Sequential defects in cardiac lineage commitment and maturation cause hypoplastic left heart syndrome

bioRxiv (Cold Spring Harbor Laboratory)

Abstract

ABSTRACT Background Complex molecular programs in specific cell lineages govern human heart development. Hypoplastic left heart syndrome (HLHS) is the most common and severe manifestation within the spectrum of left ventricular outflow tract obstruction defects occurring in association with ventricular hypoplasia. The pathogenesis of HLHS is unknown, but hemodynamic disturbances are assumed to play a prominent role. Methods To identify perturbations in gene programs controlling ventricular muscle lineage development in HLHS, we performed: i) whole-exome sequencing of 87 HLHS parent-offspring trios, ii) nuclear transcriptomics of cardiomyocytes from ventricles of 4 patients with HLHS and 15 controls at different stages of heart development, iii) single cell RNA sequencing and iv) 3D modeling in iPSCs from 3 patients with HLHS and 3 controls. Results Gene set enrichment and protein network analyses of damaging de-novo mutations and dysregulated genes from ventricles of patients with HLHS suggested alterations in specific gene programs and cellular processes critical during fetal ventricular cardiogenesis, including cell-cycle and cardiomyocyte maturation. Single-cell and 3D modeling with iPSCs demonstrated intrinsic defects in the cell-cycle/UPR/autophagy hub resulting in disrupted differentiation of early cardiac progenitor lineages leading to defective cardiomyocyte-subtype differentiation/maturation in HLHS. Additionally, premature cell-cycle exit of ventricular cardiomyocytes from HLHS patients prevented normal tissue responses to developmental signals for growth leading to multinucleation/polyploidy, accumulation of DNA damage, and exacerbated apoptosis, all potential drivers of left ventricular hypoplasia in absence of hemodynamic cues. Conclusions Our results highlight that despite genetic heterogeneity in HLHS, many mutations converge on sequential cellular processes primarily driving cardiac myogenesis, suggesting novel therapeutic approaches.

Authors 46

1. Markus Krane

   German Centre for Cardiovascular Research · Deutsches Herzzentrum München · Technical University of Munich

   Affiliation as printed

   DZHK (German Centre for Cardiovascular Research) - partner site Munich Heart Alliance, Munich, Germany

   Department of Cardiovascular Surgery, Institute Insure, German Heart Center Munich, Technical University of Munich, Lazarettstrasse 36, 80636 Munich, Germany

2. Martina Dreßen

   Deutsches Herzzentrum München · Technical University of Munich

   Affiliation as printed

   Department of Cardiovascular Surgery, Institute Insure, German Heart Center Munich, Technical University of Munich, Lazarettstrasse 36, 80636 Munich, Germany

3. Gianluca Santamaria

   TUM Klinikum · Technical University of Munich

   Affiliation as printed

   First Department of Medicine, Cardiology, Klinikum rechts der Isar - Technical University of Munich, Ismaninger Stra(e 22, 81675 Munich, Germany

4. Ilaria My

   TUM Klinikum · Technical University of Munich

   Affiliation as printed

   First Department of Medicine, Cardiology, Klinikum rechts der Isar - Technical University of Munich, Ismaninger Stra(e 22, 81675 Munich, Germany

5. Christine M. Schneider

   TUM Klinikum · Technical University of Munich

   Affiliation as printed

   First Department of Medicine, Cardiology, Klinikum rechts der Isar - Technical University of Munich, Ismaninger Stra(e 22, 81675 Munich, Germany

6. Tatjana Dorn

   TUM Klinikum · Technical University of Munich

   Affiliation as printed

   First Department of Medicine, Cardiology, Klinikum rechts der Isar - Technical University of Munich, Ismaninger Stra(e 22, 81675 Munich, Germany

7. Svenja Laue

   TUM Klinikum · Technical University of Munich

   Affiliation as printed

   First Department of Medicine, Cardiology, Klinikum rechts der Isar - Technical University of Munich, Ismaninger Stra(e 22, 81675 Munich, Germany

8. Elisa Mastantuono

   TUM Klinikum · Helmholtz Munich · Institut für Humangenetik · Technical University of Munich

   Affiliation as printed

   First Department of Medicine, Cardiology, Klinikum rechts der Isar - Technical University of Munich, Ismaninger Stra(e 22, 81675 Munich, Germany

   Institute of Human Genetics, Klinikum rechts der Isar - Technical University of Munich, Ismaninger Strasse 22, 81675 Munich, Germany, and Helmholtz Zentrum München, Ingolstaedter Landstra(e 1, 85764 Neuherberg, Germany

   Institute of Human Genetics, Klinikum rechts der Isar - Technical University of Munich, Ismaninger Strasse 22, 81675 Munich, Germany, and Helmholtz Zentrum München, Ingolstaedter Landstra(e 1, 8576 Neuherberg, Germany

9. Riccardo Berutti

   TUM Klinikum · Helmholtz Munich · Institut für Humangenetik · Technical University of Munich

   Affiliation as printed

   Institute of Human Genetics, Klinikum rechts der Isar - Technical University of Munich, Ismaninger Strasse 22, 81675 Munich, Germany, and Helmholtz Zentrum München, Ingolstaedter Landstra(e 1, 85764 Neuherberg, Germany

   Institute of Human Genetics, Klinikum rechts der Isar - Technical University of Munich, Ismaninger Strasse 22, 81675 Munich, Germany, and Helmholtz Zentrum München, Ingolstaedter Landstra(e 1, 8576 Neuherberg, Germany

10. Hilansi Rawat

    TUM Klinikum · Technical University of Munich

    Affiliation as printed

    First Department of Medicine, Cardiology, Klinikum rechts der Isar - Technical University of Munich, Ismaninger Stra(e 22, 81675 Munich, Germany

11. Ralf Gilsbach

    Goethe University Frankfurt · University of Freiburg · German Centre for Cardiovascular Research

    Affiliation as printed

    DZHK (German Centre for Cardiovascular Research) - partner site RheinMain, Frankfurt am Main, Germany

    Institute for Cardiovascular Physiology, Goethe University, Frankfurt am Main, Germany

    Institute of Experimental and Clinical Pharmacology and Toxicology University of Freiburg Albertstrasse 25, 79104 Freiburg, Germany

    Institute of Experimental and Clinical Pharmacology and Toxicology University of Freiburg Albertstrasse 2, 79104 Freiburg, Germany

12. Pedro Schneider

    University of Freiburg

    Affiliation as printed

    Institute of Experimental and Clinical Pharmacology and Toxicology University of Freiburg Albertstrasse 25, 79104 Freiburg, Germany

    Institute of Experimental and Clinical Pharmacology and Toxicology University of Freiburg Albertstrasse 2, 79104 Freiburg, Germany

13. Harald Lahm

    Deutsches Herzzentrum München · Technical University of Munich

    Affiliation as printed

    Department of Cardiovascular Surgery, Institute Insure, German Heart Center Munich, Technical University of Munich, Lazarettstrasse 36, 80636 Munich, Germany

14. Sascha Schwarz

    Munich University of Applied Sciences

    Affiliation as printed

    Center for Applied Tissue Engineering and Regenerative Medicine (CANTER), Munich University of Applied Sciences, Lothstr. 34, 80335 Munich, Germany

    Center for Applied Tissue Engineering and Regenerative Medicine (CANTER), Munich University of Applied Sciences, Lothstr. 34, 0335 Munich, Germany

15. Stefanie A. Doppler

    Deutsches Herzzentrum München · Technical University of Munich

    Affiliation as printed

    Department of Cardiovascular Surgery, Institute Insure, German Heart Center Munich, Technical University of Munich, Lazarettstrasse 36, 80636 Munich, Germany

16. Sharon L. Paige

    Stanford Medicine · Stanford Cardiovascular Institute · Stanford University

    Affiliation as printed

    Cardiovascular Institute, Stanford University School of Medicine, Stanford, CA 94305, USA

    Cardiovascular Institute, Stanford University School of Medicine, Stanford, CA 4305, USA

17. Nazan Puluca

    Deutsches Herzzentrum München · Technical University of Munich

    Affiliation as printed

    Department of Cardiovascular Surgery, Institute Insure, German Heart Center Munich, Technical University of Munich, Lazarettstrasse 36, 80636 Munich, Germany

18. Sophia Doll

    Max Planck Institute of Biochemistry

    Affiliation as printed

    Proteomics and Signal Transduction, Max-Planck Institute of Biochemistry, 82152 Martinsried, Germany

19. Irina Neb

    Deutsches Herzzentrum München · Technical University of Munich

    Affiliation as printed

    Department of Cardiovascular Surgery, Institute Insure, German Heart Center Munich, Technical University of Munich, Lazarettstrasse 36, 80636 Munich, Germany

20. Thomas Brade

    TUM Klinikum · Technical University of Munich

    Affiliation as printed

    First Department of Medicine, Cardiology, Klinikum rechts der Isar - Technical University of Munich, Ismaninger Stra(e 22, 81675 Munich, Germany

21. Zhong Zhang

    Deutsches Herzzentrum München · Technical University of Munich

    Affiliation as printed

    Department of Cardiovascular Surgery, Institute Insure, German Heart Center Munich, Technical University of Munich, Lazarettstrasse 36, 80636 Munich, Germany

22. Claudia Abou‐Ajram

    Deutsches Herzzentrum München · Technical University of Munich

    Affiliation as printed

    Department of Cardiovascular Surgery, Institute Insure, German Heart Center Munich, Technical University of Munich, Lazarettstrasse 36, 80636 Munich, Germany

23. Bernd H. Northoff

    LMU Klinikum · Institut für Laboratoriumsmedizin · Ludwig-Maximilians-Universität München

    Affiliation as printed

    Institute of Laboratory Medicine, University Hospital, LMU Munich, Marchioninistra(e 15, 81377 Munich, Germany

24. Lesca Miriam Holdt

    LMU Klinikum · Institut für Laboratoriumsmedizin · Ludwig-Maximilians-Universität München

    Affiliation as printed

    Institute of Laboratory Medicine, University Hospital, LMU Munich, Marchioninistra(e 15, 81377 Munich, Germany

25. Stefanie Sudhop

    Munich University of Applied Sciences

    Affiliation as printed

    Center for Applied Tissue Engineering and Regenerative Medicine (CANTER), Munich University of Applied Sciences, Lothstr. 34, 80335 Munich, Germany

    Center for Applied Tissue Engineering and Regenerative Medicine (CANTER), Munich University of Applied Sciences, Lothstr. 34, 0335 Munich, Germany

26. Makoto Sahara

    Karolinska Institutet

    Affiliation as printed

    Department of Cell and Molecular Biology, Karolinska Institute, Stockholm 171 77, Sweden

27. Alexander Goedel

    TUM Klinikum · Technical University of Munich

    Affiliation as printed

    First Department of Medicine, Cardiology, Klinikum rechts der Isar - Technical University of Munich, Ismaninger Stra(e 22, 81675 Munich, Germany

28. Andreas Dendorfer

    German Centre for Cardiovascular Research · Institut für Chirurgische Forschung · Ludwig-Maximilians-Universität München

    Affiliation as printed

    DZHK (German Centre for Cardiovascular Research) - partner site Munich Heart Alliance, Munich, Germany

    Walter-Brendel-Centre of Experimental Medicine, University Hospital, LMU Munich, Marchioninistr. 27, 81377 Munich, Germany

    Walter-Brendel-Centre of Experimental Medicine, University Hospital, LMU Munich, Marchioninistr. 27, 877 Munich, Germany

29. Fleur V.Y. Tjong

    Amsterdam University Medical Centers · University of Amsterdam

    Affiliation as printed

    Department of Clinical and Experimental Cardiology, Amsterdam UMC, University of Amsterdam, Heart Centre, 1105 AZ Amsterdam, The Netherlands

30. Maria E. Rijlaarsdam Aachen Department of Pediatric Cardiology

    Leiden University Medical Center

    Affiliation as printed

    Department of Pediatric Cardiology, Leiden University Medical Center, Leiden, The Netherlands

31. Julie Cleuziou

    Deutsches Herzzentrum München · Technical University of Munich

    Affiliation as printed

    Department of Congenital and Paediatric Heart Surgery, German Heart Center Munich, Technical University of Munich, Lazarettstrasse 36, 80636 Munich, Germany

32. Nora Lang

    Deutsches Herzzentrum München · Technical University of Munich

    Affiliation as printed

    Department of Paediatric Cardiology and Congenital Heart Defects, German Heart Center Munich, Technical University of Munich, Lazarettstrasse 36, 80636 Munich, Germany

33. Christian Kupatt

    TUM Klinikum · German Centre for Cardiovascular Research · Technical University of Munich

    Affiliation as printed

    DZHK (German Centre for Cardiovascular Research) - partner site Munich Heart Alliance, Munich, Germany

    First Department of Medicine, Cardiology, Klinikum rechts der Isar - Technical University of Munich, Ismaninger Stra(e 22, 81675 Munich, Germany

34. Connie R. Bezzina

    Amsterdam University Medical Centers · University of Amsterdam

    Affiliation as printed

    Department of Clinical and Experimental Cardiology, Amsterdam UMC, University of Amsterdam, Heart Centre, 1105 AZ Amsterdam, The Netherlands

35. Rüdiger Lange

    German Centre for Cardiovascular Research · Deutsches Herzzentrum München · Technical University of Munich

    Affiliation as printed

    DZHK (German Centre for Cardiovascular Research) - partner site Munich Heart Alliance, Munich, Germany

    Department of Cardiovascular Surgery, Institute Insure, German Heart Center Munich, Technical University of Munich, Lazarettstrasse 36, 80636 Munich, Germany

36. Neil E. Bowles

    University of Utah

    Affiliation as printed

    Department of Pediatrics (Division of Cardiology), University of Utah School of Medicine, Salt Lake City, UT, USA

37. Matthias Mann

    Max Planck Institute of Biochemistry

    Affiliation as printed

    Proteomics and Signal Transduction, Max-Planck Institute of Biochemistry, 82152 Martinsried, Germany

38. Bruce D. Gelb

    Icahn School of Medicine at Mount Sinai

    Affiliation as printed

    The Mindich Child Health and Development Institute, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA

39. Lia Crotti

    IRCCS Istituto Auxologico Italiano · University of Milano-Bicocca

    Affiliation as printed

    Department of Medicine and Surgery, University of Milano-Bicocca, Milan, Italy

    Istituto Auxologico Italiano, IRCCS, Cardiomyopathies Unit, Department of Cardiovascular, Neural and Metabolic Sciences, San Luca Hospital, Milan, Italy

    Istituto Auxologico Italiano, IRCCS, Center for Cardiac Arrhythmias of Genetic Origin and Laboratory of Cardiovascular Genetics, 22 Via Pier Lombardo, Milan, Italy

40. Lutz Hein

    University of Freiburg

    Affiliation as printed

    BIOSS, Center for Biological Signaling Studies, University of Freiburg, 79104 Freiburg, Germany

    Institute of Experimental and Clinical Pharmacology and Toxicology University of Freiburg Albertstrasse 25, 79104 Freiburg, Germany

    Institute of Experimental and Clinical Pharmacology and Toxicology University of Freiburg Albertstrasse 2, 79104 Freiburg, Germany

41. Thomas Meitinger

    TUM Klinikum · Helmholtz Munich · German Centre for Cardiovascular Research · Institut für Humangenetik · Technical University of Munich

    Affiliation as printed

    DZHK (German Centre for Cardiovascular Research) - partner site Munich Heart Alliance, Munich, Germany

    Institute of Human Genetics, Klinikum rechts der Isar - Technical University of Munich, Ismaninger Strasse 22, 81675 Munich, Germany, and Helmholtz Zentrum München, Ingolstaedter Landstra(e 1, 85764 Neuherberg, Germany

    Institute of Human Genetics, Klinikum rechts der Isar - Technical University of Munich, Ismaninger Strasse 22, 81675 Munich, Germany, and Helmholtz Zentrum München, Ingolstaedter Landstra(e 1, 8576 Neuherberg, Germany

42. Sean M. Wu

    Stanford Medicine · Stanford Cardiovascular Institute · Stanford University

    Affiliation as printed

    Cardiovascular Institute, Stanford University School of Medicine, Stanford, CA 94305, USA

    Cardiovascular Institute, Stanford University School of Medicine, Stanford, CA 4305, USA

43. Daniel Sinnecker

    TUM Klinikum · German Centre for Cardiovascular Research · Technical University of Munich

    Affiliation as printed

    DZHK (German Centre for Cardiovascular Research) - partner site Munich Heart Alliance, Munich, Germany

    First Department of Medicine, Cardiology, Klinikum rechts der Isar - Technical University of Munich, Ismaninger Stra(e 22, 81675 Munich, Germany

44. Peter J. Gruber

    Yale University

    Affiliation as printed

    Department of Surgery, Yale University, New Haven, CT, USA

45. Karl‐Ludwig Laugwitz

    TUM Klinikum · German Centre for Cardiovascular Research · Technical University of Munich

    Affiliation as printed

    DZHK (German Centre for Cardiovascular Research) - partner site Munich Heart Alliance, Munich, Germany

    First Department of Medicine, Cardiology, Klinikum rechts der Isar - Technical University of Munich, Ismaninger Stra(e 22, 81675 Munich, Germany

46. Alessandra Moretti

    TUM Klinikum · German Centre for Cardiovascular Research · Technical University of Munich

    Affiliation as printed

    DZHK (German Centre for Cardiovascular Research) - partner site Munich Heart Alliance, Munich, Germany

    First Department of Medicine, Cardiology, Klinikum rechts der Isar - Technical University of Munich, Ismaninger Stra(e 22, 81675 Munich, Germany