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. 2018 Aug 23;3(16):e120398. doi: 10.1172/jci.insight.120398

Figure 2. RA signaling is active in adult ASM.

Figure 2

(A) PCR analysis of mASM showing the expression of RA receptors and RA-producing enzymes (Aldh1a1 and Aldh1a2). cDNA from E12.5 lung was used as a positive control (n = 3). (B) S-gal staining of the RA activity reporter RARE-lacZ lung reveals lacZ/β-gal activity in the ASM (black arrow) (n = 3). Scale bar: 10 μm. (C) Immunostaining for lacZ/β-gal and smooth muscle–specific marker α-SMA showing expression of lacZ/β-gal in ASM (white arrow) (n = 3). (D) lacZ expression is relatively reduced in VAD (compared with VAS) and BMS (compared with CTR) RARE-lacZ mASM, revealing downregulation of RA activity in mASM with VAD and BMS treatment (n = 6 per group). (E) Expression of RARB, a known direct transcriptional target of RA signaling, is reduced in BMS- and DEAB-treated hASM compared with hASM cultured in CTR medium, indicating suppression of RA activity in hASM cultured in RA-deficient conditions (n = 3). Data represent the mean ± SEM. Student’s t test was used to calculate the P value in D and 2-way ANOVA was used for statistical analysis in E (*P < 0.05). Means with different letters in E are statistically different with Bonferroni-corrected P < 0.05.