Effect of CysLT2 receptors on CysLT1 receptor–dependent proliferation. (A) LAD2 cells and hMCs were treated for 48 hours with lentivirus encoding the indicated shRNA or with empty vector with MOIs of 10 for each. FACS analysis of permeabilized cells showing effect of shRNA treatment on-target and off-target receptor expression. The net MFI for each protein is displayed for the experiment depicted. (B) Effect of shRNA treatments on surface expression of the indicated receptors detected on nonpermeabilized LAD2 cells stained with RB34 and 1B3 Abs. Results in panels A and B are representative of 3 experiments each. (C) FACS of mBMMCs from indicated genotypes using RB34 without or with permeabilization. The curves for the isotype control of the 2 genotypes (shaded) are superimposable. The net mean MFI for CysLT1 surface expression for 5 experiments is shown (right). (D) Proliferation of hMCs in response to SCF with or without LTD4 (500 nM) and effect of CysLT1 or CysLT2 receptor knockdowns. Results are the mean (± SEM) from 3 separate experiments using cells from different donors. (E) Proliferation of mBMMCs from C57BL/6 mice of the indicated genotypes in response to SCF with LTD4. Date were normalized in each experiment to the response of each genotype to SCF only, which is set at 100%. Note the complete blockade of the LTD4-mediated increment by MK571 (1 μM). Results are the mean (± SEM) from 4 separate experiments using cells from different animals.
