Combination of apolipoprotein-A-I/apolipoprotein-A-I binding protein and anti-VEGF treatment overcomes anti-VEGF resistance in choroidal neovascularization in mice.

Many patients of choroidal neovascularization (CNV) are unresponsive to the current anti-VEGF treatment. The mechanisms for anti-VEGF resistance are poorly understood. We explore the unique property of the apolipoprotein A-I (apoA-I) binding protein (AIBP) that enhances cholesterol efflux from endothelial cells and macrophages to thereby limit angiogenesis and inflammation to tackle anti-VEGF resistance in CNV. We show that laser-induced CNV in mice with increased age showed increased resistance to anti-VEGF treatment, which correlates with increased lipid accumulation in macrophages. The combination of AIBP/apoA-I and anti-VEGF treatment overcomes anti-VEGF resistance and effectively suppresses CNV. Furthermore, macrophage depletion in old mice restores CNV sensitivity to anti-VEGF treatment and blunts the synergistic effect of combination therapy. These results suggest that cholesterol-laden macrophages play a critical role in inducing anti-VEGF resistance in CNV. Combination therapy by neutralizing VEGF and enhancing cholesterol removal from macrophages is a promising strategy to combat anti-VEGF resistance in CNV.

Effects of transgenic sterilization constructs and their repressor compounds on hatch, developmental rate and early survival of electroporated channel catfish embryos and fry.

Channel catfish (Ictalurus punctatus) embryos were electroporated with sterilization constructs targeting primordial germ cell proteins or with buffer. Some embryos then were treated with repressor compounds, cadmium chloride, copper sulfate, sodium chloride or doxycycline, to prevent expression of the transgene constructs. Promoters included channel catfish nanos and vasa, salmon transferrin (TF), modified yeast Saccharomyces cerevisiae copper transport protein (MCTR) and zebrafish racemase (RM). Knock-down systems were the Tet-off (nanos and vasa constructs), MCTR, RM and TF systems. Knock-down genes included shRNAi targeting 5' nanos (N1), 3' nanos (N2) or dead end (DND), or double-stranded nanos RNA (dsRNA) for overexpression of nanos mRNA. These constructs previously were demonstrated to knock down nanos, vasa and dead end, with the repressors having variable success. Exogenous DNA affected percentage hatch (% hatch), as all 14 constructs, except for the TF dsRNA, TF N1 (T), RM DND (C), vasa DND (C), vasa N1 (C) and vasa N2 (C), had lower % hatch than the control electroporated with buffer. The MCTR and RM DND (T) constructs resulted in delayed hatch, and the vasa and nanos constructs had minimal effects on time of hatch (P < 0.05). Cadmium chloride appeared to counteract the slow development caused by the TF constructs in two TF treatments (P < 0.05). The 4 ppt sodium chloride treatment for the RM system decreased % hatch (P < 0.05) and slowed development. In the case of nanos constructs, doxycycline greatly delayed hatch (P < 0.05). Adverse effects of the transgenes and repressors continued for several treatments for the first 6 days after hatch, but only in a few treatments during the next 10 days. Repressors and gene expression impacted the yield of putative transgenic channel catfish fry, and need to be considered and accounted for in the hatchery phase of producing transgenically sterilized catfish fry and their fertile counterparts. This fry output should be considered to ensure that sufficient numbers of transgenic fish are produced for future applications and for defining repressor systems that are the most successful.

Ranolazine, a partial fatty acid oxidation (pFOX) inhibitor, improves left ventricular function in dogs with chronic heart failure.

BACKGROUND: Abnormalities of energy metabolism are often cited as key elements in the progressive worsening of left ventricular (LV) dysfunction that characterizes the heart failure (HF) state. The present study tested the hypothesis that partial inhibition of fatty acids will ameliorate the hemodynamic abnormalities associated with HF. METHODS AND RESULTS: Chronic HF (LV ejection fraction 27 +/- 1%) was produced in 13 dogs by intracoronary microembolizations. Hemodynamic and angiographic measurements were made before and 40 minutes after intravenous administration of ranolazine, a partial fatty acid oxidation (pFOX) inhibitor. Ranolazine was administered as an intravenous bolus dose of 0.5 mg/kg followed by a continuous infusion for 40 minutes at a constant rate of 1.0 mg / kg / hr. Ranolazine significantly increased LV ejection fraction (27 +/- 1 versus 36 +/- 2, P =.0001), peak LV +dP/dt (1712 +/- 122 versus 1900 +/- 112 mm Hg/sec, P =.001), and stroke volume (20 +/- 1 versus 26 +/- 1 mL). These improvements occurred in the absence of any effects on heart rate or systemic pressure. In 8 normal healthy dogs, ranolazine had no effect on LV ejection fraction or any other index of LV function. CONCLUSIONS: In dogs with HF, acute intravenous administration of the pFOX inhibitor ranolazine improves LV systolic function. The absence of any hemodynamic effects of ranolazine in normal dogs suggests that the drug is devoid of any positive inotropic effects and acts primarily by optimizing cardiac metabolism in the setting of chronic HF.

Pharmacokinetics and pharmacodynamics of methadone enantiomers after a single oral dose of racemate.

BACKGROUND: The pharmacokinetics and dynamics of methadone are characterized by high interindividual variability. This study aimed to examine a number of factors that may contribute to this variability. METHODS: Eight healthy drug-free women were administered 0.2 mg/kg of R,S-methadone orally. The concentrations of methadone's enantiomers in plasma and urine were monitored for 96 hours. Vital signs, blood biochemical parameters, and pupillary diameter were monitored frequently during this period. Cytochrome P450 3A (CYP3A) activity and alpha1-acid glycoprotein (alpha1-AGP) concentrations and phenotypes were determined. Pharmacokinetic and pharmacodynamic modeling was used to assess the influence of the above-mentioned covariables on methadone enantiomer disposition and actions. RESULTS: The pharmacokinetic profile of the active enantiomer of methadone, R -methadone, showed a relatively normal distribution with 38% to 90% of the interindividual variability in modeled pharmacokinetic parameters being explained by their individual variability in CYP3A activity, the cumulative amount of the main CYP3A4 metabolite, 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrolidine, excreted in the urine, the fraction unbound in plasma, and the alpha1-AGP orosomucoid 2 (ORM2) variant plasma concentration. S-Methadone showed an idiosyncratic distribution with largely unpredictable pharmacokinetics. Pupillary constriction response was highly variable between individuals. CONCLUSIONS: The disposition of the active enantiomer, R -methadone, can be predicted in part by CYP3A activity and protein binding to alpha1-AGP (ORM2), whereas S-methadone disposition is not well explained by the factors examined in this study. Central nervous system effects were difficult to interpret on the basis of plasma R-methadone pharmacokinetics.