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. 2009 Apr;23(4):1102–1114. doi: 10.1096/fj.08-117812

Figure 1.

Figure 1.

Industrial isocyanates and tear gases activate heterologously expressed TRPA1 channels. A) Chemical structures of environmental and occupational irritants MITC, MIC, and HDI. B) Structures of tear gas agents CN, CS, CR, benzyl bromide and bromoacetone (bromo-2-propanone), and PS. C) Dose-response curves of isocyanate-activated Ca2+ influx into hTRPA1-transfected HEK-293T cells. [Ca2+]i induced by each dose is represented as percentage of maximal [Ca2+]i elicited by a saturating dose of mustard oil (100 μM) applied 75 s later (baseline [Ca2+]i was subtracted). hTRPA1 was activated by MIC (EC50=25±3 μM, n=28±5 cells/dose, red squares) and HDI (EC50=2.6±0.7 μM, n=29±4 cells/dose, black circles). D) Dose-response analysis of tear gas agent-activated Ca2+ influx into hTRPA1-transfected HEK-293T cells. [Ca2+]i represented as in A. hTRPA1 was activated by CN (EC50=91±12 nM, n=49±1 cells/dose, green triangles), CS (EC50=7±1 nM, n=52±5, blue squares), CR (EC50=308±150 nM, n=70±9, red circles), PS (EC50=215±62 nM, n=32±4, gray diamonds), benzyl bromide (BenzBr, EC50=12±1 μM, n=66±17, black triangles) and bromoacetone (BrAc, EC50=1.1±0.3 μM, n=78±14 cells/dose, purple stars). E) Application of MIC (100 μM, purple bar) induces an increase of single-channel openings of an excised patch in the inside-out configuration from hTRPA1-overexpressing CHO cells. Voltage was held at −40 mV; bath solution contained 0.5 mM PPPi and was devoid of Ca2+ (10 mM EGTA). F) Application of CS (10 μM, blue bar) induces an increase of single-channel openings in an excised patch in the inside-out configuration from hTRPA1-overexpressing CHO cells. Conditions were same as for E. G) Responses of hTRPA1 mutant-expressing HEK-293T cells to 100 μM MIC, 100 μM HDI, 100 μM CS, 100 μM CN, 300 μM CR, 100 μM bromoacetone (BrAc), and 100 μM benzyl bromide (BenzBr). Increase in [Ca2+]i is displayed as percentage of [Ca2+]i activated by a saturating dose of carvacrol (300 μM). Mutants tested are coded as follows: K (gray) = K708 (MIC, n=84; HDI, n=53; CS, n=157; CN, n=108; CR, n=154; BrAc, n=44; BenzBr n=64 cells); 3C (white) = C619, C639, and C663 combined (MIC, n=34; HDI, n=47; CS, n=96; CN, n=56; CR, n=90; BrAc, n=25; BenzBr, n=40 cells); 3CK (black) = C619, C639, and C663; and K708 combined (MIC, n=30; HDI, n=123; CS, n=129; CN, n=18; CR, n=17; BrAc, n=38; BenzBr, n=41).

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