Bioenergetics, mitochondria, and cardiac myocyte differentiation

Abstract

Cardiac metabolism is finely tuned, and disruption of myocardial bioenergetics can be clinically devastating. Many cardiomyopathies that present early in life are due to disruption of the maturation of these metabolic pathways. However, this bioenergetic maturation begins well before birth, when the embryonic heart is first beginning to beat, and continues into the mature animal. Thus, the changes in energy production seen after birth are actually part of a continuum that coincides with the structural and functional changes that occur as the cardiac myocyte differentiates and the heart undergoes morphogenesis. Therefore, although bioenergetics and mitochondrial biology have not been studied in great detail in the developing heart, bioenergetic maturation should be considered an important component of normal myocyte differentiation.Although events occurring after birth will be discussed, this review will focus on the changes in bioenergetics and mitochondrial biology that coincide with myocyte differentiation and cardiac morphogenesis. The relationship of these changes to the etiology and presentation of cardiomyopathies will be used as a starting point for this discussion. Then, after reviewing cardiac development and mitochondrial biology, the published data on bioenergetics and mitochondrial structure and function in the developing heart will be presented. Finally, the case will be made that mitochondria may be critical regulators of cardiac myocyte differentiation and cardiac development.

Figure 1. Gross cardiac morphology at different stages of development

Brightfield images of E9.5, E13.5, E18.5, neonatal and adult mouse hearts from roughly the anterior/ventral aspect demonstrate the changes that occur in cardiac morphology during development. Cardiac structures are labeled: Ao—aorta, LA—left atrium, LV—left ventricle, OFT—outflow tract, PA—pulmonary artery, RA—right atrium, and RV—right ventricle. The inset in the lower right corner of the “Adult” heart is the E9.5 heart at the same scale to demonstrate their relative sizes.

Figure 2. Myocardial and myocyte structure during development

Histology (A, hematoxylin and eosin stained, paraffin-embedded sections) and electron micrographs (B, EM) of ventricular myocytes in early embryonic (E9.5), late embryonic (13.5), fetal (E16.5 for Histology, 18.5 for EM), neonatal, and adult hearts demonstrate changes that occur in the general structure of the myocardium and in the composition and structure of the differentiating myocyte. In the “Histology/Late Embryonic” panel, the upper portion represents trabecular myocardium with cross striations, while the lower portion represents the morphology of the compact myocardium. The cardiac lumen (L) is indicated where appropriate.

Figure 3. Mitochondrial structure and function

A. Mitochondria undergo fission and fusion to create a dynamic network that can be fragmented or interconnected, depending on the cell type and the conditions. This example from the same cell demonstrates mitochondrial fission induced by an increase in intracellular calcium. B. Individual mitochondria are complex structures that contain an outer and inner mitochondrial membrane (OMM, IMM), an intermembrane space, and a central matrix. They generate ATP, regulate and are regulated by intracellular calcium levels, and generate reactive oxygen species as well as antioxidants. In an active mitochondrion, electrons enter the electron transport chain (ETC) at complexes I and II and are passed through the Q cycle to complex III, then cytochrome c, and finally to complex IV, where they are used to reduce molecular oxygen to water. The ETC also pumps protons (H⁺) from the matrix to the intermembrane space, creating an electrochemical gradient (Δψ_m) that is used by complex V to generate ATP from ADP and inorganic phosphate (P_i). Δψ_m is represented by the negative charges in the mitochondrial matrix and is responsible for the influx of calcium (Ca²⁺) through various channels, whose composition is not completely known. Finally, complexes I and II are also responsible for the creation of superoxide anions (O₂^•−) that are converted by Mn-superoxide dismutase (SOD) to H₂O₂, which can diffuse into the sarcoplasm.

Figure 4. Mitochondrial network morphology in cultured myocytes

Fluorescence imaging of mitochondria in cardiac myocytes from E9.5, E13.5, and adult hearts demonstrates dramatic changes in the structure of the mitochondrial network as the heart develops. E9.5 myocytes have a fragmented network, while the mitochondria in E13.5 myocytes are more filamentous and spread throughout the cell. In contrast, the mitochondria network in the adult myocyte is highly structured, with mitochondria aligning in specific bands along the myofibrils and in the perinuclear and subsarcolemmal regions. Primary myocyte cultures were labeled with MitoTracker Green and imaged using epifluorescence (E9.5, E13.5) or confocal (Adult) microscopy.