Transmembrane protein 117 knockdown protects against angiotensin-II-induced cardiac hypertrophy

Figures

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  Fig. 1. TMEM117 was increased in hypertrophic hearts and isolated NMVMs. A,B qRT-PCR and western blotting were used to determine the mRNA and protein expression levels of TMEM117 in control and hypertrophic mouse hearts, respectively. C,D The mRNA and protein levels of TMEM117 in NMVMS with PBS or Ang-II treatment were respectively detected. TMEM117, transmembrane protein 117; GAPDH, glyceraldehyde 3-phosphate dehydrogenase; PBS, phosphate-buffered saline; Ang-II, angiotensin-II. NMVMs, neonatal mouse cardiomyocytes. GAPDH was used as a loading control. Presented values are means ± SEM. N = 6–8/group. **P < 0.01 between the 2 indicated groups by two-tailed Student t test

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  Fig. 2. TMEM117 deficiency attenuated Ang-II-induced cardiac hypertrophy. A Histological analyses of heart sections stained with H&E, wheat germ agglutinin, Masson and picrosirius red from control mice and TMEM117 cKO mice after saline or Ang-II infusion. B The average cross-sectional area of cardiomyocytes was summarized after saline or Ang-II infusion in control and TMEM117 cKO mice. C,D The interstitial fibrosis and collagen deposition were quantified in the different mice, respectively. E HW/BW ratios were detected in the indicated groups. F Echocardiographic assessment of LVEDd, LVESd and FS was used to reflect cardiac function in control mice and TMEM117 cKO mice after saline or Ang-II infusion. G Representative immunofluorescence images of α-actinin staining in neonatal mice ventricular myocytes infected with siControl or siTMEM117 and treated with or without Ang-II for 48 h. H Pooled data from (G). I Western blotting was used to measure protein levels of ANP, BNP and β-MHC in the indicated groups. ANP atrial natriuretic polypeptide, BNP brain natriuretic polypeptide, β-MHC myosin chain heavy β, Col1 collagen type 1, Col3 collagen type 3, HW/BW heart weight/body weight, LVEDd left…

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  Fig. 3. TMEM117 overexpression in the heart deteriorated cardiac hypertrophy. A Heart sections were stained with H&E, wheat germ agglutinin, Masson and picrosirius red from control mice injected with Ad-EV or Ad-TMEM117 subsequently subjected to saline or Ang-II. B The average cross-sectional area of cardiomyocytes from the indicated groups was summarized. C,D The interstitial fibrosis and collagen deposition were quantified. E The rations of HW/BW in different mice were determined. F Echocardiographic assessment of LVEDd, LVESd and FS in the indicated groups. G Representative immunofluorescence images of α-actinin staining in neonatal mice ventricular myocytes infected with Ad-EV or Ad-TMEM117 and treated with or without Ang-II for 48 h. Scale bars = 10 μm. H Pooled data from (G). I Western blotting was used to measure protein levels of ANP, BNP and β-MHC in the indicated groups. Ad-EV, control adenovirus; Ad-TMEM117, recombinant adenovirus encoding TMEM117. All the data represent the means ± SEM. N = 6–8/group. *P < 0.05 and **P < 0.01. NS indicates no significance. Statistical significance was determined by 1-way ANOVA or 2-way ANOVA using Tukey post hoc test

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  Fig. 4. TMEM117 mediated cardiomyocyte hypertrophy by modulating oxidation states. A DHE staining was performed in NMVMs infected with siControl or siTMEM117 and treated with or without Ang-II. B NADPH oxidase activity was measured in the indicated groups. C–E Quantitative results of SOD activity and GSH, and plasma CAT levels in control or TMEM117 cKO mice treated with saline or Ang-II. F TMEM117 protein level was measured in control and hypertrophic myocardium with or without NAC. G Representative DHE staining in neonatal mice ventricular myocytes infected with Ad-EV or Ad-TMEM117 and treated with or without Ang-II. H NADPH oxidase activity was measured in the indicated groups. I–K Quantitative results of SOD activity and GSH, and CAT levels from control mice injected with Ad-EV or Ad-TMEM117 subsequently subjected to saline or Ang-II. Presented values are means ± SEM. N = 6–8/group. *P < 0.05 and **P < 0.01. NS indicates no significance. Statistical significance was determined by 1-way ANOVA or 2-way ANOVA using Tukey post hoc test

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  Fig. 5. TMEM117 deficiency attenuated cardiac hypertrophy-induced ERS in vivo. A NMVMs were isolated from cardiac tissue in Control and TMEM117 cKO mice and cultured in PBS and Ang-II medium. Representative western blot for p-PERK/PERK (B), e-IF2α (C) and ATF4 (D) in NMVMs. PERK, dsRNA-activated protein kinase like ER kinase; p-PERK phosphorylated dsRNA-activated protein kinase like ER kinase, eIF2α eukaryotic initiation factor 2α, ATF4 activating transcriptional factor. All the data represent the means ± SEM. N = 6–8/group. *P < 0.05 and **P < 0.01. NS indicates no significance. Statistical significance was determined by 1-way ANOVA or 2-way ANOVA using Tukey post hoc test

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  Fig. 6. TMEM117 deficiency attenuated mitochondrial disorder induced by cardiac hypertrophy. A Transmission electron microscopy was used to observe the mitochondrial morphology in control or TMEM117 cKO mice treated with saline or Ang-II. B–D Quantitative results of SDH activity, mitochondrial membrane potential and ATP production in NMVMs infected with siControl or siTMEM117 and treated with or without Ang-II. E Representative electron microscopy images from control mice injected with Ad-EV or Ad-TMEM117 subsequently subjected to saline or Ang-II. F–H Quantitative results of SDH activity, mitochondrial membrane potential and ATP production in NMVMs infected with Ad-EV or Ad-TMEM117 and treated with or without Ang-II. All the data represent the means ± SEM. N = 6–8/group. *P < 0.05 and **P < 0.01. NS indicates no significance. Statistical significance was determined by 1-way ANOVA or 2-way ANOVA using Tukey post hoc test