Overexpression of CD151 promotes cardiac hypertrophy and fibrosis.

(a) Representative gross morphologies of the hearts in mice subjected to different treatments. And the ratios of heart weight to body weight in mice with diverse treatments. (b) Echocardiography analysis of LVEF% and FS%. (c) Hemodynamic parameters were measured by the Millar cardiac catheter system. (d) H&E staining represented the areas of CMs (left). Scale bars, 50 μm. The areas of CMs were analyzed by Image Pro Plus (right). (e)Representative images of Sirius Red staining of the heart sections from mice with different treatments (left) and the quantification analysis of cardiac fibrosis (right). Scale bars, 50 μm. Sham+ NS (n = 8), TAC + NS (n = 6), TAC + rAAV2-ICAM2-GFP (n = 6), TAC + rAAV2-ICAM2-CD151 (n = 6), TAC + rAAV2-ICAM2-shCD151 (n = 8). Data are expressed as mean ± SEM.</p

The sequences of primers for mRNA detection.

BackgroundHeart failure (HF) is the last stage in the progression of various cardiovascular diseases. Although it is documented that CD151 contributes to regulate the myocardial infarction, the function of CD151 on HF and involved mechanisms are still unclear.Method and resultsIn the present study, we found that the recombinant adeno-associated virus (rAAV)-mediated endothelial cell-specific knockdown of CD151-transfected mice improved transverse aortic constriction (TAC)-induced cardiac function, attenuated myocardial hypertrophy and fibrosis, and increased coronary perfusion, whereas overexpression of the CD151 protein aggravated cardiac dysfunction and showed the opposite effects. In vitro, the cardiomyocytes hypertrophy induced by PE were significantly improved, while the proliferation and migration of cardiac fibroblasts (CFs) were significantly reduced, when co-cultured with the CD151-silenced endothelial cells (ECs). To further explore the mechanisms, the exosomes from the CD151-silenced ECs were taken by cardiomyocyte (CMs) and CFs, verified the intercellular communication. And the protective effects of CD151-silenced ECs were inhibited when exosome inhibitor (GW4869) was added. Additionally, a quantitative proteomics method was used to identify potential proteins in CD151-silenced EC exosomes. We found that the suppression of CD151 could regulate the PPAR signaling pathway via exosomes.ConclusionOur observations suggest that the downregulation of CD151 is an important positive regulator of cardiac function of heart failure, which can regulate exosome-stored proteins to play a role in the cellular interaction on the CMs and CFs. Modulating the exosome levels of ECs by reducing CD151 expression may offer novel therapeutic strategies and targets for HF treatment.</div

The expression of CD151 in heart tissue of mice.

BackgroundHeart failure (HF) is the last stage in the progression of various cardiovascular diseases. Although it is documented that CD151 contributes to regulate the myocardial infarction, the function of CD151 on HF and involved mechanisms are still unclear.Method and resultsIn the present study, we found that the recombinant adeno-associated virus (rAAV)-mediated endothelial cell-specific knockdown of CD151-transfected mice improved transverse aortic constriction (TAC)-induced cardiac function, attenuated myocardial hypertrophy and fibrosis, and increased coronary perfusion, whereas overexpression of the CD151 protein aggravated cardiac dysfunction and showed the opposite effects. In vitro, the cardiomyocytes hypertrophy induced by PE were significantly improved, while the proliferation and migration of cardiac fibroblasts (CFs) were significantly reduced, when co-cultured with the CD151-silenced endothelial cells (ECs). To further explore the mechanisms, the exosomes from the CD151-silenced ECs were taken by cardiomyocyte (CMs) and CFs, verified the intercellular communication. And the protective effects of CD151-silenced ECs were inhibited when exosome inhibitor (GW4869) was added. Additionally, a quantitative proteomics method was used to identify potential proteins in CD151-silenced EC exosomes. We found that the suppression of CD151 could regulate the PPAR signaling pathway via exosomes.ConclusionOur observations suggest that the downregulation of CD151 is an important positive regulator of cardiac function of heart failure, which can regulate exosome-stored proteins to play a role in the cellular interaction on the CMs and CFs. Modulating the exosome levels of ECs by reducing CD151 expression may offer novel therapeutic strategies and targets for HF treatment.</div

The raw blot/gel images.

BackgroundHeart failure (HF) is the last stage in the progression of various cardiovascular diseases. Although it is documented that CD151 contributes to regulate the myocardial infarction, the function of CD151 on HF and involved mechanisms are still unclear.Method and resultsIn the present study, we found that the recombinant adeno-associated virus (rAAV)-mediated endothelial cell-specific knockdown of CD151-transfected mice improved transverse aortic constriction (TAC)-induced cardiac function, attenuated myocardial hypertrophy and fibrosis, and increased coronary perfusion, whereas overexpression of the CD151 protein aggravated cardiac dysfunction and showed the opposite effects. In vitro, the cardiomyocytes hypertrophy induced by PE were significantly improved, while the proliferation and migration of cardiac fibroblasts (CFs) were significantly reduced, when co-cultured with the CD151-silenced endothelial cells (ECs). To further explore the mechanisms, the exosomes from the CD151-silenced ECs were taken by cardiomyocyte (CMs) and CFs, verified the intercellular communication. And the protective effects of CD151-silenced ECs were inhibited when exosome inhibitor (GW4869) was added. Additionally, a quantitative proteomics method was used to identify potential proteins in CD151-silenced EC exosomes. We found that the suppression of CD151 could regulate the PPAR signaling pathway via exosomes.ConclusionOur observations suggest that the downregulation of CD151 is an important positive regulator of cardiac function of heart failure, which can regulate exosome-stored proteins to play a role in the cellular interaction on the CMs and CFs. Modulating the exosome levels of ECs by reducing CD151 expression may offer novel therapeutic strategies and targets for HF treatment.</div

Exosomes are important molecules in the regulation of cardiomyocytes and cardiac fibroblasts by CD151.

(a) Relative mRNA expressions of ANP in CMs from the lower chamber with different treatments (n = 3). (b) Expression levels of markers of cardiac fibrosis in the CFs from the lower chamber with different treatments were measured by real-time PCR (n = 3). (c)exosome morphology was evaluated via electron microscopy (scale bar, 100 nm), (d) Immunoblotting for exosome markers like CD9 and CD63. (e) HUVECs-derived exosomes were labeled with the red fluorescent and then cocultured with CMs for 24 h. They were stained with F-actin to label cytoskeleton (in green) and exosomes (in red) to detect the number of recruited exosomes on CMs. (f) The cytoskeleton was labeled with F-actin stained (in red) and exosomes (in green) to detect the number of recruited exosomes on CFs. Data are presented as mean ± SEM.</p

Representative figure of LV M-mode echocardiographic tracings and pulse wave Doppler.

Representative figure of LV M-mode echocardiographic tracings and pulse wave Doppler.</p

Overexpression of CD151 increased cardiac hypertrophic and fibrosis response in vitro.

(a) Top well, ECs; lower chamber, CMs. (b) ECs in the top chamber were photographed by fluorescence microscope after being transfected with CD151. FITC staining of CMs in the lower chamber (right) and the quantitative analysis of cell sizes (left) (n = 3). Scale bar, 100 μm. (c) Top well, ECs; lower chamber, CFs. (d) ECs in the top chamber were photographed by fluorescence microscope after being transfected with CD151. Representative images of immunofluorescence staining for EdU (red), and Hoechst (blue) in CFs with different treatments (left). Scale bar, 100 μm. Quantitative analysis of EdU measured by Image J (right) (n = 3). CFs cardiac fibroblasts, CMs cardiomyocytes, TAC transverse aortic constriction, PE Phenylephrine; Data are presented as mean ± SEM.</p

Top 5 differentially expressed proteins (up and downregulated) in exosomes.

Top 5 differentially expressed proteins (up and downregulated) in exosomes.</p

Bioinformatics analysis of proteomic results.

(A). Hierarchical clustering analysis of the differentially expressed proteins in the two groups (si-CD151 vs si-NC) (n = 3) (B) The differentially expressed proteins analyzed by volcano plots between every two groups (si-CD151 vs si-NC) (C) Circus plots of GO Classification Annotation of differential protein. (D) KEGG pathways of differentially expressed proteins. OS: Organismal Systems; M: Metabolism; GIP: Genetic Information Processing; CP: Cellular Processes. (E) Western blot analysis of PPAR-α, PCG-1α and GAPDH) in CMs under PE, quantified by Image J (n = 4). (F) Western blot analysis of PPAR-γ and GAPDH) in CFs under Ang-II, quantified by Image J (n = 4). Data are expressed as mean ± SEM.</p

Silencing of CD151 inhibits cardiomyocyte hypertrophy and fibroblast proliferation.

(a) ECs in the top chamber were photographed by fluorescence microscope after being transfected with CD151 siRNA. FITC staining of CMs in the lower chamber (left) and the quantitative analysis of cell sizes (right) (n = 3). Scale bar, 100 μm. (b) ECs in the top chamber were photographed by fluorescence microscope after transfected with CD151 siRNA. Representative images of immunofluorescence staining for EdU (red), and Hoechst (blue) in CFs with different treatments (left). Scale bar, 100 μm. Quantitative analysis of EdU measured by Image J (right) (n = 3). CFs cardiac fibroblasts, CMs cardiomyocytes, TAC transverse aortic constriction, PE Phenylephrine; Data are presented as mean ± SEM.</p