Fig. 4
Effect of TRPV4 antagonist on calpain 1–NLRP3/cas-1–GSDMD pathway in the hippocampus of PISE mice. A and C. Administration of TRPV4 antagonist HC-067047 increased inactive/total calpain 1 ratio in the hippocampus of control (A, Mann–Whitney U test, P < 0.01) and PISE mice (C, independent-samples t test, t = − 17.09, P < 0.01). B and D. Administration of HC-067047 decreased NLRP3 (B, HC-067047: independent-samples t test, t = 6.49, P < 0.01; D, PISE + HC-067047: Mann–Whitney U test, P < 0.01), c-cas-1 (B, HC-067047: independent-samples t test, t = 6.53, P < 0.01; D, PISE + HC-067047: independent-samples t test, t = 9.38, P < 0.01), IL-1β (B, HC-067047: independent-samples t test, t = 4.23, P < 0.01; D, PISE + HC-067047: independent-samples t test, t = 12.31, P < 0.01), and N-GSDMD protein levels (B, HC-067047: independent-samples t test, t = 5.28, P < 0.01; D, PISE + HC-067047: Mann–Whitney U test, P < 0.01). E and F. Administration of HC-067047 decreased GSDMD+ cells (red arrow) numbers in the hippocampus of control (E-i, HC-067047: independent-samples t test, t = 4.34, P < 0.01) and PISE mice (F-i, PISE + HC-067047: Mann–Whitney U test, P < 0.01; white star). GSDMD+/NeuN+ (white arrow), GSDMD+/GFAP+ (white star) and GSDMD+/Iba-1+ (white triangle) cells in the hippocampus in control (E-ii, E-iii and E-iv) and PISE mice (F-ii, F-iii and F-iv) treated with either vehicle or HC-067047. G. HC-067047 treatment increased the numbers of surviving neurons in the hippocampal CA1 (G-i, independent-samples t test, t = 5.14, P < 0.01; G-ii, independent-samples t test, t = 10.12, P < 0.01) and CA2/3 (G-i, independent-samples t test, t = 2.35, P = 0.03; G-ii, independent-samples t test, t = 14.20, P < 0.01) area of control (G-i) and PISE mice (G-ii). Scale = 50 μm. ×4 objective for the top row in E-ii, E-iii, E-vi, F-ii, F-iii, F-vi; ×10 objective for E-i-DAPI, E-i-GSDMD, E-i-Merge, F-i-DAPI, F-i-GSDMD, F-i-Merge, and the middle columns in G-i and G-ii; ×20 objective for E-i-enlarge, F-i-enlarge, the down row in E-ii, E-iii, E-vi, F-ii, F-iii and F-vi; ×40 objective for the left and right columns in G-i, and G-ii. <P < 0.05, <<P < 0.01 vs. control (icv.), &&P < 0.01 vs. PISE + vehicle (icv.)