Cardiac transcriptional and metabolic changes following thoracotomy

Figures

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  Figure 1. Transcriptional patterns and gene enrichment analysis in cardiac tissue. (a) Heatmap of all significant genes 2 weeks (W), 4 weeks, and 6 weeks following isolated thoracotomy (ITH). Note the distinct pattern two weeks after surgery, which is not present any more 4 and 6 weeks after surgery. (b) Reactome and (c) KEGG pathway enrichment analysis two weeks after surgery shows significant downregulation of mitochondrial pathways involved in energy production and branched-chain amino acid metabolism. GO pathways enrichment analysis is reported in Table 1. Animal numbers at 2/4/6 weeks: ITH n = 7/8/10, control=5/5/5. Heatmaps were created using R and gplots9,10. Gene enrichment analysis was performed using ReactomePA and clusterProfiler12,13.

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  Figure 2. TEAD4, its targets and regulatory pathways. (a) promoter enrichment analysis showed an enrichment in the CACATTCTAT motif—a binding motif of TEAD4. (b) displays TEAD4 signaling and significant predicted (*and validated) microRNA binding. As most target genes of TEAD4 are downregulated in our dataset (c), altered upstream signaling, miR binding or inhibition of TEAD4 might have led to downregulation of mitochondrial pathways in the heart. Animal numbers at 2 weeks: ITH n = 7, control=5.

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  Figure 3. Cardiac gene expression measured by qPCR in a second cohort. Transcriptional changes of significantly deregulated genes targeted by TEAD4 measured in a second cohort 2 weeks after surgery. Expression levels are displayed relative to the mean expression of the control group. Mean and standard deviation are depicted. Key mitochondrial genes involved in the respiratory chain are downregulated 2 weeks after surgery. Animal numbers: control n = 6, ITH n = 7, TAC n = 7. See Table 2 for a direct comparison of microarray and qPCR data between the two cohorts.

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  Figure 4. MST1 and YAP1 protein expression and phosphorylation as well as oxidative phosphorylation following isolated thoracotomy. MST1 protein expression was notably reduced, while phosphorylation was significantly increased. YAP1 protein expression was also decreased and Ser127 and Ser397 phosphorylation (LATS1 phosphorylation sites) was significantly increased following isolated thoracotomy. Complexes I-III and V do not show any significant changes after thoracotomy, while decreased levels of complex IV were noted two weeks after ITH. For uncropped immunoblots see supplementary Fig. 1. ITH n = 7, controls n = 7. *p < 0.05, **p < 0.01, ***p < 0.001.

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  Figure 5. Transcriptional changes after ITH and TAC. Transcriptional changes noted after ITH are also seen 2 weeks (W) after TAC. The pattern disappears four weeks after TAC and reappears with decompensated heart failure 6 weeks after TAC. Interestingly, transcriptional changes do not lead to metabolite accumulation or depletion 2 weeks after ITH, while significant metabolic perturbations are noted following TAC. Animal numbers at 2/4/6 weeks: ITH n = 7/8/10, control=5/5/5, TAC n = 10/18/11. Heatmaps were created using R and gplots9,10.