Cardiomyocyte metabolic remodeling through the TBL1/TBLR1 co-factor complex

The heart relies on fatty acid oxidation to cover up to 70% of its energy requirements under physiological conditions, whereas the failing heart switches from fatty acid oxidation to glycolysis and downregulates key metabolic players such as PGC-1α. The mechanistic understanding of this switch during pathological remodeling and the associated regulatory pathway in physiological hypertrophy remains incomplete. A possible key regulator is the cofactor complex of transducin beta-like 1 (TBL1) and its heterodimerization partner TBL-related 1 (TBLR1), whose central role in the regulation of fatty acid oxidation and lipolysis has already been demonstrated in liver and white adipose tissue. Our preliminary data show an upregulation of the TBL1/TBLR1 complex in mice under pressure overload (transverse aortic constriction, TAC) and a downregulation under exercise conditions (swim training) indicating a regulatory effect in heart tissue as well. Therefore, the specific aim of this experimental study is to further characterize the role of TBL1/TBLR1 under baseline conditions in heart tissue.

In vivo experiments were conducted with double floxed TBL1/TBLR1 mice, which either received a Cre-expressing AAV9 to induce a sufficient TBL1/TBLR1 KO or a Luc-expressing AAV9 (control group) under the control of the TroponinT promotor. Heart function was characterized by echocardiography under baseline conditions and the heart tissue was analyzed by immunohistology (cardiomyocyte-cell-size and fibrosis). Typical metabolic targets and known gene expression patterns of cardiac hypertrophy were analyzed by qPCR and western blot (WB). Acquired in vivo results were confirmed in vitro using a siRNA mediated TBL1/TBLR1 KO in human induced pluripotent stem cell-derived cardiomyocytes (hi-PSC).

The TBL1/TBLR1 KO mice had an increased heart to body weight ratio and echocardiographic analysis showed hyperkinetic left ventricular ejection fraction, preserved stroke volume with reduced end-diastolic ventricular volume. Histological analysis of the obtained samples revealed increased cardiomyocyte cell size and fibrosis in the KO mice. WB data showed a reduced ratio of the activated form of AMP kinase (pAmpk) to AMP kinase (Ampk) in the knockout mice, indicating a shift in cardiac metabolism as described in failing hearts. Consistent with these findings, gene expression analysis by qPCR demonstrated a downregulation of PGC1α and other typical metabolic regulatory enzymes (HK2, pfk2, acadm, acadv). Furthermore, TBL1/TBLR1 KO induces a gene expression pattern of pathological hypertrophy (aMHC ↓, CITED4 ↓, PGC1α ↓, collagen 1/3 ↑, cEBPB1 ↑). In the hi-PSC model, we confirmed the downregulation of PGC1α and other typical metabolic regulatory enzymes as well as the marker for pathological hypertrophy by qPCR and the decreased pAmpk/Ampk ratio in WB.

In conclusion, these findings suggest that the TBL1/TBLR1 complex plays an important role in cardiac metabolism by attenuating the expression of genes mediating fatty acid oxidation in vivo and in vitro, suggesting energy deprivation in the cardiomyocyte. These changes correlate with more pathological remodeling, hypertrophy and fibrosis. Current experiments are investigating the effects of TBL1/TBLR1 knockdown under pressure overload (TAC model) to gain a comprehensive picture.