Figure 4. TRPA1 Increases [Ca²⁺]_i and Cancer Cell Survival in Response to Carboplatin.

(A) Representative time courses of [Ca²⁺]_i changes in the indicated HCC1569 cells upon treatment with 10 μM carboplatin in the presence or absence of 10 μM AP-18.

(B) Δ[Ca²⁺]_i evoked by carboplatin in the indicated HCC1569 cells treated with or without 10 μM AP-18. Data are shown as means ± SEM (n = 22–61). ***p < 0.001 (one-way ANOVA).

(C and D) Δ[Ca²⁺]_i evoked by 10 μM carboplatin in the indicated lung cancer (C) and MPNST cells (D) treated with or without 10 μM AP-18. Data are shown as means ± SEM (n = 12–129 for lung cancer, n = 30–72 for MPNST). *p < 0.05, **p < 0.01, and ***p < 0.001 (Student’s t test).

(E) Live cell numbers relative to carboplatin-untreated cells in the indicated HCC1569 cells treated with carboplatin for 72 hr. *p < 0.05 and ***p < 0.001 compared with shGFP (Student’s t test).

(F) Live cell numbers relative to carboplatin-untreated cells in the indicated HCC1569 cells treated with 10 μM carboplatin in the presence or absence of 10 μM AP-18 or 5 mM NAC (pH was adjusted to 7.4) for 72 hr. ***p < 0.001 (one-way ANOVA).

(G) Percentage of cell death in the indicated HCC1569 cells treated with 30 μM carboplatin in the presence or absence of 10 μM AP-18 or 5 mM NAC (pH was adjusted to 7.4) for 72 hr. ***p < 0.001 (one-way ANOVA).

(H) Live cell numbers relative to carboplatin-untreated cells in EL-12-58 cells treated with carboplatin in the presence or absence of 10 μM AP-18 for 72 hr. **p < 0.01 and ***p < 0.001 compared with vehicle control for AP-18 (AP-18 [–]) (Student’s t test).

(I) Percentage of cell death in EL-12-58 cells treated with or without 100 μM carboplatin for 72 hr in the presence or absence of 10 μM AP-18. ***p < 0.001 (Student’s t test).

(J and K) Live cell numbers relative to carboplatin-untreated cells in the indicated lung cancer (J) and MPNST cells (K) treated with carboplatin in the presence or absence of 10 μM AP-18 for 72 hr. *p < 0.05, **p < 0.01, and ***p < 0.001 compared with vehicle control for AP-18 (AP-18 [–]) (Student’s t test).

(L) Live cell numbers relative to carboplatin-untreated cells in the indicated HCC1569 cells treated with 10 μM carboplatin in the presence or absence of 10 μM BAPTA-AM for 72 hr. ***p < 0.001 (one-way ANOVA).

(M) Representative images of day-7 HCC1569 spheroids treated with or without 10 μM AP-18, 10 μM carboplatin, or both for 48 hr. The spheroids were stained with cleaved caspase-3 and DAPI. Scale bar, 50 μm.

(N) Percentage of spheroids with cleaved caspase-3 staining in the center. Data are shown as means ± SD from three independent experiments. ***p < 0.001 (one-way ANOVA).

In (E–L), data are shown as means ± SD from the sum of two or three independent experiments performed in duplicate. See also Figure S4.