Alcohol and drug use severity are independently associated with antiretroviral adherence in the current treatment era.

Substance use in people with HIV (PWH) negatively impacts antiretroviral therapy (ART) adherence. However, less is known about this in the current treatment era and the impact of specific substances or severity of substance use. We examined the associations of alcohol, marijuana, and illicit drug use (methamphetamine/crystal, cocaine/crack, illicit opioids/heroin) and their severity of use with adherence using multivariable linear regression in adult PWH in care between 2016 and 2020 at 8 sites across the US. PWH completed assessments of alcohol use severity (AUDIT-C), drug use severity (modified ASSIST), and ART adherence (visual analogue scale). Among 9400 PWH, 16% reported current hazardous alcohol use, 31% current marijuana use, and 15% current use of ≥1 illicit drugs. In multivariable analysis, current methamphetamine/crystal use, particularly common among men who had sex with men, was associated with 10.1% lower mean ART adherence (p < 0.001) and 2.6% lower adherence per 5-point higher severity of use (ASSIST score) (p < 0.001). Current and more severe use of alcohol, marijuana, and other illicit drugs were also associated with lower adherence in a dose-dependent manner. In the current HIV treatment era, individualized substance use treatment, especially for methamphetamine/crystal, and ART adherence should be prioritized.

A comparison of digital chest radiography and Xpert® MTB/RIF in active case finding for tuberculosis.

OBJECTIVE: To compare two community screening tests for TB: sputum examination using Xpert® MTB/RIF and chest radiography (CXR).METHOD: Men aged ≥15 years and women aged >45 years living in 96 sub-communes in Ca Mau, Viet Nam, were invited to provide a single sputum specimen that was tested using Xpert. Participants were also invited to attend a nearby location for digital radiography. Participants whose sputum was Xpert MTB-positive or whose CXR was reported as 'consistent with TB´ were requested to provide two further sputum specimens for culture. The sensitivities of the two tests for detecting TB (defined as sputum culture-positive for Mycobacterium tuberculosis) were compared.RESULTS: There were 72 985 eligible participants, of whom 57 597 (78.9%) participated in Xpert screening, 12 752 (17.5%) had CXR and 11 235 (15.4%) had both tests. We estimated that there were 59 cases of TB, of whom 20 were Xpert MTB-positive (programmatic sensitivity 34.0%) and 47 had CXR reported as 'consistent with TB´ (sensitivity 80.0%, P < 0.0001).CONCLUSION: In community-wide screening for TB, CXR is more sensitive than a single spontaneously expectorated sputum sample tested using Xpert, but it has a substantially lower participation rate.

Differential prognostic accuracy of right ventricular dysfunction, the Seattle heart failure model and the MAGGIC score in patients with severe mitral regurgitation undergoing the MitraClip® procedure.

BACKGROUND: MitraClip ® (MC) is an established procedure for severe mitral regurgitation (MR) in patients deemed unsuitable for surgery.Right ventricular dysfunction (RVD) is associated with a higher mortality risk. The prognostic accuracy of heart failure risk scores like the Seattle heart failure model (SHFM) and Meta-Analysis Global Group in Chronic Heart Failure (MAGGIC) score in pts undergoing MC with or without RVD has not been investigated so far. METHODS: SHFM and MAGGIC score were calculated retrospectively. RVD was determined as tricuspid annular plane systolic excursion (TAPSE) ≤15 mm. Area under receiver operating curves (AUROC) of SHFM and MAGGIC were performed for one-year all-cause mortality after MC. RESULTS: N = 103 pts with MR III° (73 ± 11 years, LVEF 37 ± 17%) underwent MC with a reduction of at least I° MR. One-year mortality was 28.2%.In Kaplan-Meier analysis, one- year mortality was significantly higher in RVD-pts (34.8% vs 2.8%, p = 0.009).Area under the Receiver Operating Characteristic (AUROC) for SHFM and MAGGIC were comparable for both scores (SHFM: 0.704, MAGGIC: 0.692). In pts without RVD, SHFM displayed a higher AUROC and therefore better diagnostic accuracy (SHFM: 0.776; MAGGIC: 0.551, p < 0.05). In pts with RVD, MAGGIC and SHFM displayed comparable AUROCs. CONCLUSION: RVD is an important prognostic marker in pts undergoing MC. SHFM and MAGGIC displayed adequate over-all prognostic power in these pts. Accuracy differed in pts with and without RVD, indicating higher predictive power of the SHFM score in pts without RVD.

Biodegradable cationic nanogels with tunable size, swelling and pKa for drug delivery.

Cationic polymers have garnered significant interest for their utility in intracellular drug delivery and gene therapy. However, due to their associated toxicities, novel synthesis approaches must be explored to develop materials that are biocompatible. The novel library of nanoparticles synthesized in this study exhibit tunable hydrodynamic diameters, composition and pH-responsive properties as a function of synthesis parameters. In addition, differences in the responsiveness of these nanoparticles under different pH conditions affords greater control over intracellular drug release.

Identification of novel and conserved microRNA and their expression in the gray mouse lemur, Microcebus murinus, a primate capable of daily torpor.

MicroRNA (miRNA) are endogenous small noncoding RNA gene products, on average 22 nt long, that play important regulatory roles in mediating gene expression by binding to and targeting mRNAs for degradation or translational repression. In this paper we identify both novel and conserved miRNA sequences present in the genome of the gray mouse lemur, Microcebus marinus. In total, 122 conserved and 44 novel miRNA were identified with high confidence from the lemur genome (Mmur_2.0) and were used for expression analysis. All conserved and novel miRNA were subjected to relative quantification by RT-qPCR in liver samples from control and torpid lemurs. A total of 26 miRNA (16 conserved and 10 novel) showed increased levels during primate torpor, whereas 31 (30 conserved and 1 novel) decreased. Additional in silico mapping of the predicted mRNA targets of torpor-responsive mature miRNA suggested that miRNA that increased during torpor were collectively involved in cell development and survival pathways, while miRNA that decreased were enriched in targeting immune function. Overall, the study suggests new regulatory mechanisms of primate torpor via miRNA action.

The role of macroscopic sputum quality assessments to optimise sputum testing for tuberculosis.

SETTING: Community-wide active case finding for tuberculosis (TB) using Xpert® MTB/RIF as the primary screening tool, Ca Mau Province, Viet Nam. OBJECTIVES: To determine whether macroscopic sputum quality characteristics (sputum colour and volume) can be used to predict Xpert MTB-negative sputum and hence exclude sputum samples from testing. DESIGN: Field staff conducted household visits to approximately 51,200 adults in 58 villages randomly selected from throughout the province. Sputum samples from all screened participants who were able to produce ⩾1 ml sputum underwent macroscopic sputum assessment and were tested with Xpert. RESULTS: Of the 21,624 sputum samples tested, 159 (0.74%) were Xpert MTB-positive; 93% of the samples were 1-2 ml and nearly all were mucoid (93%) or mucopurulent (5.7%). One salivary sample was Xpert MTB-positive (2.0% of all salivary samples). The lowest positive predictive value for any sputum volume or colour characteristic was 0.66%. This was not substantially different from the overall prevalence of positive sputum Xpert MTB (0.74%). CONCLUSION: Sputum colour and volume cannot be used to predict the presence or absence of M. tuberculosis in sputum detected using Xpert. These sputum quality parameters cannot therefore be used to exclude sputum samples from testing for TB.

Neurotrophic activity of 2,4,4-trimethyl-3-(15-hydroxypentadecyl)-2-cyclohexen-1-one in cultured central nervous system neurons.

Endogenous neurotrophic factors are essential for the development and maintenance of the nervous system. This suggests their potential utilization as therapeutic agents for neurodegenerative diseases. However, the clinical use of these proteic factors is still restricted, and brings about undesirable consequences, including adverse side effects, and bioavailability and stability difficulties. Therefore, the development of low-molecular weight, non-proteic synthetic compounds with neurotrophic properties appears as a promising approach. The aim of this study was to explore the biological activity of 2,4,4-trimethyl-3-(15-hydroxypentadecyl)-2-cyclohexen-1-one (tCFA15), a trimethyl cyclohexenonic long-chain fatty alcohol. To this end, neurons from fetal rat cerebral hemispheres were cultured in the presence of increasing doses of tCFA15 ranging from 0.1 to 1000 nM. Quantification of cell numbers after 48-h culture showed that 100 nM tCFA15 induced a significant increase in the number of surviving cells. Measurement of total neurite length in microtubule-associated protein 2-positive cells also revealed a stimulatory effect in a wider range of concentrations. The extent of this neuritogenic action was similar to that induced by dibutyryl-cyclic AMP, a well-known neurite outgrowth stimulator, but used at much higher concentration (1 mM). Analysis of structure-activity relationships with different tCFA15 analogs and derivatives corroborated the neurotrophic activity. Taken together, these findings provide strong evidence that tCFA15 exhibits neurotrophic properties in vitro.

Complexation of two proteic insect inhibitors to the active site of chymotrypsin suggests decoupled roles for binding and selectivity.

The crystal structures of two homologous inhibitors (PMP-C and PMP-D2v) from the insect Locusta migratoria have been determined in complex with bovine alpha-chymotrypsin at 2.1- and 3.0-A resolution, respectively. PMP-C is a potent bovine alpha-chymotrypsin inhibitor whereas native PMP-D2 is a weak inhibitor of bovine trypsin. One unique mutation at the P1 position converts PMP-D2 into a potent bovine alpha-chymotrypsin inhibitor. The two peptides have a similar overall conformation, which consists of a triple-stranded antiparallel beta-sheet connected by three disulfide bridges, thus defining a novel family of serine protease inhibitors. They have in common the protease interaction site, which is composed of the classical protease binding loop (position P5 to P'4, corresponding to residues 26-34) and of an internal segment (residues 15-18), held together by two disulfide bridges. Structural divergences between the two inhibitors result in an additional interaction site between PMP-D2v (position P10 to P6, residues 21-25) and the residues 172-175 of alpha-chymotrypsin. This unusual interaction may be responsible for species selectivity. A careful comparison of data on bound and free inhibitors (from this study and previous NMR studies, respectively) suggests that complexation to the protease stabilizes the flexible binding loop (from P5 to P'4).

Peroxisome proliferator-activated receptor-gamma regulates airway epithelial cell activation.

The peroxisome proliferator-activated receptors (PPARs) are nuclear hormone transcription factors that regulate genes associated with lipid and glucose metabolism. Recent evidence suggests that PPAR-gamma may also act as a negative immunomodulator. To investigate the potential role of PPAR-gamma in regulating airway inflammation, we characterized the expression and function of PPAR-gamma in airway epithelial cells. Airway epithelial cells constitutively express PPAR-gamma-specific messenger RNA and protein. Further, airway epithelial PPAR-gamma is inducible by interleukin (IL)-4 in NIH-A549 cells. Two PPAR-gamma agonists, the prostaglandin D2 metabolite 15-deoxy-(Delta)(12,14) prostaglandin J2 (15d-PGJ2) and a thiazolidinedione, ciglitazone, were used to study the effects of PPAR-gamma activation on airway epithelial cytokine expression. Activation of PPAR-gamma stimulated a PPAR-responsive reporter gene in a ligand-specific manner. In NIH-A549 cells, both ligands also blocked the cytokine-induced expression of the inducible form of nitric oxide synthase in a dose-dependent manner. In contrast, ciglitazone alone had a slight effect on cytokine-induced IL-8 secretion, but markedly inhibited IL-8 secretion from cells pretreated with IL-4. The demonstration of PPAR-gamma expression and function in airway epithelial cells expands the immunoregulatory role of PPARs and suggests a critical role for PPAR-gamma in antagonizing proinflammatory pathways in the airways.

3-(15-Hydroxypentadecyl)-2,4,4-trimethyl-2-cyclohexen-1-one and its effect on neuropeptide secretion.

The aim of the present study was to describe the synthesis of a trimethyl cyclohexenonic long chain fatty alcohol (t-CFA), and analyze its biological activity. Specifically, 3-(15-hydroxypentadecyl)-2,4,4-trimethyl-2-cyclohexen-1-one, the t-CFA containing 15 carbon atoms on the side chain (t-CFA n = 15) stimulated arginine vasopressin secretion in nerve terminals of the neurohypophysis. This effect was inhibited by extracellular calcium depletion, which suggests that t-CFA n = 15 stimulates neuropeptide secretion through a calcium-dependent exocytosis mechanism.

Apoptosis at the interface of immunosuppressive and anticancer activities: the examples of two classes of chemical inducers, oxysterols and alkylating agents.

Recent progresses in the understanding of molecular and biochemical pathways involved in apoptotic cell death offer novel perspectives for therapeutic interventions, in particular in immunosuppressive and anti-cancer therapies. In this review, we examine some chemical, biological, and mechanistic aspects of two classes of apoptosis chemical inducers: oxysterols and alkylating agents. Oxysterols represent a vast family of oxygenated derivatives of sterols. Found in both animal and vegetal kingdoms, they can be considered as ultimate products of an oxidative stress, and are chemically inert. Some of them (7beta-hydroxycholesterol, 25-hydroxycholesterol and 7, 25-dihydroxycholesterol) are cytotoxic at micromolar concentrations towards normal and tumor cells in culture, particularly lymphocytes, and reduce the growth of murine transplanted tumors. Thus, possible applications of oxysterols in medicine as immunosuppressants or as anticancer agents may be considered. Alkylating agents, on the other hand, have been widely used in cancer chemotherapy for decades. There toxicity results from their high chemical reactivity, causing lesions to macromolecules through covalent linkage. Some representative members of this class, mainly bifunctional derivatives which possess dichloroethyl groups, such as Chlormethine, Cyclophosphamide and Chlorabucil, express a pronounced cytotoxicity against lymphoid cells, and have therefore potent immunosuppressive properties. Because they triggers apoptosis via both common and distinct mechanisms, oxysterols and alkylating agents provide unique tools for exploring the initiation of this phenomenon in lymphoid cells, and may help design novel pharmacological approaches based on apoptotic modulation of these cells.

Isolation and partial characterization of immunostimulating polysaccharides from Imperata cylindrica.

The water-soluble crude extract prepared from Imperata cylindrica (Beauv.) was investigated for its immunomodulating activity. A set of polysaccharides with high molecular weights has been isolated by fractionation using gel filtration and anion-exchange chromatography. Each step of purification was monitored by bioassays. The presence of six monosaccharides has been established by chemical analysis. Quantitative analysis showed that the ratio of these monosaccharides differed from one polysaccharide to another. The crude extract as well as some of the purified polysaccharides enhance the proliferation of murine splenocytes.

Oxysterols, but not cholesterol, inhibit human immunodeficiency virus replication in vitro.

Oxysterols, oxygenated derivatives of cholesterol selected for their cytostatic activity and their inhibitory effect on cholesterol synthesis, have been investigated for their anti-human immunodeficiency virus (HIV) activity in vitro. The three oxysterols tested, 7 beta-hydroxycholesterol (7 beta-OHC), 25-hydroxycholesterol (25-OHC) and 7 beta, 25-dihydroxycholesterol (7,25-OHC), inhibit viral replication at micromolar concentrations. The selectivity indexes for 7 beta-OHC and 25-OHC are quite modest (2 to 8) but reproducible; the dihydroxycholesterol 7,25-OHC exhibited antiviral properties at concentrations 13- to 25-fold lower than the highest concentration tested at which no toxicity was measurable. Oxysterols are naturally occurring compounds, and we speculate on their physiological relevance in HIV-infected individuals.

Cytotoxic properties of a phosphoglycoconjugated derivative of 7 beta-hydroxycholesterol upon normal and tumor cells in culture.

The cytotoxic activity of a new hydrosoluble axysterol derivative, a phosphoric acid diester of 7 beta-hydroxycholesterol (7 beta-OHC, one of the most toxic oxysterol) and of galactose has been evaluated using cultured tumor cells of various origins, and compared with 7 beta-OHC. As its parent compound, XG-142 exhibits a significant cytotoxic activity against all the cell lines tested, but the IC50's were higher than those obtained with 7 beta-OHC. Moreover, the cytotoxicity was slower to appear than after a 7 beta-OHC treatment. Cell phase distribution was analysed, and revealed some differences between the two compounds. Both oxysterols induced apoptosis at micromolar concentrations, as evidenced by several methods including agarose gel electrophoresis of fragmented DNA and flow cytometry of propidium iodide labeled cells. Apoptosis was also obtained when 7 beta-OHC and XG-142 were combined at concentrations unable to induce this type of cell death when used separately. Upon normal murine spleen cells, XG-142 was found to be less toxic than 7 beta-OHC, and the capacity to respond to Con A stimulation was preserved. Therefore, XG-142 can be considered as a promising soluble analogue of 7 beta-OHC, and its application as anticancer agent should be considered.

Solution structure of PMP-C: a new fold in the group of small serine proteinase inhibitors.

The solution structure and the disulfide pairings of a 36-residue proteinase inhibitor isolated from the insect Locusta migratoria have been determined using NMR spectroscopy and simulated annealing calculations. The peptide, termed PMP-C, was previously shown to inhibit bovine alpha-chymotrypsin as well as human leukocyte elastase, and was also found to block high-voltage-activated Ca2+ currents in rat sensory neurones. PMP-C has a prolate ellipsoid shape and adopts a tertiary fold hitherto unobserved in the large group of small "canonical" proteinase inhibitors. The over-all fold consists mainly of three strands arranged in a right-handed twisted, antiparallel, beta-sheet that demarcates a cavity, together with a linear amino-terminal segment oriented almost perpendicular to the three strands of the beta-sheet. Inside the cavity a phenyl ring constitutes the centre of a hydrophobic core. The proteinase binding loop is located in the carboxy-terminal part of the molecule, between two cysteine residues involved in disulfide bridges. Its conformation resembles that found in other small canonical proteinase inhibitors. A comparison of PMP-C structure with the recently published solution structure of the related peptide PMP-D2 shows that the most significant differences are complementary changes involved in the stabilization of similar folds. This comparison led us to review the structure of PMP-D2 and to identify two salt bridges in PMP-D2.

Regioselective formation of the three disulfide bonds of a 35-residue insect peptide.

PMP-D2, a 35-residue peptide containing three disulfide bonds, was synthesized on solid-phase using 9-fluorenylmethoxy-carbonyl (Fmoc) as alpha-NH2 protection and simultaneous air oxidation of the six cysteines for formation of its disulfide bonds. The overall yield was 13%. As very little research has been done on the regioselective formation of three disulfide bonds, we decided to investigate different strategies using either trityl (Trt), acetamidomethyl (Acm) and methoxybenzyl (Mob), or methoxytrityl (Mmt), trityl and acetamidomethyl, as cysteine-protecting groups and Fmoc as alpha-NH2 protection. In the first strategy, the first disulfide bond was formed by air oxidation and the second was formed by iodine oxidation of the Cys(Acm). Then, the Cys (Mob) was deprotected using TFMSA/TFA treatment for formation of the third disulfide bond. This last step was poorly reproducible on a large scale. The overall yield was 2.5%. In the second strategy, the first disulfide was formed on the resin after removal of the methoxytrityl group, and the two remaining disulfide bonds were formed classically in solution. The overall yield was 2%. From the overall yields using these strategies, it appears clear that simultaneous oxidation of the six cysteines is particularly appropriate for the synthesis of PMP-D2.

Serine protease inhibition by insect peptides containing a cysteine knot and a triple-stranded beta-sheet.

Three insect peptides showing high sequence similarity and belonging to the same structural family incorporating a cysteine knot and a short three-stranded antiparalled beta-sheet were studied. Their inhibitory effect on two serine proteases (bovine alpha-chymotrypsin and human leukocyte elastase) is reported. One of them, PMP-C, is a strong alpha-chymotrypsin inhibitor (Ki = 0.2 nM) and interacts with leukocyte elastase with a Ki of 0.12 microM. The other two peptides, PMP-D2 and HI, interact only weakly with alpha-chymotrypsin and do not inhibit leukocyte elastase. Synthetic variants of these peptides were prepared by solid-phase synthesis, and their action toward serine proteases was evaluated. This enabled us to locate the P1 residues within the reactive sites (Leu-30 for PMP-C and Arg-29 for PMP-D2 and HI), and, interestingly, variants of PMP-D2 and HI were converted into powerful inhibitors of both alpha-chymotrypsin and leukocyte elastase, the most potent elastase inhibitor obtained in this study having a Ki of 3 nM.

Solution structure of two molecular motor domains: nonclaret disjunctional and kinesin.

The effects of selected ligands on the structure of the truncated heavy-chain chemomechanical motor domains of Drosophila ncd and human kinesin were compared using the technique of transient electric birefringence. The 366-amino acid C-terminal motor domain of Drosophila nonclaret disjunctional, ncd(335-700), and the 349-amino acid N-terminal motor domain of human kinesin, kinesin(349), were studied at 4 degrees C in neutral buffers with ionic strength of 100 mM to form complexes with either MgADP or MgADP.Vi. The rotational diffusion time adjusted to 20 degrees C and water, tau 20,W, for ncd(335-700).MgADP is 32.8 ns, and for ncd(335-700).MgADP.Vi is 34.8 ns, suggesting prolate ellipsoids with dimensions 9.40 x 3.77 nm and 9.73 x 3.70 nm, respectively. The specific Kerr constant, Ksp, of ncd is -1.65 x 10(-12) cm2V-2 for the MgADP complex and -1.15 x 10(-12) cm2V-2 for the MgADP.Vi complex. The large negative Ksp for a prolate protein suggests an unusual charge distribution with two long surfaces with opposite charge. The tau 20,W for kinesin(349).MgADP is longer than the corresponding ncd motor and shows a decrease with increased electric field. The kinesin(349).MgADP.Vi complex has a longer tau 20,W. The Ksp for kinesin(349) is 0.36 x 10(-12) cm2V-2 for each complex.

Inhibition of high voltage-activated Ca2+ currents from cultured sensory neurones by a novel insect peptide.

PMP-D2, a novel 35 amino acid peptide isolated from the brain of the locust Locusta migratoria, is localised specifically in neurosecretory cells and nerve tracts of the Pars intercerebralis. When PMP-D2 is applied onto rat sensory neurones it blocks high voltage-activated inward Ca2+ currents at concentrations ranging from 0.1 mu M to 10 mu M. The inhibitory effect of PMP-D2 is more marked on the sustained inward Ca2+ current measured at the end of 100 ms voltage step commands than on the maximum inward Ca2+ current. These results suggests that PMP-D2 may differentially inhibit the two components of the high voltage-activated inward Ca2+ currents of rat sensory neurones.

[From cholesterol to oxysterols. Current data]. / Du cholestérol aux oxystérols. Données récentes.

Oxysterols, a class of cholesterol oxidation products exhibit several important biological activities. Some of these natural compounds are potent inhibitors of the enzyme HMG-CoA reductase, a key enzyme in the cholesterol biosynthetic pathway. Many studies have been directed towards to the verification of the hypothesis that some oxysterols are endogenous intracellular regulators of cholesterol homeostasis. In adition to oxysterols derived directly from oxidation of cholesterol, several others are formed from squalene dioxide. It is presently well established that, in addition to the classical cholesterol biosynthetic pathway, there exists an alternate bifurcation from squalene oxide. The cyclisation of squalene dioxide leads to a series of new oxysterols. Thus, several types of oxysterols and several molecular targets are involved in the regulation of steroid biosynthesis. Many oxysterols, particularly those obtained from the oxidations of phytosterols and tetracyclic triterpenes are potent cytotoxic agents. They are selectively cytotoxic against tumorous cells. This cytotoxicity depends markedly on the specific structure of each oxysterol. Some structures are very cytotoxic, while their stereoisomers are inactive. The activity depends on the tumor cells which are used in the assay system: some compounds display inhibitory activity towards hepatoma cells but are inactive against lymphoma cells while others act in the opposite manner. Free oxysterols do not depress tumor growth in living animals. However, several water soluble prodrugs of oxysterols are able to depress different type of tumors in vivo. Clinical trial studies are presently conducted in order to learn the therapeutic values of these oxysterols.