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Background and Aims: The fundamental mechanisms involved in the genesis and progression of heart failure are not clearly understood. The present study was conducted to analyze the cardiac mitochondrial involvement in heart failure, the possible parallelism between cardiac and skeletal muscle and if there is a link between clinical symptoms and mitochondrial damage., Methods: Left ventricle and pectoral biopsies were obtained from patients with heart failure (n: 21) and patients with inter-auricular communication as the unique diagnosis for surgery (n: 6). Mitochondria were isolated from these tissues and studied through electron microscopy, spectrophotometry to measure the activity of respiratory complex III and immunohistochemistry to determine the presence of reactive oxygen species., Results: More than 90% of cardiac and skeletal muscle mitochondria presented structural and functional alterations in relation to an increment in the reactive oxygen species production, even in patients without the presence of any clinical Framingham criteria., Conclusions: We demonstrated some parallelism between cardiac and skeletal muscle mitochondrial alterations in patients with heart failure and that these alterations begin before the major clinical Framingham criteria are installed, pointing to mitochondria as one of the possibly responsible factors for the evolution of cardiac disease., (Copyright © 2014 IMSS. Published by Elsevier Inc. All rights reserved.)

The authors describe the case of a 50-year-old man with chronic progressive external ophthalmoplegia (CPEO), diabetes mellitus (DM), and coronary artery disease. The patient had no cardiac conduction abnormalities. During coronary artery bypass surgery, his heart and two skeletal muscles were biopsied. All three muscles showed ragged red fibers. The heart muscle showed significant glycogen accumulation. Analysis of mitochondrial DNA (mtDNA) showed a 5019-base-pair deletion, with no duplications. There were morphologically abnormal mitochondria in all 3 muscles, with clinically apparent difference in preservation of function. The combination of diabetes mellitus and mtDNA deletion is fortuitous, as they can be causally linked. The cardiac pathology allows speculation about the possible adaptive processes that may occur in the heart in DM. There are few reported cases with CPEO and excess glycogen in the heart. Most show deposition of fat and poorer clinical outcomes as compared to those with glycogen deposition. This observation may lend support to the hypothesis that in the myocardium, adaptive responses are mediated via changes in glucose handling, whereas alterations in fat metabolism likely represent maladaptation.

The authors describe the case of a 50-year-old man with chronic progressive external ophthalmoplegia (CPEO), diabetes mellitus (DM), and coronary artery disease. The patient had no cardiac conduction abnormalities. During coronary artery bypass surgery, his heart and two skeletal muscles were biopsied. All three muscles showed ragged red fibers. The heart muscle showed significant glycogen accumulation. Analysis of mitochondrial DNA (mtDNA) showed a 5019-base-pair deletion, with no duplications. There were morphologically abnormal mitochondria in all 3 muscles, with clinically apparent difference in preservation of function. The combination of diabetes mellitus and mtDNA deletion is fortuitous, as they can be causally linked. The cardiac pathology allows speculation about the possible adaptive processes that may occur in the heart in DM. There are few reported cases with CPEO and excess glycogen in the heart. Most show deposition of fat and poorer clinical outcomes as compared to those with glycogen deposition. This observation may lend support to the hypothesis that in the myocardium, adaptive responses are mediated via changes in glucose handling, whereas alterations in fat metabolism likely represent maladaptation.

Background: Previous research has demonstrated that exercise intolerance in heart failure patients is associated with significant alterations in skeletal muscle ultrastructure and oxidative metabolism that may be more consequential than cardiac output., Design: To examine the effect of exercise training on skeletal muscle mitochondrial size in chronic heart failure patients., Methods: Six heart failure patients participated in 16-weeks of supervised upper and lower extremity exercise training. At the conclusion of training, percutaneous needle biopsies of the vastus lateralis were taken and electron microscopy was used to assess mitochondrial sizes., Results: The exercise programme resulted in a significant increase in peak maximal oxygen consumption ( P< 0.05) and anaerobic threshold (P < 0.04). Knee extension muscle force increased following training ( P< 0.02). After exercise training, the average size of the mitochondria increased by 23.4% (0.036 to 0.046 mu(2), P< 0.015) and the average shape was unaltered., Conclusion: Exercise training with heart failure patients alters skeletal muscle morphology by increasing mitochondrial size, with no change in shape. This may enhance oxidative metabolism resulting in an increased exercise tolerance.

We report the polarographic analysis of subsarcolemmal and intermyofibrillar mitochondria from rat skeletal and cardiac muscle, in order to detect possible biochemical and functional differences. Electron microscopic observation of isolated mitochondria showed normal aspect, with intact membranes. Respiratory control rate with different substrates and state 3 activity did not show differences between subsarcolemmal and intermyofibrillar mitochondria in skeletal muscle and heart.

We examined the ultrastructure of right and left ventricular myocardial biopsies obtained prior to cardiopulmonary bypass in fourteen patients undergoing total correction of tetralogy of Fallot (TOF). Twelve patients were undergoing primary one-stage repair and two patients had had a previous Blalock-Taussig shunt operation. The ages of the patients ranged from 3 to 35 years. In left ventricular (LV) myocytes, the sarcolemma often formed pouches filled with mitochondria and the muscle fibers showed a disoriented arrangement. In addition, the myofibrils showed hypercontraction and the Z-bands were thickened; the degree of the myofibril contraction was more severe than that in right ventricular (RV) myocytes. These ultrastructural findings are similar to those observed in myocytes under hypoxic conditions. The number of lysosomes in LV myocytes, but not in RV myocyte increased in the older patients. While we have not established a causal relationship between the ultrastructural findings and clinical features in our cases, our observations are consistent with the possibility that early total correction of TOF may be required to prevent the development of hypoxic changes and degeneration of heart muscle.

The effects of food restriction and dietary protein restriction on the oxygen consumption of three types of muscle in the rat were compared. Control rats were fed a diet containing 27% casein. Two groups of protein-restricted rats were fed isoenergy diets containing 15% or 8% casein. Two groups of food-restricted rats were fed either the 27% or 15% casein diets in amounts equivalent to 35% of the intake of energy. Oxygen consumption was measured in papillary muscle and in fibers from the red (high oxidative) and white (low oxidative) portions of the quadriceps with pyruvate and malate as substrates. Control values (micronlO2/g dry wt/min) were 35.2 + 1.5 (papillary), 20.7+/-1.9 (red), and 7.8+/-0.7 (white). Restriction of dietary protein content failed to alter the oxygen consumption of any of the muscle types. In contrast, involuntary food restriction,combined with a reduction in protein content, caused a reduction in the oxygen consumption of papilary and white skeletal muscle, but the metabolism of red skeletal muscle was not affected. No differences were found in the appearance and distribution of mitochondria in cardiac or red skeletal muscle after examination by electron microscopy, but a depletion of subsarcolemmal mitochondria was evident in white muscle from the food-restricted rats.

Clinical and pathologic findings are presented in 14 patients (six newly reported, eight described previously), all children ranging in age from 6 to 24 months, with a clinicopathologic syndrome termed "infantile cardiomyopathy with histiocytoid change in cardiac muscle cells." This syndrome is manifested clinically by severe, eventually fatal cardiac arrhythmias, and is characterized pathologically by cardiac hypertrophy and by a distinctive type of focal degeneration of the muscle cells, which lose their myofibrils, undergo marked mitochondrial hyperplasia, become rounded in shape and enlarged, and resemble histiocytes. Evidence is presented to support the conclusions that these manifestations are those of a cardiomyopathy, that cardiac hypertrophy precedes the onset of the clinical features, that the focal degeneration is likely to be a cause rather than a consequence of the arrhythmias, and that the latter develop only in the late stages of the disorder. The etiology of this cardiomyopathy remains unclear.

After a 24 hrs' preservation of rat hearts, in 22% the viability test with allogenic perfusion showed a singular recovery of the right ventricle (57 of 257 experiments). The ultrastructural aspects correlate with the differences of the function. The micrographs of the right myocardium show--except a mild edema--ultrastructural integrity, but the micrographs of the left myocardium show marked alterations of the ultrastructure: edema, swollen mitochondria with destroyed cristal membranes, swelling of the sacrotubular system and alterations of myofibrils.

Vesicular myocardial change is a specific and common finding in the diseased hearts of humans and experimental animals. The small to large vesicles are generally bound by a double membrane. They are formed within myocardial cells and then possibly extruded into the extracellular space where they disintegrate or are phagocytosed by mononuclear cells. On the basis of our studies, most vesicles appear to be of mitochonrial origin. In humans, vesicular myocardial change appears to be most extreme in primary cardiomyopathy. It may also be present, but in lesser degree, in the apparently normal hearts of human adults. Vesicular myocardial change probably represents a specific mechanism by which myocardial cells eliminate damage mitochondria or other undesirable elements that build within them as a result of disease, aging, and perhaps normal physiological activity.

The fetal mouse heart can be maintained in organ culture for days while it retains its capacity to beat rhythmically; however, degenerative changes eventually occur. In this study, the ultrastructure of the hearts fixed immediately after removal from the fetus without incubation often revealed some swelling of organelles,suggesting that early ischemic alterations may occur during the time required for removaland fixation. Within 2 days in organ culture, most of the cells reverted to normal, indicating that the injury was reversible. However, a few cells had undergone necrosis.An interesting finding in some cells was the presence of autophagic vacuoles containing damaged mitochondria and occasionally,myrofibrils. These vacuoles were usually surrounded by a single layered membrane, but occasionally a double membrane was present. The occurrence of these vacuoles in an otherwise normal appearing cell suggests that focal cytoplasmic injury had occurred and the damaged elements were being sequestered anddigested within the vacuoles. An alternative possibility is that these cells were adjusting their cytoplasmic contents to a new level of homostasis. In as much as this phenomenon is not often seen in adult myocytes, it may suggest that fetal cells have agreater capacity to resist injury than adult cells and thus can isolate and digest damaged components without undergoing complete necrosis.

Colloidal lanthanum salts have an average particle size of 40 degrees A; consequently, this electron-opaque marker remains extracellular and does not cross the intact plasma membrane. The affinity of lanthanum for calcium-binding sites on mitochondrial membranes makes it possible to demonstrate loss of plasma membrane integrity at the cellular level in ischemic myocardium. Biopsies were obtained from infarcted, marginal and normal areas 3 1/2 hours after ischemia was produced in 9 anesthetized closed-chest dogs by electrically induced thrombosis of the left anterior descending coronary artery. The tissue was immediately fixed in 4% glutaraldehyde and 0.1 M cacodylate buffer containing 1.3% La(NO3)3, pH 7.4, for 2 hours. In normal control tissue prepared this way the lanthanum tracer, as expected, was confirmed to the extracellular spaces, including, basement membranes, gap junctions and portions of the intercalated discs. Specimens taken near the center of frank infarctions all contained intracellular as well as extracellular lanthanum. Intracellular lanthanum could be seen evenly distributed around lipid droplets and in focal deposits around mitochondria. Only when mitochondria were disrupted did lanthanum gain access to internal sites on mitochondrial membranes. Areas marginal to the infarct contained cells in varying stages of degeneration including many that appeared normal by morphologic criteria alone. Intracellular lanthanum was present in many but not all of the marginal cells in which degenerative changes could be seen. Similarly a few of the cells that appeared morphologically normal contained intracellular lanthanum. The entry of lanthanum into some of these marginal cells and its exclusion from adjacent cells demonstrated that ischemic injury affects the permeability properties of the plasma membrane and independently of other intracellular morphologic changes and that lanthanum can be a sensitive indicator of such alteration in membrane permeability.

An analytical electron microscopic study, utilizing scanning transmission electron microscopy and energy-dispersive x-ray spectroscopy, was made of two types of mitochondrial inclusions identified in canine myocardial infarcts. The data were obtained from thin sections of tissues that were fixed in aldehyde, osmicated and embedded in epoxy resin. Calcium peaks of variable intensity were detected in inclusions which contained very electron-dense spicular material and which were localized to muscle cells at the peripheries of the infarcts. These findings indicate that the spicular inclusions represent early stages in the process of mitochondrial calcification in myocardial infarcts. In contrast, calcium or other trace elements were not detected in moderately electron-dense amorphous inclusions which were present in mitochondria of muscle cells throughout the infarcts. With the tissue preparative techniques employed, the possibility cannot be excluded that the amorphous inclusions contained calcium, either in small amounts or in a readily diffusable state, in vivo. The data, however, are in accord with the previously advanced hypothesis that the amorphous inclusions represent precipitates of denatured mitochondrial protein formed during the evolution of irreversible cellular injury. This study provides further evidence that analytical electron microscopy can yield important information regarding the nature of various inclusions occurring in normal and diseased tissues.

Cardiac biopsies were taken from the hearts of 40 patients undergoing open heart surgery for acquired and congenital disease. Tissue samples were taken prior to aortic cross-clamping, at the end of the ischemic period, and 20 minutes after reperfusion. Cardiac arrest was induced with Kirsch's cardioplegic solution at mild hypothermia (28-30 degrees C). Electron-microscopy of these tissue samples showed that cardioplegic arrest allows safe recovery with a tolerable degree of cellular alterations following ischemic periods of approximately 45 minutes. This period of reversible ischemia agreed well with similar studies in dogs that were completed by metabolic studies. ATP4-time in these dogs was 60 minutes under conditions closely resembling those at operationmin spite of the similarity between ischemia tolerance times, marked discrepancies existed on the ultrastructural level. Human hearts showed, generally, a much more marked degree of cellular alterations. The good correlation between ultrastructural alterations and the metabolic status of the canine heart can, therefore, not be used to predict levels of metabolites from human electron-micrographies.

In rats the hypokinetic state obtained by restraint in special cages, produces after 30 days, a marked decrease in Na+ and K+ dependent ATP-ase in the skeletal muscle. After 15 days hypokinesia the enzymatic content of the investigated tissues did not change. In the left cardiac ventricular muscle Na+ and K+ dependent ATP-ase decreased after 15 days and particularly after 30 days hypokinesia. These changes were brought about by the hypofunctional state of both muscles, which also produced structural alterations. In the skeletal and cardiac muscles electron microscopy revealed fibrillar atrophy and mitochondrial alterations typical of an affected function. In the cardiac muscle, in which these changes were more accentuated, the dissociation of the intercalar disc was also noted, aspects partially described by others.

This is a correlative study of ultrastructural changes in the myocardium and the blood lead levels (BLLs) in acute lead toxicity in mice. No substantial ultrastructural changes were observed with BLLs of less than 20 mug/100ml. Blood lead levels of 20 mug/100ml and above were associated with nuclear chromatin clumping and nucleolar disorganization. Sarcotubular dilation and mitochondrial changes characterized by mitochondrial enlargement, cristal disorientation, and an increase in intramitochondrial matrix were observed with BLLs of 40 mug/100ml and over. Blood lead levels of 60 mug/100 ml and above were associated with local myofibrillar degeneration, focal intercalated disk disjunction, and interstitial cell changes characterized by prominent endoplasmic reticulum and the appearance of cytoplasmic lysosome-like dense bodies. Our findings indicate a differential sensitivity of various myocardial cell organelles to lead and suggest that the changes observed are directly related to the BLLs.

Morphology and energetic processes in the myocardium of rats suffering from autoimmune cardiomyopathy, and their changes under the influence of physical load were studied. Chronic cardiac insufficiency in autoimmune cardiomyopathy is explained by a decreased level of adenosin triphosphate (ATP) and creatinphosphate (CP), death of some cardiomyocytes, atrophy of contractile myofilaments in the working cells and disorders at the level of micro circulation bed. Physical load resulted in the appearance of ultrastructural signs of acute cardiac insufficiency without significant changes in the initial low content of ATP and CP, but with a high total creatinphosphokinase activity. Such a reaction of cardiomyocytes of the heart to physical load in autoimmune cardiomyopathy is due to the disorders in the process of energy transportation at the level of mitochondrial crystae.

Light and electron microscopic observations were made on left ventricular myocardium removed at operation from 16 patients with chronic aortic valve disease. In all 16 patients most cardiac muscle cells were hypertrophid, and surrounded by small amounts of fibrous tissue. In two of the six patients with pure aortic regurgitation and in four of the five patients with combined aortic stenosis and regurgitation, cardiac muscle cells with evidence of degeneration were present in addition to hypertrophied, nondegenerated cells. Degenerated cardiac muscle cells were not observed in the six patients with predominant aortic stenosis. Cardiac muscle cells with mild degeneration showed focal myofibrillar lysis, with preferential loss of thick myofilaments, and focal proliferation of tubules of sarcoplasmic reticulum. More severely degenerated muscle cells showed a marked decrease in the numbers of myofibrils and T tubules and proliferation of sarcoplasmic reticulum or mitochondria, or both. Severly degenerated cells usually were present in areas of marked fibrosis, often were atrophic, had thickened basement membranes and had lost their intercellular connections. These findings suggest that degenerated cardiac muscle cells have poor contractile function and may be responsible for impaired cardiac performance in some patients with chronic aortic valve disease.

The results of an electron microscopic histochemical investigation performed in the current study indicate that in heart cells creatine phosphokinase is localized: (1) inside mitochondria mainly on the cristae membranes, (2) on the membrane of the sarcoplasmic reticulum, (3) on myofibrils (and in cytoplasm), (4) on the plasma membrane of the cells, (5) on the membrane of the cell nuclei.

A surgical preparation that allows chemical alteration of the blood supplied to specific area of the intact heart was used to investigate the effects of lowered tissue pH on the fine structure of myocardial cells. Controlled infusion of isotonic saline acidified with HCl to a pH value between 1.0 and 2.0 produced a lowering of blood pH to values of less than 7.0. This drop in the pH of the perfusing blood resulted in a corresponding decrease in pH of the relevant area of myocardium. When this decrease occurred without a concomitant increase in tissue lactate level, the myocardial cells of the acid-perfused area showed focal glycogen depletion, moderate relaxation of myofibrils and clumping of nuclear chromatin, and mild mitochondrial change, which did not include marked swelling although intramitochondrial electron-dense inclusions were often detectable. When the fall in pH was accompanied by a large increase in lactate concentration, however, the cells showed more severe glycogen depletion and clumping of nuclear chromatin, and mitochondrial change, which did include marked swelling, together with the presence of well developed inclusions.

Mitochondria in sections of liver and heart (wall of left ventricle) from C57BL/6J mice, 8, 30 and 43 to 44 mo. old, were analyzed, using sterological techniques. The mitochondrial volume density decreased with age in both tissues: in liver the value estimated as 43 to 44 mo. of age was 65% the value estimated at 8 mo. of age; in the heart the 43 to 44 mo. value was 84% of the 8-mo. value. In both tissues, the decrease in mitochondrial volume density was the result of a decrease in the mitochondrial numerical density; the average volume of the mitochondrion remained constant throughout life.

Ultrastructural and biochemical studies of myocardial tissue from a patient with the clinical and light microscopic features of Reye's syndrome showed evidence of intramitochondrial calcium deposition. The significance to recent experimental and clinical data concerning intramitochondrial calcium deposition.

Ultrastructural changes in myocardial cells from the ischemic border of infarcts (produced in rat left ventricles by ligating the anterior coronary artery in vivo) were examined 1 to 24 hours after ligation. Twenty-four hours after ligation, irreversibly injured cells showed a selective spreading of Z-band material over the I band; disappearance of M bands, prominent N bands, and disassembly of A bands were also noteworthy. Sixty minutes after ligation the cells of the ischemic border were ultrastructurally normal except for paradoxically relaxed sarcomeres, indicative of an inability to contract in response to the calcium influx produced by osmium tetroxide; progressive vacuolization of this zone was evident after 4 to 12 hours. Paradoxical relaxation may be an ultrastructural correlate of acute ischemic "pump failure".

The ultrastructure of rat skeletal and cardiac muscles during embryogenesis was studied. Uninucleated skeletal muscle myoblasts of 13 day old embryos contain no myofibrils or discrete sarcomers, although intermediate filaments (8-13 nm) are observed. In cardiomyocytes of this stage, a formation of myofibrils from thick (15-16 nm) and thin (5-7 nm) filaments takes place. In 18-19 day old embryos, the ultrastructure of myofibrillar material is similar in skeletal and cardiac muscle cells. In cardiac muscle of 19 day old embryos, mitochondria with unusual structure were discovered: their membranes form invaginations filled up with glycogen.

The x-ray microanalysis technique was used to determine the chemical composition of intramitochondrial electron-dense deposits in ischemic myocardial cells. Semi-thin sections were cut from Araldite-embedded tissue and analyzed in a scanning electron microscope equipped with energy- and wavelength-dispersive spectrometers. The energy dispersive spectrum revealed calcium and phosphorus peaks over many mitochondrial deposits. Peak to background ratios of calcium, phosphorus and magnesium obtained with the wavelength dispersive spectrometer were 1.7, 8.8 and 1.2, respectively. There was no consistent relationship in the characteristic peaks of calcium and phosphorus in a given mitochondrial granule. Magnesium appears to be negligible, except in some mitochondrial deposits which lacked calcium, where it was present with a peak to background ratio of two. These results suggest formation of calcium or magnesium phosphate in the mitochondria during ischemia. X-ray microanalysis can provide detailed information on subcellular electrolyte distribution in normal and ischemic myocardial cells and should be attempted with improved methods of tissue preparation.

It was morphometrically shown that during 120-day hypokinesia chromatin redistribution was similar in nuclei of myocardial cells of the ventricles of the rat heart. Quantitative rearrangements in the mitochondrial apparatus of cells were of different pattern. On the 14th hypokinetic day the number of mitochondria increased and their size decreased; on the 30th day their size increased and their number decreased; on the 45 and 60th day the number and size of mitochondria returned to the normal; on the 120th hypokinetic day the number and size of mitochondria were higher than the control. Throughout the 120-day experimental the ratio of the total area of mitochondria to the total area of myofibers varied periodically and by the end of the experimental the mitochondrial area increased. Thus, it is obvious that by the 120th hypokinetic day myocardial changes increased indicating a new level of cardiac activity during prolonged hypokinesia. Quantitative changes in the organelles of the myocardium of the left ventricle were more significant than those of the right ventricle during the exposure.

The scanning electron microscope was used to study the ultrastructural alterations which occur following periods of myocardial ischemia of 30 minutes and 1, 2, 5, 10, and 24 hours. The alterations were compared with those seen with the transmission electron microscope. The early changes consist of swelling of organelles and clumping of nuclear chromatin. After 5 hours, disintegration of the transverse tubular system becomes apparent, and breakdown of the connections between the nucleus and the sarcoplasmic reticulum is seen coincident with distortion of the shape of the nucleus. It is suggested that the sarcoplasmic reticulum may be a part of the "endoskeleton" of the cell and may play a role in maintaining the shape and location of the nucleus. After 10 hours, extensive membrane alterations have occurred in many of the intracellular organelles. It is concluded that the changes seen with scanning electron microscopy are consistent with those seen with transmission electron microscopy.