Cardiac gene therapy in large animals: bridge from bench to bedside.

Several clinical trials are evaluating gene transfer as a therapeutic approach to treat cardiac diseases. Although it has just started on the path to clinical application, recent advances in gene delivery technologies with increasing knowledge of underlying mechanisms raise great expectations for the cardiac gene therapy. Although in vivo experiments using small animals provide the therapeutic potential of gene transfer, there exist many fundamental differences between the small animal and the human hearts. Before applying the therapy to clinical patients, large animal studies are a prerequisite to validate the efficacy in an animal model more relevant to the human heart. Several key factors including vector type, injected dose, delivery method and targeted cardiac disease are all important factors that determine the therapeutic efficacy. Selecting the most optimal combination of these factors is essential for successful gene therapy. In addition to the efficacy, safety profiles need to be addressed as well. In this regard, large animal studies are best suited for comprehensive evaluation at the preclinical stages of therapeutic development to ensure safe and effective gene transfer. As the cardiac gene therapy expands its potential, large animal studies will become more important to bridge the bench side knowledge to the clinical arena.

Vertical Patterns of Nitrogen Transformations during Infiltration in Two Wetland Soils.

The (sup15)N isotope dilution and pairing methods were applied to investigate the vertical distribution of nitrogen transformations during infiltration in one peaty soil and one sandy soil. Water containing (sup15)N-nitrate (99.9%; 200 (mu)M) as the only nitrogen fraction was infiltrated through cores containing homogenized soil, with lengths varying from 5.5 to 38 cm. Oxygen and nitrogen dynamics were investigated by measuring inflowing and outflowing water. The experimental design allowed determinations of vertical profiles of aerobic respiration, nitrification, and coupled and uncoupled denitrification and ammonification. In the sandy soil, all oxygen was consumed in the upper 14 cm and nitrate was subsequently consumed and removed, up to a maximum of 70% in the longest core (28 cm). In the peaty soil, oxygen was consumed in the upper 7.5 cm and all nitrate was denitrified in the top 20 cm. In both soils, nitrogen removal by denitrification was counteracted by the release of ammonium and dissolved organic nitrogen. In the sandy soil, net nitrogen removal occurred in cores of 14 cm and longer; in the longest core, 40% was removed. In the peaty soil, release was equal to removal in the top 14 cm but release exceeded removal in the deeper layers, leading to a 100% increase of total nitrogen in the effluent water from the longest core (38 cm).

Nitrogen Transformations in Wetland Soil Cores Measured by (sup15)N Isotope Pairing and Dilution at Four Infiltration Rates.

The effect of water infiltration rate (IR) on nitrogen cycling in a saturated wetland soil was investigated by applying a (sup15)N isotope dilution and pairing method. Water containing [(sup15)N]nitrate was infiltrated through 10-cm-long cores of sieved and homogenized soil at rates of 72, 168, 267, and 638 mm day(sup-1). Then the frequencies of (sup30)N(inf2), (sup29)N(inf2), (sup15)NO(inf3)(sup-), and (sup15)NH(inf4)(sup+) in the outflow water were measured. This method allowed simultaneous determination of nitrification, coupled and uncoupled denitrification, and nitrate assimilation rates. From 3% (at the highest IR) to 95% (at the lowest IR) of nitrate was removed from the water, mainly by denitrification. The nitrate removal was compensated for by the net release of ammonium and dissolved organic nitrogen. Lower oxygen concentrations in the soil at lower IRs led to a sharper decrease in the nitrification rate than in the ammonification rate, and, consequently, more ammonium leaked from the soil. The decreasing organic-carbon-to-nitrogen ratio (from 12.8 to 5.1) and the increasing light A(inf250)/A(inf365) ratio (from 4.5 to 5.2) indicated an increasing bioavailability of the outflowing dissolved organic matter with increasing IR. The efflux of nitrous oxide was also very sensitive to IR and increased severalfold when a zone of low oxygen concentration was close to the outlet of the soil cores. N(inf2)O then constituted 8% of the total gaseous N lost from the soil.

Interactions between Lugol's fixative and ethylene in the acetylene-reduction assay for nitrogenase activity in lake water.

Lugol's solution is a practical and efficient fixative for the acetylene-reduction assay of nitrogenase activity in aquatic organisms. Correction must be made, however, for the solubility of ethylene in the liquid phase and reactions between Lugol's solution and ethylene. With a vapor phase-liquid phase volume ratio of 1.9:1, the mean solubility of ethylene in mixtures of lake water and Lugol's solution was 7.2%. No correlation was found between ethylene solubility and the concentration of Lugol's solution. Storage of fixed samples for more than 1 day before gas chromatographic analysis resulted in increased loss of ethylene from the vapor phase; the loss amounted to ca. 18% after 3 days. Higher losses were noted at higher concentrations of Lugol's solution. Most probably these effects were caused by iodine addition to ethylene, as indicated by the consumption of ethylene by iodine-potassium iodide solutions. The reaction was catalyzed by the rubber septa of the incubaton vessels when the septa were in contact with the liquid phase. Loss of ethylene decreased with increased concentration of phytoplankton because the organisms absorbed iodine. By using a standardized technique and determining ethylene solubility and reaction patterns between ethylene and the mixture of water and Lugol's solution, it is possible to correct for the loss of ethylene.