Expression of protease-activated receptors in human peripheral monocytes and in monocyte-derived human macrophages. (A) Monocytes. RT-PCR analysis of PAR1, PAR2, and PAR3 mRNA expression in CD14⁺ monocytes. GAPDH was used for normalization. (B) Monocytes. Flow cytometric analysis of PAR1, PAR2, and PAR3 expression using specific mAb for PAR1 (SPAN12), PAR2 (SAM11), and polyclonal antibody for PAR3 (shaded peaks). Nonshaded peaks represent cells stained with isotype-matched control antibodies. (C) Monocyte lysates were prepared and subjected to Western blot analysis with the appropriate antibodies. (D) Macrophages. RT-PCR analysis of PAR1, PAR2, and PAR3 mRNA expression in macrophages differentiated from monocytes for 7 days in the presence of 15 ng/mL M-CSF. (E) Macrophages. Flow cytometric analysis of PAR1, PAR2, and PAR3 in human macrophages was performed as described in Figure 1B. CD14 and CD68 antigens were used as differentiation markers (shaded peaks). (F) Macrophage lysates were prepared and subjected to Western blot analysis with the appropriate antibodies. M indicates marker. Results of 1 of 6 independent experiments are shown.