Here, the ligand binding, activation, and tissue distribution of the orphan G protein-coupled receptor (GPCR) GPR92 were studied. GPR92 binds and is activated by compounds based on the lysophosphatidic acid (LPA) backbone. The binding of LPA to GPR92 was of high affinity (K D ϭ 6.4 Ϯ 0.9 nM) and led to an increase in both phosphoinositide hydrolysis and cAMP production. GPR92 is atypical in that it has a low sequence homology with the classic LPA 1-3 receptors (21-22%). Expression of GPR92 is mainly found in heart, placenta, spleen, brain, lung, and gut. Notably, GPR92 is highly expressed in the lymphocyte compartment of the gastrointestinal tract. It is the most abundant GPCR activated by LPA found in the small intestinal intraepithelial CD8 ϩ cytotoxic T cells.

No abstract

A microfluidic system for long-term real-time monitoring of dynamic cellular events of immobilized human cells was investigated. The luciferase reporter gene activity in the reporter cell line HFF11, based on HeLa cells, was used as the model system. The cells were immobilized on silicon flow-through microchips and continuously supplied with a cell medium at 2 microL/min while maintaining the chip at 37 degrees C. The HFF11 cell line was designed for high-throughput screening of ligands for seven-transmembrane receptors. When a ligand binds, the receptor is activated and a cascade of intracellular reactions starts, ending with the synthesis of the reporter protein Photinus luciferase. The major goal was to develop a microfluidic system for continuous long-term assaying of the intracellular reporter gene activity in real time and determine the conditions, which could minimize cells stress and hence unspecific expression of the reporter gene. In the resulting microfluidic system and assay protocol, the cell microchip could be kept and assayed for a period up to 30 h. The developed system and data outcome was compared with a corresponding microtiter plate performed with the same cell line to highlight the advantages obtained in the microfluidic format.

No abstract

This study, for the first time monitoring CCR5 and CXCR4 ligand activation and HIV-1 interaction concomitantly, indicates that ligands and virus use different receptor epitopes which, in turn, vary between the two receptors. One particular chimera (FC-4b), having its junctional region close to the conserved cysteine in ECL2, functioned as coreceptor for both HIV-1BaL and HIV-1IIIB, but was not activated with RANTES or SDF-1beta. The results provide a basis for tailoring drugs that block viral entry through the two major coreceptors without interfering with their physiological function.