Damaged Muscle Cells Research Articles

Structural changes in the rat bladder after acute outlet obstruction.

The urinary bladder of adult female rats was subjected to complete outlet obstruction for periods of up to 24 h. Within 2 h the obstruction led to a rise in intravesical pressure to about 80 mmH2O. Subsequently, the pressure remained high but declined slightly. After 24 h of complete obstruction the bladder was maximally distended, but its volume was similar to that of a control bladder fully distended, indicating that overstretch (or overdistension) occurs only to a very limited degree. After 6 h of obstruction there was congestion of all the intramural blood vessels and extravasation of red blood cells from some vessels of the mucosa. At 12 h and 24 h the extravasation was very substantial and there was also infiltration of erythrocytes in the muscle layer. Ultrastructurally, there were several damaged nerve endings (but no changes in the nerve trunks) and, occasionally, damaged muscle cells. Removal of the obstruction after 24 h was followed by resorption of the extravasate, partly by phagocytosis by muscle cells, a process which lasted 4-6 days, and by 7 days damaged varicosities and muscle cells became uncommon. We conclude that the changes observed in the bladder wall following complete obstruction are caused more by haemorrhage and ischaemia than by overstretch and that the changes are reversed when outlet conditions are normalized.

Increased Expression of Protease Activated Receptor-2 (PAR-2) in Balloon-Injured Rat Carotid Artery

Protease-induced cell signaling is mediated by specific receptors such as the emerging family of protease activated receptors (PARs). Since proteases are involved in various aspects of vascular injury, we assessed expression of PAR-2, a protease-activated receptor closely related to the thrombin receptor (PAR-1) but activated by an unknown protease, in vascular injury. Rat carotids were subjected to balloon-catheter injury and perfusion fixed at 1, 3, 7 or 14 days after injury. Sections of injured and normal carotid arteries were immunohistochemically labeled with a polyclonal antibody raised against the N-terminal residues 37-53 of human PAR-2. Sections were also labeled with antibodies to factor VIII-related antigen, smooth muscle actin and a proliferating cell nuclear antigen (PCNA). In normal vessels, PAR-2 labeling was diffuse and patchy in medial smooth muscle and endothelium. At one and three days after injury, before appearance of neointima, PAR-2 labeling increased in cells adjacent to damaged or necrotic smooth muscle cells. In addition, proliferating adventitial myofibroblasts labeled strongly for PAR-2. At 7 and 14 days after injury, the media and neointima of injured vessels had increased PAR-2 labeling which was most intense at the luminal edge of the neointima. Double immunohistochemical labeling confirmed the greatest expression of PAR-2 in areas with the greatest density of PCNA-positive cells. In addition, PAR-2 mRNA localization using in situ hybridization paralleled PAR-2 expression. The data suggest an upregulation of PAR-2 in response to vascular injury which is associated with medial smooth muscle damage, proliferating adventitial myofibroblasts and smooth muscle cells of the neointima, particularly those at the proliferating luminal edge of the neointima. Possible functional consequences of this receptor upregulation and its role in the response to vascular injury remain to be determined.

Localization of leukemia inhibitory factor and interleukin-6 messenger ribonucleic acids in regenerating rat skeletal muscle.

In the present study, we characterized both temporal and spatial expression patterns of leukemia inhibitory factor (LIF) and interleukin-6 (IL-6) messenger ribonucleic acids (mRNAs) in injured skeletal muscle using in situ hybridization. LIF and IL-6 mRNAs were expressed in mononucleated cells and damaged muscle cells. Further, signals for LIF mRNA were also detected in Schwann cell-like cells of intramuscular nerves. These results suggest that the earliest events involved in the repair of injured muscles and nerves may be triggered by these cytokines.

Changes in serum enzyme activities after injection of bupivacaine into rat tibialis anterior.

This study investigated the time course of changes in serum creatine kinase (CK), aspartate aminotransferase (AST), and alanine amino-transferase (ALT) activities after intramuscular injection of bupivacaine into the tibialis anterior (TA) of rats. Morphological changes in muscle cells, relationships between the amount of increase in the enzyme activities and the muscle mass damaged, and responses of serum enzymes to additional injections of bupivacaine hydrochloride (BPVC) were also examined. Adult male Wistar rats (24 wk) were placed into one of four groups. Group A (n = 7) was a control, and no injection was applied. Saline solution (0.5 ml of 0.9%) was injected into the right TA for group B (n = 5). BPVC (0.5 ml of 0.5%) was injected into the right TA for group C (n = 9) and into both the right and left TA for group D (n = 9). No increases in CK, AST, and ALT were observed for groups A and B. After BPVC injection, groups C and D showed significant (P < 0.01) increases in serum enzyme activities. CK peaked 4 h after BPVC injection, and AST and ALT peaked 12 h postinjection, then returned to the baseline by the time infiltration of mononuclear cells into the damaged muscle cells progressed. The amount of enzyme increase was significantly larger (P < 0.01) for group D compared with group C. Injection of BPVC into the right then into the left TA 4 h later displayed a bipolar response, and the second injection into the TA 12 wk after the first injection resulted in smaller increase in serum enzyme activities. It appeared that increases in serum enzyme activities reflected muscle damage; however, changes in enzymes occurred in the early stage of myonecrosis.

Changes in plasma enzyme activities in professional racing cyclists.

To reports changes in plasma levels in professional racing cyclists. Plasma levels of the intracellular enzymes aspartate aminotransferase, alanine aminotransferase, gamma glutamyl transpeptidase, lactate dehydrogenase, and alkaline phosphatase were measured resting and after exercise in professional cyclists participating in two road races: Vuelta Ciclista a Valencia (800 km, beginning of the cyclist season), and the top rank race Vuelta a España (2700 km, at the end of cyclist season). A significant increase in plasma aspartate aminotransferase, alanine aminotransferase, and alkaline phosphatase was observed at the end of the race over the corresponding paired start values (P < 0.05). The increase in plasma lactate dehydrogenase after the race was only statistically significant (P < 0.05) in the Vuelta a España. The longer the duration of the cycle tour race, the greater were the increases in aspartate aminotransferase and alanine aminotransferase levels. The high levels returned to their start values during the overnight resting period. The presence of these enzymes in the blood is probably due to mechanically damaged muscle cells leaking their contents into the interstitial fluid.

Sampling interstitial fluid from rat skeletal muscles by intermuscular wicks

A modification of the implanted wick method (K. Aukland and H. O. Fadnes. Acta Physiol. Scand. 88: 350-358, 1973) was devised to sample interstitial fluid from rat muscles. Dry nylon wicks were inserted postmortem into intermuscular spaces between leg muscles by means of a plastic catheter, which was subsequently withdrawn. Inserting the wicks postmortem avoids contaminating wick fluid with proteins extravasated as a result of local inflammatory reactions; placing them intermuscularly avoids contamination by fluid and proteins from damaged muscle cells. Wick fluid protein concentrations (mg/ml) averaged 24.1 +/- 1.1 and 28.5 +/- 1.5 (means +/- SE) in medial and lateral hindlimbs muscles, respectively. The corresponding albumin concentrations were 13.0 +/- 0.7 and 13.9 +/- 0.7 mg/ml. Total protein and albumin concentrations in plasma were 54.1 +/- 0.8 and 22.5 +/- 0.3 mg/ml. Electrophoresis of wick fluid showed a pattern of peaks similar to that of plasma, with albumin relatively high and larger molecules relatively low. Proteins from muscle cells were not detected. Isotope studies (125I-labeled albumin, 51Cr-EDTA) showed that less than 2% of the albumin in wick fluid came directly from plasma and that wick fluid was not concentrated by cell swelling postmortem. Wick fluid from intermuscular wicks implanted in anesthetized rats in vivo had nearly the same total protein concentration as fluid from postmortem wicks, but albumin-to-globulin (A/G) ratios were slightly lower (1.22 +/- 0.07 vs. 1.53 +/- 0.21 measured by gel electrophoresis), and more significantly, nearly 50% of the albumin leaked to wick fluid from plasma as a result of wick implantation.(ABSTRACT TRUNCATED AT 250 WORDS)

Ischemic Tolerance of Human Skeletal Muscle

Until now, the ischemic tolerance of muscle tissue has not been adequately understood. Even when muscle vitality is lost, the perfusion matrix of the muscle flaps is retained. Because of toxic decomposition, however, irreversibly damaged muscle cells almost certainly increase the rate of complications. The retention of the vitality of the transplanted muscle tissue is absolutely essential for the myokinetic substitute operations, currently in the development stage, involving the free transplantation of muscles. Investigations into vitality reserves were carried out on skeletal muscle specimens. Nuclear magnetic resonance spectroscopy was used to establish that, in ischemia, the ATP pool remained topped up to a large extent as long as phosphocreatine was available. As long as the ATP pool was retained, rearterialization led to the complete restoration of the essential preischemic metabolite concentrations. After the ATP had been exhausted, biochemical restitution through arterial reperfusion did not occur. The time by which the established vitality threshold was reached because of the loss of the ATP pool is called the critical ischemia time; it depends on muscle temperature. The critical ischemia time of human skeletal muscles was determined between 26 degrees and 38 degrees C. A normothermia of 34 degrees C yielded a critical ischemia time of 2.25 hours, which is shorter than that previously reported in the literature. An ischemic tolerance of 5 hours presupposes a muscle temperature of less than 26 degrees C.

Horseradish peroxidase as a permeability marker in injured rat caudal and iliac arteries

The permeability to Horseradish Peroxidase (HRP) of the rat caudal artery at the level of spontaneous lesions was evaluated by electron microscopy and compared with that of lesions experimentally induced by pinching or internal scraping of the caudal and iliac arteries. No HRP reaction product is observed in the extracellular space of the arterial wall when (i) the internal elastic lamina (IEL) and the endothelium are absent, (ii) the IEL is maintained and the endothelium is absent and (iii) the IEL is absent and the endothelium has regenerated. That HRP does enter the arterial wall in cases of gross endothelial damage is shown by its selective retention in damaged smooth muscle cells in such cases. In contrast, HRP reaction product is detected in the subendothelial space when the IEL is maintained and is covered by a regenerating or recently regenerated endothelium. Furthermore, the amount of tracer visualized under the same experimental conditions is greater in the iliac than in the caudal artery. We conclude that the detection of HRP in the subendothelial space of the artery wall requires the presence of regenerating or recently regenerated endothelial cells lying on an intact IEL. It is thus not simply related to endothelial permeability but depends also upon the retention of HRP by extracellular substances. In addition, the quantity of marker retained varies between different sites in the arterial tree.

Pharmacodynamic effects of serotonin (5-HT) receptor ligands in pigs: stimulation of 5-HT2 receptors induces malignant hyperthermia

In pigs, the serotonin-2 (5-HT2) receptor agonist 1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI), 0.8 mg/kg, induced "psychotic" behaviour (e.g., grimacing, backward locomotion, blank stare) and a muscular syndrome, which is known as malignant hyperthermia (MH) in pigs and humans. This syndrome is characterized by generalized skeletal muscle rigidity, leading to an increase in body temperature, marked acidosis, hyperkaliaemia, cyanosis and elevation of lactate, carbon dioxide and the muscle enzyme creatine kinase (CK) in plasma. In pigs which were selectively bred for susceptibility to MH induction by known triggering agents, such as halothane, the administration of DOI was fatal in 3 out of 5 animals. In genetically susceptible pigs, MH was also induced by 5-methoxy-N,N-dimethyltryptamine (5-MeO-DMT), 0.5-1.8 mg/kg, and D-lysergic acid diethylamide (LSD), 60-110 micrograms/kg. Furthermore, 5-MeO-DMT and LSD induced head shakes in the animals, which had not been observed after DOI and could not be blocked by 5-HT2-antagonists, ketanserin (0.5-5 mg/kg) and ritanserin (1-2.5 mg/kg). The psychotomimetic effects of 5-MeO-DMT could be blocked by ketanserin or ritanserin, which, depending on the dose, also reduced or totally prevented the hyperthermia and metabolic changes induced by 5-MeO-DMT in pigs. Administration of 5-MeO-DMT, 1.8 mg/kg, was fatal in 4 of 5 MH-susceptible pigs, whereas pigs injected with this dosage after pretreatment with ketanserin (0.5-5 mg/kg) or ritanserin (1-2.5 mg/kg) did not die. In pigs from MH-resistant littermates, administration of 5-MeO-DMT was not fatal. Comparison of metabolic changes in susceptible and non-susceptible pigs suggested that the marked increase in plasma potassium, which arises principally from damaged muscle cells, is primarily responsible for the fatal effect of DOI and 5-MeO-DMT in genetically susceptible individuals. In MH-susceptible pigs, which were anesthetized, relaxed and artificially ventilated, 5-MeO-DMT did not induce hyperthermia, thus substantiating that the marked hyperthermia observed in conscious pigs was a result of muscle activation and not due to effects on thermoregulation or blood pressure. The results indicate that hallucinogenic drugs with 5-HT2 agonistic effects trigger a life-threatening syndrome, MH, in genetically susceptible pigs. 5-HT2 antagonists, such as ketanserin or ritanserin, are capable of counteracting the fatality of this syndrome.

Inflammatory myopathy and acquired immunodeficiency syndrome

A 33-year-old black woman with advanced acquired immunodeficiency syndrome (AIDS) presented with rapidly progressive muscle weakness and serologic and radiologic evidence of central nervous system Toxoplasma infection. Muscle biopsy revealed an inflammatory infiltrate predominantly composed of macrophages and T suppressor/cytotoxic cells. Human immunodeficiency virus major core protein (p24) was also detected in macrophages and damaged muscle cells around the inflammatory infiltrates. The patient improved clinically with glucocorticoid therapy for polymyositis and pyrimethamine and clindamycin therapy for toxoplasmosis.

Pressure-induced myogenic activation of cat cerebral arteries is dependent on intact endothelium.

These studies were designed to determine the role of cerebral vascular endothelium in the "myogenic" depolarization and contraction observed in isolated cat middle cerebral arteries exposed to high transmural pressures. With intact endothelial cells we observed, on elevation of transmural pressure in cannulated isolated arteries, significant membrane depolarization, action potential generation, and reduction in internal diameter. After perfusion of the same vessels with collagenase and elastase for short periods of time to disrupt the endothelial layer, all previous responses to elevation of transmural pressure were no longer seen. Even though enzyme perfusion had no effect on membrane potential at "control" levels of transmural pressure, it abolished the pressure-dependent depolarization, action potential generation, and constriction. Furthermore, the contractile response to agonist stimulation was maintained after endothelial disruption via enzymes, showing that this method of endothelial disruption did not appreciably damage muscle cells. The data document a dependence of an intact endothelium in mediating the activation of isolated cat cerebral arteries in response to a changing transmural pressure. Thus, it is possible that the endothelial cell may serve as a transducer in the autoregulatory response to pressure.

Sites and mechanisms of localization of technetium-99m phosphorus radiopharmaceuticals in acute myocardial infarcts and other tissues.

This study was performed to elucidate the localization at the cellular level of technetium-99m phosphorus ((99m)Tc-P) radiopharmaceuticals in acute myocardial infarcts and the mechanisms responsible for (99m)Tc-P uptake in acute myocardial infarcts and other tissues. In 20 dogs with proximal left anterior descending coronary arterial ligation for 1-3 days, elevated calcium levels were measured at all sites of increased (99m)Tc-P uptake (acute myocardial infarcts, necrotic thoracotomy muscle, lactating breast, and normal bone); however, a consistent linear relationship between (99m)Tc-P and calcium levels was not observed. A strong correlation (r = 0.95 and 0.99, n = 2 dogs) was demonstrated between levels of (3)H-diphosphonate and (99m)Tc-P in infarcted myocardium. Autoradiographic studies with (3)H-diphosphonate revealed extensive labeling in the infarct periphery which contained necrotic muscle cells with features of severe calcium overloading, including widespread hypercontraction as well as more selective formation of mitochondrial calcific deposits. Autoradiography also demonstrated labeling of a small population of damaged border zone muscle cells which exhibited prominent accumulation of lipid droplets and focal, early mitochondrial calcification. Cell fractionation studies revealed major localization of both (99m)Tc-P and calcium in the soluble supernate and membrane-debris fractions of infarcted myocardium and less than 2% of total (99m)Tc-P and calcium in the mitochondrial fractions; however, electron microscopic examination showed that mitochondria with calcific deposits were not preserved in the mitochondrial fractions. In vitro studies evaluating the role of serum protein binding on tissue uptake of (99m)Tc-P agents demonstrated that, in spite of significant complexing with serum proteins, serum (99m)Tc-P activity retained the ability to adsorp to calcium hydroxyapatite and amorphous calcium phosphate. In vivo studies showed that concentration of human serum albumin (labeled with iodine-131) in infarcted myocardium reached a maximum of only 3.8 times normal after a circulation time of 96 h, whereas (99m)Tc-P uptake was at least 10 times normal after a circulation time as short as 1 h. It is concluded that: (a) (99m)Tc-P uptake in acutely infarcted myocardium, and possibly other types of soft tissue damage, is limited to necrotic and severely injured cells; (b) concentration of (99m)Tc-P results from selective adsorption of (99m)Tc-P with various forms of tissue calcium stores, including amorphous calcium phosphate, crystalline hydroxyapatite, and calcium complexed with myofibrils and other macromolecules, possibly supplemented by calcium-independent complexing with organic macromolecules; and (c) lack of a linear relationship between (99m)Tc-P and tissue calcium levels mainly results from local differences in composition and physicochemical properties of tissue calcium stores and from local variations in levels of blood flow for delivery of (99m)Tc-P agents.

Metabolic disturbances and plasmorrhagia into myocardial cells injured by adrenalin

Serial sections through the myocardium of rats killed 1–24 h after an injection of adrenalin were used. The results of histochemical reactions for succinate dehydrogenase (test for cell injury) were compared with the results of fibrin determination by the Coons' method. The presence and character of plasmorrhagia into irreversibly damaged muscle cells, giving the characteristic appearance in the reaction with nitro-BT, and the absence of fibrin in fibers with fatty degeneration, reflected in coarse-grained formazan deposits, were demonstrated. On the basis of the results it is postulated that the reversible and irreversible injuries to the myocardium produced by adrenalin differ in their pathogenesis.

AN ELECTORON MICROSCOPIC OBSERVATION ON THE MICROVASCULAR CHANGES CAUSED BY HABU SNAKE VENOM, WITH SPECIAL REFERENCE TO HEMORRHAGE

Crude Habu snake venom was injected in the femoral muscle of mice and the ultrastructures of the microvessels in the lesions were examined by electoron microscopy. Most of the vessels encountered between the damaged muscle cells were capillaries, partially containing postcapillary venules. The results were as follows.1. There were various injuries of the endothelial cells, such as formation of villous projections and intravascular blebs from cells, degeneration of cellular organelles, especially decrease and disappearance of pinocytotic vesicles, swelling, darkning and thinning of cells, formation of endothelial gaps and celullar necrosis.2. The fluffy changes, discontinuities and deficiency of the basement membrane were observed.3. The hemorrhage by Habu snake venom were occured by discontinuities of both the endothelium and the underlining basement membrane.4. It is considered that the increasing hemorrhage may be caused not only by the destruction of capillary wall, but also by the disturbance of coagulant activities.

Serum Enzymes in Sex-Linked (Duchenne) Muscular Dystrophy

IN MUSCULAR dystrophy, particularly of the Duchenne or sex-linked type, a number of enzymes generally have elevated levels in the serum. In diseases such as the various kinds of neurogenic muscular atrophy that may be mistaken for muscular dystrophy, enzyme levels are normal. On the other hand, diseases like hepatitis that may show high enzyme levels are readily distinguishable from muscular dystrophy by their symptoms. Thus, the serum enzymes are of great value in diagnosis whenever muscular dystrophy is a possibility. They are also very promising as a tool in research. The enzymes in the serum appear to enter the blood by leakage from damaged muscle cells. The determination of serum enzymes may provide a way of measuring the intensity of the disease process at a given time. If so, the enzymes are useful not only for diagnosis but are essential for studying the development of the disease and