A spectrophotometric technique for measuring particle ingestion by black fly larvae

A spectrophotometric technique was developed to provide insight into the feeding behavior of Simulium vittatum Zetterstedt (Diptera: Simuliidae) larvae. Larvae were exposed to water insoluble Neon Red particles (NRP) (DayGlo^®) in a controlled current. The insoluble particles were available for capture by the cephalic fans of the larvae and subsequent ingestion. The length of gut occupied by the particles after a given exposure time was determined by visual inspection and measured with the aid of a dissecting microscope. Larvae were then homogenized in acetone to solubilize the particles. After filtration, the quantity of pigmented particles in the alimentary tract of the larvae was determined using spectrophotometric analysis. The quantity of particles per unit length of the alimentary tract was calculated. Experiments were conducted to determine the ideal concentration of NRP for obtaining an accurate measure of ingestion without interfering with normal larval feeding behavior. Larval mortality following ingestion of insecticidal proteins produced by Bacillus thuringiensis ssp. israelensis was used as an indirect measure of feeding behavior for these experiments. A concentration of 15 p.p.m. of NRP in the larval medium was the highest concentration used that did not interfere with larval mortality following exposure to the insecticidal proteins. Additional experiments demonstrated that components of the experimental matrix did not interfere with NRP absorbance. The final experiment revealed that the consumption of NRP and insecticidal proteins by larvae was influenced by clay and cellulose in the larval medium.     

A randomized,phase 3 trial of naltrexone SR/bupropion SR on weight and obesity‐related risk factors (COR‐II)

Objective:

To examine the effects of naltrexone/bupropion (NB) combination therapy on weight and weight‐related risk factors in overweight and obese participants.

Design and Methods:

CONTRAVE Obesity Research‐II (COR‐II) was a double‐blind, placebo‐controlled study of 1,496 obese (BMI 30‐45 kg/m²) or overweight (27‐45 kg/m² with dyslipidemia and/or hypertension) participants randomized 2:1 to combined naltrexone sustained‐release (SR) (32 mg/day) plus bupropion SR (360 mg/day) (NB32) or placebo for up to 56 weeks. The co‐primary endpoints were percent weight change and proportion achieving ≥5% weight loss at week 28.

Results:

Significantly (P < 0.001) greater weight loss was observed with NB32 versus placebo at week 28 (?6.5% vs. ?1.9%) and week 56 (?6.4% vs. ?1.2%). More NB32‐treated participants (P < 0.001) experienced ≥5% weight loss versus placebo at week 28 (55.6% vs. 17.5%) and week 56 (50.5% vs. 17.1%). NB32 produced greater improvements in various cardiometabolic risk markers, participant‐reported weight‐related quality of life, and control of eating. The most common adverse event with NB was nausea, which was generally mild to moderate and transient. NB was not associated with increased events of depression or suicidality versus placebo.

Conclusion:

NB represents a novel pharmacological approach to the treatment of obesity, and may become a valuable new therapeutic option.

     

Integrated Delivery of Antiretroviral Treatment and Pre-exposure Prophylaxis to HIV-1–Serodiscordant Couples: A Prospective Implementation Study in Kenya and Uganda

BackgroundAntiretroviral-based interventions for HIV-1 prevention, including antiretroviral therapy (ART) to reduce the infectiousness of HIV-1 infected persons and pre-exposure prophylaxis (PrEP) to reduce the susceptibility of HIV-1 uninfected persons, showed high efficacy for HIV-1 protection in randomized clinical trials. We conducted a prospective implementation study to understand the feasibility and effectiveness of these interventions in delivery settings.ConclusionsIntegrated delivery of time-limited PrEP until sustained ART use in African HIV-1-serodiscordant couples was feasible, demonstrated high uptake and adherence, and resulted in near elimination of HIV-1 transmission, with an observed HIV incidence of <0.5% per year compared to an expected incidence of >5% per year.     

Twist and Degrade—Impact of Molecular Structure on the Photostability of Nonfullerene Acceptors and Their Photovoltaic Blends

Nonfullerene acceptors (NFAs) dominate organic photovoltaic (OPV) research due to their promising efficiencies and stabilities. However, there is very little investigation into the molecular processes of degradation, which is critical to guiding design of novel NFAs for long‐lived, commercially viable OPVs. Here, the important role of molecular structure and conformation in NFA photostability in air is investigated by comparing structurally similar but conformationally different promising NFAs: planar O‐IDTBR and nonplanar O‐IDFBR. A three‐phase degradation process is identified: i) initial photoinduced conformational change (i.e., torsion about the core–benzothiadiazole dihedral), induced by noncovalent interactions with environmental molecules, ii) followed by photo‐oxidation and fragmentation, leading to chromophore bleaching and degradation product formation, and iii) finally complete chromophore bleaching. Initial conformational change is a critical prerequisite for further degradation, providing fundamental understanding of the relative stability of IDTBR and IDFBR, where the already twisted IDFBR is more prone to degradation. When blended with the donor polymer poly(3‐hexylthiophene), both NFAs exhibit improved photostability while the photostability of the polymer itself is significantly reduced by the more miscible twisted NFA. The findings elucidate the important role of NFA molecular structure in photostability of OPV systems, and provide vital insights into molecular design rules for intrinsically photostable NFAs.     

Evaluation of α-hydroxycinnamic acids as pyruvate carboxylase inhibitors

Through a structure-based drug design project (SBDD), potent small molecule inhibitors of pyruvate carboxylase (PC) have been discovered. A series of α-keto acids (7) and α-hydroxycinnamic acids (8) were prepared and evaluated for inhibition of PC in two assays. The two most potent inhibitors were 3,3′-(1,4-phenylene)bis[2-hydroxy-2-propenoic acid] (8u) and 2-hydroxy-3-(quinoline-2-yl)propenoic acid (8v) with IC₅₀ values of 3.0 ± 1.0 μM and 4.3 ± 1.5 μM respectively. Compound 8v is a competitive inhibitor with respect to pyruvate (K_i = 0.74 μM) and a mixed-type inhibitor with respect to ATP, indicating that it targets the unique carboxyltransferase (CT) domain of PC. Furthermore, compound 8v does not significantly inhibit human carbonic anhydrase II, matrix metalloproteinase-2, malate dehydrogenase or lactate dehydrogenase.     

Regional Differences in the Contributions of TNF Reverse and Forward Signaling to the Establishment of Sympathetic Innervation

Members of the TNF and TNF receptor superfamilies acting by both forward and reverse signaling are increasingly recognized as major physiological regulators of axon growth and tissue innervation in development. Studies of the experimentally tractable superior cervical ganglion (SCG) neurons and their targets have shown that only TNF reverse signaling, not forward signaling, is a physiological regulator of sympathetic innervation. Here, we compared SCG neurons and their targets with prevertebral ganglion (PVG) neurons and their targets. Whereas all SCG targets were markedly hypoinnervated in both TNF‐deficient and TNFR1‐deficient mice, PVG targets were not hypoinnervated in these mice and one PVG target, the spleen, was significantly hyperinnervated. These in vivo regional differences in innervation density were related to in vitro differences in the responses of SCG and PVG neurons to TNF reverse and forward signaling. Though TNF reverse signaling enhanced SCG axon growth, it did not affect PVG axon growth. Whereas activation of TNF forward signaling in PVG axons inhibited growth, TNF forward signaling could not be activated in SCG axons. These latter differences in the response of SCG and PVG axons to TNF forward signaling were related to TNFR1 expression, whereas PVG axons expressed TNFR1, SCG axons did not. These results show that both TNF reverse and forward signaling are physiological regulators of sympathetic innervation in different tissues.     

CD40L reverse signaling suppresses prevertebral sympathetic axon growth and tissue innervation

CD40‐activated CD40L reverse signaling is a major physiological regulator of the growth of neural processes in the developing nervous system. Previous work on superior cervical ganglion (SCG) neurons of the paravertebral sympathetic chain has shown that CD40L reverse signaling enhances NGF‐promoted axon growth and tissue innervation. Here we show that CD40L reverse signaling has the opposite function in prevertebral ganglion (PVG) sympathetic neurons. During a circumscribed perinatal window of development, PVG neurons cultured from Cd40^–/– mice had substantially larger, more exuberant axon arbors in the presence of NGF than PVG neurons cultured from wild‐type mice. Tissues that receive their sympathetic innervation from PVG neurons were markedly hyperinnervated in Cd40^–/– mice compared with wild‐type mice. The exuberant axonal growth phenotype of cultured CD40‐deficient perinatal PVG neurons was pared back to wild‐type levels by activating CD40L reverse signaling with a CD40‐Fc chimeric protein, but not by activating CD40 forward signaling with CD40L. The co‐expression of CD40 and CD40L in PVG neurons suggests that these proteins engage in an autocrine signaling loop in these neurons. Our work shows that CD40L reverse signaling is a physiological regulator of NGF‐promoted sympathetic axon growth and tissue innervation with opposite effects in paravertebral and prevertebral neurons.     

Key features determining the specificity of aspartic proteinase inhibition by the helix-forming IA3 polypeptide

The 68-residue IA(3) polypeptide from Saccharomyces cerevisiae is essentially unstructured. It inhibits its target aspartic proteinase through an unprecedented mechanism whereby residues 2-32 of the polypeptide adopt an amphipathic alpha-helical conformation upon contact with the active site of the enzyme. This potent inhibitor (K(i) < 0.1 nm) appears to be specific for a single target proteinase, saccharopepsin. Mutagenesis of IA(3) from S. cerevisiae and its ortholog from Saccharomyces castellii was coupled with quantitation of the interaction for each mutant polypeptide with saccharopepsin and closely related aspartic proteinases from Pichia pastoris and Aspergillus fumigatus. This identified the charged K18/D22 residues on the otherwise hydrophobic face of the amphipathic helix as key selectivity-determining residues within the inhibitor and implicated certain residues within saccharopepsin as being potentially crucial. Mutation of these amino acids established Ala-213 as the dominant specificity-governing feature in the proteinase. The side chain of Ala-213 in conjunction with valine 26 of the inhibitor marshals Tyr-189 of the enzyme precisely into a position in which its side-chain hydroxyl is interconnected via a series of water-mediated contacts to the key K18/D22 residues of the inhibitor. This extensive hydrogen bond network also connects K18/D22 directly to the catalytic Asp-32 and Tyr-75 residues of the enzyme, thus deadlocking the inhibitor in position. In most other aspartic proteinases, the amino acid at position 213 is a larger hydrophobic residue that prohibits this precise juxtaposition of residues and eliminates these enzymes as targets of IA(3). The exquisite specificity exhibited by this inhibitor in its interaction with its cognate folding partner proteinase can thus be readily explained.