Figure 1. 5-HT induces rolling of human Eos and alters cell shape and morphology.

(A) Expression of 5-HT receptors by human Eos from allergic donors (D1-D5) by RT-PCR. Expression of β-actin is shown as the internal control. (B) Rolling of human Eos on rh VCAM-1-coated cover-slips under conditions of flow in vitro. Cells were treated with 5-HT or the 5-HT2A/2C receptor agonist DOI (both at 10 µM) for 5 min prior to infusion into the flow chamber. Cells treated with DMSO (vehicle) alone served as a control. (C) FSC versus side scatter (SSC) dot plots for untreated (control) and 5-HT (10 µM)- or C5a (1 nM, positive control)-treated human Eos after 1 min by GAFS assay. (D) Human Eos shape change measured as an increase in FSC after treatment with 5-HT or C5a for 10 min at 37°C measured. Results are expressed as percent change in mean FSC of stimulated cells compared to control cells. (E) Confocal microscopy of FITC-phalloidin stained human Eos initially allowed to attach to rh VCAM-1 on cover-slips for 20 min in PBS followed by an additional 5 min in the presence of 5-HT or DOI (both at 10 µM) or vehicle. Magnification ×600. Combined data (Mean ± SEM) of Eos from 4–6 donors is shown for B and D. *p<0.01 for 5-HT and <0.03 for DOI in B for comparison with DMSO-treated Eos and <0.01 in D for comparison with control Eos. Representative data of six independent experiments in C and three independent experiments in E with Eos from different donors is shown.