Pathogenic Role of mTORC1 and mTORC2 in Pulmonary Hypertension
Haiyang Tang, Kang Wu, Jian Wang, Sujana Vinjamuri, Yali Gu, Shanshan Song, Ziyi Wang, Qian Zhang, Angela Balistrieri, Ramon J. Ayon, Franz Rischard, Rebecca Vanderpool, Jiwang Chen, Guofei Zhou, Ankit A. Desai, Stephen M. Black, Joe G.N. Garcia, Jason X.-J. Yuan and Ayako Makino
mTOR (mTORC1/mTORC2) in Smooth Muscle Cells and Pulmonary Hypertension
Smooth muscle (SM)-specific conditional and inducible knock-out (KO) of mTOR attenuates hypoxia-induced pulmonary hypertension in mTORSM−/− mice. (A) Schematic strategy for the generation of mTORSM−/− mice (a) and the timeline indicating the time for injection of tamoxifen (Tam) (to induce mTOR KO), hypoxic exposure (for inducing pulmonary hypertension) and experimental measurements (b). (B) Representative immunofluorescence images showing cell nuclei (4′,6′-diamidino-2-phenylindole [DAPI]; blue), smooth muscle cells (smooth muscle actin [SMA]; red), and mammalian target of rapamycin (mTOR; green) in the cross-section of small pulmonary artery (PA) in lung tissues from wild-type (WT) (mTOR-Oil) and mTORSM−/− (mTOR-Tam) mice (a). Summarized data (mean ± SE; n = 5 in each group) for DAPI, SMA, and mTOR fluorescence intensity are shown in panels b. It is noted that the mTOR (green) expression is almost abolished in the SMA-positive PA wall in mTOR-Tam mice but preserved in the mTOR-Oil mice. Student’s t-test (DAPI and SMA level) and Welch’s t-test (mTOR level), **p < 0.01 and ***p < 0.001 versus mTOR-Oil. (C) Representative record of right ventricular pressure (RVP) in WT and mTORSM−/− mice exposed to normoxia (room air, 21% oxygen) and hypoxia (10% oxygen for 3 weeks) (a). Summarized data (mean ± SE) showing the peak value of right ventricular systolic pressure (RVSP) (b) (Kruskal-Wallis test, p < 0.001) and the Fulton index (the ratio of weight of the right ventricle divided by weight of the left ventricle plus the septum [RV/(LV + S)]) (c) (Kruskal-Wallis test, p = 0.005) in WT and mTORSM−/− mice exposed to normoxia and hypoxia. Dunn test, *p < 0.05, ***p < 0.001 versus Normoxia-WT; ##p < 0.01 versus Hypoxia-WT. (D) Representative hematoxylin and eosin images (a) of the cross-section of small PA and summarized data (mean ± SE) (b) showing the PA wall thickness in WT and mTORSM−/− mice under normoxic and hypoxic conditions. Kruskal-Wallis test, p < 0.001; Dunn test, **p < 0.01, *p < 0.05 versus Normoxia-WT, ##p < 0.01 versus Hypoxia-WT. (E) Summarized data (mean ± SE) showing the number of red blood cells (RBC) (Kruskal-Wallis test, p = 0.04), hemoglobin concentration (HGB) (Kruskal-Wallis test, p = 0.01), and hematocrit percentage (HCT) (Kruskal-Wallis test, p = 0.01) in WT and mTORSM−/− mice exposed to normoxia and hypoxia. Analysis of variance, **p < 0.01, *p < 0.05 versus Normoxia-WT; #p < 0.05 versus Hypoxia-WT. The numbers of experiments (n) for each group are indicated in each bar.