Alex Veithen

Stroma Cell-Derived Factor-1α Signaling Enhances Calcium Transients and Beating Frequency in Rat Neonatal Cardiomyocytes

Stroma cell-derived factor-1α (SDF-1α) is a cardioprotective chemokine, acting through its G-protein coupled receptor CXCR4. In experimental acute myocardial infarction, administration of SDF-1α induces an early improvement of systolic function which is difficult to explain solely by an anti-apoptotic and angiogenic effect. We wondered whether SDF-1α signaling might have direct effects on calcium transients and beating frequency. Primary rat neonatal cardiomyocytes were culture-expanded and characterized by immunofluorescence staining. Calcium sparks were studied by fluorescence microscopy after calcium loading with the Fluo-4 acetoxymethyl ester sensor. The cardiomyocyte enriched cellular suspension expressed troponin I and CXCR4 but was vimentin negative. Addition of SDF-1α in the medium increased cytoplasmic calcium release. The calcium response was completely abolished by using a neutralizing anti-CXCR4 antibody and partially suppressed and delayed by preincubation with an inositol triphosphate receptor (IP3R) blocker, but not with a ryanodine receptor (RyR) antagonist. Calcium fluxes induced by caffeine, a RyR agonist, were decreased by an IP3R blocker. Treatment with forskolin or SDF-1α increased cardiomyocyte beating frequency and their effects were additive. In vivo, treatment with SDF-1α increased left ventricular dP/dtmax. These results suggest that in rat neonatal cardiomyocytes, the SDF-1α/CXCR4 signaling increases calcium transients in an IP3-gated fashion leading to a positive chronotropic and inotropic effect.

The human bitter taste receptor T2R38 is broadly tuned for bacterial compounds.

T2R38 has been shown to be a specific bacterial detector implicated in innate immune defense mechanism of human upper airway. Several clinical studies have demonstrated that this receptor is associated with the development of chronic rhinosinusitis (CRS). T2R38 was previously reported to bind to homoserine lactones (HSL), quorum sensing molecules specific of Pseudomonas Aeruginosa and other gram negative species. Nevertheless, these bacteria are not the major pathogens found in CRS. Here we report on the identification of bacterial metabolites acting as new agonists of T2R38 based on a single cell calcium imaging study. Two quorum sensing molecules (Agr D1 thiolactone from Staphylococcus Aureus and CSP-1 from Streptococcus Pneumoniae) and a list of 32 bacterial metabolites from pathogens frequently implicated in CRS were tested. First, we observed that HSL failed to activate T2R38 in our experimental system, but that the dimethylsulfoxide (DMSO), used as a solvent for these lactones may, by itself, account for the agonistic effect previously described. Secondly, we showed that both Agr D1 thiolactone and CSP-1 are inactive but that at least 7 bacterial metabolites (acetone, 2-butanone, 2-pentanone, 2-methylpropanal, dimethyl disulfide, methylmercaptan, γ-butyrolactone) are able to specifically trigger this receptor. T2R38 is thus much more broadly tuned for bacterial compounds than previously thought.

Deorphanization and characterization of human olfactory receptors in heterologous cells.

Olfaction plays an indispensable role in human and animals in self and environmental recognition, as well as intra‐ and interspecific communication. Following the discovery of a family of olfactory receptors (ORs) by Buck and Axel in 1991, it has been established that the sense of smell begins with the molecular recognition of a chemical odorant by one or more ORs expressed in the olfactory sensory neurons. Therefore, characterization of the molecular interactions between odorant molecules and ORs is a key step in the elucidation of the general properties of the olfactory system and in the development of applications, i.e., design of new odorants, search for blockers, etc. The process putted in place at ChemCom to improve the expression of ORs at the cytoplasmic membrane of the HEK293 cell and assays enabling large‐scale deorphanization, and to characterize the interaction between chemical odorants and ORs is described. The family of human ORs includes ca. 400 putatively functional ORs which are GPCRs (G protein‐coupled receptors); to date over 100 human ORs have been deorphanized.

High Throughput Receptor Screening Assays

The recent technical advances in functional expression of olfactory receptors (OR s) make full deorphanization of human OR repertoire a realistic objective. Such a global knowledge of the precise mechanisms of odorant/receptor pairings will represent a crucial step for the development of an accurate model of how human nose perceives its chemical environment. Beyond its interest for basic science, it will also lead to the development of industrial applications such as receptor-based odorant design, development of selective odor blockers or enhancers and represents therefore an interesting opportunity for players active in the field of flavors and fragrances. Here, we will describe and discuss a high-throughput screening approach that aims at the objective of human OR mass deorphanization. However, the completion of this ambitious task is not a prerequisite to the development of commercial applications. With the expanding number of deorphanized ORs, structure–activity relationship studies on OR responding to an odorant of interest has already started. Likewise, the use of the screening approach to identify either blockers for malodor-responding ORs or positive enhancers of fine fragrance-tuned ORs is underway. These different aspects will also be discussed. Finally, beyond the human ORs, other classes of human chemoreceptors for volatiles as well as animal chemoreceptors may also represent industrial opportunities that will be briefly reviewed.