Brain tissue transcriptomic analysis of SIV-infected macaques identifies several altered metabolic pathways linked to neuropathogenesis and poly (ADP-ribose) polymerases (PARPs) as potential therapeutic targets.

Despite improvements in antiretroviral therapy, human immunodeficiency virus type 1 (HIV-1)-associated neurocognitive disorders (HAND) remain prevalent in subjects undergoing therapy. HAND significantly affects individuals' quality of life, as well as adherence to therapy, and, despite the increasing understanding of neuropathogenesis, no definitive diagnostic or prognostic marker has been identified. We investigated transcriptomic profiles in frontal cortex tissues of Simian immunodeficiency virus (SIV)-infected Rhesus macaques sacrificed at different stages of infection. Gene expression was compared among SIV-infected animals (n = 11), with or without CD8+ lymphocyte depletion, based on detectable (n = 6) or non-detectable (n = 5) presence of the virus in frontal cortex tissues. Significant enrichment in activation of monocyte and macrophage cellular pathways was found in animals with detectable brain infection, independently from CD8+ lymphocyte depletion. In addition, transcripts of four poly (ADP-ribose) polymerases (PARPs) were up-regulated in the frontal cortex, which was confirmed by real-time polymerase chain reaction. Our results shed light on involvement of PARPs in SIV infection of the brain and their role in SIV-associated neurodegenerative processes. Inhibition of PARPs may provide an effective novel therapeutic target for HIV-related neuropathology.

Machine Learning Prediction and Phyloanatomic Modeling of Viral Neuroadaptive Signatures in the Macaque Model of HIV-Mediated Neuropathology.

In human immunodeficiency virus (HIV) infection, virus replication in and adaptation to the central nervous system (CNS) can result in neurocognitive deficits in approximately 25% of patients with unsuppressed viremia. While no single viral mutation can be agreed upon as distinguishing the neuroadapted population, earlier studies have demonstrated that a machine learning (ML) approach could be applied to identify a collection of mutational signatures within the virus envelope glycoprotein (Gp120) predictive of disease. The S[imian]IV-infected macaque is a widely used animal model of HIV neuropathology, allowing in-depth tissue sampling infeasible for human patients. Yet, translational impact of the ML approach within the context of the macaque model has not been tested, much less the capacity for early prediction in other, noninvasive tissues. We applied the previously described ML approach to prediction of SIV-mediated encephalitis (SIVE) using gp120 sequences obtained from the CNS of animals with and without SIVE with 97% accuracy. The presence of SIVE signatures at earlier time points of infection in non-CNS tissues indicated these signatures cannot be used in a clinical setting; however, combined with protein structural mapping and statistical phylogenetic inference, results revealed common denominators associated with these signatures, including 2-acetamido-2-deoxy-beta-d-glucopyranose structural interactions and high rate of alveolar macrophage (AM) infection. AMs were also determined to be the phyloanatomic source of cranial virus in SIVE animals, but not in animals that did not develop SIVE, implicating a role for these cells in the evolution of the signatures identified as predictive of both HIV and SIV neuropathology. IMPORTANCE HIV-associated neurocognitive disorders remain prevalent among persons living with HIV (PLWH) owing to our limited understanding of the contributing viral mechanisms and ability to predict disease onset. We have expanded on a machine learning method previously used on HIV genetic sequence data to predict neurocognitive impairment in PLWH to the more extensively sampled SIV-infected macaque model in order to (i) determine the translatability of the animal model and (ii) more accurately characterize the predictive capacity of the method. We identified eight amino acid and/or biochemical signatures in the SIV envelope glycoprotein, the most predominant of which demonstrated the potential for aminoglycan interaction characteristic of previously identified HIV signatures. These signatures were not isolated to specific points in time or to the central nervous system, limiting their use as an accurate clinical predictor of neuropathogenesis; however, statistical phylogenetic and signature pattern analyses implicate the lungs as a key player in the emergence of neuroadapted viruses.

Effects of ß-caryophyllene and oxygen availability on cholesterol and fatty acids in breast cancer cells.

Hypoxia is a common feature of most solid tumors, one that favors tumor progression and limits treatment effectiveness. Targeting hypoxia has long been a goal in cancer therapy, by identifying factors that reverse or ameliorate the effects of hypoxia on cancer cells. We, and others, have shown that ß-caryophyllene (BCP) exhibits anti-proliferative properties in cancer cells. We have further shown that non-cytotoxic concentrations of BCP affect cholesterol and lipid biosynthesis in hypoxic hBrC cells at both transcriptional and translational levels. This led us to hypothesize that BCP may reverse the hypoxic phenotype in hBrC cells. To test this, we determined the effect of BCP on hypoxic sensitive pathways, including oxygen consumption, glycolysis, oxidative stress, cholesterol and fatty acid biosynthesis, and ERK activation. While each of these studies revealed new information on the regulation by hypoxia and BCP, only the lipidomic studies showed reversal of hypoxic-dependent effects by BCP. These later studies showed that hypoxia-treated samples lowered monounsaturated fatty acid levels, shifting the saturation ratios of the fatty acid pools. This signature was ameliorated by sub-lethal concentrations of BCP, possibly through an effect on the C:16 fatty acid saturation ratios. This is consistent with BCP-induced upregulation of the stearoyl-CoA desaturase (SCD) gene, observed previously. This suggests that BCP may interfere with the lipid signature modulated by hypoxia which could have consequences for membrane biosynthesis or composition, both of which are important for cell replication.

Differing impacts of global and regional responses on SARS-CoV-2 transmission cluster dynamics.

Although the global response to COVID-19 has not been entirely unified, the opportunity arises to assess the impact of regional public health interventions and to classify strategies according to their outcome. Analysis of genetic sequence data gathered over the course of the pandemic allows us to link the dynamics associated with networks of connected individuals with specific interventions. In this study, clusters of transmission were inferred from a phylogenetic tree representing the relationships of patient sequences sampled from December 30, 2019 to April 17, 2020. Metadata comprising sampling time and location were used to define the global behavior of transmission over this earlier sampling period, but also the involvement of individual regions in transmission cluster dynamics. Results demonstrate a positive impact of international travel restrictions and nationwide lockdowns on global cluster dynamics. However, residual, localized clusters displayed a wide range of estimated initial secondary infection rates, for which uniform public health interventions are unlikely to have sustainable effects. Our findings highlight the presence of so-called "super-spreaders", with the propensity to infect a larger-than-average number of people, in countries, such as the USA, for which additional mitigation efforts targeting events surrounding this type of spread are urgently needed to curb further dissemination of SARS-CoV-2.

ß-caryophyllene enhances the transcriptional upregulation of cholesterol biosynthesis in breast cancer cells.

ß-caryophyllene (BCP) exhibits anti-proliferative properties in cancer cells. Here, we examine the hypothesis that BCP induces membrane remodeling. Our data show that high concentrations of BCP increase membrane permeability of human breast cells (hBrC) causing detachment and cell death. At a sub-lethal concentration of BCP, we show that BCP induces a striking upregulation of genes involved in cholesterol biosynthesis, including the gene that encodes for HMGCoA reductase (HMGCR), the rate-determining step in cholesterol biosynthesis. In addition, stearoyl-CoA desaturase (SCD) is also upregulated which would lead to the enhanced formation of monounsaturated fatty acids, specifically oleate and palmitoleate from stearoyl CoA and palmitoyl CoA, respectively. These fatty acids are major components of membrane phospholipids and cholesterol esters. Together, these data suggest that cells respond to BCP by increasing the synthesis of components found in membranes. These responses could be viewed as a repair mechanism and/or as a mechanism to mount resistance to the cytotoxic effect of BCP. Blocking HMGCR activity enhances the cytotoxicity of BCP, suggesting that BCP may provide an additional therapeutic tool in controlling breast cancer cell growth.

Toxin B Variants from Clostridium difficile Strains VPI 10463 and NAP1/027 Share Similar Substrate Profile and Cellular Intoxication Kinetics but Use Different Host Cell Entry Factors.

Clostridium difficile induces antibiotic-associated diarrhea due to the release of toxin A (TcdA) and toxin B (TcdB), the latter being its main virulence factor. The epidemic strain NAP1/027 has an increased virulence attributed to different factors. We compared cellular intoxication by TcdBNAP1 with that by the reference strain VPI 10463 (TcdBVPI). In a mouse ligated intestinal loop model, TcdBNAP1 induced higher neutrophil recruitment, cytokine release, and epithelial damage than TcdBVPI. Both toxins modified the same panel of small GTPases and exhibited similar in vitro autoprocessing kinetics. On the basis of sequence variations in the frizzled-binding domain (FBD), we reasoned that TcdBVPI and TcdBNAP1 might have different receptor specificities. To test this possibility, we used a TcdB from a NAP1 variant strain (TcdBNAP1v) unable to glucosylate RhoA but with the same receptor-binding domains as TcdBNAP1. Cells were preincubated with TcdBNAP1v to block cellular receptors, prior to intoxication with either TcdBVPI or TcdBNAP1. Preincubation with TcdBNAP1v blocked RhoA glucosylation by TcdBNAP1 but not by TcdBVPI, indicating that the toxins use different host factors for cell entry. This crucial difference might explain the increased biological activity of TcdBNAP1 in the intestine, representing a contributing factor for the increased virulence of the NAP1/027 strain.