Optimization of Xenografting Methods for Generating Human Skeletal Muscle in Mice.

Xenografts of human skeletal muscle generated in mice can be used to study muscle pathology and to test drugs designed to treat myopathies and muscular dystrophies for their efficacy and specificity in human tissue. We previously developed methods to generate mature human skeletal muscles in immunocompromised mice starting with human myogenic precursor cells (hMPCs) from healthy individuals and individuals with facioscapulohumeral muscular dystrophy (FSHD). Here, we examine a series of alternative treatments at each stage in order to optimize engraftment. We show that (i) X-irradiation at 25Gy is optimal in preventing regeneration of murine muscle while supporting robust engraftment and the formation of human fibers without significant murine contamination; (ii) hMPC lines differ in their capacity to engraft; (iii) some hMPC lines yield grafts that respond better to intermittent neuromuscular electrical stimulation (iNMES) than others; (iv) some lines engraft better in male than in female mice; (v) coinjection of hMPCs with laminin, gelatin, Matrigel, or Growdex does not improve engraftment; (vi) BaCl2 is an acceptable replacement for cardiotoxin, but other snake venom preparations and toxins, including the major component of cardiotoxin, cytotoxin 5, are not; and (vii) generating grafts in both hindlimbs followed by iNMES of each limb yields more robust grafts than housing mice in cages with running wheels. Our results suggest that replacing cardiotoxin with BaCl2 and engrafting both tibialis anterior muscles generates robust grafts of adult human muscle tissue in mice.

µ-Crystallin in Mouse Skeletal Muscle Promotes a Shift from Glycolytic toward Oxidative Metabolism.

µ-Crystallin, encoded by the CRYM gene, binds the thyroid hormones, T3 and T4. Because T3 and T4 are potent regulators of metabolism and gene expression, and CRYM levels in human skeletal muscle can vary widely, we investigated the effects of overexpression of Crym. We generated transgenic mice, Crym tg, that expressed Crym protein specifically in skeletal muscle at levels 2.6-147.5 fold higher than in controls. Muscular functions, Ca2+ transients, contractile force, fatigue, running on treadmills or wheels, were not significantly altered, although T3 levels in tibialis anterior (TA) muscle were elevated ~190-fold and serum T4 was decreased 1.2-fold. Serum T3 and thyroid stimulating hormone (TSH) levels were unaffected. Crym transgenic mice studied in metabolic chambers showed a significant decrease in the respiratory exchange ratio (RER) corresponding to a 13.7% increase in fat utilization as an energy source compared to controls. Female but not male Crym tg mice gained weight more rapidly than controls when fed high fat or high simple carbohydrate diets. Although labeling for myosin heavy chains showed no fiber type differences in TA or soleus muscles, application of machine learning algorithms revealed small but significant morphological differences between Crym tg and control soleus fibers. RNA-seq and gene ontology enrichment analysis showed a significant shift towards genes associated with slower muscle function and its metabolic correlate, ß-oxidation. Protein expression showed a similar shift, though with little overlap. Our study shows that µ-crystallin plays an important role in determining substrate utilization in mammalian muscle and that high levels of µ-crystallin are associated with a shift toward greater fat metabolism.

Keratin 18 is an integral part of the intermediate filament network in murine skeletal muscle.

Intermediate filaments (IFs) contribute to force transmission, cellular integrity, and signaling in skeletal muscle. We previously identified keratin 19 (Krt19) as a muscle IF protein. We now report the presence of a second type I muscle keratin, Krt18. Krt18 mRNA levels are about half those for Krt19 and only 1:1,000th those for desmin; the protein was nevertheless detectable in immunoblots. Muscle function, measured by maximal isometric force in vivo, was moderately compromised in Krt18-knockout (Krt18-KO) or dominant-negative mutant mice (Krt18 DN), but structure was unaltered. Exogenous Krt18, introduced by electroporation, was localized in a reticulum around the contractile apparatus in wild-type muscle and to a lesser extent in muscle lacking Krt19 or desmin or both proteins. Exogenous Krt19, which was either reticular or aggregated in controls, became reticular more frequently in Krt19-null than in Krt18-null, desmin-null, or double-null muscles. Desmin was assembled into the reticulum normally in all genotypes. Notably, all three IF proteins appeared in overlapping reticular structures. We assessed the effect of Krt18 on susceptibility to injury in vivo by electroporating siRNA into tibialis anterior (TA) muscles of control and Krt19-KO mice and testing 2 wk later. Results showed a 33% strength deficit (reduction in maximal torque after injury) compared with siRNA-treated controls. Conversely, electroporation of siRNA to Krt19 into Krt18-null TA yielded a strength deficit of 18% after injury compared with controls. Our results suggest that Krt18 plays a complementary role to Krt19 in skeletal muscle in both assembling keratin-based filaments and transducing contractile force.

Muscle xenografts reproduce key molecular features of facioscapulohumeral muscular dystrophy.

Aberrant expression of DUX4, a gene unique to humans and primates, causes Facioscapulohumeral Muscular Dystrophy-1 (FSHD), yet the pathogenic mechanism is unknown. As transgenic overexpression models have largely failed to replicate the genetic changes seen in FSHD, many studies of endogenously expressed DUX4 have been limited to patient biopsies and myogenic cell cultures, which never fully differentiate into mature muscle fibers. We have developed a method to xenograft immortalized human muscle precursor cells from patients with FSHD and first-degree relative controls into the tibialis anterior muscle compartment of immunodeficient mice, generating human muscle xenografts. We report that FSHD cells mature into organized and innervated human muscle fibers with minimal contamination of murine myonuclei. They also reconstitute the satellite cell niche within the xenografts. FSHD xenografts express DUX4 and DUX4 downstream targets, retain the 4q35 epigenetic signature of their original donors, and express a novel protein biomarker of FSHD, SLC34A2. Ours is the first scalable, mature in vivo human model of FSHD. It should be useful for studies of the pathogenic mechanism of the disease as well as for testing therapeutic strategies targeting DUX4 expression.

Dynamic flood modeling essential to assess the coastal impacts of climate change.

Coastal inundation due to sea level rise (SLR) is projected to displace hundreds of millions of people worldwide over the next century, creating significant economic, humanitarian, and national-security challenges. However, the majority of previous efforts to characterize potential coastal impacts of climate change have focused primarily on long-term SLR with a static tide level, and have not comprehensively accounted for dynamic physical drivers such as tidal non-linearity, storms, short-term climate variability, erosion response and consequent flooding responses. Here we present a dynamic modeling approach that estimates climate-driven changes in flood-hazard exposure by integrating the effects of SLR, tides, waves, storms, and coastal change (i.e. beach erosion and cliff retreat). We show that for California, USA, the world's 5th largest economy, over $150 billion of property equating to more than 6% of the state's GDP and 600,000 people could be impacted by dynamic flooding by 2100; a three-fold increase in exposed population than if only SLR and a static coastline are considered. The potential for underestimating societal exposure to coastal flooding is greater for smaller SLR scenarios, up to a seven-fold increase in exposed population and economic interests when considering storm conditions in addition to SLR. These results highlight the importance of including climate-change driven dynamic coastal processes and impacts in both short-term hazard mitigation and long-term adaptation planning.

Effect of Ibuprofen on Skeletal Muscle of Dysferlin-Null Mice.

Ibuprofen, a nonsteroidal anti-inflammatory drug, and nitric oxide (NO) donors have been reported to reduce the severity of muscular dystrophies in mice associated with the absence of dystrophin or α-sarcoglycan, but their effects on mice that are dystrophic due to the absence of dysferlin have not been examined. We have tested ibuprofen, as well as isosorbide dinitrate (ISDN), a NO donor, to learn whether used alone or together they protect dysferlin-null muscle in A/J mice from large strain injury (LSI) induced by a series of high strain lengthening contractions. Mice were maintained on chow containing ibuprofen and ISDN for 4 weeks. They were then subjected to LSI and maintained on the drugs for 3 additional days. We measured loss of torque immediately following injury and at day 3 postinjury, fiber necrosis, and macrophage infiltration at day 3 postinjury, and serum levels of the drugs at the time of euthanasia. Loss of torque immediately after injury was not altered by the drugs. However, the torque on day 3 postinjury significantly decreased as a function of ibuprofen concentration in the serum (range, 0.67-8.2 µg/ml), independent of ISDN. The effects of ISDN on torque loss at day 3 postinjury were not significant. In long-term studies of dysferlinopathic BlAJ mice, lower doses of ibuprofen had no effects on muscle morphology, but reduced treadmill running by 40%. Our results indicate that ibuprofen can have deleterious effects on dysferlin-null muscle and suggest that its use at pharmacological doses should be avoided by individuals with dysferlinopathies.

Neuromuscular electrical stimulation promotes development in mice of mature human muscle from immortalized human myoblasts.

BACKGROUND: Studies of the pathogenic mechanisms underlying human myopathies and muscular dystrophies often require animal models, but models of some human diseases are not yet available. Methods to promote the engraftment and development of myogenic cells from individuals with such diseases in mice would accelerate such studies and also provide a useful tool for testing therapeutics. Here, we investigate the ability of immortalized human myogenic precursor cells (hMPCs) to form mature human myofibers following implantation into the hindlimbs of non-obese diabetic-Rag1 (null) IL2rγ (null) (NOD-Rag)-immunodeficient mice. RESULTS: We report that hindlimbs of NOD-Rag mice that are X-irradiated, treated with cardiotoxin, and then injected with immortalized control hMPCs or hMPCs from an individual with facioscapulohumeral muscular dystrophy (FSHD) develop mature human myofibers. Furthermore, intermittent neuromuscular electrical stimulation (iNMES) of the peroneal nerve of the engrafted limb enhances the development of mature fibers in the grafts formed by both immortal cell lines. With control cells, iNMES increases the number and size of the human myofibers that form and promotes closer fiber-to-fiber packing. The human myofibers in the graft are innervated, fully differentiated, and minimally contaminated with murine myonuclei. CONCLUSIONS: Our results indicate that control and FSHD human myofibers can form in mice engrafted with hMPCs and that iNMES enhances engraftment and subsequent development of mature human muscle.

Myopathic changes in murine skeletal muscle lacking synemin.

Diseases of striated muscle linked to intermediate filament (IF) proteins are associated with defects in the organization of the contractile apparatus and its links to costameres, which connect the sarcomeres to the cell membrane. Here we study the role in skeletal muscle of synemin, a type IV IF protein, by examining mice null for synemin (synm-null). Synm-null mice have a mild skeletal muscle phenotype. Tibialis anterior (TA) muscles show a significant decrease in mean fiber diameter, a decrease in twitch and tetanic force, and an increase in susceptibility to injury caused by lengthening contractions. Organization of proteins associated with the contractile apparatus and costameres is not significantly altered in the synm-null. Elastimetry of the sarcolemma and associated contractile apparatus in extensor digitorum longus myofibers reveals a reduction in tension consistent with an increase in sarcolemmal deformability. Although fatigue after repeated isometric contractions is more marked in TA muscles of synm-null mice, the ability of the mice to run uphill on a treadmill is similar to controls. Our results suggest that synemin contributes to linkage between costameres and the contractile apparatus and that the absence of synemin results in decreased fiber size and increased sarcolemmal deformability and susceptibility to injury. Thus synemin plays a moderate but distinct role in fast twitch skeletal muscle.

Physiotherapy extended-role practitioner for individuals with hip and knee arthritis: patient perspectives of a rural/urban partnership.

PURPOSE: To explore the perspectives of people with hip and knee arthritis regarding a physiotherapy extended-role practitioner (ERP) model of care in a rural setting. METHOD: Using semi-structured interviews, a qualitative descriptive case study was undertaken with 13 participants from a rural family practice located in the province of Ontario, Canada, who had all been assessed by an ERP. Transcribed interviews were analyzed for emergent themes. RESULTS: Three main themes were identified: (1) timely access to care, (2) distance as a factor in seeking care, and (3) perceptions of the ERP model of care. CONCLUSIONS: Participants reported many positive experiences with the physiotherapy ERP rural model. Processes related to minimizing travel required to access care are important for those in rural communities. An ERP model of care offers competent care that includes musculoskeletal diagnosis as well as time for educating patients and addressing questions.

Objectif : Explorer les perspectives des personnes qui ont de l'arthrite dans la hanche et le genou en ce qui concerne un modèle de soins basé sur le rôle élargi du professionnel de la physiothérapie (REP) en contexte rural. Méthode   : Nous avons entrepris une étude de cas descriptive qualitative basée sur des entrevues structurées menées auprès de 13 participants d'un cabinet de médecine familiale en milieu rural dans la province d'Ontario, au Canada, qui avaient tous été évalués par un professionnel à rôle élargi. Nous avons analysé les entrevues transcrites pour en dégager des thèmes émergents. Résultats : Nous avons dégagé trois grands thèmes : (1) l'accès rapide aux soins; (2) la distance comme facteur de la recherche de soins; (3) les perceptions du modèle de soins basé sur le rôle élargi du professionnel. Conclusions : Des participants ont signalé de nombreuses expériences positives du modèle rural REP en physiothérapie. Les moyens de réduire au minimum les déplacements pour avoir accès aux soins sont importants pour les membres des communautés rurales. Un modèle de soins basé sur le rôle élargi du professionnel offre des soins compétents qui incluent la capacité de poser un diagnostic de l'appareil locomoteur et du temps pour l'éducation et pour répondre aux questions.

Influences of desmin and keratin 19 on passive biomechanical properties of mouse skeletal muscle.

In skeletal muscle fibers, forces must be transmitted between the plasma membrane and the intracellular contractile lattice, and within this lattice between adjacent myofibrils. Based on their prevalence, biomechanical properties and localization, desmin and keratin intermediate filaments (IFs) are likely to participate in structural connectivity and force transmission. We examined the passive load-bearing response of single fibers from the extensor digitorum longus (EDL) muscles of young (3 months) and aged (10 months) wild-type, desmin-null, K19-null, and desmin/K19 double-null mice. Though fibers are more compliant in all mutant genotypes compared to wild-type, the structural response of each genotype is distinct, suggesting multiple mechanisms by which desmin and keratin influence the biomechanical properties of myofibers. This work provides additional insight into the influences of IFs on structure-function relationships in skeletal muscle. It may also have implications for understanding the progression of desminopathies and other IF-related myopathies.

Unmasking potential intracellular roles for dysferlin through improved immunolabeling methods.

Mutations in the DYSF gene that severely reduce the levels of the protein dysferlin are implicated in muscle-wasting syndromes known as dysferlinopathies. Although studies of its function in skeletal muscle have focused on its potential role in repairing the plasma membrane, dysferlin has also been found, albeit inconsistently, in the sarcoplasm of muscle fibers. The aim of this article is to study the localization of dysferlin in skeletal muscle through optimized immunolabeling methods. We studied the localization of dysferlin in control rat skeletal muscle using several different methods of tissue collection and subsequent immunolabeling. We then applied our optimized immunolabeling methods on human cadaveric muscle, control and dystrophic human muscle biopsies, and control and dysferlin-deficient mouse muscle. Our data suggest that dysferlin is present in a reticulum of the sarcoplasm, similar but not identical to those containing the dihydropyridine receptors and distinct from the distribution of the sarcolemmal protein dystrophin. Our data illustrate the importance of tissue fixation and antigen unmasking for proper immunolocalization of dysferlin. They suggest that dysferlin has an important function in the internal membrane systems of skeletal muscle, involved in calcium homeostasis and excitation-contraction coupling.

Daily multidisciplinary rounds to implement the ventilator bundle decreases ventilator-associated pneumonia in trauma patients: but does it affect outcome?

BACKGROUND: The incidence of ventilator-associated pneumonia (VAP) in trauma patients can be decreased with use of the ventilator bundle (VAPB). Our VAP rate remained high despite the adoption of the VAPB. To better implement the VAPB, a multidisciplinary team composed of the surgical intensive care unit (SICU) nursing staff, physician, and respiratory therapist reviewed briefly a checklist of VAPB goals for each patient before morning attending rounds. We hypothesized that such daily goal rounds (GR) focused on the VAPB would decrease the VAP rate. METHODS: A pre-GR ten-month period (November 2006 to August 2007) was compared with the ten-month period (September 2007 to June 2008) with daily GRs. The occurrence of VAPs was tallied prospectively in all intubated trauma patients using the National Nosocomial Infection Surveillance criteria. Patient characteristics and outcome data were obtained from our trauma registry and medical records. Patient characteristics were similar in the 85 pre-GR patients and the 89 GR patients. RESULTS: The number of VAPs decreased 67% in the GR patients (15 pre-GR vs. 5 GR; p=0.02); however, the all-cause mortality rate remained similar (16.5% vs. 21.3%; p=0.41). When patients were divided into those with and without VAP, there was a significant increase in mean ventilator, SICU, and hospital days in patients with VAP (p=0.01 for all). There were only two deaths among trauma patients with VAP. CONCLUSION: Daily multidisciplinary GRs focused on the VAPB can decrease the incidence of VAP significantly in trauma patients. Ventilator-associated pneumonia correlated with extended mean ventilator, SICU, and hospital days. Interestingly, despite a significant decrease in VAP, a decrease in the mortality rate was not observed. Given the small number of deaths in the VAP cohort, this study has insufficient statistical power to elucidate the true impact of GR intervention or VAP on the mortality rate in trauma patients.

Physiology, structure, and susceptibility to injury of skeletal muscle in mice lacking keratin 19-based and desmin-based intermediate filaments.

Intermediate filaments, composed of desmin and of keratins, play important roles in linking contractile elements to each other and to the sarcolemma in striated muscle. Our previous results show that the tibialis anterior (TA) muscles of mice lacking keratin 19 (K19) lose costameres, accumulate mitochondria under the sarcolemma, and generate lower specific tension than controls. Here we compare the physiology and morphology of TA muscles of mice lacking K19 with muscles lacking desmin or both proteins [double knockout (DKO)]. K19-/- mice and DKO mice showed a threefold increase in the levels of creatine kinase (CK) in the serum. The absence of desmin caused a larger change in specific tension (-40%) than the absence of K19 (-19%) and played the predominant role in contractile function (-40%) and decreased tolerance to exercise in the DKO muscle. By contrast, the absence of both proteins was required to obtain a significantly greater loss of contractile torque after injury (-48%) compared with wild type (-39%), as well as near-complete disruption of costameres. The DKO muscle also showed a significantly greater misalignment of myofibrils than either mutant alone. In contrast, large subsarcolemmal gaps and extensive accumulation of mitochondria were only seen in K19-null TA muscles, and the absence of both K19 and desmin yielded milder phenotypes. Our results suggest that keratin filaments containing K19- and desmin-based intermediate filaments can play independent, complementary, or antagonistic roles in the physiology and morphology of fast-twitch skeletal muscle.

Absence of keratin 19 in mice causes skeletal myopathy with mitochondrial and sarcolemmal reorganization.

Intermediate filaments, composed of desmin and of keratins, play important roles in linking contractile elements to each other and to the sarcolemma in striated muscle. We examined the contractile properties and morphology of fast-twitch skeletal muscle from mice lacking keratin 19. Tibialis anterior muscles of keratin-19-null mice showed a small but significant decrease in mean fiber diameter and in the specific force of tetanic contraction, as well as increased plasma creatine kinase levels. Costameres at the sarcolemma of keratin-19-null muscle, visualized with antibodies against spectrin or dystrophin, were disrupted and the sarcolemma was separated from adjacent myofibrils by a large gap in which mitochondria accumulated. The costameric dystrophin-dystroglycan complex, which co-purified with gamma-actin, keratin 8 and keratin 19 from striated muscles of wild-type mice, co-purified with gamma-actin but not keratin 8 in the mutant. Our results suggest that keratin 19 in fast-twitch skeletal muscle helps organize costameres and links them to the contractile apparatus, and that the absence of keratin 19 disrupts these structures, resulting in loss of contractile force, altered distribution of mitochondria and mild myopathy. This is the first demonstration of a mammalian phenotype associated with a genetic perturbation of keratin 19.

Public health lessons learned from analysis of New York City subway injuries.

Serious subway injuries are devastating to their young victims and have high rates of mortality and amputation. We identified the urban population at greatest risk for subway injuries and investigated the influence of local economies on injury rates. We propose using changes in social conditions as a "trigger" for increased vigilance and protective measures at times of higher risk.

Devastating consequences of subway accidents: traumatic amputations.

The efficiencies of the subway system are tempered by the occurrence of accidents, some with devastating injuries. The purpose of this study is to examine our experience with traumatic amputations after subway accidents. A retrospective trauma registry review (1989-2003) of 41 patients who presented to Bellevue Hospital, New York City, with amputations from subway accidents was undertaken to examine the following end points: age, sex, Injury Severity Score, time and mechanism of accident, history of psychiatric disorders and alcohol use, admission vital signs, Glasgow Coma Scale score, amputation type, associated injuries, limb salvage rate, operative procedures, mortality, and disposition. Elevated alcohol levels and prior psychiatric diagnoses were present in 39 per cent and 17 per cent of the patients, respectively. Patients were stable on admission with a mean systolic blood pressure of 114 mmHg, hematocrit of 32, and Glasgow Coma Scale score range of 13 to 15. The most common amputation was below knee, and patients underwent an average of three operative procedures. Limb salvage was attempted in eight patients with no successes. Amputation wound infection rate was 32 per cent and mortality rate was 5 per cent. Victims of subway trauma who arrive at the hospital with devastating amputations have an excellent chance of surviving to discharge.

Association of small ankyrin 1 with the sarcoplasmic reticulum.

Small ankyrin 1, or sAnk1, is a small, alternatively spliced product of the erythroid ankyrin gene, ANK1, that is expressed in striated muscle and concentrated in the network sarcoplasmic reticulum (SR) surrounding the Z disks and M lines. We have characterized sAnk1 in muscle homogenates and SR vesicles, and have identified the region that targets it to the network SR. Selective extractions and partitioning into Triton X-114 show that sAnk1 behaves like the SR Ca-ATPase and so is an integral protein of the SR membrane. Mild proteolytic treatment of isolated SR vesicles indicates that sAnk1 is oriented with its hydrophilic, C-terminal sequence exposed to the solution, which is equivalent to the cytoplasmic face of the SR membrane in situ. SDS-PAGE in non-reducing gels suggests that sAnk1 is present as dimers and larger oligomers in the native SR. These results suggest that sAnk1 is oligomeric and oriented with its C-terminus exposed to the cytoplasm, where it may interact with proteins of the contractile apparatus. The N-terminal 29 amino acid hydrophobic sequence of sAnk1, which is predicted to span the SR membrane, is sufficient to target proteins to and anchor them in internal membranes of HEK 293 cells. It also targets reporter proteins to the network SR of skeletal myofibers and is thus the first example of a sequence that targets proteins to a particular compartment of the SR.

Specific interaction of the actin-binding domain of dystrophin with intermediate filaments containing keratin 19.

Cytokeratins 8 and 19 concentrate at costameres of striated muscle and copurify with the dystrophin-glycoprotein complex, perhaps through the interaction of the cytokeratins with the actin-binding domain of dystrophin. We overexpressed dystrophin's actin-binding domain (Dys-ABD), K8 and K19, as well as closely related proteins, in COS-7 cells to assess the basis and specificity of their interaction. Dys-ABD alone associated with actin microfilaments. Expressed with K8 and K19, which form filaments, Dys-ABD associated preferentially with the cytokeratins. This interaction was specific, as the homologous ABD of betaI-spectrin failed to interact with K8/K19 filaments, and Dys-ABD did not associate with desmin or K8/K18 filaments. Studies in COS-7 cells and in vitro showed that Dys-ABD binds directly and specifically to K19. Expressed in muscle fibers in vivo, K19 accumulated in the myoplasm in structures that contained dystrophin and spectrin and disrupted the organization of the sarcolemma. K8 incorporated into sarcomeres, with no effect on the sarcolemma. Our results show that dystrophin interacts through its ABD with K19 specifically and are consistent with the idea that cytokeratins associate with dystrophin at the sarcolemma of striated muscle.

Cloning and characterization of cytokeratins 8 and 19 in adult rat striated muscle. Interaction with the dystrophin glycoprotein complex.

We used degenerate primers for the amino- and carboxyl-terminal ends of the rod domains of intermediate filament proteins in reverse transcriptase-PCR experiments to identify and clone cytokeratins 8 and 19 (K8 and K19) from cardiac muscle of the adult rat. Northern blots showed that K8 has a 2.2-kb transcript and K19 has a 1.9-kb transcript in both adult cardiac and skeletal muscles. Immunolocalization of the cytokeratins in adult cardiac muscle with isoform-specific antibodies for K8 and K19 showed labeling at Z-lines within the muscle fibers and at Z-line and M-line domains at costameres at the sarcolemmal membrane. Dystrophin and K19 could be co-immunoprecipitated and co-purified from extracts of cardiac muscle, suggesting a link between the cytokeratins and the dystrophin-based cytoskeleton at the sarcolemma. Furthermore, transfection experiments indicate that K8 and K19 may associate with dystrophin through a specific interaction with its actin-binding domain. Consistent with this observation, the cytokeratins are disrupted at the sarcolemmal membrane of skeletal muscle of the mdx mouse that lacks dystrophin. Together these results indicate that at least two cytokeratins are expressed in adult striated muscle, where they may contribute to the organization of both the myoplasm and sarcolemma.