fails under others and possibly may even shift to negative avoidance. [ABSTRACT FROM AUTHOR]

Zebrafish larvae (Danio rerio) are among the most used model species to test biological effects of different substances in biomedical research, neuroscience and ecotoxicology. Most tests are based on changes in swimming activity of zebrafish larvae by using commercially available high-throughput screening systems. These systems record and analyse behaviour patterns using visible (VIS) and near-infrared (NIR) light sources, to simulate day (VIS) and night (NIR) phases, which allow continuous recording of the behaviour using a NIR sensitive camera. So far, however, the sensitivity of zebrafish larvae to NIR has never been tested experimentally, although being a critical piece of information for interpreting their behaviour under experimental conditions. Here, we investigated the swimming activity of 96 hpf (hours post fertilization) and 120 hpf zebrafish larvae under light sources of NIR at 860 nm and at 960 nm wavelength and under VIS light. A thermal source was simultaneously presented opposite to one of the light sources as control. We found that zebrafish larvae of both larval stages showed a clear negative phototactic response towards 860 nm NIR light and to VIS light, but not to 960 nm NIR light. Our results demonstrated that zebrafish larvae are able to perceive NIR at 860 nm, which is almost identical to the most commonly used light source in commercial screening systems (NIR at 850 nm) to create a dark environment. These tests, however, are not performed in the dark from the zebrafish´s point of view. We recommend testing sensitivity of the used test organism before assuming no interaction with the applied light source of commonly used biosensor test systems. Previous studies on biological effects of substances to zebrafish larvae should be interpreted with caution. [ABSTRACT FROM AUTHOR]

Perinatal asphyxia is a severe medical condition resulting from oxygen deficiency (hypoxia) at the time of birth, causing worldwide approximately 680,000 newborn deaths every year. Better prediction of severity of damages including early biomarkers is highly demanded. Elevated levels of circulating cell-free DNA (cfDNA) in blood have been reported for a range of different diseases and conditions, including cancer and prematurity. The objective of this study was to validate methods for assessing cfDNA in blood and cerebrospinal fluid (CSF) and to explore temporal variations in a piglet model of neonatal hypoxia-reoxygenation. Different cfDNA extraction methods in combination with cfDNA detection systems were tested, including a fluorescent assay using SYBR Gold and a qRT-PCR-based technique. Newborn piglets (n = 55) were exposed to hypoxia-reoxygenation, hypoxia-reoxygenation and hypothermia, or were part of the sham-operated control group. Blood was sampled at baseline and at post-intervention, further at 30, 270, and 570 minutes after the end of hypoxia. Applying the fluorescent method, cfDNA concentration in piglets exposed to hypoxia (n = 32) increased from 36.8±27.6 ng/ml prior to hypoxia to a peak level of 61.5±54.9 ng/ml after the intervention and deceased to 32.3±19.1 ng/ml at 570 minutes of reoxygenation, whereas the group of sham-operated control animals (n = 11) revealed a balanced cfDNA profile. Animals exposed to hypoxia and additionally treated with hypothermia (n = 12) expressed a cfDNA concentration of 54.4±16.9 ng/ml at baseline, 39.2±26.9 ng/ml at the end of hypoxia, and of 41.1±34.2 ng/ml at 570 minutes post-intervention. Concentrations of cfDNA in the CSF of piglets exposed to hypoxia revealed at post-intervention higher levels in comparison to the controls. However, these observations were only tendencies and not significant. In a first methodological proof-of-principle study exploring cfDNA using a piglet model of hypoxia-reoxygenation variations in the temporal patterns suggest that cfDNA might be an early indicator for damages caused by perinatal asphyxia. [ABSTRACT FROM AUTHOR]

Stellaria dichotoma var. lanceolata (SdLv), a member of the Caryophyllaceae, is a traditional herbal medicine that has been used to treat fever, night sweats, and malaria in East Asia. Inflammation plays an essential role in both host defense and pathogenesis during infection by diverse intracellular pathogens. Herein, we showed that an herbal extract from SdLv effectively attenuated inflammatory responses from infection of Mycobacterium abscessus (Mab), but not Toxoplasma gondii (T. gondii). In primary murine macrophages, Mab infection resulted in the rapid activation of nuclear factor (NF)-κB and mitogen-activated protein kinase (MAPK), as well as in the generation of proinflammatory cytokines, such as tumor necrosis factor α and interleukin-6, which were all significantly inhibited by pretreatment with SdLv. However, herbal extracts from Bupleurum chinense DC. (Buch) or Bupleurum falcatum L. (Bufa) did not affect M. abs-induced activation of proinflammatory responses. Importantly, we demonstrated that generation of intracellular reactive oxygen species, which are important signaling intermediaries in the activation of NF-κB and the MAPK signaling pathway, was rapidly increased in Mab-infected macrophages, and this was effectively suppressed by pretreatment with SdLv, but not Buch and Bufa. We further found that the treatment of Buch and Bufa, but not SdLv, led to the activation of NF-κB and the MAPK signaling pathway and the generation of intracellular reactive oxygen species. Moreover, oral administration of SdLv significantly reduced lethality in Mab-infected mice. Collectively, these results suggest the possible use of SdLv as an effective treatment for Mab infection. [ABSTRACT FROM AUTHOR]

Background: Zika virus (ZIKV) is an emerging pathogen with no approved therapeutics and only limited diagnostics available. To address this gap, six mouse single-chain antibodies (scFvs) to ZIKV envelope (E) protein were isolated rapidly and efficiently from a ribosome-displayed antibody library constructed from the spleens of five immunized mice. Methodology/Results: In this report, we have generated a panel of mouse scFvs to ZIKV E protein using ribosome display. The six scFvs demonstrated no cross-reactivity with DENV2 NGC envelope protein, suggesting specificity for ZIKV E protein. These scFvs showed differences in their affinity: two (scFv45-3, scFv63-1) of them were dominant after four rounds of panning, and showed higher affinity (an apparent Kd values from 19 to 27 nM) than the other four (scFv5-1, scFv7-2, scFv38-1, and scFv51-2). All six scFvs showed ZIKV-neutralizing activity in the plaque reduction neutralization test (PRNT) assay and their neutralizing activity was positively correlated with their affinities. Conclusions/Significance: The scFvs (45–3 and 63–1) with highest affinity may have dual utility as diagnostics capable of recognizing ZIKV E subtypes and may be further developed to treat ZIKV infection. Our approach has the added advantage of generating Fc receptor-deficient antibodies, minimizing concern of antibody-dependent enhancement (ADE) of infection. [ABSTRACT FROM AUTHOR]

Prevalence of depression is higher in patients with cancer than in the general population. Sustained systemic inflammation has been associated with depressive behavior and it has been reported that depressed patients commonly display alterations in their immune system. We previously showed that cancer in mice induces a systemic environment that promotes neutrophil activation and leukocytosis. We thus hypothesized that the peripheral systemic response to a solid tumor leads to endothelial activation, which may promote inflammatory changes in the brain with behavioral consequences. Using the Lewis lung carcinoma (LLC) model, we show that tumor growth induces a progressive increase in peripheral inflammation as observed by elevated interleukin-6 (IL-6). In behavioral studies, tumor-bearing mice showed no sign of motor, coordination or short term working memory deficits as assessed by rotarod, balance-beam, and novel object recognition tests. However, there was an impairment in the grip strength test and interestingly, an anxious and despair-like phenotype in the elevated plus-maze, and tail suspension tests, respectively. Immunostaining of perfused brains revealed fibrin accumulation in the vasculature with some leakage into the parenchyma, a process known to activate endothelial cells. Taken together, our results suggest that the inflamed and prothrombotic systemic environment created by the growth of a peripherally-located solid tumor induces endothelial activation, accumulation of fibrin in the brain and astrocyte activation, perhaps leading to depressive-like behavior. [ABSTRACT FROM AUTHOR]

Endometriosis is a disorder in which endometrial tissue is found outside the uterus causing pain, infertility and stress. Finding effective, non-hormonal and long-term treatments for endometriosis still remains one of the most significant challenges in the field. Corticotropin releasing hormone (CRH) is one of the main signaling peptides within the hypothalamic pituitary adrenal (HPA) axis released in response to stress. CRH can affect nervous and visceral tissues such as the uterus and gut via activation of two types of CRH receptors: CRHR1 and CRHR2. Our aim was to determine if blocking CRHR1 with antalarmin will reduce endometriosis progression. In experiment 1 we induced endometriosis in female rats by suturing uterine horn tissue next to the intestinal mesentery and allowed to progress for 7 days. We determined that after 7 days, there was a significant increase in CRHR1 within endometriotic vesicles as compared to normal uterus. In Experiment 2, we induced endometriosis and administered either antalarmin (20 mg/kg, i.p.) or vehicle during the first 7 days after surgery. A separate group of sham surgery rats served as non-endometriosis controls. Endometriosis was allowed to progress until 60 days after surgery, at which time rats were tested for anxiety behaviors. At the time of sacrifice, endometriotic vesicles, uterus and blood were collected. Treatment with antalarmin significantly reduced the size (67% decrease) and number (30% decrease) of endometriotic vesicles. Antalarmin also prevented the increase in CRH and CRHR1 mRNA within endometriotic vesicles but not of glucocorticoid receptor. Endometriosis did not change anxiety behaviors in the open field and zero-maze tests and prior antalarmin administration did not modify this. Our data provides the first in-vivo demonstration for use of CRHR1 antagonist for the treatment of endometriosis opening the possibility for further exploring CRH signaling as a treatment target for this debilitating disease. [ABSTRACT FROM AUTHOR]

Platelet activation has been reported to play a major role in inflammatory response and thrombocytopenia during dengue viral infection. Cells expressing FcϒR2Aand DC-SIGN receptors are reported to be involved in dengue virulence. The present study is designed to assess the expression level of these two receptors on platelet surface collected from dengue patients and to study its association in patients with platelet RNA positive for dengue virus. This was an analytical cross-sectional study carried out in JIPMER hospital, Puducherry. Forty-four patients with dengue infection as cases and 44 patients with non dengue acute other febrile illness(OFI) as controls were recruited. Peripheral venous blood was withdrawn on day of admission, day 3 post admission and day of discharge and serological tests for NS1 dengue antigen and anti IgM antibody were analyzed for diagnosis of dengue infection. Platelet rich plasma was assessed for DC SIGN, FcϒR2A levels and platelets separated from dengue patients were subjected to RNA extraction and detection of presence of viral RNA. The study observed a decreased expression of DC-SIGN and FcϒR2A on platelets in dengue patients compared to OFI group on all the time points. Further, cells expressing DC-SIGN and FcϒR2A were found to be decreased on platelets in dengue patients who were positive for NS1 antigen. DC-SIGN and FcϒR2A expression was also found to be notably decreased in patients positive for platelet DENV RNA when compared with patients negative for platelet DENV RNA. Our results suggest that DC-SIGN and FcϒR2A, which are receptors for viral capture and immune mediated clearance respectively, might be down regulated on platelets in patients with dengue infection. The decreased receptor expression diminishes platelet activation and subsequently has protective action on the host from the ongoing conflict between immune system and dengue virus. [ABSTRACT FROM AUTHOR]

Leishmaniasis is amongst the most important neglected diseases, afflicting more than 12 million people in 88 countries. There is an urgent need for safe orally bioavailable and cost-effective drugs for the treatment of leishmaniasis. It has recently been shown that Leishmania activates host macrophage serine/threonine kinase Akt, to promote survival of both parasites and infected cells. Here, we sought to evaluate a compound, Miransertib (ARQ 092), an orally bioavailable and selective allosteric Akt inhibitor currently in clinical trials for patients with PI3K/Akt-driven tumors or Proteus syndrome. Miransertib was tested against Leishmania donovani and Leishmania amazonensis, causative agents of visceral and cutaneous leishmaniasis, respectively. Cultured promastigotes were susceptible to Miransertib. In addition, Miransertib was markedly effective against intracellular amastigotes of L. donovani or L. amazonensis-infected macrophages. Miransertib also enhanced mTOR dependent autophagy in Leishmania-infected macrophages, which may represent one mechanism of Miransertib-mediated killing of intracellular Leishmania. Whereas parasite clearance in the spleen of mice infected with L. donovani and treated with Miransertib was comparable to that when treated with miltefosine, Miransertib caused a greater reduction in the parasite load in the liver. In the cutaneous leishmaniasis infection model, lesions were reduced by 40% as compared to mock treated mice. Together, these results provide direct evidence to support the conclusion that Miransertib is an excellent lead compound for the development of a new oral drug therapy for visceral and cutaneous leishmaniasis. [ABSTRACT FROM AUTHOR]

Cellular immune responses were tested to determine the effect of fenbendazole on the function of lymphocytes from Bolivian squirrel monkeys (Samiri boliviensis boliviensis). Giardia-infected squirrel monkeys were treated with commercially available fenbendazole (FBZ)-medicated monkey chow. Immune responses were compared between historical controls (Giardia naïve, untreated with FBZ (control animals)) and Giardia-infected, FBZ-treated squirrel monkeys (study animals). Peripheral blood lymphocytes from study monkeys had significantly lower stimulation indices compared to control animals when cultured in vitro with concanavalin A (Con A) (p<0.0001), phytohaemagglutinin (PHA) (p<0.0001) and lipopolysaccharide (LPS) (p<0.0001). PBMCs were also analyzed for IFN-γ producing cells in response to stimulation with Con A, PHA, PWM, and LPS by the cytokine ELISPOT assay. Significantly higher responses to Con A- (p<0.0001), and PHA- (p<0.001) stimulated cultures from Giardia-infected and fenbendazole treated compared to controls. Flow cytometric analysis for expression of cell surface markers revealed a significant increase in B- and NKT-lymphocytes and significant decrease in CD14+CD16+ monocytes after FBZ treatment. Also, circulating plasma cytokines IFN-γ, TNF-α, IL-12p40, IL-1β, IL-10, IL-13, IL-1ra, IL-6 and IL-4 were significantly decreased after FBZ treatment. Comparison of hematologic parameters between controls and FBZ-treated squirrel monkeys revealed significantly lower numbers of total leukocytes, neutrophils, monocytes, and eosinophils compared to controls. However, erythrocyte indices (red cell count, hemoglobin and hematocrit were significantly higher in FBZ-treated monkeys. Our findings suggest that fenbendazole treatment may alter sensitive immune and molecular measures of inflammation. Postponing the experimental use of squirrel monkeys until at least 6 weeks after FBZ treatment should be considered. [ABSTRACT FROM AUTHOR]

The calcium carbonate skeletons of corals provide the underlying structure of coral reefs; however, the cellular mechanisms responsible for coral calcification remain poorly understood. In osteoblasts from vertebrate animals, a Na+/Ca2+ exchanger (NCX) present in the plasma membrane transports Ca2+ to the site of bone formation. The aims of this study were to establish whether NCX exists in corals and its localization within coral cells, which are essential first steps to investigate its potential involvement in calcification. Data mining identified genes encoding for NCX proteins in multiple coral species, a subset of which were more closely related to NCXs from vertebrates (NCXA). We cloned NCXA from Acropora yongei (AyNCXA), which, unexpectedly, contained a peptide signal that targets proteins to vesicles from the secretory pathway. AyNCXA subcellular localization was confirmed by heterologous expression of fluorescently tagged AyNCXA protein in sea urchin embryos, which localized together with known markers of intracellular vesicles. Finally, immunolabeling of coral tissues with specific antibodies revealed AyNCXA was present throughout coral tissue. AyNCXA was especially abundant in calcifying cells, where it exhibited a subcellular localization pattern consistent with intracellular vesicles. Altogether, our results demonstrate AyNCXA is present in vesicles in coral calcifying cells, where potential functions include intracellular Ca2+ homeostasis and Ca2+ transport to the growing skeleton as part of an intracellular calcification mechanism. [ABSTRACT FROM AUTHOR]

We herein examined the importance of H2-Eb1 and H2-Ab1 in the susceptibility of mice to allergic rhinitis (AR) by developing double-gene (H2-Eb1+H2-Ab1) knockout mice. The mice were randomly grouped into different sensitization and excitation treatments, then their behavioral scores; nasal mucosa HE staining; thymus tissue toluidine blue staining; levels of ovalbumin (OVA)-specific IgE, IL-2 and IL-13 in the serum; and expression of IL-2 and IL-13 in the nasal mucosa were observed. H2-Ab1 and H2-Eb1 were both successfully knocked out in the study group (KO-OVA). Compared with the control group (WT-OVA), the nasal mucosal tissue in the KO-OVA mice showed fewer histological changes, reduced numbers of eosinophilic granulocytes, fewer mast cells in the thymus tissue, reduced concentrations of OVA-specific IgE and IL-13 in the serum, and reduced expression of IL-13 in the nasal mucosa. The behavior of the mice was also improved. In addition, the IL-2 concentration in the serum and IL-2 expression in the nasal mucosa were increased. There were two important findings of this study: (1) The H2-Ab1 and H2-Eb1 double knockout model of allergic rhinitis was successfully constructed, and the Th1/Th2 cell factors were in imbalance in these mice compared to WT mice; (2) the AR susceptibility of the dual knockout mice was reduced, confirming that H2-Ab1 and H2-Eb1 contribute to allergic rhinitis, at least in mice. [ABSTRACT FROM AUTHOR]

Optogenetics has become an important tool for perturbing neural circuitry with unparalleled temporal precision and cell-type specificity. However, direct activation of a specific subpopulation of neurons can rapidly modulate the activity of other neurons within the network and may lead to unexpected and complex downstream effects. Here, we have developed a biologically-constrained computational model that exploits these non-intuitive network responses in order to gain insight into underlying properties of the network. We apply this model to data recorded during optogenetic stimulation in the primary visual cortex of the alert macaque. In these experiments, we found that optogenetic depolarization of excitatory neurons often suppressed neuronal responses, consistent with engagement of normalization circuitry. Our model suggests that the suppression seen in these responses may be mediated by slow excitatory and inhibitory conductance channels. Furthermore, the model predicted that the response of the network to optogenetic perturbation depends critically on the relationship between inherent temporal properties of the network and the temporal properties of the opsin. Consistent with model predictions, stimulation of the C1V1TT opsin, an opsin with a fast time constant (tau = 45 ms), caused faster and stronger suppressive effects after laser offset, as compared to stimulation of the slower C1V1T opsin (tau = 60ms). This work illustrates how the non-intuitive network responses that result from optogenetic stimulation can be exploited to gain insight regarding network properties that underlie fundamental neuronal computations, such as normalization. This novel hybrid opto-theoretical approach can thus enhance the power of optogenetics to dissect complex neural circuits. [ABSTRACT FROM AUTHOR]

Human Immunodeficiency Virus-1 (HIV-1) entry is dependent on the envelope glycoprotein (Env) that is present on the virion and facilitates fusion between the envelope and the cellular membrane. The protein consists of two subunits, gp120 and gp41, with the former required for binding the CD4 receptor and either the CXCR4 or CCR5 coreceptor, and the latter for mediating fusion. The requirement of fusion for infection has made Env an attractive target for HIV therapy development and led to the FDA approval of enfuvirtide, a fusion inhibitor. Continued development of entry inhibitors is warranted because enfuvirtide resistant HIV-1 strains have emerged. In this study, a novel HIV-1 fusion assay was validated using neutralizing antibodies and then used to investigate the mechanism of action of eCD4-Igmim2, an HIV-1 inhibitor proposed to cooperatively bind the CD4 binding site and the sulfotyrosine-binding pocket of gp120. Greater reduction in fusion levels was observed with eCD4-Igmim2 in the fusion assay than all of the gp120 antibodies evaluated. Lab adapted isolates, HIV-1HXB2 and HIV-1YU2, were sensitive to eCD4-Igmim2 in the fusion assay, while primary isolates, HIV-1BG505 and HIV-1ZM651 were resistant. These results correlated with greater IC50 values for primary isolates compared to the lab adapted isolates observed in a virus neutralization assay. Analysis of gp120 models identified differences in the V1 and V2 domains that are associated with eCD4-Igmim2 sensitivity. This study highlights the use of a fusion assay to identify key areas for improving the potency of eCD4-Igmim2. [ABSTRACT FROM AUTHOR]

Background: Human infants frequently acquire human cytomegalovirus (HCMV) through breastfeeding, resulting in persistent high-level viral shedding in saliva and urine and infectivity to others, including pregnant women. Thus, vaccination to interrupt postnatal HCMV transmission is an attractive strategy to prevent HCMV spread and congenital infection. Rhesus CMV (RhCMV) in nonhuman primates is a valuable model for the study of immune strategies to prevent CMV transmission. Although rhesus monkeys typically acquire RhCMV before 1 year of age, the timing and mode of natural infant RhCMV transmission remain unknown. Methods: We followed 5 RhCMV-seropositive dams and their infants from birth until weaning, approximately 6 months later. RhCMV DNA levels in plasma, breast milk, saliva, and urine were measured every 2 weeks by quantitative PCR. RhCMV-specific T cell responses in peripheral blood and breast milk were measured by interferon gamma ELISpot assays. Serum IgG antibody levels were measured by ELISA. Results: Four of five postpartum RhCMV-seropositive mothers had intermittent, low-level RhCMV shedding in breast milk, whereas all had high-magnitude RhCMV shedding in saliva and urine. The kinetics of maternal blood RhCMV-specific T cell responses and viral shedding in urine and saliva did not strongly associate, though dams with consistently high systemic RhCMV-specific T cell responses tended to have undetectable RhCMV shedding in breast milk. All RhCMV-exposed infants had intermittent, low-level RhCMV shedding in saliva during the lactation period, with minimal systemic RhCMV-specific T cell responses. Conclusions: Despite exposure to RhCMV shedding in breast milk and other maternal fluids, postnatal mother-to-child RhCMV transmission appears to be less efficient than that of HCMV. A greater understanding of the determinants of RhCMV transmission and its usefulness as a model of HCMV mucosal acquisition may provide insight into strategies to prevent HCMV infections in humans. [ABSTRACT FROM AUTHOR]

Human papillomaviruses (HPVs) are a group of circular double-stranded DNA viruses, showing severe tropism to mucosal tissues. A subset of HPVs, especially HPV16 and 18, are the primary etiological cause for several epithelial cell malignancies, causing about 5.2% of all cancers worldwide. Due to the high prevalence and mortality, HPV-associated cancers have remained as a significant health problem in human society, making an urgent need to develop an effective therapeutic vaccine against them. Achieving this goal is primarily dependent on the identification of efficient tumor-associated epitopes, inducing a robust cell-mediated immune response. Previous information has shown that E5, E6, and E7 early proteins are responsible for the induction and maintenance of HPV-associated cancers. Therefore, the prediction of major histocompatibility complex (MHC) class I T cell epitopes of HPV16, 18, 31 and 45 oncoproteins was targeted in this study. For this purpose, a two-step plan was designed to identify the most probable CD8+ T cell epitopes. In the first step, MHC-I and II binding, MHC-I processing, MHC-I population coverage and MHC-I immunogenicity prediction analyses, and in the second step, MHC-I and II protein-peptide docking, epitope conservation, and cross-reactivity with host antigens’ analyses were carried out successively by different tools. Finally, we introduced five probable CD8+ T cell epitopes for each oncoprotein of the HPV genotypes (60 epitopes in total), which obtained better scores by an integrated approach. These predicted epitopes are valuable candidates for in vitro or in vivo therapeutic vaccine studies against the HPV-associated cancers. Additionally, this two-step plan that each step includes several analyses to find appropriate epitopes provides a rational basis for DNA- or peptide-based vaccine development. [ABSTRACT FROM AUTHOR]

Drosophila melanogaster are known to live in a social but cryptic world of touch and odours, but the extent to which they can perceive and integrate static visual information is a hotly debated topic. Some researchers fixate on the limited resolution of D. melanogaster’s optics, others on their seemingly identical appearance; yet there is evidence of individual recognition and surprising visual learning in flies. Here, we apply machine learning and show that individual D. melanogaster are visually distinct. We also use the striking similarity of Drosophila’s visual system to current convolutional neural networks to theoretically investigate D. melanogaster’s capacity for visual understanding. We find that, despite their limited optical resolution, D. melanogaster’s neuronal architecture has the capability to extract and encode a rich feature set that allows flies to re-identify individual conspecifics with surprising accuracy. These experiments provide a proof of principle that Drosophila inhabit a much more complex visual world than previously appreciated. [ABSTRACT FROM AUTHOR]

Mechanically ventilated surgical patients have a variety of bacterial flora that are often undetectable by traditional culture methods. The source of infection in many of these patients remains unclear. To address this clinical problem, the microbiome profile and host inflammatory response in bronchoalveolar lavage samples from the surgical intensive care unit were examined relative to clinical pathology diagnoses. The hypothesis was tested that clinical diagnosis of respiratory tract flora were similar to culture positive lavage samples in both microbiome and inflammatory profile. Bronchoalveolar lavage samples were collected in the surgical intensive care unit as standard of care for intubated individuals with a clinical pulmonary infection score of >6 or who were expected to be intubated for >48 hours. Cytokine analysis was conducted with the Bioplex Pro Human Th17 cytokine panel. The microbiome of the samples was sequenced for the 16S rRNA region using the Ion Torrent. Microbiome diversity analysis showed the culture-positive samples had the lowest levels of diversity and culture negative with the highest based upon the Shannon-Wiener index (culture positive: 0.77 ± 0.36, respiratory tract flora: 2.06 ± 0.73, culture negative: 3.97 ± 0.65). Culture-negative samples were not dominated by a single bacterial genera. Lavages classified as respiratory tract flora were more similar to the culture-positive in the microbiome profile. A comparison of cytokine expression between groups showed increased levels of cytokines (IFN-g, IL-17F, IL-1B, IL-31, TNF-a) in culture-positive and respiratory tract flora groups. Culture-positive samples exhibited a more robust immune response and reduced diversity of bacterial genera. Lower cytokine levels in culture-negative samples, despite a greater number of bacterial species, suggest a resident nonpathogenic bacterial community may be indicative of a normal pulmonary environment. Respiratory tract flora samples were most similar to the culture-positive samples and may warrant classification as culture-positive when considering clinical treatment. [ABSTRACT FROM AUTHOR]

GSK3532795 (formerly known as BMS-955176) is a second-generation maturation inhibitor targeting a specific Gag cleavage site between capsid p24 and spacer peptide 1 of HIV-1. Study 205891 (previously AI468038) investigated the efficacy, safety, and dose response of GSK3532795 in treatment-naive, HIV-1-infected participants. Study 205891 (NCT02415595) was a Phase IIb, randomized, active-controlled, double-blind, international trial. Participants were randomized 1:1:1:1 to one of three GSK3532795 arms at doses 60 mg, 120 mg or 180 mg once daily (QD), or to efavirenz (EFV) at 600 mg QD, each in combination with tenofovir disoproxil fumarate and emtricitabine (TDF/FTC) (300/200 mg QD). Primary endpoint was proportion of participants with plasma HIV-1 RNA <40 copies/mL at Week 24. Between May 2015 and May 2016, 206 participants received treatment. At Week 24, 76–83% participants receiving GSK3532795 and 77% receiving EFV achieved HIV-1 RNA <40 copies/mL. Fifteen participants receiving GSK3532795 and one receiving EFV met resistance testing criteria; 10/15 receiving GSK3532795 had emergent substitutions at reverse transcriptase positions M184, and one at position K65, while the participant receiving EFV did not have any nucleoside reverse transcriptase inhibitor (NRTI)/non-NRTI mutations. EFV, relative to GSK3532795, had more serious adverse events (9% versus 5%) and adverse events leading to discontinuation (17% versus 5%). However, 3–4-fold higher rates of gastrointestinal adverse events were observed with GSK3532795 relative to EFV. GSK3532795 combined with TDF/FTC is efficacious with 24 weeks of therapy. However, GSK3532795 showed a higher rate of gastrointestinal intolerability and treatment-emergent resistance to the NRTI backbone relative to EFV. Trial registration: ClinicalTrials.gov . [ABSTRACT FROM AUTHOR]