[Ultrastructural and morphometrical analysis of apoptosis stages in cardiomyocytes of MDX mice]. 2001

V M Mikhaĭlov, and S A Komarov, and V K Nilova, and G I Shteĭn, and V S Baranov
Institute of Cytology RAS, St. Petersburg.

Our previous study of apoptosis in mdx mouse myocardium cells demonstrated the presence of middle-sized DNA fragments (60-65 kbp) in extracts of myocardium DNA, and irregular shape of membrane enveloped nuclei in cardiomyocytes. The DNA fragmentation (DNA laddering) was observed after biomechanical stress (5 min sweeming) only. Based on these results we concluded that the majority of cardiomyocytes were at the first stage of apoptosis. The purpose of this work was to provide some morphometrical quantitive characteristics of ultrastructural properties of the nuclei and mitochondria, and to determine morphological patterns of apoptosis in cardiomyocytes of mdx and C57B1 mice. To resolve the task, we made a morphometrical analysis of the electron microscope images of nuclei and mitochondria. First of all, we divided all nuclear images into three categories: normal, semi-pathological, and pathological forms according to the extent of nuclear membrane invaginations and that of condensed chromatin spreading. The most part of C57B1 cardiomyocyte nuclei belonged to the normal form (88.9 +/- 4.3%), while the smaller part (11.1 +/- 4.3%) was regarded as semi-pathological forms. Just a reverse was observed in mdx mice: the largest part of cardiomyocytes fell into category of semi-pathological (54.6 +/- 4.4%) and pathological (31.5 +/- 4.1%) forms while, the smallest part belonged to the normal form (13.8 +/- 3.0%). 24 h after biodynamic stress, the quantity of normal nuclei of C57B1 cardiomyocytes decreased to 61 +/- 5%, the number of semi-pathological nuclei increased to 39.0 +/- 4.4% (P < 0.05). The number of pathological nuclei of mdx, cardiomyocytes fell to 15.4 +/- 3.0% (P < 0.05). It means that mdx cardiomyocytes with pathological form of their nuclei disappear because of emerging the second, destructive stage of apoptosis. To estimate the degree of ultrastructural changes in the nuclei of all three forms of cardiomyocytes we counted the square/perimeter ratio in each nucleus (circle shape factor; CSF). The value of CSF for normal nuclei of all the forms of cardiomyocytes varied between 0.65 +/- 0.02 and 0.71 +/- 0.04. In semi-pathological and pathological nuclei a significant decrease in CSF to 0.56 +/- 0.02 and 0.56 +/- 0.03 was observed, respectively (P < 0.05). The biodynamical stress did not reduce the CSF value below this level. We also counted the ratio of the square to the product of a long and a short axes (ellipse shape factor; ESF). The ESF value for normal nuclei of all forms of cardiomyocytes varied between 0.97 +/- 0.01 and 0.99 +/- 0.01. In the case of mdx mice the biodynamical stress reduced ESF to 0.95 +/- 0.01 (P < 0.05) for pathological form of nuclei. The specific density of mitochondria in mdx cardiomyocytes (0.274 +/- 0.016) was less than that in C57B1 cardiomyocytes (0.329 +/- 0.018). At the destructive stage of apoptosis, the nuclei of cardiomyocytes were round in shape, the nuclear chromatin being hypercondensed, and mitochondria swollen. The cardiomyocyte morphology was in agreement with the definition of the final stage of apoptosis as secondary necrosis. Morphometrical results show that as many as 86-90% of nuclei of mdx cardiomyocytes have abnormal structure that confirms our conclusion that mdx cardiomyocytes were at the first stage of apoptosis. The final stage of apoptosis is rarely observed by biochemical or morphological methods. It suggests the presence of some inner mechanisms regulating the initiation of the final (destructive) stage of mdx cardiomyocyte apoptosis.

UI MeSH Term Description Entries
D008810 Mice, Inbred C57BL One of the first INBRED MOUSE STRAINS to be sequenced. This strain is commonly used as genetic background for transgenic mouse models. Refractory to many tumors, this strain is also preferred model for studying role of genetic variations in development of diseases. Mice, C57BL,Mouse, C57BL,Mouse, Inbred C57BL,C57BL Mice,C57BL Mice, Inbred,C57BL Mouse,C57BL Mouse, Inbred,Inbred C57BL Mice,Inbred C57BL Mouse
D008928 Mitochondria Semiautonomous, self-reproducing organelles that occur in the cytoplasm of all cells of most, but not all, eukaryotes. Each mitochondrion is surrounded by a double limiting membrane. The inner membrane is highly invaginated, and its projections are called cristae. Mitochondria are the sites of the reactions of oxidative phosphorylation, which result in the formation of ATP. They contain distinctive RIBOSOMES, transfer RNAs (RNA, TRANSFER); AMINO ACYL T RNA SYNTHETASES; and elongation and termination factors. Mitochondria depend upon genes within the nucleus of the cells in which they reside for many essential messenger RNAs (RNA, MESSENGER). Mitochondria are believed to have arisen from aerobic bacteria that established a symbiotic relationship with primitive protoeukaryotes. (King & Stansfield, A Dictionary of Genetics, 4th ed) Mitochondrial Contraction,Mitochondrion,Contraction, Mitochondrial,Contractions, Mitochondrial,Mitochondrial Contractions
D009206 Myocardium The muscle tissue of the HEART. It is composed of striated, involuntary muscle cells (MYOCYTES, CARDIAC) connected to form the contractile pump to generate blood flow. Muscle, Cardiac,Muscle, Heart,Cardiac Muscle,Myocardia,Cardiac Muscles,Heart Muscle,Heart Muscles,Muscles, Cardiac,Muscles, Heart
D009685 Nuclear Envelope The membrane system of the CELL NUCLEUS that surrounds the nucleoplasm. It consists of two concentric membranes separated by the perinuclear space. The structures of the envelope where it opens to the cytoplasm are called the nuclear pores (NUCLEAR PORE). Nuclear Membrane,Envelope, Nuclear,Envelopes, Nuclear,Membrane, Nuclear,Membranes, Nuclear,Nuclear Envelopes,Nuclear Membranes
D002467 Cell Nucleus Within a eukaryotic cell, a membrane-limited body which contains chromosomes and one or more nucleoli (CELL NUCLEOLUS). The nuclear membrane consists of a double unit-type membrane which is perforated by a number of pores; the outermost membrane is continuous with the ENDOPLASMIC RETICULUM. A cell may contain more than one nucleus. (From Singleton & Sainsbury, Dictionary of Microbiology and Molecular Biology, 2d ed) Cell Nuclei,Nuclei, Cell,Nucleus, Cell
D002843 Chromatin The material of CHROMOSOMES. It is a complex of DNA; HISTONES; and nonhistone proteins (CHROMOSOMAL PROTEINS, NON-HISTONE) found within the nucleus of a cell. Chromatins
D000818 Animals Unicellular or multicellular, heterotrophic organisms, that have sensation and the power of voluntary movement. Under the older five kingdom paradigm, Animalia was one of the kingdoms. Under the modern three domain model, Animalia represents one of the many groups in the domain EUKARYOTA. Animal,Metazoa,Animalia
D013312 Stress, Physiological The unfavorable effect of environmental factors (stressors) on the physiological functions of an organism. Prolonged unresolved physiological stress can affect HOMEOSTASIS of the organism, and may lead to damaging or pathological conditions. Biotic Stress,Metabolic Stress,Physiological Stress,Abiotic Stress,Abiotic Stress Reaction,Abiotic Stress Response,Biological Stress,Metabolic Stress Response,Physiological Stress Reaction,Physiological Stress Reactivity,Physiological Stress Response,Abiotic Stress Reactions,Abiotic Stress Responses,Abiotic Stresses,Biological Stresses,Biotic Stresses,Metabolic Stress Responses,Metabolic Stresses,Physiological Stress Reactions,Physiological Stress Responses,Physiological Stresses,Reaction, Abiotic Stress,Reactions, Abiotic Stress,Response, Abiotic Stress,Response, Metabolic Stress,Stress Reaction, Physiological,Stress Response, Metabolic,Stress Response, Physiological,Stress, Abiotic,Stress, Biological,Stress, Biotic,Stress, Metabolic
D017209 Apoptosis A regulated cell death mechanism characterized by distinctive morphologic changes in the nucleus and cytoplasm, including the endonucleolytic cleavage of genomic DNA, at regularly spaced, internucleosomal sites, i.e., DNA FRAGMENTATION. It is genetically programmed and serves as a balance to mitosis in regulating the size of animal tissues and in mediating pathologic processes associated with tumor growth. Apoptosis, Extrinsic Pathway,Apoptosis, Intrinsic Pathway,Caspase-Dependent Apoptosis,Classic Apoptosis,Classical Apoptosis,Programmed Cell Death,Programmed Cell Death, Type I,Apoptoses, Extrinsic Pathway,Apoptoses, Intrinsic Pathway,Apoptosis, Caspase-Dependent,Apoptosis, Classic,Apoptosis, Classical,Caspase Dependent Apoptosis,Cell Death, Programmed,Classic Apoptoses,Extrinsic Pathway Apoptoses,Extrinsic Pathway Apoptosis,Intrinsic Pathway Apoptoses,Intrinsic Pathway Apoptosis
D051379 Mice The common name for the genus Mus. Mice, House,Mus,Mus musculus,Mice, Laboratory,Mouse,Mouse, House,Mouse, Laboratory,Mouse, Swiss,Mus domesticus,Mus musculus domesticus,Swiss Mice,House Mice,House Mouse,Laboratory Mice,Laboratory Mouse,Mice, Swiss,Swiss Mouse,domesticus, Mus musculus

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