Mast cell activation occurs in sickle cell anemia. (A) ir pixels for tryptase, SP, and CGRP in the dorsal skin. *P < .05, **P < .01 vs corresponding HbAA-BERK (ANOVA, with Bonferroni). (B) Representative confocal images of dorsal skin sections showing mast cell–specific costaining for c-kit/CD117 (red), FcεRI (green), and tryptase (blue). (C) Skin mast cells in culture stained for c-kit/CD117 (red), FcεRI (green), and tryptase (blue). (D-F) Gene expression of c-kit/CD117 (D), FcεRI (E), and TLR-4 (F) in mast cells derived from skin, measured by reverse-transcription quantitative polymerase chain reaction (RT-qPCR), normalized to GAPDH mRNA, and shown relative to HbAA-BERK, which was given an arbitrary value of 1. ***P < .001 (Student t test). (G) Tryptase levels in the supernatant of skin-derived mast cells after incubation with vehicle (Veh) or SP for indicated time. Black bars, HbAA-BERK; red bars, HbSS-BERK. *P < .05, **P < .01 vs corresponding HbAA-BERK; #P < .05 vs vehicle HbSS-BERK (ANOVA, with Bonferroni). (H-I) Skin-derived mast cells incubated with imatinib mesylate (Imat, 10 μM) and/or morphine sulfate (MS, 1 μM) for 24 hours. Culture medium showing tryptase (H) and SP (I) levels. *P < .05, **P < .01, ***P < .001 vs corresponding HbAA-BERK, #P < .05, ##P < .01 vs HbSS-BERK vehicle, $P < .05, $$P < .01 vs HbAA-BERK vehicle (Student t test). Each value is the mean ± SEM of 5 mice of each type, and each image represents images from the skin or skin-derived mast cells from 5 different mice.