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. 2022 Feb 11;21(3):e13566. doi: 10.1111/acel.13566

FIGURE 6.

FIGURE 6

Systematic TPM1 functions by phosphorylating PKA in aging retinas. (a) ELISA analysis of cAMP in BV2 cells after different treatments. Following exposure to DMSO, rTPM1, FSK, SQ, or FSK + SQ for 24 h, BV2 cells were collected to detect the protein level of cAMP. Data are presented as mean ± SEM and analyzed by one‐way ANOVA with Tukey's multiple comparison test (compared to FSK, *p < 0.05). Five independent experiments were performed. (b–e) Western blot analysis (b) and quantification of TPM1, p‐PKA, and PKA (c–e) in BV2 cells after different treatments. Data are presented as mean ±SEM and analyzed by one‐way ANOVA with Tukey's multiple comparison test (compared with rTPM1, *p < 0.05, **p < 0.01; compared with FSK, #p < 0.05, ##p < 0.01; SQ vs. FSK + SQ, $p < 0.05). Five independent experiments were performed. (f–g) Western blot analysis (f) and quantification of TPM1 (g) in BV2 cells after rTPM1 and H89 treatments. Data are presented as mean ± SEM and analyzed by one‐way ANOVA with Tukey's multiple comparison test (compared with rTPM1, **p < 0.01). (h) ELISA analysis of cAMP in young retinas after YMP or OMP treatment. Data are presented as mean ± SEM, n = 5 mice in each group. (i–n) Western blot analysis (i) and quantification of Adcy2, p‐PKA, PKA, FABP5, and TPM1 (j–n) in young retinas after YMP or OMP treatment. Data are presented as mean ± SEM, n = 4 mice in each group, one‐way ANOVA with Tukey's multiple comparison test. (o) ELISA analysis of cAMP in young retinas after rTPM1 protein or PBS treatment. Data are presented as mean ± SEM, n = 4 mice in each group. (p–u) Western blot analysis (p) and quantification of Adcy2, p‐PKA, PKA, FABP5, and TPM1 proteins (q–u) in young retinas after rTPM1 protein or PBS treatment. Data are presented as mean ± SEM, n = 5 mice in each group, unpaired two‐tailed Student's t test. (v–w) qPCR analysis of TPM1.5, TPM1.9, and TPM1.10 in young (v) or aged (w) retinas after YMP or OMP treatment. Data are presented as mean ± SEM, n = 4 mice in each group, one‐way ANOVA with Tukey's multiple comparison test