Intercostal Biopsies Research Articles

Inter-alpha-trypsin inhibitor heavy chain H3 is a potential biomarker for disease activity in myasthenia gravis

Myasthenia gravis is a chronic antibody-mediated autoimmune disease disrupting neuromuscular synaptic transmission. Informative biomarkers remain an unmet need to stratify patients with active disease requiring intensified monitoring and therapy; their identification is the primary objective of this study. We applied mass spectrometry-based proteomic serum profiling for biomarker discovery. We studied an exploration and a prospective validation cohort consisting of 114 and 140 anti-acetylcholine receptor antibody (AChR-Ab)-positive myasthenia gravis patients, respectively. For downstream analysis, we applied a machine learning approach. Protein expression levels were confirmed by ELISA and compared to other myasthenic cohorts, in addition to myositis and neuropathy patients. Anti-AChR-Ab levels were determined by a radio receptor assay. Immunohistochemistry and immunofluorescence of intercostal muscle biopsies were employed for validation in addition to interactome studies of inter-alpha-trypsin inhibitor heavy chain H3 (ITIH3). Machine learning identified ITIH3 as potential serum biomarker reflective of disease activity. Serum levels correlated with disease activity scores in the exploration and validation cohort and were confirmed by ELISA. Lack of correlation between anti-AChR-Ab levels and clinical scores underlined the need for biomarkers. In a subgroup analysis, ITIH3 was indicative of treatment responses. Immunostaining of muscle specimens from these patients demonstrated ITIH3 localization at the neuromuscular endplates in myasthenia gravis but not in controls, thus providing a structural equivalent for our serological findings. Immunoprecipitation of ITIH3 and subsequent proteomics lead to identification of its interaction partners playing crucial roles in neuromuscular transmission. This study provides data on ITIH3 as a potential pathophysiological-relevant biomarker of disease activity in myasthenia gravis. Future studies are required to facilitate translation into clinical practice.

Impaired sarcoplasmic reticulum Ca2+ release is the major cause of fatigue-induced force loss in intact single fibres from human intercostal muscle.

Changes in intramuscular Ca2+ handling contribute to development of fatigue and disease-related loss of muscle mass and function. To date, no data on human intact living muscle fibres have been described. We manually dissected intact single fibres from human intercostal muscle and simultaneously measured force and myoplasmic free [Ca2+ ] at physiological temperature. Based on their fatigue resistance, two distinct groups of fibres were distinguished: fatigue sensitive and fatigue resistant. Force depression in fatigue and during recovery was due to impaired sarcoplasmic reticulum Ca2+ release in both groups of fibres. Acidification did not affect force production in unfatigued fibres and did not affect fatigue development in fatigue-resistant fibres. The current study provides novel insight into the mechanisms of fatigue in human intercostal muscle. Changes in intracellular Ca2+ handling of individual skeletal muscle fibres cause a force depression following physical activity and are also implicated in disease-related loss of function. The relation of intracellular Ca2+ handling with muscle force production and fatigue tolerance is best studied in intact living single fibres that allow continuous measurements of force and myoplasmic free [Ca2+ ] during repeated contractions. To this end, manual dissections of human intercostal muscle biopsies were performed to isolate intact single fibres. Based on the ability to maintain tetanic force at >40% of the initial value during 500 fatiguing contractions, fibres were classified as either fatigue sensitive or fatigue resistant. Following fatigue all fibres demonstrated a marked reduction in sarcoplasmic reticulum Ca2+ release, while myofibrillar Ca2+ sensitivity was either unaltered or increased. In unfatigued fibres, acidosis caused a reduction in myofibrillar Ca2+ sensitivity that was offset by increased tetanic myoplasmic free [Ca2+ ] so that force remained unaffected. Acidification did not affect the fatigue tolerance of fatigue-resistant fibres, whereas uncertainties remain whether or not fatigue-sensitive fibres were affected. Following fatigue, a prolonged force depression at preferentially low-frequency stimulation was evident in fatigue-sensitive fibres and this was caused exclusively by an impaired sarcoplasmic reticulum Ca2+ release. We conclude that impaired sarcoplasmic reticulum Ca2+ release is the predominant mechanism of force depression both in the development of, and recovery from, fatigue in human intercostal muscle.

Severe damage of respiratory muscles in polymyositis

Objectives - to present a clinical case of polymyositis in a 55-year-old man. The patient was admitted with a diagnosis "transient ischemic attack", quickly followed by an acute respiratory failure, which required the continuous use of mechanical ventilation. Other clinical symptoms included: a high diaphragm position up to the 4-th ribs, the concurrent purulent endobronchitis, bilateral pneumonia, pleurisy, multiple atelectasis, myocardial dysfunction with a decrease in ejection fraction to 46%, local hypokinesia according to EchoCG data. Despite the severe condition of the patient, there was not registered any distinct impairment of the proximal limb muscles and an increase in creatinphosphokinase blood level. The antinuclear myositis-specific antibodies, the signs of primary muscular damage according to the limb muscles ENMG data were also absent. The paraneoplastic process was excluded. The results of clinical examination made the diagnosis of polymyositis uncertain. To clarify the cause of the respiratory muscles damage, an intercostal muscle biopsy was performed.It revealed expressed changes in the muscle, with massive necrosis areas, and perivascular lymphohistiocytic infiltration. Conclusion. The described case of polymyositis is extremely rare, has a severe course and is complicated for diagnosis and treatment. In this case, the most informative examination method can be a biopsy of the intercostal muscles, which allows to identify the inflammatory origine of the disease and to determine the pathogenetic therapy.

Complement activation at the motor end-plates in amyotrophic lateral sclerosis.

BackgroundAmyotrophic lateral sclerosis (ALS) is a fatal progressive neurodegenerative disease with no available therapy. Components of the innate immune system are activated in the spinal cord and central nervous system of ALS patients. Studies in the SOD1G93A mouse show deposition of C1q and C3/C3b at the motor end-plate before neurological symptoms are apparent, suggesting that complement activation precedes neurodegeneration in this model. To obtain a better understanding of the role of complement at the motor end-plates in human ALS pathology, we analyzed post-mortem tissue of ALS donors for complement activation and its regulators.MethodsPost-mortem intercostal muscle biopsies were collected at autopsy from ALS (n = 11) and control (n = 6) donors. The samples were analyzed for C1q, membrane attack complex (MAC), CD55, and CD59 on the motor end-plates, using immunofluorescence or immunohistochemistry.ResultsHere, we show that complement activation products and regulators are deposited on the motor end-plates of ALS patients. C1q co-localized with neurofilament in the intercostal muscle of ALS donors and was absent in controls (P = 0.001). In addition, C1q was found deposited on the motor end-plates in the intercostal muscle. MAC was also found deposited on motor end-plates that were innervated by nerves in the intercostal muscle of ALS donors but not in controls (P = 0.001).High levels of the regulators CD55 and CD59 were detected at the motor end-plates of ALS donors but not in controls, suggesting an attempt to counteract complement activation and prevent MAC deposition on the end-plates before they are lost.ConclusionsThis study provides evidence that complement activation products are deposited on innervated motor end-plates in the intercostal muscle of ALS donors, indicating that complement activation may precede end-plate denervation in human ALS. This study adds to the understanding of ALS pathology in man and identifies complement as a potential modifier of the disease process.Electronic supplementary materialThe online version of this article (doi:10.1186/s12974-016-0538-2) contains supplementary material, which is available to authorized users.

Intracellular Ca(2+)-handling differs markedly between intact human muscle fibers and myotubes.

BackgroundIn skeletal muscle, intracellular Ca2+ is an important regulator of contraction as well as gene expression and metabolic processes. Because of the difficulties to obtain intact human muscle fibers, human myotubes have been extensively employed for studies of Ca2+-dependent processes in human adult muscle. Despite this, it is unknown whether the Ca2+-handling properties of myotubes adequately represent those of adult muscle fibers.MethodsTo enable a comparison of the Ca2+-handling properties of human muscle fibers and myotubes, we developed a model of dissected intact single muscle fibers obtained from human intercostal muscle biopsies. The intracellular Ca2+-handling of human muscle fibers was compared with that of myotubes generated by the differentiation of primary human myoblasts obtained from vastus lateralis muscle biopsies.ResultsThe intact single muscle fibers all demonstrated strictly regulated cytosolic free [Ca2+] ([Ca2+]i) transients and force production upon electrical stimulation. In contrast, despite a more mature Ca2+-handling in myotubes than in myoblasts, myotubes lacked fundamental aspects of adult Ca2+-handling and did not contract. These functional differences were explained by discrepancies in the quantity and localization of Ca2+-handling proteins, as well as ultrastructural differences between muscle fibers and myotubes.ConclusionsIntact single muscle fibers that display strictly regulated [Ca2+]i transients and force production upon electrical stimulation can be obtained from human intercostal muscle biopsies. In contrast, human myotubes lack important aspects of adult Ca2+-handling and are thus an inappropriate model for human adult muscle when studying Ca2+-dependent processes, such as gene expression and metabolic processes.Electronic supplementary materialThe online version of this article (doi:10.1186/s13395-015-0050-x) contains supplementary material, which is available to authorized users.

S53 Studying fibre specific gene expression in COPD using laser capture micro-dissection in human skeletal muscle

BackgroundApproximately 30% of patients with Chronic Obstructive Pulmonary Disease (COPD) exhibit peripheral skeletal muscle dysfunction and a shift towards type II glycolytic fibres in the quadriceps compared to healthy controls...

Dok‐7 myasthenia: Phenotypic and molecular genetic studies in 16 patients

Detailed analysis of phenotypic and molecular genetic aspects of Dok-7 myasthenia in 16 patients. We assessed our patients by clinical and electromyographic studies, by intercostal muscle biopsies for in vitro microelectrode analysis of neuromuscular transmission and quantitative electron microscopy EM of 409 end plates (EPs), and by mutation analysis, and expression studies of the mutants. The clinical spectrum varied from mild static limb-girdle weakness to severe generalized progressive disease. The synaptic contacts were single or multiple, and some, but not all, were small. In vitro microelectrode studies indicated variable decreases of the number of released quanta and of the synaptic response to acetylcholine; acetylcholine receptor (AChR) channel kinetics were normal. EM analysis demonstrated widespread and previously unrecognized destruction and remodeling of the EPs. Each patient carries 2 or more heteroallelic mutations: 11 in genomic DNA, 7 of which are novel; and 6 identifiable only in complementary DNA or cloned complementary DNA, 3 of which are novel. The pathogenicity of the mutations was confirmed by expression studies. Although the functions of Dok-7 include AChR beta-subunit phosphorylation and maintaining AChR site density, patient EPs showed normal AChR beta-subunit phosphorylation, and the AChR density on the remaining junctional folds appeared normal. First, the clinical features of Dok-7 myasthenia are highly variable. Second, some mutations are complex and identifiable only in cloned complementary DNA. Third, Dok-7 is essential for maintaining not only the size but also the structural integrity of the EP. Fourth, the profound structural alterations at the EPs likely contribute importantly to the reduced safety margin of neuromuscular transmission.

Neuromuscular junction autoimmune disease: muscle specific kinase antibodies and treatments for myasthenia gravis

Some of the 20% of myasthenia gravis patients who do not have antibodies to acetylcholine receptors (AChRs) have antibodies to muscle specific kinase (MuSK), but a full understanding of their frequency, the associated clinical phenotype and the mechanisms of action of the antibodies has not yet been achieved. Moreover, some patients do not respond well to conventional corticosteroid therapy. Here we review recent clinical and experimental studies on MuSK antibody associated myasthenia gravis, and summarize the results of newer treatments for myasthenia gravis. MuSK antibodies are found in a variable proportion of AChR antibody negative myasthenia gravis patients who are often, but not exclusively, young adult females, with bulbar, neck, or respiratory muscle weakness. The thymus histology is normal or only very mildly abnormal. Surprisingly, limb or intercostal muscle biopsies exhibit no reduction in AChR numbers or complement deposition. However, patients without AChR or MuSK antibodies appear to be similar to those with AChR antibodies and may have low-affinity AChR antibodies. A variety of treatments, often intended to enable corticosteroid doses to be reduced, have been used in all types of myasthenia gravis with some success, but they have not been subjected to randomized clinical trials. MuSK antibodies define a form of myasthenia gravis that can be difficult to diagnose, can be life threatening and may require additional treatments. An improved AChR antibody assay may be helpful in patients without AChR or MuSK antibodies. Clinical trials of drugs in other neuroimmunological diseases may help to guide the treatment of myasthenia gravis.

Inheritance of recurrent exertional rhabdomyolysis in Thoroughbreds

To develop a diagnostic test for recurrent exertional rhabdomyolysis (RER) in Thoroughbreds that relied on in vitro contracture of muscle biopsy specimens and determine whether the inheritance pattern of RER diagnosed on the basis of this contracture test was consistent with an autosomal dominant trait. Clinical trial. 8 adult horses with RER and 16 control adult horses for development of the contracture test; 23 foals for inheritance of RER. External intercostal muscle biopsy specimens from the 24 adult horses were tested for contracture in response to halothane and caffeine, and criteria for a positive test result were determined. These criteria were then applied to results for the 23 foals to determine whether they had RER. Simple segregation analysis was performed to determine whether results were consistent with a dominant pattern of inheritance. Results of the contracture test were positive for 5 of the 12 colts and 4 of the 11 fillies. Results of segregation analysis were consistent with an auto-. somal dominant pattern of inheritance. Two sires with RER produced colts with RER, supporting the hypothesis that RER had an autosomal, rather than an X-linked, inheritance pattern. In addition, in 1 instance, an unaffected colt was produced by 2 affected parents, which was not consistent with a recessive mode of inheritance. Although the expression of the RER trait is influenced by sex, temperament, and diet, among other factors, results from the in vitro muscle contracture test and this breeding trial suggest that RER in Thoroughbreds can be modeled as a genetic trait with an autosomal dominant pattern of inheritance.

Novel truncating RAPSN mutations causing congenital myasthenic syndrome responsive to 3,4-diaminopyridine

Rapsyn is essential for clustering the acetylcholine receptor at the postsynaptic membrane of the neuromuscular junction. Direct sequencing of RAPSN in two children with congenital myasthenic syndromes with no mutation in any of the AChR subunits identified two heterozygous recessive mutations in each: a previously characterized N88K mutation in both, and a second frameshifting mutation in Patient (Pt) 1 and a nonsense mutation in Pt 2. An intercostal muscle biopsy in Pt 1 revealed decreased AChRs per endplate and decreased amplitude of the miniature endplate potential, predicted consequences of rapsyn deficiency. Clinically, both children manifested with hypomotility in utero, fatigable ocular and limb weakness since birth, decreased strength during viral illness, decremental response on electromyography, and absence of AChR antibodies. Pt 1, however, had a more severe clinical course with recurrent episodes of respiratory failure, contractures, and craniofacial malformations. In both patients, treatment with pyridostigmine was of some benefit, but the addition of 3,4-diaminopyridine led to significant clinical improvement. Thus, rapsyn deficiency predicting similar consequences at the cellular level can result in phenotypes with marked differences in severity of symptoms, risk of respiratory failure, and presence of contractures and craniofacial malformations.

Recent Advances in Understanding Congenital Myasthenic Syndromes

Congenital myasthenic syndromes (CMS) are rare genetic disorders that continue to challenge the skills of clinical electrophysiologists, neuromuscular specialists, and general neurologists. Their pathogenesis remained elusive until the early 1990s, when advances in molecular genetics and in vitro neurophysiology of intercostal muscle biopsies provided major insights into disease mechanisms and created new ways to confirm …

Hypophosphatemia and Phosphorus Depletion in Respiratory and Peripheral Muscles of Patients With Respiratory Failure due to COPD

In 22 patients (19 men, 3 women; mean [+/- SD] age, 63 +/- 6 years) with chronic obstructive pulmonary disease (COPD), phosphorus content was measured by spectrophotometric methods on muscle fragments of both peripheral (quadriceps femoris needle biopsy in 22 patients) and respiratory muscles (external intercostal muscle surgical biopsy in 14 patients). Thirty age- and sex-matched subjects were used as controls (19 for quadriceps femoris muscle biopsy and 11 for intercostal muscle biopsy). Serum phosphorus levels, as well as the main determinants of overall phosphorus metabolism (dietary intake of phosphorus and renal phosphate handling), were also obtained in all patients and control subjects. Muscle phosphorus content of both respiratory and peripheral muscles was significantly reduced in the COPD patient group, no matter what reference index was used (fat-free dry muscle weight or muscle fragment DNA content); muscle phosphorus depletion was present in about 50 percent of patients with COPD. In the same patient group, a significant relationship between muscle and serum phosphorus levels was demonstrable in the case of peripheral muscles only. No relationship was found between phosphorus content of both types of skeletal muscles and dietary phosphorus intake levels or with nutritional status, even though patients with COPD had significantly reduced anthropometric, biochemical, and immunologic indices as compared with controls. Renal phosphorus handling indices of the COPD patient group were compatible with a condition of inadequacy of the renal compensatory mechanism to hypophosphatemia and phosphorus depletion (low percent tubular reabsorption of phosphorus, low renal threshold concentration values). Our study suggests that phosphorus depletion occurs frequently in COPD, but in this clinical condition serum phosphorus levels are not representative of cellular phosphorus levels. Phosphorus depletion, which is equally severe in respiratory and peripheral muscles, could depend, at least in part, on malnutrition and a condition of renal phosphorus wasting possibly linked to some drugs commonly used in patients with COPD (xanthine derivatives, diuretics, etc).

Secondary changes of the motor endplate in Lambert-Eaton myasthenic syndrome: a quantitative study.

Underlying the Lambert-Eaton myasthenic syndrome (LEMS) is a decrease in the release of the neurotransmitter acetylcholine. Only few reports on light and transmission electron microscopical observations of motor endplates in LEMS are available and changes reported so far differ from those found in experimental blocking of acetylcholine release. We performed a quantitative study on intercostal muscle biopsies of five patients. The main light microscopical finding was an enlargement of the area of contact between nerve and muscle, which was interpreted as a compensatory phenomenon. At the ultrastructural level we found the mean postsynaptic area and membrane length of the neuromuscular junctions to be decreased, putatively due to small postsynaptic regions of newly created neuromuscular junctions. The secondary changes in LEMS endplates as seen in this study are similar to those reported in experimental blocking of acetylcholine release.

Chloride channel regulation in the skeletal muscle of normal and myotonic goats

External intercostal muscle biopsies from normal and congenitally myotonic goats were studied in vitro at 30 degrees C using a two-microelectrode square-pulse cable analysis assisted by computer. The resting chloride conductance (Gcl) was estimated from the difference between the mean membrane conductance in chloride-containing and chloride-free bathing media. The protein kinase C (PKC) activator, 4-beta-phorbol-12,13-dibutyrate. (0.1-2.0 microM) blocks a maximum of 76% of Gcl in normal goat fibers and induces myotonic hyperexcitability similar to that of congenitally myotonic goat fibers. The Gcl block was partially antagonized by pretreatment with the PKC inhibitor, staurosporine (10 microM). The "inactive" 4-alpha-phorbol-12,13-didecanoate had no effect at 50 microM, whereas the "active" 4-beta isomer blocked 41% Gcl at 1 microM. The nearly absent Gcl of congenitally myotonic goat fibers was not restored by treatment with high concentrations of the PKC inhibitors staurosporine, 1-(5-isoquinolinesulfonyl)-2-methylpiperazine (H7), or tetrahydropapaveralone (THP). Also, forskolin and cholera toxin, which may increase cyclic adenosine monophosphate (cAMP) levels, or the R(+) clofibric acid enantiomers and taurine, which increase Gcl in normal fibers, were also unable to restore Gcl in myotonic goat fibers. The data suggest that PKC may be a chloride channel regulator in normal goat skeletal muscle fibers, however the molecular defect of congenitally myotonic fiber does not appear to be due to excessive activity of PKC.

Responses of intercostal muscle biopsies from normal subjects and patients with myasthenia gravis

In order to evaluate the mechanisms of weakness in muscles of patients with myasthenia gravis (MG), intercostal muscle biopsies were obtained from 9 normal subjects and 6 MG patients, and the compound muscle action potential (AP) and tension responses to nerve and muscle stimulation, and contracture responses on exposure to caffeine, were monitored in vitro. In normal muscle, on stimulation of the nerve or muscle at 30 to 100 Hz, the AP responses showed decrement in amplitude, one-third of which was attributable to failure of neuromuscular transmission and two-thirds to failure of muscle membrane excitation. On stimulation at 1 to 5 Hz, the AP responses showed very little decrement, while the contractile responses showed significant fade in tension, due to failure of E-C coupling or contractility. In muscle from patients with generalized MG, stimulation of the nerve at all frequencies (1 to 100 Hz) caused much greater decrement in APs and fade in tension responses than in normal muscle, due mainly to failure of neuromuscular transmission. However, at 100 Hz, 40% of the decrement in APs was due to failure of muscle membrane excitation, and at 1 to 5 Hz, 40% of the fade in tension was due to failure of E-C coupling or contractility, as in normal muscle. On direct stimulation the contraction and half-relaxation times were slower and the tetanic tension was smaller than in normal muscle, especially in the MG patient with thymoma. Caffeine-induced contractures were smaller in MG muscle than in normal muscle. These results indicate that while the weakness of MG muscle is due mainly to failure of neuromuscular transmission, it is also partly due to reduced E-C coupling or contractility.

Respiratory Muscle Fiber Morphometry: Correlation with Pulmonary Function and Nutrition

To examine the relationship between nutrition, pulmonary function, respiratory muscle strength, and respiratory muscle morphometry, we compared physiologic data and muscle morphometry obtained from internal intercostal, external intercostal, and latissimus dorsi muscle biopsies in 68 patients who were having a thoracotomy. We stained the biopsies for myosin ATPase and measured the proportions and diameters of the type 1 and type 2 fibers. There were more qualitative changes in the external intercostal muscles than in the other two, and some of these changes related to the incidence of malignancy. There were more type 1 fibers in the external intercostal (64 +/- 10 percent) and internal intercostal muscles (59 +/- 12 percent) than in the latissimus dorsi (44 +/- 13 percent) (p less than 0.005). The mean diameter of the type 2 fibers in the external intercostal muscles was less (44 mu +/- 7 mu) than the diameter in the latissimus dorsi (51 mu +/- 9 mu) and the internal intercostal muscles (52 mu +/- 8 mu) (p less than 0.01). The diameters of both fiber types were greater in men than in women. There was no significant relationship between measures of pulmonary function or respiratory muscle strength and muscle fiber proportions and diameters. There were significant correlations between the percentage of ideal body weight and type 1 and type 2 fiber diameters. We conclude that sex and nutrition influence respiratory muscle morphometry.

Dantrolene sodium is able to reduce the resting ionic [Ca2+]i in muscle from humans with malignant hyperthermia.

Malignant hyperthermia (MH) is a hereditary myopathy, triggered when susceptible patients are exposed to a depolarizing muscle relaxant and/or potent volatile anesthetics. We have studied the effects of dantrolene on the free [Ca2+]i of intercostal muscle biopsies obtained from two MH-susceptible patients before and after administration of dantrolene orally (2.5 mg/kg for 3 days) and intravenously (1.0 mg/kg 2 hours before the biopsy). The free [Ca2+]i was measured by Ca2+-selective microelectrodes. The mean resting free [Ca2+]i in the MH-susceptible muscle before dantrolene treatment was 0.42 +/- 0.01 microM (mean +/- SEM, n = 12). The administration of dantrolene reduced this value to 0.27 +/- 0.01 microM (n = 14). There was no detectable difference in the resting membrane potential after dantrolene. These results represent the first direct demonstration that dantrolene is able to reduce the resting free [Ca2+]i in skeletal muscle of MH-susceptible patients.

Intracellular ionized calcium concentration in muscles from humans with malignant hyperthermia

Ca2+ selective microelectrodes have been used to determine the free myoplasmic [Ca2+] in human skeletal muscle obtained from patients who had developed early signs associated with malignant hyperthermia (MH) during anesthesia. Intercostal muscle biopsies were performed under local anesthesia in four MH patients 15 days to 4 months after developing the MH crisis and in three control subjects. We used only microelectrodes that showed a Nernstian response between pCa3 and pCa7 (30.5 mV per decade at 37 degrees C). Membrane resting potential (V(m)) and calcium potential (V(Ca)) were obtained from superficial fibers. The free cytosolic [Ca2+] was 0.39 +/- 0.1 microM (mean +/- SEM, n = 18) in muscle fibers obtained from malignant hyperthermic patients, whereas in control subjects it was 0.11 +/- 0.02 microM (n = 10). These results suggest that this syndrome might be related to an abnormally high myoplasmic free resting calcium concentration, probably due to a defective function of the plasma membrane or the sarcoplasmic reticulum.

Intercostal muscle biopsy studies in myasthenia gravis: clinical correlations and the direct effects of drugs and myasthenic serum.

We have found a wide range of mean MEPP amplitude in intercostal muscle biopsies from 43 patients with MG, including several values in the normal range. There was no correlation between MEPP amplitude and the severity of clinical disease as assessed by manual muscle testing or by single-fiber EMG measurements of jitter in arm muscles. Through most of these patients were in a state of clinical remission or marked improvement after treatment with prednisone, we could not attribute the difference between our results and those of others to this factor alone. The application of morphine, meperidine and aminoglycoside antibiotics to intercostal muscle in vitro confirms effects previously demonstrated in rat muscle: (1) At equal therapeutic concentrations, meperidine has greater neuromuscular blocking effects than does morphine, but neither has significant effects at concentrations achieved in the serum clinically. (2) Tobramycin, netilmicin and neomycin have varying severity and sites of action, but their effects are the same in human myasthenic muscle as in normal rat muscle. Bath application of serum from myasthenic patients produces an acute, reversible worsening of neuromuscular blockade in myasthenic muscle. Electrophysiologic measurements in intercostal biopsies from patients with MG can provide information about the basic abnormality of neuromuscular transmission in this disease and can confirm the relevance of studies made in animal muscle.

Content of acetylcholine receptor and antibodies bound to receptor in myasthenia gravis, experimental autoimmune myasthenia gravis, and Eaton‐Lambert syndrome

The acetylcholine receptor content of intercostal muscle biopsies from patients with myasthenia gravis is reduced, and many of the remaining receptors have antibodies bound to them. Decreased receptor content and inhibition of receptor activity by bound antibodies can account for the reduced amplitude of the miniature endplate potential in these patients. Patients with Eaton-Lambert syndrome, who release reduced numbers of acetylcholine quanta, have normal receptor content and miniature endplate potential amplitude, and do not have antibodies bound to their receptors. As the severity of myasthenia gravis increases, the content of receptor remaining unbound by antibody decreases, as does miniature endplate potential amplitude. Similar decreases in receptor content and similar concentrations of antibody-labeled receptor are found in rats with experimental autoimmune myasthenia gravis. Regardless of the severity of their myasthenia, receptor content in rats does not decrease much below 40 percent of normal, and increasing weakness is associated with the binding of antibodies to an increasing fraction of the remaining receptors. In both myasthenia gravis and its animal model, a decrease in the content of fully active receptor is probably the most important factor impairing neuromuscular transmission. Pathologic mechanisms responsible for decreased receptor content and alteration of postsynaptic membrane morphology are discussed.