Supplementary MaterialsSupplementary material mmc1. Unlike LysoPS, only 5 or 10?g injection of 2-deoxy-1-C3-pH- em p /em -O-C11-LysoPT induced dramatic hypothermia. The rank order matched to that observed for in vitro degranulation-stimulating activity (Fig. 4). Again, 2-deoxy-1-C3-pH- em p /em -O-C11-LysoPT was induced the greatest hypothermic action. Open in a separate window Fig. 4 Hypothermic effect of LysoPS analogs C57BL/6 mice were injected intravenously with 5?g (A) or 10?g (B) of 2-deoxy-1-C3-pH- em p /em -O-C11-LysoPT and related compounds (LysoPS (18:1), 2-deoxy-1-LysoPS (18:1), 2-deoxy-1-LysoPT (18:1) and 2-deoxy-1-C3-pH- em p /em -O-C11-LysoPS for comparison) at indicated dosage and rectal heat was monitored every 5 or 10?min. Data are representative of three experiments, each in triplicate. Each sign represents; closed circle (?) 2-deoxy-1-C3-pH- em p /em -O-C11-LysoPT, open circle () 2-deoxy-1-LysoPT (18:1), closed triangle () 2-deoxy-1-C3-pH- em p /em -O-C11-LysoPS, open triangle () 2-deoxy-1-LysoPS (18:1) and closed square () LysoPS (18:1). 3.4. 2-deoxy-1-C3-pH-p-O-C11-LysoPT did not activate LPS1-3 To examine if 2-deoxy-1-C3-pH- em p /em -O-C11-LysoPT activates LPS1C3, we used TGF shedding assay. While LysoPS activated all the three LysoPS receptors in TGF shedding assay, 2-deoxy-1-C3-pH- em p /em -O-C11-LysoPT didnt (Fig. 5). These results suggest that the putative LysoPS receptor on MCs is different from your cloned GPCR-type LysoPS receptors. Open in a separate windows Fig. 5 2-deoxy-1-C3-pH- em p /em -O-C11-LysoPT did not Gemcitabine HCl distributor activate LPS1, LPS2 and LPS3 (A, B) TGF shedding responses of HEK293A cells expressing LPS1 (A) and LPS2 (B) induced by LysoPS (18:1) (closed circles) or 2-deoxy-1-C3-pH- em p /em -O-C11-LysoPT (open circles). Data are representative of three experiments. Error bars are SD (standard deviation) for three assay replicates for one experiment. (C) TGF shedding response of HEK293FT cells expressing LPS3 to LysoPS (closed circles) or 2-deoxy-1-C3-pH- em p /em -O-C11-LysoPT (open circles). Data are representative of three experiments. Error bars are SD for three assay replicates for one experiment. Lysophosphatidylserine (LysoPS) is composed of four modules, i.e., a serine, a phosphate, a glycerol and a fatty acid, which are chemically linked by phosphodiester or ester bonds. We previously examined structure-activity relationship (SAR) of LysoPS by synthesizing a number of LysoPS analogs with modifications of the four modules, and by evaluating Gemcitabine HCl distributor them for both MC degranulation and activation of the cloned GPCR-type LysoPS receptors (LPS1, LPS2 and LPS3) , , . In the beginning our modification was focused exclusively on serine and glycerol modules . As a result, threonine was found to be superior to serine for MC degranulation. In this study, to identify a preferable fatty acid module, we used a new set of LysoPS analogs with modification in the fatty acid module . We first recognized C3-pH- em p /em -O-C11 Gemcitabine HCl distributor as the Gemcitabine HCl distributor fatty acid module that conferred the greatest MC degranulation activity (Fig. 2 and Table 1). Introducing the fatty acid surrogate into the structure of a potent ligand, 2-deoxy-1-LysoPT, made it possible to identify a super agonist, i.e., 2-deoxy-1-C3-pH- em p /em -O-C11-LysoPT. Indeed, the activities of the resulting LysoPS analogs showed 100 times higher than LysoPS (18:1) both in vitro (Fig. 3 and Table 2) and in vivo (Fig. 4). The SAR for MC degranulation obtained in this study was different from the SAR for the LysoPS receptors (LPS1, LPS2 and LPS3) . It should be noted that 2-deoxy-1-C3-pH em -o /em -O-C11-LysoPS, a poor inducer of MC degranulation (Fig. 2 and Table 1), Rabbit Polyclonal to MED24 was actually a potent agonist for cloned GPCR-type LysoPS receptors . We previously showed that 1-stearoyl (18:0)-LysoPS is a potent agonist for MC degranulation , while it was poor agonist for all the cloned GPCR-type LysoPS receptors. By contrast, LysoPS with unsaturated fatty acid such as oleic acid (18:1) and arachidonic acid (20:4) was preferable ligand for cloned GPCR-type LysoPS receptors. Thus, it is reasonable to assume Gemcitabine HCl distributor that the ligand recognizing pocket of putative MC LysoPS receptor is quite different from those of cloned GPCR-type LysoPS receptors. Taking account of the fact that the structure of C3-pH- em p /em -O-C11 is more linear than C3-pH- em o /em -O-C11 (Fig. 1), it.