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Models of cardiac repair are needed to understand mechanisms underlying failure to regenerate in human cardiac tissue. Such studies are currently dominated by the use of zebrafish and mice. Remarkably, it is between these two evolutionary separated species that the adult cardiac regenerative capacity is thought to be lost, but causes of this difference remain largely unknown. Amphibians, evolutionary positioned between these two models, are of particular interest to help fill this lack of knowledge. We thus developed an endoscopy-based resection method to explore the consequences of cardiac injury in adult Xenopus laevis. This method allowed in situ live heart observation, standardised tissue amputation size and reproducibility. During the first week following amputation, gene expression of cell proliferation markers remained unchanged, whereas those relating to sarcomere organisation decreased and markers of inflammation, fibrosis and hypertrophy increased. One-month post-amputation, fibrosis and hypertrophy were evident at the injury site, persisting through 11 months. Moreover, cardiomyocytesarcomere organisation deteriorated early following amputation, and was not completely recovered as far as 11 months later. We conclude that the adult Xenopus heart is unable to regenerate, displaying cellular and molecular marks of scarring. Our work suggests that, contrary to urodeles and teleosts, with the exception of medaka, adult anurans share a cardiac injury outcome similar to adult mammals. This observation is at odds with current hypotheses that link loss of cardiac regenerative capacity with acquisition of homeothermy.
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???displayArticle.pmcLink???PMC5344503 ???displayArticle.link???PLoS One
Fig 1. Ventricular resection endoscopic biopsy and animal survival in adult Xenopus laevis.
(A-C) Key steps of the heart ventricular biopsy procedure using an endoscope. After an abdominal incision, the endoscope is inserted into the pleuroperitoneal cavity (A), using a xenon light source and light cable (A’) to search for the heart and observe the procedure internally. Biopsy forceps are used to break the falciform ligament, open the pericardial sac (B), then collect a calibrated piece of cardiac tissue from the apical region of the ventricle (see also S1 File). The surgery is ended (C) by performing a single suture to close the incision. (D-E) Comparison of a control non-operated heart (D) with an operated heart, displaying a large blood clot 1 day post-amputation (E), the size of biopsy tissue (E’) collected corresponds to approximately 4% of the ventricle. (F) Graphical representation of survival after the procedure, detailing total number of animals used for the full amputation procedure (n = 34), and animals ethically sacrificed (n = 2) or experimentally sacrificed at the end of the planned protocol (n = 32). Scale bars, 2 mm (D, E and E’).
Fig 2. Long-term monitoring reveals persisting fibrotic scar in adult frog hearts after cardiac amputation.
Top: Schematic timing of heart collection following endoscopic cardiac biopsy in adult frogs from 1 dpa up to 11 mpa. (A-D) Histology of adult heart after picrosirius (PSR) staining to label cytoplasm (i.e. cardiomyocytes) in blue and fibrous matrix (i.e. pericardium, epicardium and endocardium as well as fibrotic scar tissue) in red, for a control non-operated heart (A, CTRL), compared to heart sections at one day (B, 1 dpa), one month (C, 1 mpa) and six months (D, 6 mpa) post amputation. Black arrows indicate amputation sites; faint blue staining formed a clot around the ventricle at 1 dpa; note the presence of a low to intense red-stained scar at the site of amputation for 1 mpa and 6 mpa respectively. (EE’-LL’) Magnifications of sections stained with PSR and adjacent sections immuno-labelled for tropomyosin (CH1, red), fibronectin (fn, green) and counterstained with DAPI (nuclei, blue), for control (E-E’), and operated hearts at 1 dpa (F-F’), 7 dpa (G-G’), 1 mpa (H-H’), 2 mpa (I-I’), 3 mpa (J-J’), 6 mpa (K-K’), and 11 mpa (L-L’). PRS labelling allowed the observation of scars on each amputated section, the red labelling displaying an increase of intensity with time (E to L): a low and diffuse red staining was detectable at 1 dpa and 7 dpa around the site of amputation (*) and intensifies from 1 mpa to 11 mpa. Likewise, fibronectin immunolabelling shares a similar pattern as PSR with an increasing accumulation at the site of amputation from 1 mpa to 11 mpa (E’ to L’). White arrows: pe, pericardium; ep, epicardium. Animals: CTRL, n = 2; 1dpa, n = 1; 7dpa, n = 2; 1mpa, n = 3; 2mpa, n = 3; 3mpa, n = 2; 6mpa, n = 2, 11mpa, n = 2. Scale bars, 1 mm (A–D, E-L and E’-L’).
Fig 4. Increased cardiomyocyte size persists eleven months post amputation.
(A) Using the WGA labelled heart pictures, the cell-membrane labelled boundaries of orthogonal view cardiomyocytes (left panel) was delineated in red (middle panel) and filled with blue colour (right panel) for non-amputated control hearts (CTRL) and different time-points post-amputation (7 dpa, 1, 3, 6 and 11 mpa). (B) The cross-sectional area of each cell was automatically calculated using ImageJ software. Quantification was performed on 2 or 3 independent heart sections, corresponding to minima of 300 cell areas counted for each group, and samples were compared to the CTRL. Animals: CTRL, n = 1; 7dpa, n = 1; 1mpa, n = 1; 3mpa, n = 1; 6mpa, n = 1, 11mpa, n = 1. An unpaired non-parametric t-test (Mann Whitney) was performed: ****, p<0.0001.
Fig 8. Endoscopic ventricle biopsy procedure in adult frogs induces a fibrous scar with absence of heart regeneration.
Endoscopic procedure in anesthetised frog allows “in situ†live visualisation of the operating field and collection ventricletissue in the apical region using biopsy forceps. The resulting outcome for the frog heart is the induction of scar related gene expression during the first week post-amputation (SHORT TERM) and the establishment of a persistent fibrous scar, cardiac hypertrophy and sarcomere disorganisation at the amputation site, without the capacity for the cardiac tissue to regenerate, even after almost one year (LONG TERM)
Andersen,
Persistent scarring and dilated cardiomyopathy suggest incomplete regeneration of the apex resected neonatal mouse myocardium--A 180 days follow up study.
2016, Pubmed
Andersen,
Persistent scarring and dilated cardiomyopathy suggest incomplete regeneration of the apex resected neonatal mouse myocardium--A 180 days follow up study.
2016,
Pubmed
Andersen,
Normalization of real-time quantitative reverse transcription-PCR data: a model-based variance estimation approach to identify genes suited for normalization, applied to bladder and colon cancer data sets.
2004,
Pubmed
Andersen,
Do neonatal mouse hearts regenerate following heart apex resection?
2014,
Pubmed
Bao,
ACTL6a enforces the epidermal progenitor state by suppressing SWI/SNF-dependent induction of KLF4.
2013,
Pubmed
Becker,
Regeneration of the ventricular myocardium in amphibians.
1974,
Pubmed
Becker,
Human cardiomyopathy mutations induce myocyte hyperplasia and activate hypertrophic pathways during cardiogenesis in zebrafish.
2011,
Pubmed
Bersell,
Neuregulin1/ErbB4 signaling induces cardiomyocyte proliferation and repair of heart injury.
2009,
Pubmed
Boppart,
Noninvasive assessment of the developing Xenopus cardiovascular system using optical coherence tomography.
1997,
Pubmed
,
Xenbase
Borchardt,
Cardiovascular regeneration in non-mammalian model systems: what are the differences between newts and man?
2007,
Pubmed
Bryant,
A systematic analysis of neonatal mouse heart regeneration after apical resection.
2015,
Pubmed
Burchfield,
Pathological ventricular remodeling: mechanisms: part 1 of 2.
2013,
Pubmed
Burggren,
Amphibians as animal models for laboratory research in physiology.
2007,
Pubmed
,
Xenbase
Cano-Martínez,
Functional and structural regeneration in the axolotl heart (Ambystoma mexicanum) after partial ventricular amputation.
2010,
Pubmed
Chai,
Surgery in Amphibians.
2016,
Pubmed
Chai,
Endoscopy in Amphibians.
2015,
Pubmed
Chen,
Impaired cardiovascular function caused by different stressors elicits a common pathological and transcriptional response in zebrafish embryos.
2013,
Pubmed
Coen,
Caspase-9 regulates apoptosis/proliferation balance during metamorphic brain remodeling in Xenopus.
2007,
Pubmed
,
Xenbase
Dickover,
Zebrafish cardiac injury and regeneration models: a noninvasive and invasive in vivo model of cardiac regeneration.
2013,
Pubmed
Gamba,
Cardiac regeneration in model organisms.
2014,
Pubmed
Gardner,
Defining Akt actions in muscle differentiation.
2012,
Pubmed
Hein,
Advanced echocardiography in adult zebrafish reveals delayed recovery of heart function after myocardial cryoinjury.
2015,
Pubmed
Heinz-Taheny,
Cardiovascular physiology and diseases of amphibians.
2009,
Pubmed
Hempel,
A Matter of the Heart: The African Clawed Frog Xenopus as a Model for Studying Vertebrate Cardiogenesis and Congenital Heart Defects.
2016,
Pubmed
,
Xenbase
Hill,
Cardiac plasticity.
2008,
Pubmed
Holterhoff,
Sequence and expression of the zebrafish alpha-actinin gene family reveals conservation and diversification among vertebrates.
2009,
Pubmed
Huang,
Myofibrillogenesis in the developing zebrafish heart: A functional study of tnnt2.
2009,
Pubmed
Ito,
Differential reparative phenotypes between zebrafish and medaka after cardiac injury.
2014,
Pubmed
Jaworska-Wilczynska,
Pericardium: structure and function in health and disease.
2016,
Pubmed
Jaźwińska,
Regeneration versus scarring in vertebrate appendages and heart.
2016,
Pubmed
Jopling,
Zebrafish heart regeneration occurs by cardiomyocyte dedifferentiation and proliferation.
2010,
Pubmed
Karmon,
Biological causal links on physiological and evolutionary time scales.
2016,
Pubmed
Klocke,
Surgical animal models of heart failure related to coronary heart disease.
2007,
Pubmed
Konfino,
The type of injury dictates the mode of repair in neonatal and adult heart.
2015,
Pubmed
Krasteva,
The BAF53a subunit of SWI/SNF-like BAF complexes is essential for hemopoietic stem cell function.
2012,
Pubmed
Liu,
Research proceedings on amphibian model organisms.
2016,
Pubmed
,
Xenbase
Livak,
Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method.
2001,
Pubmed
MacArthur,
A gene for speed? The evolution and function of alpha-actinin-3.
2004,
Pubmed
Mercer,
A dynamic spatiotemporal extracellular matrix facilitates epicardial-mediated vertebrate heart regeneration.
2013,
Pubmed
Nakamura,
Expression analysis of Baf60c during heart regeneration in axolotls and neonatal mice.
2016,
Pubmed
Nishii,
Targeted disruption of the cardiac troponin T gene causes sarcomere disassembly and defects in heartbeat within the early mouse embryo.
2008,
Pubmed
Oberpriller,
Response of the adult newt ventricle to injury.
1974,
Pubmed
Polizzotti,
A cryoinjury model in neonatal mice for cardiac translational and regeneration research.
2016,
Pubmed
Porrello,
Transient regenerative potential of the neonatal mouse heart.
2011,
Pubmed
Porrello,
A neonatal blueprint for cardiac regeneration.
2014,
Pubmed
Poss,
Heart regeneration in zebrafish.
2002,
Pubmed
Poss,
Advances in understanding tissue regenerative capacity and mechanisms in animals.
2010,
Pubmed
RUMIANTSEV,
[Demonstration of regeneration of significant portions of the myocardial fibers of the frog after trauma].
1961,
Pubmed
Rumyantsev,
Post-injury DNA synthesis, mitosis and ultrastructural reorganization of adult frog cardiac myocytes. An electron microscopic-autoradiographic study.
1973,
Pubmed
Rumyantsev,
Interrelations of the proliferation and differentiation processes during cardiact myogenesis and regeneration.
1977,
Pubmed
Sallin,
Acute stress is detrimental to heart regeneration in zebrafish.
2016,
Pubmed
Sehnert,
Cardiac troponin T is essential in sarcomere assembly and cardiac contractility.
2002,
Pubmed
Seto,
Deficiency of α-actinin-3 is associated with increased susceptibility to contraction-induced damage and skeletal muscle remodeling.
2011,
Pubmed
Simon,
Assessing cardiomyocyte proliferative capacity in the newt heart and primary culture.
2015,
Pubmed
Sussman,
Myocardial AKT: the omnipresent nexus.
2011,
Pubmed
Szibor,
Remodeling and dedifferentiation of adult cardiomyocytes during disease and regeneration.
2014,
Pubmed
Teramoto,
Experimental pig model of old myocardial infarction with long survival leading to chronic left ventricular dysfunction and remodeling as evaluated by PET.
2011,
Pubmed
Uygur,
Mechanisms of Cardiac Regeneration.
2016,
Pubmed
Vargas-González,
[Myocardial regeneration in Ambystoma mexicanum after surgical injury].
2005,
Pubmed
Vivien,
Evolution, comparative biology and ontogeny of vertebrate heart regeneration.
2016,
Pubmed
Warkman,
Xenopus as a model system for vertebrate heart development.
2007,
Pubmed
,
Xenbase
Witman,
Recapitulation of developmental cardiogenesis governs the morphological and functional regeneration of adult newt hearts following injury.
2011,
Pubmed
Xiang,
Endogenous Mechanisms of Cardiac Regeneration.
2016,
Pubmed
Yang,
alpha-actinin-3 and performance.
2009,
Pubmed
Ye,
Primer-BLAST: a tool to design target-specific primers for polymerase chain reaction.
2012,
Pubmed
Zhou,
Akt1/protein kinase B enhances transcriptional reprogramming of fibroblasts to functional cardiomyocytes.
2015,
Pubmed
