Cyclic Nucleotide Phosphodiesterases and Compartmentation in Normal and Diseased Heart

International audienceCyclic nucleotide phosphodiesterases (PDEs) degrade the second messengers cAMP and cGMP, thereby regulating multiple aspects of cardiac function. This highly diverse class of enzymes encoded by 21 genes encompasses 11 families which are not only responsible for the termination of cyclic nucleotide signalling, but are also involved in the generation of dynamic microdomains of cAMP and cGMP controlling specific cell functions in response to various neurohormonal stimuli. In myocardium, the PDE3 and PDE4 families are predominant to degrade cAMP and thereby regulate cardiac excitation-contraction coupling. PDE3 inhibitors are positive inotropes and vasodilators in human, but their use is limited to acute heart failure and intermittent claudication. PDE5 is particularly important to degrade cGMP in vascular smooth muscle, and PDE5 inhibitors are used to treat erectile dysfunction and pulmonary hypertension. However, these drugs do not seem efficient in heart failure with preserved ejection fraction. There is experimental evidence that these PDEs as well as other PDE families including PDE1, PDE2 and PDE9 may play important roles in cardiac diseases such as hypertrophy and heart failure. After a brief presentation of the cyclic nucleotide pathways in cardiac cells and the major characteristics of the PDE superfamily, this chapter will present their role in cyclic nucleotide compartmentation and the current use of PDE inhibitors in cardiac diseases together with the recent research progresses that could lead to a better exploitation of the therapeutic potential of these enzymes in the future

Survival Tests for Leptospira spp.

International audienceMeasuring viability is an important and necessary assessment in studying microorganisms. Several methods can be applied to Leptospira spp, each with advantages and inconveniencies. Here, we describe the traditional colony-forming unit method, together with two other methods based respectively on the reducing capacity of live cells (Alamar Blue Assay) and differential staining of live and dead cells (LIVE/DEAD BacLight). The Alamar Blue Assay uses the blue reagent resazurin, which can be reduced into the pink reagent resorufin by live cell oxidoreductases. Production of resorufin can be quantified by absorbance or fluorescence reading. The LIVE/DEAD BacLight assay uses a mixture of two nucleic acid dyes (Syto9 and propidium iodide) that differentially penetrate and stain nucleic acid of cells with decreased membrane integrity. The colony-forming unit method is labor-intensive but the most sensitive and linear method. The two other methods are not laborious and well-adapted to high throughput studies but the range of detection and linearity is limited