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Sample records for mdx mouse duodenum

  1. The Molecular Basis of Muscular Dystrophy in the mdx Mouse: A Point Mutation

    NASA Astrophysics Data System (ADS)

    Sicinski, Piotr; Geng, Yan; Ryder-Cook, Allan S.; Barnard, Eric A.; Darlison, Mark G.; Barnard, Pene J.

    1989-06-01

    The mdx mouse is an X-linked myopathic mutant, an animal model for human Duchenne muscular dystrophy. In both mouse and man the mutations lie within the dystrophin gene, but the phenotypic differences of the disease in the two species confer much interest on the molecular basis of the mdx mutation. The complementary DNA for mouse dystrophin has been cloned, and the sequence has been used in the polymerase chain reaction to amplify normal and mdx dystrophin transcripts in the area of the mdx mutation. Sequence analysis of the amplification products showed that the mdx mouse has a single base substitution within an exon, which causes premature termination of the polypeptide chain.

  2. Caspase-12 ablation preserves muscle function in the mdx mouse

    PubMed Central

    Moorwood, Catherine; Barton, Elisabeth R.

    2014-01-01

    Duchenne muscular dystrophy (DMD) is a devastating muscle wasting disease caused by mutations in dystrophin. Several downstream consequences of dystrophin deficiency are triggers of endoplasmic reticulum (ER) stress, including loss of calcium homeostasis, hypoxia and oxidative stress. During ER stress, misfolded proteins accumulate in the ER lumen and the unfolded protein response (UPR) is triggered, leading to adaptation or apoptosis. We hypothesized that ER stress is heightened in dystrophic muscles and contributes to the pathology of DMD. We observed increases in the ER stress markers BiP and cleaved caspase-4 in DMD patient biopsies, compared with controls, and an increase in multiple UPR pathways in muscles of the dystrophin-deficient mdx mouse. We then crossed mdx mice with mice null for caspase-12, the murine equivalent of human caspase-4, which are resistant to ER stress. We found that deleting caspase-12 preserved mdx muscle function, resulting in a 75% recovery of both specific force generation and resistance to eccentric contractions. The compensatory hypertrophy normally found in mdx muscles was normalized in the absence of caspase-12; this was found to be due to decreased fibre sizes, and not to a fibre type shift or a decrease in fibrosis. Fibre central nucleation was not significantly altered in the absence of caspase-12, but muscle fibre degeneration found in the mdx mouse was reduced almost to wild-type levels. In conclusion, we have identified heightened ER stress and abnormal UPR signalling as novel contributors to the dystrophic phenotype. Caspase-4 is therefore a potential therapeutic target for DMD. PMID:24879640

  3. Disease course in mdx:utrophin+/− mice: comparison of three mouse models of Duchenne muscular dystrophy

    PubMed Central

    McDonald, Abby A; Hebert, Sadie L; Kunz, Matthew D; Ralles, Steven J; McLoon, Linda K

    2015-01-01

    The mdx mouse model of Duchenne muscular dystrophy (DMD) is used to study disease mechanisms and potential treatments, but its pathology is less severe than DMD patients. Other mouse models were developed to more closely mimic the human disease based on knowledge that upregulation of utrophin has a protective effect in mdx muscle. An mdx:utrophin−/− (dko) mouse was created, which had a severe disease phenotype and a shortened life span. An mdx:utrophin+/− mouse was also created, which had an intermediate disease phenotype compared to the mdx and dko mice. To determine the usefulness of mdx:utrophin+/− mice for long-term DMD studies, limb muscle pathology and function were assessed across the life span of wild-type, mdx, mdx:utrophin+/−, and dko mice. Muscle function assessment, specifically grip duration and rotarod performance, demonstrated that mdx:utrophin+/− mice were weaker for a longer time than mdx mice. Mean myofiber area was smaller in mdx:utrophin+/− mice compared to mdx mice at 12 months. Mdx:utrophin+/− mice had a higher percentage of centrally nucleated myofibers compared to mdx mice at 6 and 12 months. Collagen I and IV density was significantly higher in mdx:utrophin+/− muscle compared to mdx at most ages examined. Generally, mdx:utrophin+/− mice showed an intermediate disease phenotype over a longer time course compared to the mdx and dko mice. While they do not genetically mirror human DMD, mdx:utrophin+/− mice may be a more useful animal model than mdx or dko mice for investigating long-term efficacy of potential treatments when fibrosis or muscle function is the focus. PMID:25921779

  4. X-irradiation improves mdx mouse muscle as a model of myofiber loss in DMD

    SciTech Connect

    Wakeford, S.; Watt, D.J.; Partridge, T.A. )

    1991-01-01

    The mdx mouse, although a genetic and biochemical homologue of human Duchenne muscular dystrophy (DMD), presents a comparatively mild histopathological and clinical phenotype. These differences are partially attributable to the greater efficacy of regeneration in the mdx mouse than in DMD muscle. To lessen this disparity, we have used a single dose of X-irradiation (16 Gy) to inhibit regeneration in one leg of mdx mice. The result is an almost complete block of muscle fiber regeneration leading to progressive loss of muscle fibers and their replacement by loose connective tissue. Surviving fibers are mainly peripherally nucleated and, surprisingly, of large diameter. Thus, X-irradiation converts mdx muscle to a model system in which the degenerative process can be studied in isolation from the complicating effect of myofiber regeneration. This system should be of use for testing methods of alleviating the myofiber degeneration which is common to mdx and DMD.

  5. Defective regulation of energy metabolism in mdx-mouse skeletal muscles.

    PubMed

    Even, P C; Decrouy, A; Chinet, A

    1994-12-01

    Our previous finding of a reduced energy metabolism in slow- and fast-twitch skeletal muscle fibres from the murine model of Duchenne muscular dystrophy (the mdx mouse) led us to examine the importance of intracellular glucose availability for a normal energy turnover. To this end, basal and KCl-stimulated (20.9 mM total extracellular K+) rates of glucose uptake (GUP) and heat production were measured in isolated, glucose-incubated (5 mM) soleus and extensor digitorum longus muscles from mdx and control C57B1/10 mice, in the presence and in the absence of insulin (1.7 nM). Under all conditions and for both muscle types, glucose uptake values for mdx and control muscles were similar although heat production was lower in mdx muscles. The marked stimulation of GUP by insulin in both mdx and control muscles had only minor effects on heat production. In contrast, glucose deprivation or inhibition of glycolysis with 2-deoxy-D-glucose (5 mM) significantly decreased heat production in control muscles only, which attenuated, although did not suppress, the difference in basal heat production between mdx and control muscles. Stimulation of heat production by a short-chain fatty acid salt (octanoate, 2 mM) was significantly less marked in mdx than in control muscles. Increased cytoplasmic synthesis of CoA by addition of 5 mM pantothenate (vitamin B5) increased the thermogenic response to glucose more in mdx than in control muscles. We conclude that the low energy turnover in mdx-mouse muscle fibres is not due to a decrease of intracellular glucose availability, but rather to a decreased oxidative utilization of glucose and free fatty acids. We suggest that some enzyme complex of the tricarboxylic acid cycle or inefficiency of CoA transport in the mitochondria could be involved. PMID:7999003

  6. Increased metallothionein in mouse liver, kidneys, and duodenum during lactation.

    PubMed

    Solaiman, D; Jonah, M M; Miyazaki, W; Ho, G; Bhattacharyya, M H

    2001-03-01

    Lactation-induced increases in cadmium absorption and retention have been demonstrated in mid-lactating mice, but no systematic measurements of endogenous metal-binding protein concentrations during lactation have been reported. Using Cd/hemoglobin radioassay, this study detected significant increases in metallothionein (MT) concentrations in liver (4-fold), kidneys (2-fold), and duodenum (2-fold), but not jejunum, of mouse dams on days 13 and 20 of lactation. These increases occurred in the absence of cadmium exposure and were specific to the lactation period; dams 5 days after weaning showed MT levels that were similar to those of nonpregnant (NP) mice. Similarly, Northern blot analyses of livers from lactating mice demonstrated that MT mRNA concentrations in maternal liver during mid-lactation were 6-fold higher than those observed 5 days after pups were weaned. Gel filtration of final supernatants from the Cd/hemoglobin assay confirmed that the Cd-binding molecule induced during lactation was indeed metallothionein. In addition, chromatographic analyses of cytosols from tissues taken from dams administered small amounts of Cd (66 ng/mouse) showed that the trace amounts of Cd absorbed through the maternal gastrointestinal tract during mid-lactation were also bound to the MT. These results indicate MT induction in mouse dams occurs as a physiological consequence of lactation, requiring no external stimulus. This induced MT participates in binding low levels of dietary cadmium consumed by the dam. During lactation, elevated maternal MT may affect pathways for essential trace metals as well as sequester toxic metals harmful to the neonate. Multiparous humans may have increased risk of accumulating environmental Cd. PMID:11222885

  7. Assessing functional performance in the mdx mouse model.

    PubMed

    Aartsma-Rus, Annemieke; van Putten, Maaike

    2014-01-01

    Duchenne muscular dystrophy (DMD) is a severe and progressive muscle wasting disorder for which no cure is available. Nevertheless, several potential pharmaceutical compounds and gene therapy approaches have progressed into clinical trials. With improvement in muscle function being the most important end point in these trials, a lot of emphasis has been placed on setting up reliable, reproducible, and easy to perform functional tests to pre clinically assess muscle function, strength, condition, and coordination in the mdx mouse model for DMD. Both invasive and noninvasive tests are available. Tests that do not exacerbate the disease can be used to determine the natural history of the disease and the effects of therapeutic interventions (e.g. forelimb grip strength test, two different hanging tests using either a wire or a grid and rotarod running). Alternatively, forced treadmill running can be used to enhance disease progression and/or assess protective effects of therapeutic interventions on disease pathology. We here describe how to perform these most commonly used functional tests in a reliable and reproducible manner. Using these protocols based on standard operating procedures enables comparison of data between different laboratories. PMID:24747372

  8. Assessing Functional Performance in the Mdx Mouse Model

    PubMed Central

    Aartsma-Rus, Annemieke; van Putten, Maaike

    2014-01-01

    Duchenne muscular dystrophy (DMD) is a severe and progressive muscle wasting disorder for which no cure is available. Nevertheless, several potential pharmaceutical compounds and gene therapy approaches have progressed into clinical trials. With improvement in muscle function being the most important end point in these trials, a lot of emphasis has been placed on setting up reliable, reproducible, and easy to perform functional tests to pre clinically assess muscle function, strength, condition, and coordination in the mdx mouse model for DMD. Both invasive and noninvasive tests are available. Tests that do not exacerbate the disease can be used to determine the natural history of the disease and the effects of therapeutic interventions (e.g. forelimb grip strength test, two different hanging tests using either a wire or a grid and rotarod running). Alternatively, forced treadmill running can be used to enhance disease progression and/or assess protective effects of therapeutic interventions on disease pathology. We here describe how to perform these most commonly used functional tests in a reliable and reproducible manner. Using these protocols based on standard operating procedures enables comparison of data between different laboratories. PMID:24747372

  9. Ventilatory chemosensory drive is blunted in the mdx mouse model of Duchenne Muscular Dystrophy (DMD).

    PubMed

    Mosqueira, Matias; Baby, Santhosh M; Lahiri, Sukhamay; Khurana, Tejvir S

    2013-01-01

    Duchenne Muscular Dystrophy (DMD) is caused by mutations in the DMD gene resulting in an absence of dystrophin in neurons and muscle. Respiratory failure is the most common cause of mortality and previous studies have largely concentrated on diaphragmatic muscle necrosis and respiratory failure component. Here, we investigated the integrity of respiratory control mechanisms in the mdx mouse model of DMD. Whole body plethysmograph in parallel with phrenic nerve activity recordings revealed a lower respiratory rate and minute ventilation during normoxia and a blunting of the hypoxic ventilatory reflex in response to mild levels of hypoxia together with a poor performance on a hypoxic stress test in mdx mice. Arterial blood gas analysis revealed low PaO2 and pH and high PaCO2 in mdx mice. To investigate chemosensory respiratory drive, we analyzed the carotid body by molecular and functional means. Dystrophin mRNA and protein was expressed in normal mice carotid bodies however, they are absent in mdx mice. Functional analysis revealed abnormalities in Dejours test and the early component of the hypercapnic ventilatory reflex in mdx mice. Together, these results demonstrate a malfunction in the peripheral chemosensory drive that would be predicted to contribute to the respiratory failure in mdx mice. These data suggest that investigating and monitoring peripheral chemosensory drive function may be useful for improving the management of DMD patients with respiratory failure. PMID:23922741

  10. Ventilatory Chemosensory Drive Is Blunted in the mdx Mouse Model of Duchenne Muscular Dystrophy (DMD)

    PubMed Central

    Mosqueira, Matias; Baby, Santhosh M.; Khurana, Tejvir S.

    2013-01-01

    Duchenne Muscular Dystrophy (DMD) is caused by mutations in the DMD gene resulting in an absence of dystrophin in neurons and muscle. Respiratory failure is the most common cause of mortality and previous studies have largely concentrated on diaphragmatic muscle necrosis and respiratory failure component. Here, we investigated the integrity of respiratory control mechanisms in the mdx mouse model of DMD. Whole body plethysmograph in parallel with phrenic nerve activity recordings revealed a lower respiratory rate and minute ventilation during normoxia and a blunting of the hypoxic ventilatory reflex in response to mild levels of hypoxia together with a poor performance on a hypoxic stress test in mdx mice. Arterial blood gas analysis revealed low PaO2 and pH and high PaCO2 in mdx mice. To investigate chemosensory respiratory drive, we analyzed the carotid body by molecular and functional means. Dystrophin mRNA and protein was expressed in normal mice carotid bodies however, they are absent in mdx mice. Functional analysis revealed abnormalities in Dejours test and the early component of the hypercapnic ventilatory reflex in mdx mice. Together, these results demonstrate a malfunction in the peripheral chemosensory drive that would be predicted to contribute to the respiratory failure in mdx mice. These data suggest that investigating and monitoring peripheral chemosensory drive function may be useful for improving the management of DMD patients with respiratory failure. PMID:23922741

  11. Biochemical and Functional Comparisons of mdx and Sgcg−/− Muscular Dystrophy Mouse Models

    PubMed Central

    Roberts, Nathan W.; Holley-Cuthrell, Jenan; Gonzalez-Vega, Magdalis; Mull, Aaron J.; Heydemann, Ahlke

    2015-01-01

    Mouse models have provided an essential platform to investigate facets of human diseases, from etiology, diagnosis, and prognosis, to potential treatments. Muscular dystrophy (MD) is the most common human genetic disease occurring in approximately 1 in 2500 births. The mdx mouse, which is dystrophin-deficient, has long been used to model this disease. However, this mouse strain displays a rather mild disease course compared to human patients. The mdx mice have been bred to additional genetically engineered mice to worsen the disease. Alternatively, other genes which cause human MD have been genetically disrupted in mice. We are now comparing disease progression from one of these alternative gene disruptions, the γ-sarcoglycan null mouse Sgcg−/− on the DBA2/J background, to the mdx mouse line. This paper aims to assess the time-course severity of the disease in the mouse models and determine which is best for MD research. The Sgcg−/− mice have a more severe phenotype than the mdx mice. Muscle function was assessed by plethysmography and echocardiography. Histologically the Sgcg−/− mice displayed increased fibrosis and variable fiber size. By quantitative Evan's blue dye uptake and hydroxyproline content two key disease determinants, membrane permeability and fibrosis respectively, were also proven worse in the Sgcg−/− mice. PMID:26064876

  12. Decrease in Prosaposin in the Dystrophic mdx Mouse Brain

    PubMed Central

    Gao, Hui-ling; Li, Cheng; Nabeka, Hiroaki; Shimokawa, Tetsuya; Kobayashi, Naoto; Saito, Shouichiro; Wang, Zhan-You; Cao, Ya-ming; Matsuda, Seiji

    2013-01-01

    Background Duchenne muscular dystrophy caused by a mutation in the X-linked dystrophin gene induces metabolic and structural disorders in the brain. A lack of dystrophin in brain structures is involved in impaired cognitive function. Prosaposin (PS), a neurotrophic factor, is abundant in the choroid plexus and various brain regions. We investigated whether PS serves as a link between dystrophin loss and gross and/or ultrastructural brain abnormalities. Methodology/Principal Findings The distribution of PS in the brains of juvenile and adult mdx mice was investigated by immunochemistry, Western blotting, and in situ hybridization. Immunochemistry revealed lower levels of PS in the cytoplasm of neurons of the cerebral cortex, hippocampus, cerebellum, and choroid plexus in mdx mice. Western blotting confirmed that PS levels were lower in these brain regions in both juveniles and adults. Even with low PS production in the choroids plexus, there was no significant PS decrease in cerebrospinal fluid (CSF). In situ hybridization revealed that the primary form of PS mRNA in both normal and mdx mice was Pro+9, a secretory-type PS, and the hybridization signals for Pro+9 in the above-mentioned brain regions were weaker in mdx mice than in normal mice. We also investigated mitogen-activated protein kinase signalling. Stronger activation of ERK1/2 was observed in mdx mice, ERK1/2 activity was positively correlated with PS activity, and exogenous PS18 stimulated both p-ERK1/2 and PS in SH-SY5Y cells. Conclusions/Significance Low levels of PS and its receptors suggest the participation of PS in some pathological changes in the brains of mdx mice. PMID:24244600

  13. Metabolic profile of dystrophic mdx mouse muscles analyzed with in vitro magnetic resonance spectroscopy (MRS).

    PubMed

    Martins-Bach, Aurea B; Bloise, Antonio C; Vainzof, Mariz; Rahnamaye Rabbani, Said

    2012-10-01

    Duchenne muscular dystrophy (DMD) is a recessive X-linked form of muscular dystrophy characterized by progressive and irreversible degeneration of the muscles. The mdx mouse is the classical animal model for DMD, showing similar molecular and protein defects. The mdx mouse, however, does not show significant muscle weakness, and the diaphragm muscle is significantly more degenerated than skeletal muscles. In this work, (1)H magnetic resonance spectroscopy (MRS) was used to study the metabolic profile of quadriceps and diaphragm muscles from mdx and control mice. Using principal components analysis (PCA), the animals were separated into groups according to age and lineages. The classification was compared to histopathological analysis. Among the 24 metabolites identified from the nuclear MR spectra, only 19 were used by the PCA program for classification purposes. These can be important key biomarkers associated with the progression of degeneration in mdx muscles and with natural aging in control mice. Glutamate, glutamine, succinate, isoleucine, acetate, alanine and glycerol were increased in mdx samples as compared to control mice, in contrast to carnosine, taurine, glycine, methionine and creatine that were decreased. These results suggest that MRS associated with pattern recognition analysis can be a reliable tool to assess the degree of pathological and metabolic alterations in the dystrophic tissue, thereby affording the possibility of evaluation of beneficial effects of putative therapies. PMID:22673895

  14. Dystropathology increases energy expenditure and protein turnover in the Mdx mouse model of Duchenne muscular dystrophy

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The skeletal muscles in Duchenne muscular dystrophy and the mdx mouse model lack functional dystrophin and undergo repeated bouts of necrosis, regeneration, and growth. These processes have a high metabolic cost. However, the consequences for whole body energy and protein metabolism, and on the diet...

  15. Cardiomyocyte Regeneration in the mdx Mouse Model of Nonischemic Cardiomyopathy

    PubMed Central

    Laval, Steven; Owens, William Andrew

    2015-01-01

    Endogenous regeneration has been demonstrated in the mammalian heart after ischemic injury. However, approximately one-third of cases of heart failure are secondary to nonischemic heart disease and cardiac regeneration in these cases remains relatively unexplored. We, therefore, aimed at quantifying the rate of new cardiomyocyte formation at different stages of nonischemic cardiomyopathy. Six-, 12-, 29-, and 44-week-old mdx mice received a 7 day pulse of BrdU. Quantification of isolated cardiomyocyte nuclei was undertaken using cytometric analysis to exclude nondiploid nuclei. Between 6–7 and 12–13 weeks, there was a statistically significant increase in the number of BrdU-labeled nuclei in the mdx hearts compared with wild-type controls. This difference was lost by the 29–30 week time point, and a significant decrease in cardiomyocyte generation was observed in both the control and mdx hearts by 44–45 weeks. Immunohistochemical analysis demonstrated BrdU-labeled nuclei exclusively in mononucleated cardiomyocytes. This study demonstrates cardiomyocyte regeneration in a nonischemic model of mammalian cardiomyopathy, controlling for changes in nuclear ploidy, which is lost with age, and confirms a decrease in baseline rates of cardiomyocyte regeneration with aging. While not attempting to address the cellular source of regeneration, it confirms the potential utility of innate regeneration as a therapeutic target. PMID:25749191

  16. Dystropathology increases energy expenditure and protein turnover in the mdx mouse model of duchenne muscular dystrophy.

    PubMed

    Radley-Crabb, Hannah G; Marini, Juan C; Sosa, Horacio A; Castillo, Liliana I; Grounds, Miranda D; Fiorotto, Marta L

    2014-01-01

    The skeletal muscles in Duchenne muscular dystrophy and the mdx mouse model lack functional dystrophin and undergo repeated bouts of necrosis, regeneration, and growth. These processes have a high metabolic cost. However, the consequences for whole body energy and protein metabolism, and on the dietary requirements for these macronutrients at different stages of the disease, are not well-understood. This study used juvenile (4- to 5- wk-old) and adult (12- to 14-wk-old) male dystrophic C57BL/10ScSn-mdx/J and age-matched C57BL/10ScSn/J control male mice to measure total and resting energy expenditure, food intake, spontaneous activity, body composition, whole body protein turnover, and muscle protein synthesis rates. In juvenile mdx mice that have extensive muscle damage, energy expenditure, muscle protein synthesis, and whole body protein turnover rates were higher than in age-matched controls. Adaptations in food intake and decreased activity were insufficient to meet the increased energy and protein needs of juvenile mdx mice and resulted in stunted growth. In (non-growing) adult mdx mice with less severe dystropathology, energy expenditure, muscle protein synthesis, and whole body protein turnover rates were also higher than in age-matched controls. Food intake was sufficient to meet their protein and energy needs, but insufficient to result in fat deposition. These data show that dystropathology impacts the protein and energy needs of mdx mice and that tailored dietary interventions are necessary to redress this imbalance. If not met, the resultant imbalance blunts growth, and may limit the benefits of therapies designed to protect and repair dystrophic muscles. PMID:24586653

  17. A Mathematical Model of Skeletal Muscle Disease and Immune Response in the mdx Mouse

    PubMed Central

    Evans, Nicholas P.; Grange, Robert W.

    2014-01-01

    Duchenne muscular dystrophy (DMD) is a genetic disease that results in the death of affected boys by early adulthood. The genetic defect responsible for DMD has been known for over 25 years, yet at present there is neither cure nor effective treatment for DMD. During early disease onset, the mdx mouse has been validated as an animal model for DMD and use of this model has led to valuable but incomplete insights into the disease process. For example, immune cells are thought to be responsible for a significant portion of muscle cell death in the mdx mouse; however, the role and time course of the immune response in the dystrophic process have not been well described. In this paper we constructed a simple mathematical model to investigate the role of the immune response in muscle degeneration and subsequent regeneration in the mdx mouse model of Duchenne muscular dystrophy. Our model suggests that the immune response contributes substantially to the muscle degeneration and regeneration processes. Furthermore, the analysis of the model predicts that the immune system response oscillates throughout the life of the mice, and the damaged fibers are never completely cleared. PMID:25013809

  18. Early manifestation of alteration in cardiac function in dystrophin deficient mdx mouse using 3D CMR tagging

    PubMed Central

    Li, Wei; Liu, Wei; Zhong, Jia; Yu, Xin

    2009-01-01

    Background Duchenne muscular dystrophy (DMD) is caused by the absence of the cytoskeletal protein, dystrophin. In DMD patients, dilated cardiomyopathy leading to heart failure may occur during adolescence. However, early cardiac dysfunction is frequently undetected due to physical inactivity and generalized debilitation. The objective of this study is to determine the time course of cardiac functional alterations in mdx mouse, a mouse model of DMD, by evaluating regional ventricular function with CMR tagging. Methods In vivo myocardial function was evaluated by 3D CMR tagging in mdx mice at early (2 months), middle (7 months) and late (10 months) stages of disease development. Global cardiac function, regional myocardial wall strains, and ventricular torsion were quantified. Myocardial lesions were assessed with Masson's trichrome staining. Results Global contractile indexes were similar between mdx and C57BL/6 mice in each age group. Histology analysis showed that young mdx mice were free of myocardial lesions. Interstitial fibrosis was present in 7 month mdx mice, with further development into patches or transmural lesions at 10 months of age. As a result, 10 month mdx mice showed significantly reduced regional strain and torsion. However, young mdx mice showed an unexpected increase in regional strain and torsion, while 7 month mdx mice displayed similar regional ventricular function as the controls. Conclusion Despite normal global ventricular function, CMR tagging detected a biphasic change in myocardial wall strain and torsion, with an initial increase at young age followed by progressive decrease at older ages. These results suggest that CMR tagging can provide more sensitive measures of functional alterations than global functional indexes in dystrophin-related cardiomyopathies. PMID:19849858

  19. Fast skeletal myofibers of mdx mouse, model of Duchenne muscular dystrophy, express connexin hemichannels that lead to apoptosis.

    PubMed

    Cea, Luis A; Puebla, Carlos; Cisterna, Bruno A; Escamilla, Rosalba; Vargas, Aníbal A; Frank, Marina; Martínez-Montero, Paloma; Prior, Carmen; Molano, Jesús; Esteban-Rodríguez, Isabel; Pascual, Ignacio; Gallano, Pía; Lorenzo, Gustavo; Pian, Héctor; Barrio, Luis C; Willecke, Klaus; Sáez, Juan C

    2016-07-01

    Skeletal muscles of patients with Duchenne muscular dystrophy (DMD) show numerous alterations including inflammation, apoptosis, and necrosis of myofibers. However, the molecular mechanism that explains these changes remains largely unknown. Here, the involvement of hemichannels formed by connexins (Cx HCs) was evaluated in skeletal muscle of mdx mouse model of DMD. Fast myofibers of mdx mice were found to express three connexins (39, 43 and 45) and high sarcolemma permeability, which was absent in myofibers of mdx Cx43(fl/fl)Cx45(fl/fl):Myo-Cre mice (deficient in skeletal muscle Cx43/Cx45 expression). These myofibers did not show elevated basal intracellular free Ca(2+) levels, immunoreactivity to phosphorylated p65 (active NF-κB), eNOS and annexin V/active Caspase 3 (marker of apoptosis) but presented dystrophin immunoreactivity. Moreover, muscles of mdx Cx43(fl/fl)Cx45(fl/fl):Myo-Cre mice exhibited partial decrease of necrotic features (big cells and high creatine kinase levels). Accordingly, these muscles showed similar macrophage infiltration as control mdx muscles. Nonetheless, the hanging test performance of mdx Cx43(fl/fl)Cx45(fl/fl):Myo-Cre mice was significantly better than that of control mdx Cx43(fl/fl)Cx45(fl/fl) mice. All three Cxs found in skeletal muscles of mdx mice were also detected in fast myofibers of biopsy specimens from patients with muscular dystrophy. Thus, reduction of Cx expression and/or function of Cx HCs may be potential therapeutic approaches to abrogate myofiber apoptosis in DMD. PMID:26803842

  20. Towards developing standard operating procedures for pre-clinical testing in the mdx mouse model of Duchenne muscular dystrophy

    PubMed Central

    Grounds, Miranda D.; Radley, Hannah G.; Lynch, Gordon S.; Nagaraju, Kanneboyina; De Luca, Annamaria

    2008-01-01

    This review discusses various issues to consider when developing standard operating procedures for pre-clinical studies in the mdx mouse model of Duchenne muscular dystrophy (DMD). The review describes and evaluates a wide range of techniques used to measure parameters of muscle pathology in mdx mice and identifies some basic techniques that might comprise standardised approaches for evaluation. While the central aim is to provide a basis for the development of standardised procedures to evaluate efficacy of a drug or a therapeutic strategy, a further aim is to gain insight into pathophysiological mechanisms in order to identify other therapeutic targets. The desired outcome is to enable easier and more rigorous comparison of pre-clinical data from different laboratories around the world, in order to accelerate identification of the best pre-clinical therapies in the mdx mouse that will fast-track translation into effective clinical treatments for DMD. PMID:18499465

  1. A PCR-based assay for the wild-type dystrophin gene transferred into the mdx mouse.

    PubMed

    Shrager, J B; Naji, A; Kelly, A M; Stedman, H H

    1992-10-01

    Myoblast transfer has emerged as a promising treatment for inherited myopathies such as Duchenne muscular dystrophy (DMD). Further development of the technique's therapeutic potential requires an experimental system in which issues of graft rejection can be clearly discriminated from those related to myoblast biology. Here we report the development and initial application of a quantitative assay for myogenic cells bearing a wild-type dystrophin gene following transfer into the mdx mouse. The technique relies upon the ability of a mutagenizing polymerase chain reaction (PCR) primer to create a new restriction site in the amplification production of the wild-type, but not the mdx dystrophin gene. The ratio of host to donor cells can be determined from muscle biopsies as small as 1 mg, regardless of donor H-2 background. This simple technique should allow a number of basic questions related to myoblast and direct gene transfer to be addressed using the mdx mouse model. PMID:1357549

  2. Recombinase-mediated reprogramming and dystrophin gene addition in mdx mouse induced pluripotent stem cells.

    PubMed

    Zhao, Chunli; Farruggio, Alfonso P; Bjornson, Christopher R R; Chavez, Christopher L; Geisinger, Jonathan M; Neal, Tawny L; Karow, Marisa; Calos, Michele P

    2014-01-01

    A cell therapy strategy utilizing genetically-corrected induced pluripotent stem cells (iPSC) may be an attractive approach for genetic disorders such as muscular dystrophies. Methods for genetic engineering of iPSC that emphasize precision and minimize random integration would be beneficial. We demonstrate here an approach in the mdx mouse model of Duchenne muscular dystrophy that focuses on the use of site-specific recombinases to achieve genetic engineering. We employed non-viral, plasmid-mediated methods to reprogram mdx fibroblasts, using phiC31 integrase to insert a single copy of the reprogramming genes at a safe location in the genome. We next used Bxb1 integrase to add the therapeutic full-length dystrophin cDNA to the iPSC in a site-specific manner. Unwanted DNA sequences, including the reprogramming genes, were then precisely deleted with Cre resolvase. Pluripotency of the iPSC was analyzed before and after gene addition, and ability of the genetically corrected iPSC to differentiate into myogenic precursors was evaluated by morphology, immunohistochemistry, qRT-PCR, FACS analysis, and intramuscular engraftment. These data demonstrate a non-viral, reprogramming-plus-gene addition genetic engineering strategy utilizing site-specific recombinases that can be applied easily to mouse cells. This work introduces a significant level of precision in the genetic engineering of iPSC that can be built upon in future studies. PMID:24781921

  3. Quantitative T2 Combined with Texture Analysis of Nuclear Magnetic Resonance Images Identify Different Degrees of Muscle Involvement in Three Mouse Models of Muscle Dystrophy: mdx, Largemyd and mdx/Largemyd

    PubMed Central

    Martins-Bach, Aurea B.; Malheiros, Jackeline; Matot, Béatrice; Martins, Poliana C. M.; Almeida, Camila F.; Caldeira, Waldir; Ribeiro, Alberto F.; Loureiro de Sousa, Paulo; Azzabou, Noura; Tannús, Alberto; Carlier, Pierre G.; Vainzof, Mariz

    2015-01-01

    Quantitative nuclear magnetic resonance imaging (MRI) has been considered a promising non-invasive tool for monitoring therapeutic essays in small size mouse models of muscular dystrophies. Here, we combined MRI (anatomical images and transverse relaxation time constant—T2—measurements) to texture analyses in the study of four mouse strains covering a wide range of dystrophic phenotypes. Two still unexplored mouse models of muscular dystrophies were analyzed: The severely affected Largemyd mouse and the recently generated and worst double mutant mdx/Largemyd mouse, as compared to the mildly affected mdx and normal mice. The results were compared to histopathological findings. MRI showed increased intermuscular fat and higher muscle T2 in the three dystrophic mouse models when compared to the wild-type mice (T2: mdx/Largemyd: 37.6±2.8 ms; mdx: 35.2±4.5 ms; Largemyd: 36.6±4.0 ms; wild-type: 29.1±1.8 ms, p<0.05), in addition to higher muscle T2 in the mdx/Largemyd mice when compared to mdx (p<0.05). The areas with increased muscle T2 in the MRI correlated spatially with the identified histopathological alterations such as necrosis, inflammation, degeneration and regeneration foci. Nevertheless, muscle T2 values were not correlated with the severity of the phenotype in the 3 dystrophic mouse strains, since the severely affected Largemyd showed similar values than both the mild mdx and worst mdx/Largemyd lineages. On the other hand, all studied mouse strains could be unambiguously identified with texture analysis, which reflected the observed differences in the distribution of signals in muscle MRI. Thus, combined T2 intensity maps and texture analysis is a powerful approach for the characterization and differentiation of dystrophic muscles with diverse genotypes and phenotypes. These new findings provide important noninvasive tools in the evaluation of the efficacy of new therapies, and most importantly, can be directly applied in human translational research

  4. Quantitative T2 combined with texture analysis of nuclear magnetic resonance images identify different degrees of muscle involvement in three mouse models of muscle dystrophy: mdx, Largemyd and mdx/Largemyd.

    PubMed

    Martins-Bach, Aurea B; Malheiros, Jackeline; Matot, Béatrice; Martins, Poliana C M; Almeida, Camila F; Caldeira, Waldir; Ribeiro, Alberto F; Loureiro de Sousa, Paulo; Azzabou, Noura; Tannús, Alberto; Carlier, Pierre G; Vainzof, Mariz

    2015-01-01

    Quantitative nuclear magnetic resonance imaging (MRI) has been considered a promising non-invasive tool for monitoring therapeutic essays in small size mouse models of muscular dystrophies. Here, we combined MRI (anatomical images and transverse relaxation time constant-T2-measurements) to texture analyses in the study of four mouse strains covering a wide range of dystrophic phenotypes. Two still unexplored mouse models of muscular dystrophies were analyzed: The severely affected Largemyd mouse and the recently generated and worst double mutant mdx/Largemyd mouse, as compared to the mildly affected mdx and normal mice. The results were compared to histopathological findings. MRI showed increased intermuscular fat and higher muscle T2 in the three dystrophic mouse models when compared to the wild-type mice (T2: mdx/Largemyd: 37.6±2.8 ms; mdx: 35.2±4.5 ms; Largemyd: 36.6±4.0 ms; wild-type: 29.1±1.8 ms, p<0.05), in addition to higher muscle T2 in the mdx/Largemyd mice when compared to mdx (p<0.05). The areas with increased muscle T2 in the MRI correlated spatially with the identified histopathological alterations such as necrosis, inflammation, degeneration and regeneration foci. Nevertheless, muscle T2 values were not correlated with the severity of the phenotype in the 3 dystrophic mouse strains, since the severely affected Largemyd showed similar values than both the mild mdx and worst mdx/Largemyd lineages. On the other hand, all studied mouse strains could be unambiguously identified with texture analysis, which reflected the observed differences in the distribution of signals in muscle MRI. Thus, combined T2 intensity maps and texture analysis is a powerful approach for the characterization and differentiation of dystrophic muscles with diverse genotypes and phenotypes. These new findings provide important noninvasive tools in the evaluation of the efficacy of new therapies, and most importantly, can be directly applied in human translational research. PMID

  5. Discovery of serum protein biomarkers in the mdx mouse model and cross-species comparison to Duchenne muscular dystrophy patients

    PubMed Central

    Hathout, Yetrib; Marathi, Ramya L.; Rayavarapu, Sree; Zhang, Aiping; Brown, Kristy J.; Seol, Haeri; Gordish-Dressman, Heather; Cirak, Sebahattin; Bello, Luca; Nagaraju, Kanneboyina; Partridge, Terry; Hoffman, Eric P.; Takeda, Shin'ichi; Mah, Jean K.; Henricson, Erik; McDonald, Craig

    2014-01-01

    It is expected that serum protein biomarkers in Duchenne muscular dystrophy (DMD) will reflect disease pathogenesis, progression and aid future therapy developments. Here, we describe use of quantitative in vivo stable isotope labeling in mammals to accurately compare serum proteomes of wild-type and dystrophin-deficient mdx mice. Biomarkers identified in serum from two independent dystrophin-deficient mouse models (mdx-Δ52 and mdx-23) were concordant with those identified in sera samples of DMD patients. Of the 355 mouse sera proteins, 23 were significantly elevated and 4 significantly lower in mdx relative to wild-type mice (P-value < 0.001). Elevated proteins were mostly of muscle origin: including myofibrillar proteins (titin, myosin light chain 1/3, myomesin 3 and filamin-C), glycolytic enzymes (aldolase, phosphoglycerate mutase 2, beta enolase and glycogen phosphorylase), transport proteins (fatty acid-binding protein, myoglobin and somatic cytochrome-C) and others (creatine kinase M, malate dehydrogenase cytosolic, fibrinogen and parvalbumin). Decreased proteins, mostly of extracellular origin, included adiponectin, lumican, plasminogen and leukemia inhibitory factor receptor. Analysis of sera from 1 week to 7 months old mdx mice revealed age-dependent changes in the level of these biomarkers with most biomarkers acutely elevated at 3 weeks of age. Serum analysis of DMD patients, with ages ranging from 4 to 15 years old, confirmed elevation of 20 of the murine biomarkers in DMD, with similar age-related changes. This study provides a panel of biomarkers that reflect muscle activity and pathogenesis and should prove valuable tool to complement natural history studies and to monitor treatment efficacy in future clinical trials. PMID:25027324

  6. Discovery of serum protein biomarkers in the mdx mouse model and cross-species comparison to Duchenne muscular dystrophy patients.

    PubMed

    Hathout, Yetrib; Marathi, Ramya L; Rayavarapu, Sree; Zhang, Aiping; Brown, Kristy J; Seol, Haeri; Gordish-Dressman, Heather; Cirak, Sebahattin; Bello, Luca; Nagaraju, Kanneboyina; Partridge, Terry; Hoffman, Eric P; Takeda, Shin'ichi; Mah, Jean K; Henricson, Erik; McDonald, Craig

    2014-12-15

    It is expected that serum protein biomarkers in Duchenne muscular dystrophy (DMD) will reflect disease pathogenesis, progression and aid future therapy developments. Here, we describe use of quantitative in vivo stable isotope labeling in mammals to accurately compare serum proteomes of wild-type and dystrophin-deficient mdx mice. Biomarkers identified in serum from two independent dystrophin-deficient mouse models (mdx-Δ52 and mdx-23) were concordant with those identified in sera samples of DMD patients. Of the 355 mouse sera proteins, 23 were significantly elevated and 4 significantly lower in mdx relative to wild-type mice (P-value < 0.001). Elevated proteins were mostly of muscle origin: including myofibrillar proteins (titin, myosin light chain 1/3, myomesin 3 and filamin-C), glycolytic enzymes (aldolase, phosphoglycerate mutase 2, beta enolase and glycogen phosphorylase), transport proteins (fatty acid-binding protein, myoglobin and somatic cytochrome-C) and others (creatine kinase M, malate dehydrogenase cytosolic, fibrinogen and parvalbumin). Decreased proteins, mostly of extracellular origin, included adiponectin, lumican, plasminogen and leukemia inhibitory factor receptor. Analysis of sera from 1 week to 7 months old mdx mice revealed age-dependent changes in the level of these biomarkers with most biomarkers acutely elevated at 3 weeks of age. Serum analysis of DMD patients, with ages ranging from 4 to 15 years old, confirmed elevation of 20 of the murine biomarkers in DMD, with similar age-related changes. This study provides a panel of biomarkers that reflect muscle activity and pathogenesis and should prove valuable tool to complement natural history studies and to monitor treatment efficacy in future clinical trials. PMID:25027324

  7. Long-term treatment with naproxcinod significantly improves skeletal and cardiac disease phenotype in the mdx mouse model of dystrophy.

    PubMed

    Uaesoontrachoon, Kitipong; Quinn, James L; Tatem, Kathleen S; Van Der Meulen, Jack H; Yu, Qing; Phadke, Aditi; Miller, Brittany K; Gordish-Dressman, Heather; Ongini, Ennio; Miglietta, Daniela; Nagaraju, Kanneboyina

    2014-06-15

    In Duchenne muscular dystrophy (DMD) patients and the mouse model of DMD, mdx, dystrophin deficiency causes a decrease and mislocalization of muscle-specific neuronal nitric oxide synthase (nNOSμ), leading to functional impairments. Previous studies have shown that nitric oxide (NO) donation associated with anti-inflammatory action has beneficial effects in dystrophic mouse models. In this study, we have systematically investigated the effects of naproxcinod, an NO-donating naproxen derivative, on the skeletal and cardiac disease phenotype in mdx mice. Four-week-old mdx and C57BL/10 mice were treated with four different concentrations (0, 10, 21 and 41 mg/kg) of naproxcinod and 0.9 mg/kg of prednisolone in their food for 9 months. All mice were subjected to twice-weekly treadmill sessions, and functional and behavioral parameters were measured at 3, 6 and 9 months of treatment. In addition, we evaluated in vitro force contraction, optical imaging of inflammation, echocardiography and blood pressure (BP) at the 9-month endpoint prior to sacrifice. We found that naproxcinod treatment at 21 mg/kg resulted in significant improvement in hindlimb grip strength and a 30% decrease in inflammation in the fore- and hindlimbs of mdx mice. Furthermore, we found significant improvement in heart function, as evidenced by improved fraction shortening, ejection fraction and systolic BP. In addition, the long-term detrimental effects of prednisolone typically seen in mdx skeletal and heart function were not observed at the effective dose of naproxcinod. In conclusion, our results indicate that naproxcinod has significant potential as a safe therapeutic option for the treatment of muscular dystrophies. PMID:24463621

  8. Long-term treatment with naproxcinod significantly improves skeletal and cardiac disease phenotype in the mdx mouse model of dystrophy

    PubMed Central

    Uaesoontrachoon, Kitipong; Quinn, James L; Tatem, Kathleen S; Van Der Meulen, Jack H; Yu, Qing; Phadke, Aditi; Miller, Brittany K; Gordish-Dressman, Heather; Ongini, Ennio; Miglietta, Daniela; Nagaraju, Kanneboyina

    2014-01-01

    In Duchenne muscular dystrophy (DMD) patients and the mouse model of DMD, mdx, dystrophin deficiency causes a decrease and mislocalization of muscle-specific neuronal nitric oxide synthase (nNOSμ), leading to functional impairments. Previous studies have shown that nitric oxide (NO) donation associated with anti-inflammatory action has beneficial effects in dystrophic mouse models. In this study, we have systematically investigated the effects of naproxcinod, an NO-donating naproxen derivative, on the skeletal and cardiac disease phenotype in mdx mice. Four-week-old mdx and C57BL/10 mice were treated with four different concentrations (0, 10, 21 and 41 mg/kg) of naproxcinod and 0.9 mg/kg of prednisolone in their food for 9 months. All mice were subjected to twice-weekly treadmill sessions, and functional and behavioral parameters were measured at 3, 6 and 9 months of treatment. In addition, we evaluated in vitro force contraction, optical imaging of inflammation, echocardiography and blood pressure (BP) at the 9-month endpoint prior to sacrifice. We found that naproxcinod treatment at 21 mg/kg resulted in significant improvement in hindlimb grip strength and a 30% decrease in inflammation in the fore- and hindlimbs of mdx mice. Furthermore, we found significant improvement in heart function, as evidenced by improved fraction shortening, ejection fraction and systolic BP. In addition, the long-term detrimental effects of prednisolone typically seen in mdx skeletal and heart function were not observed at the effective dose of naproxcinod. In conclusion, our results indicate that naproxcinod has significant potential as a safe therapeutic option for the treatment of muscular dystrophies. PMID:24463621

  9. A new immuno-, dystrophin-deficient model, the NSG-mdx(4Cv) mouse, provides evidence for functional improvement following allogeneic satellite cell transplantation.

    PubMed

    Arpke, Robert W; Darabi, Radbod; Mader, Tara L; Zhang, Yu; Toyama, Akira; Lonetree, Cara-Lin; Nash, Nardina; Lowe, Dawn A; Perlingeiro, Rita C R; Kyba, Michael

    2013-08-01

    Transplantation of a myogenic cell population into an immunodeficient recipient is an excellent way of assessing the in vivo muscle-generating capacity of that cell population. To facilitate both allogeneic and xenogeneic transplantations of muscle-forming cells in mice, we have developed a novel immunodeficient muscular dystrophy model, the NSG-mdx(4Cv) mouse. The IL2Rg mutation, which is linked to the Dmd gene on the X chromosome, simultaneously depletes NK cells and suppresses thymic lymphomas, issues that limit the utility of the SCID/mdx model. The NSG-mdx(4Cv) mouse presents a muscular dystrophy of similar severity to the conventional mdx mouse. We show that this animal supports robust engraftment of both pig and dog muscle mononuclear cells. The question of whether satellite cells prospectively isolated by flow cytometry can confer a functional benefit upon transplantation has been controversial. Using allogeneic Pax7-ZsGreen donors and NSG-mdx(4Cv) recipients, we demonstrate definitively that as few as 900 FACS-isolated satellite cells can provide functional regeneration in vivo, in the form of an increased mean maximal force-generation capacity in cell-transplanted muscles, compared to a sham-injected control group. These studies highlight the potency of satellite cells to improve muscle function and the utility of the NSG-mdx(4Cv) model for studies on muscle regeneration and Duchenne muscular dystrophy therapy. PMID:23606600

  10. Multivariate Data EXplorer (MDX)

    SciTech Connect

    Steed, Chad Allen

    2012-08-01

    The MDX toolkit facilitates exploratory data analysis and visualization of multivariate datasets. MDX provides and interactive graphical user interface to load, explore, and modify multivariate datasets stored in tabular forms. MDX uses an extended version of the parallel coordinates plot and scatterplots to represent the data. The user can perform rapid visual queries using mouse gestures in the visualization panels to select rows or columns of interest. The visualization panel provides coordinated multiple views whereby selections made in one plot are propagated to the other plots. Users can also export selected data or reconfigure the visualization panel to explore relationships between columns and rows in the data.

  11. Myofibrillar misalignment correlated to triad disappearance of mdx mouse gastrocnemius muscle probed by SHG microscopy.

    PubMed

    Rouède, Denis; Coumailleau, Pascal; Schaub, Emmanuel; Bellanger, Jean-Jacques; Blanchard-Desce, Mireille; Tiaho, François

    2014-03-01

    We show that the canonical single frequency sarcomeric SHG intensity pattern (SHG-IP) of control muscles is converted to double frequency sarcomeric SHG-IP in preserved mdx mouse gastrocnemius muscles in the vicinity of necrotic fibers. These double frequency sarcomeric SHG-IPs are often spatially correlated to double frequency sarcomeric two-photon excitation fluorescence (TPEF) emitted from Z-line and I-bands and to one centered spot SHG angular intensity pattern (SHG-AIP) suggesting that these patterns are signature of myofibrillar misalignement. This latter is confirmed with transmission electron microscopy (TEM). Moreover, a good spatial correlation between SHG signature of myofibrillar misalignment and triad reduction is established. Theoretical simulation of sarcomeric SHG-IP is used to demonstrate the correlation between change of SHG-IP and -AIP and myofibrillar misalignment. The extreme sensitivity of SHG microscopy to reveal the submicrometric organization of A-band thick filaments is highlighted. This report is a first step toward future studies aimed at establishing live SHG signature of myofibrillar misalignment involving excitation contraction defects due to muscle damage and disease. PMID:24688819

  12. Effective detection of corrected dystrophin loci in mdx mouse myogenic precursors.

    PubMed

    Todaro, Marian; Quigley, Anita; Kita, Magdalena; Chin, Judy; Lowes, Kym; Kornberg, Andrew J; Cook, Mark J; Kapsa, Robert

    2007-08-01

    Targeted corrective gene conversion (TCGC) holds much promise as a future therapy for many hereditary diseases in humans. Mutation correction frequencies varying between 0.0001% and 40% have been reported using chimeraplasty, oligoplasty, triplex-forming oligonucleotides, and small corrective PCR amplicons (CPA). However, PCR technologies used to detect correction events risk either falsely indicating or greatly exaggerating the presence of corrected loci. This is a problem that is considerably exacerbated by attempted improvement of the TCGC system using high corrective nucleic acid (CNA) to nuclear ratios. Small fragment homologous replacement (SFHR)-mediated correction of the exon 23 dystrophin (DMD) gene mutation in the mdx mouse model of DMD has been used in this study to evaluate the effect of increasing CPA amounts. In these experiments, we detected extremely high levels of apparently corrected loci and determined that at higher CNA to nuclear ratios the extent of locus correction was highly exaggerated by residual CNA species in the nucleic acids extracted from the treated cells. This study describes a generic locus-specific detection protocol designed to eradicate residual CNA species and avoid the artifactual or exaggerated detection of gene correction. PMID:17394239

  13. Reduction in mdx mouse muscle degeneration by low-intensity endurance exercise: a proteomic analysis in quadriceps muscle of exercised compared with sedentary mdx mice

    PubMed Central

    Fontana, Simona; Schillaci, Odessa; Frinchi, Monica; Giallombardo, Marco; Morici, Giuseppe; Liberto, Valentina Di; Alessandro, Riccardo; De Leo, Giacomo; Perciavalle, Vincenzo; Belluardo, Natale; Mudò, Giuseppa

    2015-01-01

    In our recent study was shown a significant recovery of damaged skeletal muscle of mice with X-linked muscular dystrophy (mdx) following low-intensity endurance exercise, probably by reducing the degeneration of dystrophic muscle. Consequently, in the present work, we aimed to identify proteins involved in the observed reduction in degenerating fibres. To this end, we used proteomic analysis to evaluate changes in the protein profile of quadriceps dystrophic muscles of exercised compared with sedentary mdx mice. Four protein spots were found to be significantly changed and were identified as three isoforms of carbonic anhydrase 3 (CA3) and superoxide dismutase [Cu-Zn] (SODC). Protein levels of CA3 isoforms were significantly up-regulated in quadriceps of sedentary mdx mice and were completely restored to wild–type (WT) mice values, both sedentary and exercised, in quadriceps of exercised mdx mice. Protein levels of SODC were down-regulated in quadriceps of sedentary mdx mice and were significantly restored to WT mice values, both sedentary and exercised, in quadriceps of exercised mdx mice. Western blot data were in agreement with those obtained using proteomic analysis and revealed the presence of one more CA3 isoform that was significantly changed. Based on data found in the present study, it seems that low-intensity endurance exercise may in part contribute to reduce cell degeneration process in mdx muscles, by counteracting oxidative stress. PMID:26182375

  14. Galectin-1 Protein Therapy Prevents Pathology and Improves Muscle Function in the mdx Mouse Model of Duchenne Muscular Dystrophy.

    PubMed

    Van Ry, Pam M; Wuebbles, Ryan D; Key, Megan; Burkin, Dean J

    2015-08-01

    Duchenne muscular dystrophy (DMD) is a fatal neuromuscular disease caused by mutations in the dystrophin gene, leading to the loss of a critical component of the sarcolemmal dystrophin glycoprotein complex. Galectin-1 is a small 14 kDa protein normally found in skeletal muscle and has been shown to be a modifier of immune response, muscle repair, and apoptosis. Galectin-1 levels are elevated in the muscle of mouse and dog models of DMD. Together, these findings led us to hypothesize that Galectin-1 may serve as a modifier of disease progression in DMD. To test this hypothesis, recombinant mouse Galectin-1 was produced and used to treat myogenic cells and the mdx mouse model of DMD. Here we show that intramuscular and intraperitoneal injections of Galectin-1 into mdx mice prevented pathology and improved muscle function in skeletal muscle. These improvements were a result of enhanced sarcolemmal stability mediated by elevated utrophin and α7β1 integrin protein levels. Together our results demonstrate for the first time that Galectin-1 may serve as an exciting new protein therapeutic for the treatment of DMD. PMID:26050991

  15. Diaphragm degeneration and cardiac structure in mdx mouse: potential clinical implications for Duchenne muscular dystrophy.

    PubMed

    Barbin, Isabel Cristina Chagas; Pereira, Juliano Alves; Bersan Rovere, Matheus; de Oliveira Moreira, Drielen; Marques, Maria Julia; Santo Neto, Humberto

    2016-05-01

    We examined the effects of exercise on diaphragm degeneration and cardiomyopathy in dystrophin-deficient mdx mice. Mdx mice (11 months of age) were exercised (swimming) for 2 months to worsen diaphragm degeneration. Control mdx mice were kept sedentary. Morphological evaluation demonstrated increased fibrosis in the diaphragm of exercised mdx mice (33.3 ± 6.0% area of fibrosis) compared with control mdx mice (20.9 ± 1.7% area of fibrosis). Increased (26%) activity of MMP-2, a marker of fibrosis, was detected in the diaphragms from exercised mdx mice. Morphological evaluation of the heart demonstrated a 45% increase in fibrosis in the right ventricle (8.3 ± 0.6% in sedentary vs. 12.0 ± 0.6% of fibrosis in exercised) and in the left ventricle (35% increase) in the exercised mdx mice. The density of inflammatory cells-degenerating cardiomyocytes increased 95% in the right ventricle (2.3 ± 0.6 in sedentary vs. 4.5 ± 0.8 in exercised) and 71% in the left ventricle (1.4 ± 0.6 sedentary vs. 2.4 ± 0.5 exercised). The levels of both active MMP-2 and the pro-fibrotic factor transforming growth factor beta were elevated in the hearts of exercised compared with sedentary mdx mice. The wall thickness to lumen diameter ratio of the pulmonary trunk was significantly increased in the exercised mdx mice (0.11 ± 0.04 in sedentary vs. 0.28 ± 0.12 in exercised), as was the thickness of the right ventricle wall, which suggests the occurrence of pulmonary hypertension in those animals. It is suggested that diaphragm degeneration is a main contributor to right ventricle dystrophic pathology. These findings may be relevant for future interventional studies for Duchenne muscular dystrophy-associated cardiomyopathy. PMID:26822140

  16. Introduction of a human-specific deletion in mouse Cmah increases disease severity in the mdx model of Duchenne muscular dystrophy

    PubMed Central

    Chandrasekharan, Kumaran; Yoon, Jung Hae; Xu, Ying; deVries, Sarah; Camboni, Marybeth; Janssen, Paulus M.L.; Varki, Ajit; Martin, Paul T.

    2010-01-01

    The evolution of humans included introduction of an inactivating deletion in the CMAH gene, which eliminated biosynthesis of N-glycolylneuraminic acid from all human cells. Here we show that this human-specific sialylation change contributes to the marked discrepancy in phenotype between the mdx mouse model for Duchenne muscular dystrophy (DMD) and the human disease. Despite lacking dystrophin protein in almost all muscle cells, mdx mice show slower development, relative to overall lifespan, or reduced severity of a number of clinically relevant disease phenotypes compared to DMD patients. This is especially true for loss of ambulation, cardiac and respiratory muscle weakness, and loss of lifespan, all major phenotypes contributing to DMD morbidity and mortality. All these phenotypes occur at an earlier age or to a greater degree in mdx mice bearing a human-like mutation in the mouse Cmah gene. Altered phenotypes correlate with changes in two mechanisms; reduced strength and expression of the dystrophin-associated glycoprotein complex and increased activation of complement. Activation of complement may be driven by the increased expression of anti-Neu5Gc antibodies in Cmah−/−mdx animals and ultimately by uptake of N-glycolylneuraminic acid, a foreign glycan in humans and Cmah-deficient mice, from dietary sources. Cmah-deficient mdx mice represent a new small animal model for DMD that better approximates the human glycome and its contributions to muscular dystrophy. PMID:20668298

  17. How much dystrophin is enough: the physiological consequences of different levels of dystrophin in the mdx mouse.

    PubMed

    Godfrey, Caroline; Muses, Sofia; McClorey, Graham; Wells, Kim E; Coursindel, Thibault; Terry, Rebecca L; Betts, Corinne; Hammond, Suzan; O'Donovan, Liz; Hildyard, John; El Andaloussi, Samir; Gait, Michael J; Wood, Matthew J; Wells, Dominic J

    2015-08-01

    Splice modulation therapy has shown great clinical promise in Duchenne muscular dystrophy, resulting in the production of dystrophin protein. Despite this, the relationship between restoring dystrophin to established dystrophic muscle and its ability to induce clinically relevant changes in muscle function is poorly understood. In order to robustly evaluate functional improvement, we used in situ protocols in the mdx mouse to measure muscle strength and resistance to eccentric contraction-induced damage. Here, we modelled the treatment of muscle with pre-existing dystrophic pathology using antisense oligonucleotides conjugated to a cell-penetrating peptide. We reveal that 15% homogeneous dystrophin expression is sufficient to protect against eccentric contraction-induced injury. In addition, we demonstrate a >40% increase in specific isometric force following repeated administrations. Strikingly, we show that changes in muscle strength are proportional to dystrophin expression levels. These data define the dystrophin restoration levels required to slow down or prevent disease progression and improve overall muscle function once a dystrophic environment has been established in the mdx mouse model. PMID:25935000

  18. Tissue distribution of the dystrophin-related gene product and expression in the mdx and dy mouse

    SciTech Connect

    Love, D.R.; Marsden, R.F.; Bloomfield, J.F.; Davies, K.E. ); Morris, G.E.; Ellis, J.M. ); Fairbrother, U.; Edwards, Y.H. ); Slater, C.P. ); Parry, D.J. )

    1991-04-15

    The authors have previously reported a dystrophin-related locus (DMDL for Duchenne muscular dystrophy-like) on human chromosome 6 that maps close to the dy mutation on mouse chromosome 10. Here they show that this gene is expressed in a wide range of tissues at varying levels. The transcript is particularly abundant in several human fetal tissues, including heart, placenta, and intestine. Studies with antisera raised against a DMDL fusion protein identify a 400,000 M{sub r} protein in all mouse tissues tested, including those of mdx and dy mice. Unlike the dystrophin gene, the DMDL gene transcript is not differentially spliced at the 3{prime} end in either fetal muscle or brain.

  19. Identification of muscle necrosis in the mdx mouse model of Duchenne muscular dystrophy using three-dimensional optical coherence tomography

    NASA Astrophysics Data System (ADS)

    Klyen, Blake R.; Shavlakadze, Thea; Radley-Crabb, Hannah G.; Grounds, Miranda D.; Sampson, David D.

    2011-07-01

    Three-dimensional optical coherence tomography (3D-OCT) was used to image the structure and pathology of skeletal muscle tissue from the treadmill-exercised mdx mouse model of human Duchenne muscular dystrophy. Optical coherence tomography (OCT) images of excised muscle samples were compared with co-registered hematoxylin and eosin-stained and Evans blue dye fluorescence histology. We show, for the first time, structural 3D-OCT images of skeletal muscle dystropathology well correlated with co-located histology. OCT could identify morphological features of interest and necrotic lesions within the muscle tissue samples based on intrinsic optical contrast. These findings demonstrate the utility of 3D-OCT for the evaluation of small-animal skeletal muscle morphology and pathology, particularly for studies of mouse models of muscular dystrophy.

  20. Rescue of a dystrophin-like protein by exon skipping normalizes synaptic plasticity in the hippocampus of the mdx mouse.

    PubMed

    Dallérac, Glenn; Perronnet, Caroline; Chagneau, Carine; Leblanc-Veyrac, Pascale; Samson-Desvignes, Nathalie; Peltekian, Elise; Danos, Olivier; Garcia, Luis; Laroche, Serge; Billard, Jean-Marie; Vaillend, Cyrille

    2011-09-01

    Duchenne muscular dystrophy (DMD) is caused by the absence of dystrophin, a protein that fulfills important functions in both muscle and brain. The mdx mouse model of DMD, which also lacks dystrophin, shows a marked reduction in γ-aminobutyric acid type A (GABA(A))-receptor clustering in central inhibitory synapses and enhanced long-term potentiation (LTP) at CA3-CA1 synapses of the hippocampus. We have recently shown that U7 small nuclear RNAs modified to encode antisense sequences and expressed from recombinant adeno-associated viral (rAAV) vectors are able to induce skipping of the mutated exon 23 and to rescue expression of a functional dystrophin-like product both in the muscle and nervous tissue in vivo. In the brain, this rescue was accompanied by restoration of both the size and number of hippocampal GABA(A)-receptor clustering. Here, we report that 25.2±8% of re-expression two months after intrahippocampal injection of rAAV reverses the abnormally enhanced LTP phenotype at CA3-CA1 synapses of mdx mice. These results suggests that dystrophin expression indirectly influences synaptic plasticity through modulation of GABA(A)-receptor clustering and that re-expression of the otherwise deficient protein in the adult can significantly alleviate alteration of neural functions in DMD. PMID:21624465

  1. Global Transcriptional Response to Hfe Deficiency and Dietary Iron Overload in Mouse Liver and Duodenum

    PubMed Central

    Rodriguez, Alejandra; Luukkaala, Tiina; Fleming, Robert E.; Britton, Robert S.; Bacon, Bruce R.; Parkkila, Seppo

    2009-01-01

    Iron is an essential trace element whose absorption is usually tightly regulated in the duodenum. HFE-related hereditary hemochromatosis (HH) is characterized by abnormally low expression of the iron-regulatory hormone, hepcidin, which results in increased iron absorption. The liver is crucial for iron homeostasis as it is the main production site of hepcidin. The aim of this study was to explore and compare the genome-wide transcriptome response to Hfe deficiency and dietary iron overload in murine liver and duodenum. Illumina™ arrays containing over 47,000 probes were used to study global transcriptional changes. Quantitative RT-PCR (Q-RT-PCR) was used to validate the microarray results. In the liver, the expression of 151 genes was altered in Hfe−/− mice while dietary iron overload changed the expression of 218 genes. There were 173 and 108 differentially expressed genes in the duodenum of Hfe−/− mice and mice with dietary iron overload, respectively. There was 93.5% concordance between the results obtained by microarray analysis and Q-RT-PCR. Overexpression of genes for acute phase reactants in the liver and a strong induction of digestive enzyme genes in the duodenum were characteristic of the Hfe-deficient genotype. In contrast, dietary iron overload caused a more pronounced change of gene expression responsive to oxidative stress. In conclusion, Hfe deficiency caused a previously unrecognized increase in gene expression of hepatic acute phase proteins and duodenal digestive enzymes. PMID:19787063

  2. The use of urinary and kidney SILAM proteomics to monitor kidney response to high dose morpholino oligonucleotides in the mdx mouse

    PubMed Central

    Zhang, Aiping; Uaesoontrachoon, Kitipong; Shaughnessy, Conner; Das, Jharna R.; Rayavarapu, Sree; Brown, Kristy J; Ray, Patricio E.; Nagaraju, Kanneboyina; van den Anker, John N.; Hoffman, Eric P; Hathout, Yetrib

    2015-01-01

    Phosphorodiamidate morpholino oligonucleotides (PMO) are used as a promising exon-skipping gene therapy for Duchenne Muscular Dystrophy (DMD). One potential complication of high dose PMO therapy is its transient accumulation in the kidneys. Therefore new urinary biomarkers are needed to monitor this treatment. Here, we carried out a pilot proteomic profiling study using stable isotope labeling in mammals (SILAM) strategy to identify new biomarkers to monitor the effect of PMO on the kidneys of the dystrophin deficient mouse model for DMD (mdx-23). We first assessed the baseline renal status of the mdx-23 mouse compared to the wild type (C57BL10) mouse, and then followed the renal outcome of mdx-23 mouse treated with a single high dose intravenous PMO injection (800 mg/kg). Surprisingly, untreated mdx-23 mice showed evidence of renal injury at baseline, which was manifested by albuminuria, increased urine output, and changes in established urinary biomarker of acute kidney injury (AKI). The PMO treatment induced further transient renal injury, which peaked at 7 days, and returned to almost the baseline status at 30 days post-treatment. In the kidney, the SILAM approach followed by western blot validation identified changes in Meprin A subunit alpha at day 2, then returned to normal levels at day 7 and 30 after PMO injection. In the urine, SILAM approach identified an increase in Clusterin and γ-glutamyl transpeptidase 1 as potential candidates to monitor the transient renal accumulation of PMO. These results, which were confirmed by Western blots or ELISA, demonstrate the value of the SILAM approach to identify new candidate biomarkers of renal injury in mdx-23 mice treated with high dose PMO. Chemical compounds studied in this article: Phosphorodiamidate morpholino (PubChem CID: 22140692); isoflurane (PubChem CID: 3763); formic acid (PubChem CID: 284); acetonitrile (PubChem CID: 6342); acetone (PubChem CID: 180); methanol (PubChem CID: 887) PMID:26213685

  3. Multivariate Data EXplorer (MDX)

    Energy Science and Technology Software Center (ESTSC)

    2012-08-01

    The MDX toolkit facilitates exploratory data analysis and visualization of multivariate datasets. MDX provides and interactive graphical user interface to load, explore, and modify multivariate datasets stored in tabular forms. MDX uses an extended version of the parallel coordinates plot and scatterplots to represent the data. The user can perform rapid visual queries using mouse gestures in the visualization panels to select rows or columns of interest. The visualization panel provides coordinated multiple views wherebymore » selections made in one plot are propagated to the other plots. Users can also export selected data or reconfigure the visualization panel to explore relationships between columns and rows in the data.« less

  4. Granulocyte-Colony Stimulating Factor Improves MDX Mouse Response to Peripheral Nerve Injury

    PubMed Central

    Simões, Gustavo Ferreira; de Oliveira, Alexandre Leite Rodrigues

    2012-01-01

    Background G-CSF has been shown to increase neuronal survival, which may positively influence the spinal cord microenvironment during the course of muscular dystrophies. Methodology/Principal Findings Male MDX mice that were six weeks of age received a left sciatic nerve transection and were treated with intraperitoneal injections of 200 µg/kg/day of G-CSF 7 days before and 7 days after the transection. The axotomy was performed after the cycles of muscular degeneration/regeneration, consistent with previous descriptions of this model of muscular dystrophy. C57BL/10 mice were used as control subjects. Seven days after the surgery, the animals were sacrificed and their lumbar spinal cords were processed for immunohistochemistry (anti-MHC I, anti-Synaptophysin, anti-GFAP and anti-IBA-1) and transmission electron microscopy. MHC I expression increased in both strains of mice after the axotomy. Nevertheless, the MDX mice displayed a significantly smaller MHC I upregulation than the control mice. Regarding GFAP expression, the MDX mice showed a stronger astrogliosis compared with the C57BL/10 mice across all groups. Both groups that were treated with G-CSF demonstrated preservation of synaptophysin expression compared with the untreated and placebo groups. The quantitative analysis of the ultrastructural level showed a preservation of the synaptic covering for the both groups that were treated with G-CSF and the axotomized groups showed a smaller loss of synaptic contact in relation to the treated groups after the lesion. Conclusions/Significance The reduction of active inputs to the alpha-motoneurons and increased astrogliosis in the axotomized and control groups may be associated with the cycles of muscle degeneration/regeneration that occur postnatally. The G-CSF treated group showed a preservation of the spinal cord microenvironment after the lesion. Moreover, the increase of MHC I expression in the MDX mice that were treated with G-CSF may indicate that this drug

  5. Alteration of excitation-contraction coupling mechanism in extensor digitorum longus muscle fibres of dystrophic mdx mouse and potential efficacy of taurine

    PubMed Central

    De Luca, Annamaria; Pierno, Sabata; Liantonio, Antonella; Cetrone, Michela; Camerino, Claudia; Simonetti, Simonetta; Papadia, Francesco; Camerino, Diana Conte

    2001-01-01

    No clear data is available about functional alterations in the calcium-dependent excitation-contraction (e-c) coupling mechanism of dystrophin-deficient muscle of mdx mice. By means of the intracellular microelectrode ‘point' voltage clamp method, we measured the voltage threshold for contraction (mechanical threshold; MT) in intact extensor digitorum longus (EDL) muscle fibres of dystrophic mdx mouse of two different ages: 8–12 weeks, during the active regeneration of hind limb muscles, and 6–8 months, when regeneration is complete. The EDL muscle fibres of 8–12-week-old wildtype animals had a more negative rheobase voltage (potential of equilibrium for contraction- and relaxation-related calcium movements) with respect to control mice of 6–8 months. However, at both ages, the EDL muscle fibres of mdx mice contracted at more negative potentials with respect to age-matched controls and had markedly slower time constants to reach the rheobase. The in vitro application of 60 mM taurine, whose normally high intracellular muscle levels play a role in e-c coupling, was without effect on 6–8-month-old wildtype EDL muscle, while it significantly ameliorated the MT of mdx mouse. HPLC determination of taurine content at 6–8 months showed a significant 140% rise of plasma taurine levels and a clear trend toward a decrease in amino acid levels in hind limb muscles, brain and heart, suggesting a tissue difficulty in retaining appropriate levels of the amino acid. The data is consistent with a permanent alteration of e-c coupling in mdx EDL muscle fibres. The alteration could be related to the proposed increase in intracellular calcium, and can be ameliorated by taurine, suggesting a potential therapeutic role of the amino acid. PMID:11226135

  6. Nitric oxide donors improve prednisone effects on muscular dystrophy in the mdx mouse diaphragm.

    PubMed

    Mizunoya, Wataru; Upadhaya, Ritika; Burczynski, Frank J; Wang, Guqi; Anderson, Judy E

    2011-05-01

    In Duchenne muscular dystrophy (DMD), palliative glucocorticoid therapy can produce myopathy or calcification. Since increased nitric oxide synthase activity in dystrophic mice promotes regeneration, the outcome of two nitric oxide (NO) donor drugs, MyoNovin (M) and isosorbide dinitrate (I), on the effectiveness of the anti-inflammatory drug prednisone (P) in alleviating progression of dystrophy was tested. Dystrophic mdx mice were treated (18 days) as controls or with an NO donor ± P. Fiber permeability and DNA synthesis were labeled by Evans blue dye (EBD) and bromodeoxyuridine uptake, respectively. P decreased body weight gain, M increased quadriceps mass, and I increased heart mass. P increased fiber permeability (%EBD+ fibers) and calcification in diaphragm. Treatment with NO donors + P (M+P, I+P) reduced %EBD+ fibers and calcification vs. P alone. %EBD+ fibers in M+P diaphragm did not differ from control. NO donor treatment reduced proliferation and the population of c-met+ cells and accelerated fiber regeneration. Concurrent with P, NO donor treatment suppressed two important detrimental effects of P in mice, possibly by accelerating regeneration, rebalancing satellite cell quiescence and activation in dystrophy, and/or increasing perfusion. Results suggest that NO donors could improve current therapy for DMD. PMID:21270295

  7. The effect of respiratory muscle training with CO2 breathing on cellular adaptation of mdx mouse diaphragm

    PubMed Central

    Matécki, Stefan; Rivier, François; Hugon, Gerald; Koechlin, Christelle; Michel, Alain; Préfaut, Christian; Mornet, Dominique; Ramonatxo, Michèle

    2005-01-01

    The aim of our study was to investigate the cellular mechanisms induced by hypercapnic stimulation of ventilation, during 6 weeks/30 min per day, in 10 mdx and 8 C57BL10 mice (10G0.2 months old). Ten mdx and eight C57BL10 mice served as control group. This respiratory training increases in vitro maximal tetanic tension of the diaphragm only in mdx mice. Western blot analysis of diaphragm showed: (1) an over-expression of a-dystrobrevin in mdx and C57BL10 training group compared to control group (8100G710 versus 6100G520 and 2800G400 versus 2200G250 arbitrary units); (2) a decrease in utrophin expression only in mdx training group compared to control group (2100G320 versus 3100G125 arbitrary units). Daily respiratory muscle training in mdx mice, induces a beneficial effect on diaphragm strength, with an over-expression of a-dystrobrevin. Further studies are needed to determine if, in absence of dystrophin, the over-expression of a-dystrobrevin could be interpreted as a possible pathway to improve function of dystrophic muscle. PMID:15907290

  8. The mdx mouse model as a surrogate for Duchenne muscular dystrophy

    PubMed Central

    Partridge, Terence A.

    2014-01-01

    Research into fundamental principles and the testing of therapeutic hypotheses for treatment of human disease is commonly conducted on mouse models of human diseases. Although this is often the only practicable approach, it carries a number of caveats arising from differences between the two species. This article is centred on the example of skeletal muscle disease, in particular muscular dystrophy, to identify some of the principal classes of obstacle to the translation of data from mouse to man. Of these, the difference in scale is one of the most commonly ignored and is of particular interest because it has quite major repercussions for evaluation of some classes of intervention and of assessment criteria while having comparatively little bearing on others. Likewise, interspecies differences and similarities in cell and molecular biological mechanisms underlying development, growth and response to pathological processes should be considered on an individual basis. An awareness of such distinctions is crucial if we are to avoid misjudgement of the likely efficacy in man of results obtained on mouse models. PMID:23551987

  9. Effect of nuclear factor κB inhibition on serotype 9 adeno-associated viral (AAV9) minidystrophin gene transfer to the mdx mouse.

    PubMed

    Reay, Daniel P; Niizawa, Gabriela A; Watchko, Jon F; Daood, Molly; Reay, Ja'Nean C; Raggi, Eugene; Clemens, Paula R

    2012-01-01

    Gene therapy studies for Duchenne muscular dystrophy (DMD) have focused on viral vector-mediated gene transfer to provide therapeutic protein expression or treatment with drugs to limit dystrophic changes in muscle. The pathological activation of the nuclear factor (NF)-κB signaling pathway has emerged as an important cause of dystrophic muscle changes in muscular dystrophy. Furthermore, activation of NF-κB may inhibit gene transfer by promoting inflammation in response to the transgene or vector. Therefore, we hypothesized that inhibition of pathological NF-κB activation in muscle would complement the therapeutic benefits of dystrophin gene transfer in the mdx mouse model of DMD. Systemic gene transfer using serotype 9 adeno-associated viral (AAV9) vectors is promising for treatment of preclinical models of DMD because of vector tropism to cardiac and skeletal muscle. In quadriceps of C57BL/10ScSn-Dmd(mdx)/J (mdx) mice, the addition of octalysine (8K)-NF-κB essential modulator (NEMO)-binding domain (8K-NBD) peptide treatment to AAV9 minidystrophin gene delivery resulted in increased levels of recombinant dystrophin expression suggesting that 8K-NBD treatment promoted an environment in muscle tissue conducive to higher levels of expression. Indices of necrosis and regeneration were diminished with AAV9 gene delivery alone and to a greater degree with the addition of 8K-NBD treatment. In diaphragm muscle, high-level transgene expression was achieved with AAV9 minidystoophin gene delivery alone; therefore, improvements in histological and physiological indices were comparable in the two treatment groups. The data support benefit from 8K-NBD treatment to complement gene transfer therapy for DMD in muscle tissue that receives incomplete levels of transduction by gene transfer, which may be highly significant for clinical applications of muscle gene delivery. PMID:22231732

  10. Prophylactic pamidronate partially protects from glucocorticoid-induced bone loss in the mdx mouse model of Duchenne muscular dystrophy.

    PubMed

    Yoon, Sung-Hee; Chen, Jinghan; Grynpas, Marc D; Mitchell, Jane

    2016-09-01

    Glucocorticoids are extensively used to treat patients with Duchenne muscular dystrophy because of their ability to delay muscle damage, prolong ambulation and extend life. However, use of glucocorticoids significantly increases bone loss, fragility and fractures. To determine if antiresorptive bisphosphonates could prevent the effects of glucocorticoids on bone quality, we used dystrophic mdx mice treated with the glucocorticoid prednisone during 8weeks of rapid bone growth from 5 to 13weeks of age and treated some mice with the bisphosphonate pamidronate during the first two weeks of prednisone administration. Prednisone reduced long bone growth, decreased cortical bone thickness and area and decreased the strength of the femurs. Pamidronate treatment protected mice from cortical bone loss but did not increase bone strength. The combination of prednisone and pamidronate inhibited remodeling of metaphyseal trabecular bone with large numbers of trabeculae containing remnants of calcified cartilage. Prednisone improved muscle strength in the mdx mice and decreased serum creatine kinase with evidence of improved muscle histology and these effects were maintained in mice treated with pamidronate. PMID:27373502

  11. Read-through compound 13 restores dystrophin expression and improves muscle function in the mdx mouse model for Duchenne muscular dystrophy

    PubMed Central

    Kayali, Refik; Ku, Jin-Mo; Khitrov, Gregory; Jung, Michael E.; Prikhodko, Olga; Bertoni, Carmen

    2012-01-01

    Molecules that induce ribosomal read-through of nonsense mutations in mRNA and allow production of a full-length functional protein hold great therapeutic potential for the treatment of many genetic disorders. Two such read-through compounds, RTC13 and RTC14, were recently identified by a luciferase-independent high-throughput screening assay and were shown to have potential therapeutic functions in the treatment of nonsense mutations in the ATM and the dystrophin genes. We have now tested the ability of RTC13 and RTC14 to restore dystrophin expression into skeletal muscles of the mdx mouse model for Duchenne muscular dystrophy (DMD). Direct intramuscular injection of compound RTC14 did not result in significant read-through activity in vivo and demonstrated the levels of dystrophin protein similar to those detected using gentamicin. In contrast, significant higher amounts of dystrophin were detected after intramuscular injection of RTC13. When administered systemically, RTC13 was shown to partially restore dystrophin protein in different muscle groups, including diaphragm and heart, and improved muscle function. An increase in muscle strength was detected in all treated animals and was accompanied by a significant decrease in creatine kinase levels. These studies establish the therapeutic potential of RTC13 in vivo and advance this newly identified compound into preclinical application for DMD. PMID:22692682

  12. Systemic Trans-splicing adeno-associated viral delivery efficiently transduces the heart of adult mdx mouse, a model for duchenne muscular dystrophy.

    PubMed

    Ghosh, Arkasubhra; Yue, Yongping; Shin, Jin-Hong; Duan, Dongsheng

    2009-11-01

    Trans-splicing adeno-associated viral (tsAAV) vectors hold great promise for delivering large therapeutic genes. One potential application is in the treatment of Duchenne muscular dystrophy (DMD). In this case, it is necessary to transduce whole body muscle. We demonstrated body-wide AAV-9 tsAAV transduction in normal neonatal mice. However, it was not clear whether such an approach would work in diseased mice. In this study we delivered the AAV-9 alkaline phosphatase (AP) tsAAV vector (3 x 10(12) vector genome particles per vector per mouse, tail vein injection) to 2-month-old mdx mice, the most widely used DMD model. Four months later, we observed widespread AP expression in the heart. It reached the same level as we have seen in normal neonatal puppy. Interestingly, myocardial transduction correlated with beta-myosin heavy chain expression but not with LamR, the putative AAV-9 receptor. AP expression was also detected in various skeletal muscles but at levels much lower than in normal newborn mice. Despite the existing inflammatory milieu, we did not see any appreciable increase in CD4(+) and CD8(+) T cells and macrophages in striated muscles after systemic tsAAV infection. In summary, our results have paved the way for tsAAV-mediated gene therapy for Duchenne cardiomyopathy. PMID:19627234

  13. Read-through compound 13 restores dystrophin expression and improves muscle function in the mdx mouse model for Duchenne muscular dystrophy.

    PubMed

    Kayali, Refik; Ku, Jin-Mo; Khitrov, Gregory; Jung, Michael E; Prikhodko, Olga; Bertoni, Carmen

    2012-09-15

    Molecules that induce ribosomal read-through of nonsense mutations in mRNA and allow production of a full-length functional protein hold great therapeutic potential for the treatment of many genetic disorders. Two such read-through compounds, RTC13 and RTC14, were recently identified by a luciferase-independent high-throughput screening assay and were shown to have potential therapeutic functions in the treatment of nonsense mutations in the ATM and the dystrophin genes. We have now tested the ability of RTC13 and RTC14 to restore dystrophin expression into skeletal muscles of the mdx mouse model for Duchenne muscular dystrophy (DMD). Direct intramuscular injection of compound RTC14 did not result in significant read-through activity in vivo and demonstrated the levels of dystrophin protein similar to those detected using gentamicin. In contrast, significant higher amounts of dystrophin were detected after intramuscular injection of RTC13. When administered systemically, RTC13 was shown to partially restore dystrophin protein in different muscle groups, including diaphragm and heart, and improved muscle function. An increase in muscle strength was detected in all treated animals and was accompanied by a significant decrease in creatine kinase levels. These studies establish the therapeutic potential of RTC13 in vivo and advance this newly identified compound into preclinical application for DMD. PMID:22692682

  14. Coronary adventitial cells are linked to perivascular cardiac fibrosis via TGFβ1 signaling in the mdx mouse model of Duchenne Muscular Dystrophy

    PubMed Central

    Ieronimakis, Nicholas; Hays, Aislinn L.; Janebodin, Kajohnkiart; Mahoney, William M.; Duffield, Jeremy S.; Majesky, Mark W.; Reyes, Morayma

    2013-01-01

    In Duchenne Muscular Dystrophy (DMD), progressive accumulation of cardiac fibrosis promotes heart failure. While the cellular origins of fibrosis in DMD hearts remain enigmatic, fibrotic tissue conspicuously forms near the coronary adventitia. Therefore, we sought to characterize the role of coronary adventitial cells in the formation of perivascular fibrosis. Utilizing the mdx model of DMD, we have identified a population of Sca1+, PDGFRα+, CD31−, CD45− coronary adventitial cells responsible for perivascular fibrosis. Histopathology of dystrophic hearts revealed Sca1+ cells extend from the adventitia and occupy regions of perivascular fibrosis. The number of Sca1+ adventitial cells increased two-fold in fibrotic mdx hearts vs. age matched wild-type hearts. Moreover, relative to Sca1−, PDGFRα+, CD31−, CD45− cells and endothelial cells, Sca1+ adventitial cells FACS-sorted from mdx hearts expressed the highest level of Collagen1α1 and 3α1, Connective tissue growth factor, and Tgfβr1 transcripts. Surprisingly, mdx endothelial cells expressed the greatest level of the Tgfβ1 ligand. Utilizing Collagen1α1-GFP reporter mice, we confirmed that the majority of Sca1+ adventitial cells expressed type I collagen, an abundant component of cardiac fibrosis, in both wt (71% ±4.1) and mdx (77% ±3.5) hearts. In contrast, GFP+ interstitial fibroblasts were PDGFRα+ but negative for Sca1. Treatment of cultured Collagen1α1-GFP+ adventitial cells with TGFβ1 resulted in increased collagen synthesis, whereas pharmacological inhibition of TGFβR1 signaling reduced the fibrotic response. Therefore, perivascular cardiac fibrosis by coronary adventitial cells may be mediated by TGFβ1 signaling. Our results implicate coronary endothelial cells in mediating cardiac fibrosis via transmural TGFβ signaling, and suggest that the coronary adventitia is a promising target for developing novel anti-fibrotic therapies. PMID:23911435

  15. Dystrophin-deficient mdx mice display a reduced life span and are susceptible to spontaneous rhabdomyosarcoma.

    PubMed

    Chamberlain, Jeffrey S; Metzger, Joseph; Reyes, Morayma; Townsend, DeWayne; Faulkner, John A

    2007-07-01

    Duchenne muscular dystrophy (DMD) is the most common, lethal genetic disorder of children. A number of animal models of muscular dystrophy exist, but the most effective model for characterizing the structural and functional properties of dystrophin and therapeutic interventions has been the mdx mouse. Despite the approximately 20 years of investigations of the mdx mouse, the impact of the disease on the life span of mdx mice and the cause of death remain unresolved. Consequently, a life span study of the mdx mouse was designed that included cohorts of male and female mdx and wild-type C57BL/10 mice housed under specific pathogen-free conditions with deaths restricted to natural causes and with examination of the carcasses for pathology. Compared with wild-type mice, both mdx male and female mice had reduced life spans and displayed a progressively dystrophic muscle histopathology. Surprisingly, old mdx mice were prone to develop muscle tumors that resembled the human form of alveolar rhabdomyosarcoma, a cancer associated with poor prognosis. Rhabdomyosarcomas have not been observed previously in nontransgenic mice. The results substantiate the mdx mouse as an important model system for studies of the pathogenesis of and potential remedies for DMD. PMID:17360850

  16. Targeted gene correction in the mdx mouse using short DNA fragments: towards application with bone marrow-derived cells for autologous remodeling of dystrophic muscle.

    PubMed

    Kapsa, R M; Quigley, A F; Vadolas, J; Steeper, K; Ioannou, P A; Byrne, E; Kornberg, A J

    2002-06-01

    In muscle, mutant genes can be targeted and corrected directly by intramuscular (i.m.) injection of corrective DNA, or by ex vivo delivery of DNA to myogenic cells, followed by cell transplantation. Short fragment homologous replacement (SFHR) has been used to repair the exon 23 nonsense transition at the Xp21.1 dys locus in cultured cells and also, directly in tibialis anterior from male mdx mice. Whilst mdx dys locus correction can be achieved in up to 20% of cells in culture, much lower efficiency is evident by i.m. injection. The major consideration for application of targeted gene correction to muscle is delivery throughout relevant tissues. Systemically injected bone marrow (BM)-derived cells from wt C57BL/10 ScSn mice are known to remodel mdx muscle when injected into the systemic route. Provided that non muscle-derived cell types most capable of muscle remodeling activity can be more specifically identified, isolated and expanded, cell therapy seems presently the most favorable vehicle by which to deliver gene correction throughout muscle tissues. Using wt bone marrow as a model, this study investigates systemic application of bone marrow-derived cells as potential vehicles to deliver corrected (ie wt) dys locus to dystrophic muscle. Intravenous (i.v.) and intraperitoneal (i.p.) injections of wt BM were given to lethally and sub-lethally irradiated mdx mice. Despite both i.v. and surviving i.p. groups containing wt dys loci in 100% and less than 1% of peripheral blood nuclei, respectively, both groups displayed equivalent levels of wt dys transcript in muscle RNA. These results suggest that the muscle remodeling activity observed in systemically injected BM cells is not likely to be found in the hemopoietic fraction. PMID:12032690

  17. Absence of Dp71 in mdx3cv mouse spermatozoa alters flagellar morphology and the distribution of ion channels and nNOS

    PubMed Central

    Hernández-González, Enrique O; Mornet, Dominique; Rendon, Alvaro; Martínez-Rojas, Dalila

    2005-01-01

    Summary In muscle, the absence of dystrophin alters the dystrophin associated protein complex (DAPC), which is involved in the clustering and anchoring of signaling proteins and ion and water channels. Here we show that mice spermatozoa express only dystrophin Dp71 and utrophin Up71. The purpose of this study was to explore the effect of the absence of Dp71 on the morphology and membrane distribution of members of the DAPC, ion channels and signaling proteins of spermatozoa obtained from dystrophic mutant mdx3cv mice. Our work indicates that although the absence of Dp71 results in a dramatic decrease in β-dystroglycan, it induces membrane redistribution and an increase in the total level of a-syntrophin, voltage dependent Na+ (μ1) and K+ (Kv1.1) channels and neural nitric oxide synthase (nNOS). The short utrophin (Up71) was upregulated and redistributed in the spermatozoa of mdx3cv mice. A significant increase in abnormal flagella morphology was observed in the absence of Dp71, which was partially corrected when the plasma membrane was eliminated by detergent treatment. Our observations point to a new phenotype associated with the absence of Dp71. Abnormal flagellar structure and altered distribution of ion channels and signaling proteins may be responsible for the fertility problems of mdx3cv mice. PMID:15601658

  18. Evidence of hypoxic tolerance in weak upper airway muscle from young mdx mice.

    PubMed

    Burns, David P; O'Halloran, Ken D

    2016-06-01

    Duchenne muscular dystrophy (DMD) is a genetic disease characterised by deficiency in the protein dystrophin. The respiratory system is weakened and patients suffer from sleep disordered breathing and hypoventilation culminating in periods of hypoxaemia. We examined the effects of an acute (6h) hypoxic stress on sternohyoid muscle function (representative pharyngeal dilator). 8 week old male, wild-type (WT; C57BL/10ScSnJ; n=18) and mdx (C57BL/10ScSn-Dmd(mdx)/J; n=16) mice were exposed to sustained hypoxia (FIO2=0.10) or normoxia. Muscle functional properties were examined ex vivo. Additional WT (n=5) and mdx (n=5) sternohyoid muscle was exposed to an anoxic challenge. Sternohyoid dysfunction was observed in mdx mice with significant reductions in force and power. Following exposure to the acute in vivo hypoxic stress, WT sternohyoid muscle showed evidence of functional impairment (reduced force, work and power). Conversely, mdx sternohyoid showed an apparent tolerance to the acute hypoxic stress. This tolerance was not maintained for mdx following a severe hypoxic stress. A dysfunctional upper airway muscle phenotype is present at 8 weeks of age in the mdx mouse, which may have implications for the control of airway patency in DMD. Hypoxic tolerance in mdx respiratory muscle is suggestive of adaptation to chronic hypoxia, which could be present due to respiratory morbidity. We speculate a role for hypoxia in mdx respiratory muscle morbidity. PMID:26691169

  19. Chronic Dosing with Membrane Sealant Poloxamer 188 NF Improves Respiratory Dysfunction in Dystrophic Mdx and Mdx/Utrophin-/- Mice

    PubMed Central

    Markham, Bruce E.; Kernodle, Stace; Nemzek, Jean; Wilkinson, John E.; Sigler, Robert

    2015-01-01

    Poloxamer 188 NF (national formulary (NF) grade of P-188) improves cardiac muscle function in the mdx mouse and golden retriever muscular dystrophy models. However in vivo effects on skeletal muscle have not been reported. We postulated that P-188 NF might protect diaphragm muscle membranes from contraction-induced injury in mdx and mdx/utrophin-/- (dko) muscular dystrophy models. In the first study 7-month old mdx mice were treated for 22 weeks with subcutaneous (s.c.) injections of saline or P-188 NF at 3 mg/Kg. In the second, dkos were treated with saline or P-188 NF (1 mg/Kg) for 8 weeks beginning at age 3 weeks. Prednisone was the positive control in both studies. Respiratory function was monitored using unrestrained whole body plethysmography. P-188 NF treatment affected several respiratory parameters including tidal volume/BW and minute volume/BW in mdx mice. In the more severe dko model, P-188 NF (1 mg/Kg) significantly slowed the decline in multiple respiratory parameters compared with saline-treated dko mice. Prednisone’s effects were similar to those seen with P-188 NF. Diaphragms from P-188 NF or prednisone treated mdx and dko mice showed signs of muscle fiber protection including less centralized nuclei, less variation in fiber size, greater fiber density, and exhibited a decreased amount of collagen deposition. P-188 NF at 3 mg/Kg s.c. also improved parameters of systolic and diastolic function in mdx mouse hearts. These results suggest that P-188 NF may be useful in treating respiratory and cardiac dysfunction, the leading causes of death in Duchenne muscular dystrophy patients. PMID:26248188

  20. Functional and Molecular Effects of Arginine Butyrate and Prednisone on Muscle and Heart in the mdx Mouse Model of Duchenne Muscular Dystrophy

    PubMed Central

    Guerron, Alfredo D.; Rawat, Rashmi; Sali, Arpana; Spurney, Christopher F.; Pistilli, Emidio; Cha, Hee-Jae; Pandey, Gouri S.; Gernapudi, Ramkishore; Francia, Dwight; Farajian, Viken; Escolar, Diana M.; Bossi, Laura; Becker, Magali; Zerr, Patricia; de la Porte, Sabine; Gordish-Dressman, Heather; Partridge, Terence; Hoffman, Eric P.; Nagaraju, Kanneboyina

    2010-01-01

    Background The number of promising therapeutic interventions for Duchenne Muscular Dystrophy (DMD) is increasing rapidly. One of the proposed strategies is to use drugs that are known to act by multiple different mechanisms including inducing of homologous fetal form of adult genes, for example utrophin in place of dystrophin. Methodology/Principal Findings In this study, we have treated mdx mice with arginine butyrate, prednisone, or a combination of arginine butyrate and prednisone for 6 months, beginning at 3 months of age, and have comprehensively evaluated the functional, biochemical, histological, and molecular effects of the treatments in this DMD model. Arginine butyrate treatment improved grip strength and decreased fibrosis in the gastrocnemius muscle, but did not produce significant improvement in muscle and cardiac histology, heart function, behavioral measurements, or serum creatine kinase levels. In contrast, 6 months of chronic continuous prednisone treatment resulted in deterioration in functional, histological, and biochemical measures. Arginine butyrate-treated mice gene expression profiling experiments revealed that several genes that control cell proliferation, growth and differentiation are differentially expressed consistent with its histone deacetylase inhibitory activity when compared to control (saline-treated) mdx mice. Prednisone and combination treated groups showed alterations in the expression of genes that control fibrosis, inflammation, myogenesis and atrophy. Conclusions/Significance These data indicate that 6 months treatment with arginine butyrate can produce modest beneficial effects on dystrophic pathology in mdx mice by reducing fibrosis and promoting muscle function while chronic continuous treatment with prednisone showed deleterious effects to skeletal and cardiac muscle. Our results clearly indicate the usefulness of multiple assays systems to monitor both beneficial and toxic effects of drugs with broad range of in vivo

  1. Exclusive skeletal muscle correction does not modulate dystrophic heart disease in the aged mdx model of Duchenne cardiomyopathy

    PubMed Central

    Wasala, Nalinda B.; Bostick, Brian; Yue, Yongping; Duan, Dongsheng

    2013-01-01

    Duchenne muscular dystrophy (DMD) is characterized by severe degeneration and necrosis of both skeletal and cardiac muscle. While many experimental therapies have shown great promise in treating skeletal muscle disease, an effective therapy for Duchenne cardiomyopathy remains a challenge in large animal models and human patients. The current views on cardiac consequences of skeletal muscle-centered therapy are controversial. Studies performed in young adult mdx mice (a mild DMD mouse model) have yielded opposing results. Since mdx mice do not develop dystrophic cardiomyopathy until ≥21 months of age, we reasoned that old mdx mice may represent a better model to assess the impact of skeletal muscle rescue on dystrophic heart disease. Here, we aged skeletal muscle-specific micro-dystrophin transgenic mdx mice to 23 months and examined the cardiac phenotype. As expected, transgenic mdx mice had minimal skeletal muscle disease and they also outperformed original mdx mice on treadmill running. On cardiac examination, the dystrophin-null heart of transgenic mdx mice displayed severe cardiomyopathy matching that of non-transgenic mdx mice. Specifically, both the strains showed similar heart fibrosis and cardiac function deterioration in systole and diastole. Cardiac output and ejection fraction were also equally compromised. Our results suggest that skeletal muscle rescue neither aggravates nor alleviates cardiomyopathy in aged mdx mice. These findings underscore the importance of treating both skeletal and cardiac muscles in DMD therapy. PMID:23459935

  2. Delayed Cardiomyopathy in Dystrophin Deficient mdx Mice Relies on Intrinsic Glutathione Resource

    PubMed Central

    Khouzami, Lara; Bourin, Marie-Claude; Christov, Christo; Damy, Thibaud; Escoubet, Brigitte; Caramelle, Philippe; Perier, Magali; Wahbi, Karim; Meune, Christophe; Pavoine, Catherine; Pecker, Françoise

    2010-01-01

    Oxidative stress contributes to the pathogenesis of Duchenne muscular dystrophy (DMD). Although they have been a model for DMD, mdx mice exhibit slowly developing cardiomyopathy. We hypothesized that disease process was delayed owing to the development of an adaptive mechanism against oxidative stress, involving glutathione synthesis. At 15 to 20 weeks of age, mdx mice displayed a 33% increase in blood glutathione levels compared with age-matched C57BL/6 mice. In contrast, cardiac glutathione content was similar in mdx and C57BL/6 mice as a result of the balanced increased expression of glutamate cysteine ligase catalytic and regulatory subunits ensuring glutathione synthesis in the mdx mouse heart, as well as increased glutathione peroxidase-1 using glutathione. Oral administration from 10 weeks of age of the glutamate cysteine ligase inhibitor, l-buthionine(S,R)-sulfoximine (BSO, 5 mmol/L), led to a 33% and 50% drop in blood and cardiac glutathione, respectively, in 15- to 20-week-old mdx mice. Moreover, 20-week-old BSO-treated mdx mice displayed left ventricular hypertrophy associated with diastolic dysfunction, discontinuities in β-dystroglycan expression, micronecrosis and microangiopathic injuries. Examination of the glutathione status in four DMD patients showed that three displayed systemic glutathione deficiency as well. In conclusion, low glutathione resource hastens the onset of cardiomyopathy linked to a defect in dystrophin in mdx mice. This is relevant to the glutathione deficiency that DMD patients may suffer. PMID:20696779

  3. Acute relaxation of mouse duodenum [correction of duodenun] by estrogens. Evidence for an estrogen receptor-independent modulation of muscle excitability.

    PubMed

    Díaz, Mario; Ramírez, Cristina M; Marin, Raquel; Marrero-Alonso, Jorge; Gómez, Tomás; Alonso, Rafael

    2004-10-01

    17-beta-Estradiol, the stereoisomer 17-alpha-estradiol and the synthetic estrogen diethylstilbestrol (DES), all caused a rapid (<3 min) dose-dependent reversible relaxation of mouse duodenal spontaneous activity, reduced basal tone and depressed the responses to CaCl(2) and KCl. The steroidal antiestrogen 7alpha-[9-[(4,4,5,5,5,-pentafluoropenty)sulphinyl]nonyl]-estra-1,3,5(19)-triene-3,17beta-diol (ICI182,780) failed to either mimic or prevent the effect of 17-beta-estradiol. The effect of estrogens was unrelated to activation of nitric oxide (NO), mitogen-activated protein kinase (MAPK), protein kinase A (PKA), protein kinase G (PKG) or protein kinase C (PKC). Estrogen-induced relaxation was partially reversed by 1,4-dihydro-2,6-dimethyl-5-nitro-4-[2-(trifluoromethyl)phenyl]-pyridine-3-carboxilic acid methyl ester (BAY-K8644), depolarization, or by application of tetraethylammonium or 4-aminopyridine, but not by glibenclamide, apamin, charybdotoxin, paxilline or verruculogen. The effects of BAY-K8644 and K(+) channel blockers were synergistic, and allowed relaxed tissues to recover spontaneous activity and basal tone. We hypothesize that the rapid non-genomic spasmolytic effect of estrogens on mouse duodenal muscle might be triggered by an estrogen-receptor-independent mechanism likely involving activation of tetraethylamonium- and 4-aminopyridine-sensitive K(+) channels and inhibition of L-type Ca2(+) channels on the smooth muscle cells. PMID:15464075

  4. Transient receptor potential cation channels in normal and dystrophic mdx muscle.

    PubMed

    Krüger, Jana; Kunert-Keil, Christiane; Bisping, Frederike; Brinkmeier, Heinrich

    2008-06-01

    To investigate the defective calcium regulation of dystrophin-deficient muscle fibres we studied gene expression and localization of non-voltage gated cation channels in normal and mdx mouse skeletal muscle. We found TRPC3, TRPC6, TRPV4, TRPM4 and TRPM7 to be the most abundant isoforms. Immunofluorescent staining of muscle cross-sections with antibodies against TRP proteins showed sarcolemmal localization of TRPC6 and TRPM7, both, for mdx and control. TRPV4 was found only in a fraction of fibres at the sarcolemma and around myonuclei, while TRPC3 staining revealed intracellular patches, preferentially in mdx muscle. Transcripts of low abundance coding for TRPC5, TRPA1 and TRPM1 channels were increased in mdx skeletal muscle at certain stages. The increased Ca(2+)-influx into dystrophin-deficient mdx fibres cannot be explained by increased gene expression of major TRP channels. However, a constant TRP channel expression in combination with the well described weaker Ca(2+)-handling system of mdx fibres may indicate an imbalance between Ca(2+)-influx and cellular Ca(2+)-control. PMID:18504127

  5. A canine minidystrophin is functional and therapeutic in mdx mice.

    PubMed

    Wang, B; Li, J; Qiao, C; Chen, C; Hu, P; Zhu, X; Zhou, L; Bogan, J; Kornegay, J; Xiao, X

    2008-08-01

    Duchenne muscular dystrophy (DMD) is the most common and lethal genetic muscle disorder lacking a curative treatment. We wish to use the dystrophin-deficient golden retriever muscular dystrophy (GRMD) dog, a canine model of DMD, to investigate adeno-associated virus (AAV) vector-mediated minidystrophin gene therapy. The dog model is useful in evaluating vector dose requirement and immunological consequences owing to its large size and outbred nature. In this study, we have cloned and constructed a canine minidystrophin gene vector. Owing to limited availability of the GRMD dogs, here we first examined the functions and therapeutic effects of the canine minidystrophin in the mdx mouse model. We observed efficient minigene expression without cellular immune responses in mdx mice after AAV1-cMinidys vector intramuscular injection. We also observed restoration of the missing dystrophin-associated protein complex (DPC) onto the sarcolemma, including sarcoglycans and dystrobrevin, and a partial restoration of alpha-syntrophin and neural nitric oxide synthase (nNOS). In addition, minidystrophin treatment ameliorated dystrophic pathology, such as fibrosis and myofiber central nucleation (CN). CN remained minimal (<2%) after AAV injection in the neonatal mdx mice and was reduced from more than 75% to about 25% after AAV injection in adult mdx mice. Finally, in vivo cell membrane leakage test with Evans blue dye showed that the canine minidystrophin could effectively protect the myofiber plasma membrane integrity. Our results, thus, demonstrated the functionality and therapeutic potential of the canine minidystrophin and paved its way for further testing in the GRMD dog model. PMID:18432277

  6. Muscle genome-wide expression profiling during disease evolution in mdx mice.

    PubMed

    Marotta, Mario; Ruiz-Roig, Claudia; Sarria, Yaris; Peiro, Jose Luis; Nuñez, Fatima; Ceron, Julian; Munell, Francina; Roig-Quilis, Manuel

    2009-04-10

    Mdx mice show a milder phenotype than Duchenne patients despite bearing an analogous genetic defect. Our aim was to sort out genes, differentially expressed during the evolution of skeletal muscle mdx mouse disease, to elucidate the mechanisms by which these animals overcome the lack of dystrophin. Genome-wide microarray-based gene expression analysis was carried out at 3 wk and 1.5 and 3 mo of life. Candidate genes were selected by comparing: 1) mdx vs. controls at each point in time, and 2) mdx mice and 3) control mice among the three points in time. The first analysis showed a strong upregulation (96%) of inflammation-related genes and in >75% of genes related to cell adhesion, muscle structure/regeneration, and extracellular matrix remodeling during mdx disease evolution. Lgals3, Postn, Ctss, and Sln genes showed the strongest variations. The analysis performed among points in time demonstrated significant changes in Ecm1, Spon1, Thbs1, Csrp3, Myo10, Pde4b, and Adamts-5 exclusively during mdx mice lifespan. RT-PCR analysis of Postn, Sln, Ctss, Thbs1, Ecm1, and Adamts-5 expression from 3 wk to 9 mo, confirmed microarray data and demonstrated variations beyond 3 mo of age. A high-confidence functional network analysis demonstrated a strong relationship between them and showed two main subnetworks, having Dmd-Utrn-Myo10 and Adamts5-Thbs1-Spon1-Postn as principal nodes, which are functionally linked to Abca1, Actn4, Crebbp, Csrp3, Lama1, Lama3, Mical2, Mical3, Myf6, Pxn, and Sparc genes. Candidate genes may participate in the decline of muscle necrosis in mdx mice and could be considered potential therapeutic targets for Duchenne patients. PMID:19223608

  7. Activation of Wnt3a signaling promotes myogenic differentiation of mesenchymal stem cells in mdx mice

    PubMed Central

    Shang, Yan-chang; Wang, Shu-hui; Xiong, Fu; Peng, Fu-ning; Liu, Zhen-shan; Geng, Jia; Zhang, Cheng

    2016-01-01

    Aim: Duchenne muscular dystrophy (DMD) is an X-linked genetic muscular disorder with no effective treatment at present. Mesenchymal stem cell (MSC) transplantation has been used to treat DMD, but the efficiency is low. Our previous studies show that activation of Wnt3a signaling promotes myogenic differentiation of MSCs in vitro. Here we report an effective MSC transplantation therapy in mdx mice by activation of Wnt3a signaling. Methods: MSCs were isolated from mouse bone marrow, and pretreated with Wnt3a-conditioned medium (Wnt3a-CM), then transplanted into mdx mice. The recipient mice were euthanized at 4, 8, 12, 16 weeks after the transplantation, and muscle pathological changes were examined. The expression of dystrophin in muscle was detected using immunofluorescence staining, RT-PCR and Western blotting. Results: Sixteen weeks later, transplantation of Wnt3a-pretreated MSCs in mdx mice improved the characteristics of dystrophic muscles evidenced by significant reductions in centrally nucleated myofibers, the variability range of cross-sectional area (CSA) and the connective tissue area of myofibers. Furthermore, transplantation of Wnt3a-pretreated MSCs in mdx mice gradually and markedly increased the expression of dystrophin in muscle, and improved the efficiency of myogenic differentiation. Conclusion: Transplantation of Wnt3a-pretreated MSCs in mdx mice results in long-term amelioration of the dystrophic phenotype and restores dystrophin expression in muscle. The results suggest that Wnt3a may be a promising candidate for the treatment of DMD. PMID:27133298

  8. Extensive but Coordinated Reorganization of the Membrane Skeleton in Myofibers of Dystrophic (mdx) Mice

    PubMed Central

    Williams, McRae W.; Bloch, Robert J.

    1999-01-01

    We used immunofluorescence techniques and confocal imaging to study the organization of the membrane skeleton of skeletal muscle fibers of mdx mice, which lack dystrophin. β-Spectrin is normally found at the sarcolemma in costameres, a rectilinear array of longitudinal strands and elements overlying Z and M lines. However, in the skeletal muscle of mdx mice, β-spectrin tends to be absent from the sarcolemma over M lines and the longitudinal strands may be disrupted or missing. Other proteins of the membrane and associated cytoskeleton, including syntrophin, β-dystroglycan, vinculin, and Na,K-ATPase are also concentrated in costameres, in control myofibers, and mdx muscle. They also distribute into the same altered sarcolemmal arrays that contain β-spectrin. Utrophin, which is expressed in mdx muscle, also codistributes with β-spectrin at the mutant sarcolemma. By contrast, the distribution of structural and intracellular membrane proteins, including α-actinin, the Ca-ATPase and dihydropyridine receptors, is not affected, even at sites close to the sarcolemma. Our results suggest that in myofibers of the mdx mouse, the membrane- associated cytoskeleton, but not the nearby myoplasm, undergoes widespread coordinated changes in organization. These changes may contribute to the fragility of the sarcolemma of dystrophic muscle. PMID:10087268

  9. The mdx Mutation in the 129/Sv Background Results in a Milder Phenotype: Transcriptome Comparative Analysis Searching for the Protective Factors

    PubMed Central

    Calyjur, Priscila Clara; Almeida, Camila de Freitas; Ayub-Guerrieri, Danielle; Ribeiro, Antonio Fernando; Fernandes, Stephanie de Alcântara; Ishiba, Renata; dos Santos, Andre Luis Fernandes; Onofre-Oliveira, Paula; Vainzof, Mariz

    2016-01-01

    The mdx mouse is a good genetic and molecular murine model for Duchenne Muscular Dystrophy (DMD), a progressive and devastating muscle disease. However, this model is inappropriate for testing new therapies due to its mild phenotype. Here, we transferred the mdx mutation to the 129/Sv strain with the aim to create a more severe model for DMD. Unexpectedly, functional analysis of the first three generations of mdx129 showed a progressive amelioration of the phenotype, associated to less connective tissue replacement, and more regeneration than the original mdxC57BL. Transcriptome comparative analysis was performed to identify what is protecting this new model from the dystrophic characteristics. The mdxC57BL presents three times more differentially expressed genes (DEGs) than the mdx129 (371 and 137 DEGs respectively). However, both models present more overexpressed genes than underexpressed, indicating that the dystrophic and regenerative alterations are associated with the activation rather than repression of genes. As to functional categories, the DEGs of both mdx models showed a predominance of immune system genes. Excluding this category, the mdx129 model showed a decreased participation of the endo/exocytic pathway and homeostasis categories, and an increased participation of the extracellular matrix and enzymatic activity categories. Spp1 gene overexpression was the most significant DEG exclusively expressed in the mdx129 strain. This was confirmed through relative mRNA analysis and osteopontin protein quantification. The amount of the 66 kDa band of the protein, representing the post-translational product of the gene, was about 4,8 times higher on western blotting. Spp1 is a known DMD prognostic biomarker, and our data indicate that its upregulation can benefit phenotype. Modeling the expression of the DEGs involved in the mdx mutation with a benign course should be tested as a possible therapeutic target for the dystrophic process. PMID:26954670

  10. Increased levels of interleukin-6 exacerbate the dystrophic phenotype in mdx mice

    PubMed Central

    Pelosi, Laura; Berardinelli, Maria Grazia; Forcina, Laura; Spelta, Elisa; Rizzuto, Emanuele; Nicoletti, Carmine; Camilli, Carlotta; Testa, Erika; Catizone, Angela; De Benedetti, Fabrizio; Musarò, Antonio

    2015-01-01

    Duchenne muscular dystrophy (DMD) is characterized by progressive lethal muscle degeneration and chronic inflammatory response. The mdx mouse strain has served as the animal model for human DMD. However, while DMD patients undergo extensive necrosis, the affected muscles of adult mdx mice rapidly regenerates and regains structural and functional integrity. The basis for the mild effects observed in mice compared with the lethal consequences in humans remains unknown. In this study, we provide evidence that interleukin-6 (IL-6) is causally linked to the pathogenesis of muscular dystrophy. We report that forced expression of IL-6, in the adult mdx mice, recapitulates the severe phenotypic characteristics of DMD in humans. Increased levels of IL-6 exacerbate the dystrophic muscle phenotype, sustaining inflammatory response and repeated cycles of muscle degeneration and regeneration, leading to exhaustion of satellite cells. The mdx/IL6 mouse closely approximates the human disease and more faithfully recapitulates the disease progression in humans. This study promises to significantly advance our understanding of the pathogenic mechanisms that lead to DMD. PMID:26251044

  11. Fetal muscle-derived cells can repair dystrophic muscles in mdx mice

    SciTech Connect

    Auda-Boucher, Gwenola; Rouaud, Thierry; Lafoux, Aude; Levitsky, Dmitri; Huchet-Cadiou, Corinne; Feron, Marie; Guevel, Laetitia; Talon, Sophie; Fontaine-Perus, Josiane; Gardahaut, Marie-France . E-mail: Marie-France.Gardahaut@univ-nantes.fr

    2007-03-10

    We have previously reported that CD34{sup +} cells purified from mouse fetal muscles can differentiate into skeletal muscle in vitro and in vivo when injected into muscle tissue of dystrophic mdx mice. In this study, we investigate the ability of such donor cells to restore dystrophin expression, and to improve the functional muscle capacity of the extensor digitorum longus muscle (EDL) of mdx mice. For this purpose green fluorescent-positive fetal GFP{sup +}/CD34{sup +} cells or desmin{sup +}/{sup -}LacZ/CD34{sup +} cells were transplanted into irradiated or non-irradiated mdx EDL muscle. Donor fetal muscle-derived cells predominantly fused with existing fibers. Indeed more than 50% of the myofibers of the host EDL contained donor nuclei delivering dystrophin along 80-90% of the length of their sarcolemma. The presence of significant amounts of dystrophin (about 60-70% of that found in a control wild-type mouse muscle) was confirmed by Western blot analyses. Dystrophin expression also outcompeted that of utrophin, as revealed by a spatial shift in the distribution of utrophin. At 1 month post-transplant, the recipient muscle appeared to have greater resistance to fatigue than control mdx EDL muscle during repeated maximal contractions.

  12. Regional Enteritis of the Duodenum

    PubMed Central

    Edwards, A. M.; Michalyshyn, B.; Sherbaniuk, R. W.; Costopoulos, L. B.

    1965-01-01

    Forty-three cases of regional enteritis of the duodenum were found in the world literature. Regional duodenitis is relatively uncommon; in one large series of 600 cases of regional enteritis only three involved the duodenum. At the University of Alberta Hospital, in a three-year period (1962 to 1965), the authors encountered five patients with regional duodenitis, demonstrating a spectrum of clinical, radiologic and pathologic characteristics of this disease. The description of these patients brings the world's total to 48 reported cases. Two of these patients had symptoms of severe duodenal obstruction and were relieved by bypass procedures and vagotomy; one required surgery because of co-existent obstructive ileal disease: and two patients have improved on corticoids and salicylazosulfapyridine without surgery. In our experience treatment with corticoids and salicylazosulfapyridine is beneficial. Four of the five patients remain in a state of mild to moderate nutritional impairment and have evidence of intestinal malabsorption. In the fifth case the period of followup is too short to permit assessment. ImagesFig. 1Fig. 2Fig. 3Fig. 4Fig. 5Figs. 6 (X 50) and 7 (X 450)Fig. 8Fig. 9Fig. 10Fig. 11Fig. 12Figs. 13 and 14 (both X 100)Fig. 15Fig. 16Fig. 17Fig. 18Fig. 19Fig. 20Fig. 21 PMID:5843869

  13. Preservation of Muscle Force in Mdx3cv Mice Correlates with Low-Level Expression of a Near Full-Length Dystrophin Protein

    PubMed Central

    Li, Dejia; Yue, Yongping; Duan, Dongsheng

    2008-01-01

    The complete absence of dystrophin causes Duchenne muscular dystrophy. Its restoration by greater than 20% is needed to reduce muscle pathology and improve muscle force. Dystrophin levels lower than 20% are considered therapeutically irrelevant but are associated with a less severe phenotype in certain Becker muscular dystrophy patients. To understand the role of low-level dystrophin expression, we compared muscle force and pathology in mdx3cv and mdx4cv mice. Dystrophin was eliminated in mdx4cv mouse muscle but was expressed in mdx3cv mice as a near full-length protein at ∼5% of normal levels. Consistent with previous reports, we found dystrophic muscle pathology in both mouse strains. Surprisingly, mdx3cv extensor digitorium longus muscle showed significantly higher tetanic force and was also more resistant to eccentric contraction-induced injury than mdx4cv extensor digitorium longus muscle. Furthermore, mdx3cv mice had stronger forelimb grip strength than mdx4cv mice. Immunostaining revealed utrophin up-regulation in both mouse strains. The dystrophin-associated glycoprotein complex was also restored in the sarcolemma in both strains although at levels lower than those in normal mice. Our results suggest that subtherapeutic expression levels of near full-length, membrane-bound dystrophin, possibly in conjunction with up-regulated utrophin levels, may help maintain minimal muscle force but not arrest muscle degeneration or necrosis. Our findings provide valuable insight toward understanding delayed clinical onset and/or slow disease progression in certain Becker muscular dystrophy patients. PMID:18385524

  14. Short Telomeres and Stem Cell Exhaustion Model Duchenne Muscular Dystrophy in mdx/mTR Mice

    PubMed Central

    Sacco, Alessandra; Mourkioti, Foteini; Tran, Rose; Choi, Jinkuk; Llewellyn, Michael; Kraft, Peggy; Shkreli, Marina; Delp, Scott; Pomerantz, Jason H.; Artandi, Steven E.; Blau, Helen M.

    2010-01-01

    Summary In Duchenne muscular dystrophy (DMD), dystrophin mutation leads to progressive lethal skeletal muscle degeneration. For unknown reasons, dystrophin deficiency does not recapitulate DMD in mice (mdx), which have mild skeletal muscle defects and potent regenerative capacity. We postulated that human DMD progression is a consequence of loss of functional muscle stem cells (MuSC) and the mild mouse mdx phenotype results from greater MuSC reserve fueled by longer telomeres. We report that mdx mice lacking the RNA component of telomerase (mdx/mTR) have shortened telomeres in muscle cells and severe muscular dystrophy that progressively worsens with age. Muscle wasting severity parallels a decline in MuSC regenerative capacity, and is ameliorated histologically by transplantation of wild-type MuSC. These data show that DMD progression results in part from a cell-autonomous failure of MuSC to maintain the damage-repair cycle initiated by dystrophin deficiency. The essential role of MuSC function has therapeutic implications for DMD. PMID:21145579

  15. Duodenum identification mechanism for capsule endoscopy.

    PubMed

    Woo, Sang Hyo; Mohy-Ud-Din, Zia; Cho, Jin Ho

    2011-04-01

    The aim of this study is to implement a duodenum identification mechanism for capsule endoscopes because commercially available capsule endoscopes sometimes present a false negative diagnosis of the duodenum. One reason for the false negative diagnosis is that the duodenum is the fastest moving part within the gastrointestinal tract and the current frame rate of the capsule is not fast enough. When the capsule can automatically identify that it is in the duodenum, the frame rate of the capsule can be temporarily increased to reduce the possibility of a false negative diagnosis. This study proposes a mechanism to identify the duodenum using capacitive proximity sensors that can distinguish the surrounding tissue and transmit data using RF communication. The implemented capsule (D11 mm × L22 mm) was smaller than the commercially available capsule endoscopes, and power consumption was as low as 0.642 mW. Preexperiments were conducted to select an appropriate electrode width in order to increase the signal-to-noise ratio (SNR), and in vitro experiments were conducted to verify whether the implemented capsule could identify the duodenum within 3 s. The experiment showed that the identification rate of duodenum was 93% when the velocity of the capsule was less than 1 cm/s. PMID:21134813

  16. Gastrointestinal stromal tumor (gist) of the duodenum.

    PubMed

    Ghazanfar, Shahriyar; Sial, Khadim S; Quraishy, M S

    2007-06-01

    This is a report of a rare gastrointestinal stromal tumor of the duodenum in a 75 years old man who presented with recurrent episodes of intestinal obstruction and melena. The patient underwent successful Whipple's procedure. PMID:17623589

  17. SCFA transport in rat duodenum

    PubMed Central

    Kaji, Izumi; Iwanaga, Toshihiko; Watanabe, Masahiko; Guth, Paul H.; Engel, Eli; Akiba, Yasutada

    2014-01-01

    Bacterial or ingested food-derived short-chain fatty acids (SCFAs) are present in the duodenal lumen. Acetate, the most abundant SCFA in the foregut lumen, is absorbed immediately after ingestion, although the mechanism by which this absorption occurs is not fully understood. We investigated the distribution and function of candidate SCFA transporters in rat duodenum. The Na+-coupled monocarboxylate transporter-1 (SMCT1) was localized to the brush border, whereas the pH-dependent monocarboxylate transporter (MCT) 1 and MCT4 were localized to the duodenocyte basolateral membrane. In Ussing chambered duodenal mucosa, luminal acetate dose-dependently increased short-circuit current (Isc) in the presence of serosal bumetanide and indomethacin by a luminal Na+-dependent, ouabain-sensitive mechanism. The Isc response was inhibited dose-dependently by the SMCT1 nonsubstrate inhibitor ibuprofen, consistent with net electrogenic absorption of acetate via SMCT1. Other SCFAs and lactate also increased Isc. Furthermore, duodenal loop perfusion of acetate increased portal venous acetate concentration, inhibited by coperfusion of ibuprofen or a MCT inhibitor. Luminal acetate perfusion increased duodenal HCO3− secretion via capsaicin-sensitive afferent nerve activation and cyclooxygenase activity, consistent with absorption-mediated HCO3− secretion. These results suggest that absorption of luminal SCFA via SMCT1 and MCTs increases duodenal HCO3− secretion. In addition to SCFA sensing via free fatty acid receptors, the presence of rapid duodenal SCFA absorption may be important for the suppression of luminal bacterial colonization and implicated in the generation of functional dyspepsia due to bacterial overgrowth. PMID:25394661

  18. Utrophins compensate for Dp71 absence in mdx3cv in adhered platelets.

    PubMed

    Cerecedo, Doris; Mondragón, Ricardo; Candelario, Aurora; García-Sierra, Francisco; Mornet, Dominique; Rendón, Alvaro; Martínez-Rojas, Dalila

    2008-01-01

    Platelet adhesion is a critical step due to its hemostatic role in stopping bleeding after vascular damage. Short dystrophins are the most abundant dmd gene products in nonmuscle tissues, and in association with cytoskeleton proteins contribute to their intrinsic function; while utrophins are dystrophin-homologous related family proteins with structural and functional similarities. We previously demonstrated the presence of Dp71 isoforms, utrophins, and various dystrophin-associated proteins and their participation in cytoskeleton re-organization, filopodia and lamellipodia extension, and in centralizing cytoplasmic granules during the adhesion process of human platelets. To evaluate the morphologic changes and actin-based structures of mdx(3cv) platelets during the adhesion process, we compared the topographic distribution of Dp71d/Dp71Delta110(m) and dystrophin-associated protein in adhered platelets from dystrophic mdx(3cv) mouse. By confocal microscopy, we showed that absence of Dp71 isoforms in platelets from this animal model disrupted dystrophin-associated protein expression and distribution without modifying the platelet morphology displayed during the glass-adhesion process. By immunoprecipitation assays, we proved that up-regulated utrophins were associated with dystrophin-associated proteins to conform the dystrophin-associated protein complex corresponding to utrophins, which might compensate for Dp71 absence in mdx(3cv) platelets. PMID:18180614

  19. Effects of Dantrolene Therapy on Disease Phenotype in Dystrophin Deficient mdx Mice.

    PubMed

    Quinn, James L; Huynh, Tony; Uaesoontrachoon, Kitipong; Tatem, Kathleen; Phadke, Aditi; Van der Meulen, Jack H; Yu, Qing; Nagaraju, Kannaboyina

    2013-01-01

    Dystrophin deficiency causes contraction-induced injury and damage to the muscle fiber, resulting in sustained increase in intracellular calcium levels, activation of calcium-dependent proteases and cell death. It is known that the Ryanodine receptor (RyR1) on the sarcoplasmic reticular (SR) membrane controls calcium release. Dantrolene, an FDA approved skeletal muscle relaxant, inhibits the release of calcium from the SR during excitation-contraction and suppresses uncontrolled calcium release by directly acting on the RyR complex to limit its activation. This study examines whether Dantrolene can reduce the disease phenotype in the mdx mouse model of muscular dystrophy. We treated mdx mice (4 weeks old) with daily intraperitoneal injections of 40mg/kg of Dantrolene for 6 weeks and measured functional (grip strength, in vitro force contractions), behavioral (open field digiscan), imagining (optical imaging for inflammation), histological (H&E), and molecular (protein and RNA) endpoints in a blinded fashion. We found that treatment with Dantrolene resulted in decreased grip strength and open field behavioral activity in mdx mice. There was no significant difference in inflammation either by optical imaging analysis of cathepsin activity or histological (H&E) analysis. In vitro force contraction measures showed no changes in EDL muscle-specific force, lengthening-contraction force deficit, or fatigue resistance. We found Dantrolene treatment significantly reduces serum CK levels. Further, Dantrolene-treated mice showed decreased SERCA1 but not RyR1 expression in skeletal muscle. These results suggest that Dantrolene treatment alone has no significant beneficial effects at the tested doses in young mdx mice. PMID:24270550

  20. Effects of Dantrolene Therapy on Disease Phenotype in Dystrophin Deficient mdx Mice

    PubMed Central

    Quinn, James L; Huynh, Tony; Uaesoontrachoon, Kitipong; Tatem, Kathleen; Phadke, Aditi; Van der Meulen, Jack H; Yu, Qing; Nagaraju, Kannaboyina

    2013-01-01

    Dystrophin deficiency causes contraction-induced injury and damage to the muscle fiber, resulting in sustained increase in intracellular calcium levels, activation of calcium-dependent proteases and cell death. It is known that the Ryanodine receptor (RyR1) on the sarcoplasmic reticular (SR) membrane controls calcium release. Dantrolene, an FDA approved skeletal muscle relaxant, inhibits the release of calcium from the SR during excitation-contraction and suppresses uncontrolled calcium release by directly acting on the RyR complex to limit its activation. This study examines whether Dantrolene can reduce the disease phenotype in the mdx mouse model of muscular dystrophy. We treated mdx mice (4 weeks old) with daily intraperitoneal injections of 40mg/kg of Dantrolene for 6 weeks and measured functional (grip strength, in vitro force contractions), behavioral (open field digiscan), imagining (optical imaging for inflammation), histological (H&E), and molecular (protein and RNA) endpoints in a blinded fashion. We found that treatment with Dantrolene resulted in decreased grip strength and open field behavioral activity in mdx mice. There was no significant difference in inflammation either by optical imaging analysis of cathepsin activity or histological (H&E) analysis. In vitro force contraction measures showed no changes in EDL muscle-specific force, lengthening-contraction force deficit, or fatigue resistance. We found Dantrolene treatment significantly reduces serum CK levels. Further, Dantrolene-treated mice showed decreased SERCA1 but not RyR1 expression in skeletal muscle. These results suggest that Dantrolene treatment alone has no significant beneficial effects at the tested doses in young mdx mice. PMID:24270550

  1. Novel adeno-associated viral vector delivering the utrophin gene regulator jazz counteracts dystrophic pathology in mdx mice.

    PubMed

    Strimpakos, Georgios; Corbi, Nicoletta; Pisani, Cinzia; Di Certo, Maria Grazia; Onori, Annalisa; Luvisetto, Siro; Severini, Cinzia; Gabanella, Francesca; Monaco, Lucia; Mattei, Elisabetta; Passananti, Claudio

    2014-09-01

    Over-expression of the dystrophin-related gene utrophin represents a promising therapeutic strategy for Duchenne muscular dystrophy (DMD). The strategy is based on the ability of utrophin to functionally replace defective dystrophin. We developed the artificial zinc finger transcription factor "Jazz" that up-regulates both the human and mouse utrophin promoter. We observed a significant recovery of muscle strength in dystrophic Jazz-transgenic mdx mice. Here we demonstrate the efficacy of an experimental gene therapy based on the systemic delivery of Jazz gene in mdx mice by adeno-associated virus (AAV). AAV serotype 8 was chosen on the basis of its high affinity for skeletal muscle. Muscle-specific expression of the therapeutic Jazz gene was enhanced by adding the muscle α-actin promoter to the AAV vector (mAAV). Injection of mAAV8-Jazz viral preparations into mdx mice resulted in muscle-specific Jazz expression coupled with up-regulation of the utrophin gene. We show a significant recovery from the dystrophic phenotype in mAAV8-Jazz-treated mdx mice. Histological and physiological analysis revealed a reduction of fiber necrosis and inflammatory cell infiltration associated with functional recovery in muscle contractile force. The combination of ZF-ATF technology with the AAV delivery can open a new avenue to obtain a therapeutic strategy for treatment of DMD. PMID:24469912

  2. In vivo and in vitro correction of the mdx dystrophin gene nonsense mutation by short-fragment homologous replacement.

    PubMed

    Kapsa, R; Quigley, A; Lynch, G S; Steeper, K; Kornberg, A J; Gregorevic, P; Austin, L; Byrne, E

    2001-04-10

    Targeted genetic correction of mutations in cells is a potential strategy for treating human conditions that involve nonsense, missense, and transcriptional splice junction mutations. One method of targeted gene repair, single-stranded short-fragment homologous replacement (ssSFHR), has been successful in repairing the common deltaF508 3-bp microdeletion at the cystic fibrosis transmembrane conductance regulator (CFTR) locus in 1% of airway epithelial cells in culture. This study investigates in vitro and in vivo application of a double-stranded method variant of SFHR gene repair to the mdx mouse model of Duchenne muscular dystrophy (DMD). A 603-bp wild-type PCR product was used to repair the exon 23 C-to-T mdx nonsense transition at the Xp21.1 dys locus in cultured myoblasts and in tibialis anterior (TA) from male mdx mice. Multiple transfection and variation of lipofection reagent both improved in vitro SFHR efficiency, with successful conversion of mdx to wild-type nucleotide at the dys locus achieved in 15 to 20% of cultured loci and in 0.0005 to 0.1% of TA. The genetic correction of mdx myoblasts was shown to persist for up to 28 days in culture and for at least 3 weeks in TA. While a high frequency of in vitro gene repair was observed, the lipofection used here appeared to have adverse effects on subsequent cell viability and corrected cells did not express dystrophin transcript. With further improvements to in vitro and in vivo gene repair efficiencies, SFHR may find some application in DMD and other genetic neuromuscular disorders in humans. PMID:11426463

  3. Lack of Dystrophin Affects Bronchial Epithelium in mdx Mice.

    PubMed

    Morici, Giuseppe; Rappa, Francesca; Cappello, Francesco; Pace, Elisabetta; Pace, Andrea; Mudò, Giuseppa; Crescimanno, Grazia; Belluardo, Natale; Bonsignore, Maria R

    2016-10-01

    Mild exercise training may positively affect the course of Duchenne Muscular Dystrophy (DMD). Training causes mild bronchial epithelial injury in both humans and mice, but no study assessed the effects of exercise in mdx mice, a well known model of DMD. The airway epithelium was examined in mdx (C57BL/10ScSn-Dmdmdx) mice, and in wild type (WT, C57BL/10ScSc) mice either under sedentary conditions (mdx-SD, WT-SD) or during mild exercise training (mdx-EX, WT-EX). At baseline, and after 30 and 45 days of training (5 d/wk for 6 weeks), epithelial morphology and markers of regeneration, apoptosis, and cellular stress were assessed. The number of goblet cells in bronchial epithelium was much lower in mdx than in WT mice under all conditions. At 30 days, epithelial regeneration (PCNA positive cells) was higher in EX than SD animals in both groups; however, at 45 days, epithelial regeneration decreased in mdx mice irrespective of training, and the percentage of apoptotic (TUNEL positive) cells was higher in mdx-EX than in WT-EX mice. Epithelial expression of HSP60 (marker of stress) progressively decreased, and inversely correlated with epithelial apoptosis (r = -0.66, P = 0.01) only in mdx mice. Lack of dystrophin in mdx mice appears associated with defective epithelial differentiation, and transient epithelial regeneration during mild exercise training. Hence, lack of dystrophin might impair repair in bronchial epithelium, with potential clinical consequences in DMD patients. J. Cell. Physiol. 231: 2218-2223, 2016. © 2016 Wiley Periodicals, Inc. PMID:26868633

  4. SERCA1 overexpression minimizes skeletal muscle damage in dystrophic mouse models

    PubMed Central

    Mázala, Davi A. G.; Pratt, Stephen J. P.; Chen, Dapeng; Molkentin, Jeffery D.; Lovering, Richard M.

    2015-01-01

    Duchenne muscular dystrophy (DMD) is characterized by progressive muscle wasting secondary to repeated muscle damage and inadequate repair. Elevations in intracellular free Ca2+ have been implicated in disease progression, and sarcoplasmic/endoplasmic reticulum Ca2+-ATPase 1 (SERCA1) overexpression has been shown to ameliorate the dystrophic phenotype in mdx mice. The purpose of this study was to assess the effects of SERCA1 overexpression in the more severe mdx/Utr−/− mouse model of DMD. Mice overexpressing SERCA1 were crossed with mdx/Utr+/− mice to generate mdx/Utr−/−/+SERCA1 mice and compared with wild-type (WT), WT/+SERCA1, mdx/+SERCA1, and genotype controls. Mice were assessed at ∼12 wk of age for changes in Ca2+ handling, muscle mass, quadriceps torque, markers of muscle damage, and response to repeated eccentric contractions. SERCA1-overexpressing mice had a two- to threefold increase in maximal sarcoplasmic reticulum Ca2+-ATPase activity compared with WT which was associated with normalization in body mass for both mdx/+SERCA1 and mdx/Utr−/−/+SERCA1. Torque deficit in the quadriceps after eccentric injury was 2.7-fold greater in mdx/Utr−/− vs. WT mice, but only 1.5-fold greater in mdx/Utr−/−/+SERCA1 vs. WT mice, an attenuation of 44%. Markers of muscle damage (% centrally nucleated fibers, necrotic area, and serum creatine kinase levels) were higher in both mdx and mdx/Utr−/− vs. WT, and all were attenuated by overexpression of SERCA1. These data indicate that SERCA1 overexpression ameliorates functional impairments and cellular markers of damage in a more severe mouse model of DMD. These findings support targeting intracellular Ca2+ control as a therapeutic approach for DMD. PMID:25652448

  5. CRISPR-mediated Genome Editing Restores Dystrophin Expression and Function in mdx Mice.

    PubMed

    Xu, Li; Park, Ki Ho; Zhao, Lixia; Xu, Jing; El Refaey, Mona; Gao, Yandi; Zhu, Hua; Ma, Jianjie; Han, Renzhi

    2016-03-01

    Duchenne muscular dystrophy (DMD) is a degenerative muscle disease caused by genetic mutations that lead to the disruption of dystrophin in muscle fibers. There is no curative treatment for this devastating disease. Clustered regularly interspaced short palindromic repeat/Cas9 (CRISPR/Cas9) has emerged as a powerful tool for genetic manipulation and potential therapy. Here we demonstrate that CRIPSR-mediated genome editing efficiently excised a 23-kb genomic region on the X-chromosome covering the mutant exon 23 in a mouse model of DMD, and restored dystrophin expression and the dystrophin-glycoprotein complex at the sarcolemma of skeletal muscles in live mdx mice. Electroporation-mediated transfection of the Cas9/gRNA constructs in the skeletal muscles of mdx mice normalized the calcium sparks in response to osmotic shock. Adenovirus-mediated transduction of Cas9/gRNA greatly reduced the Evans blue dye uptake of skeletal muscles at rest and after downhill treadmill running. This study provides proof evidence for permanent gene correction in DMD. PMID:26449883

  6. Whole Body Periodic Acceleration Is an Effective Therapy to Ameliorate Muscular Dystrophy in mdx Mice

    PubMed Central

    Altamirano, Francisco; Perez, Claudio F.; Liu, Min; Widrick, Jeffrey; Barton, Elisabeth R.; Allen, Paul D.; Adams, Jose A.; Lopez, Jose R.

    2014-01-01

    Duchenne muscular dystrophy (DMD) is a genetic disorder caused by the absence of dystrophin in both skeletal and cardiac muscles. This leads to severe muscle degeneration, and dilated cardiomyopathy that produces patient death, which in most cases occurs before the end of the second decade. Several lines of evidence have shown that modulators of nitric oxide (NO) pathway can improve skeletal muscle and cardiac function in the mdx mouse, a mouse model for DMD. Whole body periodic acceleration (pGz) is produced by applying sinusoidal motion to supine humans and in standing conscious rodents in a headward-footward direction using a motion platform. It adds small pulses as a function of movement frequency to the circulation thereby increasing pulsatile shear stress to the vascular endothelium, which in turn increases production of NO. In this study, we examined the potential therapeutic properties of pGz for the treatment of skeletal muscle pathology observed in the mdx mouse. We found that pGz (480 cpm, 8 days, 1 hr per day) decreased intracellular Ca2+ and Na+ overload, diminished serum levels of creatine kinase (CK) and reduced intracellular accumulation of Evans Blue. Furthermore, pGz increased muscle force generation and expression of both utrophin and the carboxy-terminal PDZ ligand of nNOS (CAPON). Likewise, pGz (120 cpm, 12 h) applied in vitro to skeletal muscle myotubes reduced Ca2+ and Na+ overload, diminished abnormal sarcolemmal Ca2+ entry and increased phosphorylation of endothelial NOS. Overall, this study provides new insights into the potential therapeutic efficacy of pGz as a non-invasive and non-pharmacological approach for the treatment of DMD patients through activation of the NO pathway. PMID:25181488

  7. Whole body periodic acceleration is an effective therapy to ameliorate muscular dystrophy in mdx mice.

    PubMed

    Altamirano, Francisco; Perez, Claudio F; Liu, Min; Widrick, Jeffrey; Barton, Elisabeth R; Allen, Paul D; Adams, Jose A; Lopez, Jose R

    2014-01-01

    Duchenne muscular dystrophy (DMD) is a genetic disorder caused by the absence of dystrophin in both skeletal and cardiac muscles. This leads to severe muscle degeneration, and dilated cardiomyopathy that produces patient death, which in most cases occurs before the end of the second decade. Several lines of evidence have shown that modulators of nitric oxide (NO) pathway can improve skeletal muscle and cardiac function in the mdx mouse, a mouse model for DMD. Whole body periodic acceleration (pGz) is produced by applying sinusoidal motion to supine humans and in standing conscious rodents in a headward-footward direction using a motion platform. It adds small pulses as a function of movement frequency to the circulation thereby increasing pulsatile shear stress to the vascular endothelium, which in turn increases production of NO. In this study, we examined the potential therapeutic properties of pGz for the treatment of skeletal muscle pathology observed in the mdx mouse. We found that pGz (480 cpm, 8 days, 1 hr per day) decreased intracellular Ca(2+) and Na(+) overload, diminished serum levels of creatine kinase (CK) and reduced intracellular accumulation of Evans Blue. Furthermore, pGz increased muscle force generation and expression of both utrophin and the carboxy-terminal PDZ ligand of nNOS (CAPON). Likewise, pGz (120 cpm, 12 h) applied in vitro to skeletal muscle myotubes reduced Ca(2+) and Na(+) overload, diminished abnormal sarcolemmal Ca(2+) entry and increased phosphorylation of endothelial NOS. Overall, this study provides new insights into the potential therapeutic efficacy of pGz as a non-invasive and non-pharmacological approach for the treatment of DMD patients through activation of the NO pathway. PMID:25181488

  8. Laryngeal Muscles Are Spared in the Dystrophin Deficient "mdx" Mouse

    ERIC Educational Resources Information Center

    Thomas, Lisa B.; Joseph, Gayle L.; Adkins, Tracey D.; Andrade, Francisco H.; Stemple, Joseph C.

    2008-01-01

    Purpose: "Duchenne muscular dystrophy (DMD)" is caused by the loss of the cytoskeletal protein, dystrophin. The disease leads to severe and progressive skeletal muscle wasting. Interestingly, the disease spares some muscles. The purpose of the study was to determine the effects of dystrophin deficiency on 2 intrinsic laryngeal muscles, the…

  9. Laparoscopic duodenum-preserving pancreatic head resection

    PubMed Central

    Zhou, Jiayu; Zhou, Yucheng; Mou, Yiping; Xia, Tao; Xu, Xiaowu; Jin, Weiwei; Zhang, Renchao; Lu, Chao; Chen, Ronggao

    2016-01-01

    Abstract Background: Solid pseudopapillary neoplasms (SPNs) of the pancreas are uncommon neoplasms and are potentially malignant. Complete resection is advised due to rare recurrence and metastasis. Duodenum-preserving pancreatic head resection (DPPHR) is indicated for SPNs located in the pancreatic head and is only performed using the open approach. To the best of our knowledge, there are no reports describing laparoscopic DPPHR (LDPPHR) for SPNs. Methods: Herein, we report a case of 41-year-old female presented with a 1-week history of epigastric abdominal discomfort, and founded an SPN of the pancreatic head by abdominal computed tomography/magnetic resonance, who was treated by radical LDPPHR without complications, such as pancreatic fistula and bile leakage. Histological examination of the resected specimen confirmed the diagnosis of SPN. Results: The patient was discharged 1 week after surgery following an uneventful postoperative period. She was followed up 3 months without readmission and local recurrence according to abdominal ultrasound. Conclusion: LDPPHR is a safe, feasible, and effective surgical procedure for SPNs. PMID:27512859

  10. Overview of MDX-A System for Medical Diagnosis

    PubMed Central

    Mittal, S.; Chandrasekaran, B.; Smith, J.

    1979-01-01

    We describe the design and performance of MDX, an experimental medical diagnosis system, which currently diagnoses in the syndrome called Cholestasis. The needed medical knowledge is represented in a scheme called conceptual structures, which can be viewed as a collection of conceptual experts interacting according to certain well-defined principles. MDX has three components: the diagnostic system, a patient data base and a radiology consultant. We describe these components, the inter-expert communication system and the query language used by these components. The system is illustrated by means of its performance on a real case.

  11. D-Amino Acid Substitution of Peptide-Mediated NF-κB Suppression in mdx Mice Preserves Therapeutic Benefit in Skeletal Muscle, but Causes Kidney Toxicity.

    PubMed

    Reay, Daniel P; Bastacky, Sheldon I; Wack, Kathryn E; Stolz, Donna B; Robbins, Paul D; Clemens, Paula R

    2015-01-01

    In Duchenne muscular dystrophy (DMD) patients and the mdx mouse model of DMD, chronic activation of the classical nuclear factor-κB (NF-κB) pathway contributes to the pathogenesis that causes degeneration of muscle fibers, inflammation and fibrosis. Prior studies demonstrate that inhibition of inhibitor of κB kinase (IKK)-mediated NF-κB activation using L-isomer NF-κB essential modulator (NEMO)-binding domain (NBD) peptide-based approaches reduce muscle pathology in the mdx mouse. For our studies, the NBD peptide is synthesized as a fusion peptide with an eight-lysine (8K) protein transduction domain to facilitate intracellular delivery. We hypothesized that the d-isoform peptide could have a greater effect than the naturally occurring L-isoform peptide due to the longer persistence of the D-isoform peptide in vivo. In this study, we compared systemic treatment with low (1 mg/kg) and high (10 mg/kg) doses of L- and D-isomer 8K-wild-type-NBD peptide in mdx mice. Treatment with both L- or D-isoform 8K-wild-type-NBD peptide resulted in decreased activation of NF-κB and improved histology in skeletal muscle of the mdx mouse. However, we observed kidney toxicity (characterized by proteinuria), increased serum creatinine, activation of NF-κB and pathological changes in kidney cortex that were most severe with treatment with the D-isoform of 8K-wild-type-NBD peptide. The observed toxicity was also seen in normal mice. PMID:26018805

  12. Removal of Duodenum Elicits GLP-1 Secretion

    PubMed Central

    Muscogiuri, Giovanna; Mezza, Teresa; Prioletta, Annamaria; Sorice, Gian Pio; Clemente, Gennaro; Sarno, Gerardo; Nuzzo, Gennaro; Pontecorvi, Alfredo; Holst, Jens J.; Giaccari, Andrea

    2013-01-01

    OBJECTIVE To evaluate the effect of removal of the duodenum on the complex interplay between incretins, insulin, and glucagon in nondiabetic subjects. RESEARCH DESIGN AND METHODS For evaluation of hormonal secretion and insulin sensitivity, 10 overweight patients without type 2 diabetes (age 61 ± 19.3 years and BMI 27.9 ± 5.3 kg/m2) underwent a mixed-meal test and a hyperinsulinemic-euglycemic clamp before and after pylorus-preserving pancreatoduodenectomy for ampulloma. RESULTS All patients experienced a reduction in insulin (P = 0.002), C-peptide (P = 0.0002), and gastric inhibitory peptide (GIP) secretion (P = 0.0004), while both fasting and postprandial glucose levels increased (P = 0.0001); GLP-1 and glucagon responses to the mixed meal increased significantly after surgery (P = 0.02 and 0.031). While changes in GIP levels did not correlate with insulin, glucagon, and glucose levels, the increase in GLP-1 secretion was inversely related to the postsurgery decrease in insulin secretion (R2 = 0.56; P = 0.012) but not to the increased glucagon secretion, which correlated inversely with the reduction of insulin (R2 = 0.46; P = 0.03) and C-peptide (R2 = 0.37; P = 0.04). Given that the remaining pancreas presumably has preserved intraislet anatomy, insulin secretory capacity, and α- and β-cell interplay, our data suggest that the increased glucagon secretion is related to decreased systemic insulin. CONCLUSIONS Pylorus-preserving pancreatoduodenectomy was associated with a decrease in GIP and a remarkable increase in GLP-1 levels, which was not translated into increased insulin secretion. Rather, the hypoinsulinemia may have caused an increase in glucagon secretion. PMID:23393218

  13. Physiological Characterization of Muscle Strength With Variable Levels of Dystrophin Restoration in mdx Mice Following Local Antisense Therapy

    PubMed Central

    Sharp, Paul S; Bye-a-Jee, Hema; Wells, Dominic J

    2011-01-01

    Antisense-induced exon skipping can restore the open reading frame, and thus correct the dystrophin deficiency that causes Duchenne muscular dystrophy (DMD), a lethal muscle wasting condition. Successful proof-of-principle in preclinical models has led to human clinical trials. However, it is still not known what percentage of dystrophin-positive fibers and what level of expression is necessary for functional improvement. This study directly address these key questions in the mdx mouse model of DMD. To achieve a significant variation in dystrophin expression, we locally administered into tibialis anterior muscles various doses of a phosphorodiamidate morpholino oligomer (PMO) designed to skip the mutated exon 23 from the mRNA of murine dystrophin. We found a highly significant correlation between the number of dystrophin-positive fibers and resistance to contraction-induced injury, with a minimum of 20% of dystrophin-positive fibers required for meaningful improvement. Furthermore, our results also indicate that a relatively low level of dystrophin expression in muscle fibers may have significant clinical benefits. In contrast, improvements in muscle force were not correlated with either the number of positive fibers or total dystrophin levels, which highlight the need to conduct appropriate functional assessments in preclinical testing using the mdx mouse. PMID:20924363

  14. Electroretinographic genotype-phenotype correlations for mouse and man at the dmd/DMD locus

    SciTech Connect

    Millers, D.M.; Weleber, R.G.; Woodward, W.R.

    1994-09-01

    Reduced or absent b-waves in the dark-adapted electroretinogram (ERG) of Duchenne and Becker muscular dystrophy (DMD/BMD) patients led to the identification of dystrophin in human retina and the proposal that it plays a role in retinal electrophysiology. Study of a large group of Duchenne and Becker muscular dystrophy males to determine their ocular characteristics indicated that there were position-specific effects of deletions, with 3{prime} defects associated with severe electroretinographic changes, whereas some 5{prime} patients demonstrated less severe, or even normal, ERGs. We studied the mdx mouse, a model with X-linked muscular dystrophy and defective full-length dystrophin, which failed to show any ERG abnormalities. Given the presence of alternate isoforms of dystrophin in retina, and the 5{prime} deletion DMD/BMD patients with normal ERGs, we studied mouse models with differing dystrophin mutations (mdx{sup Cv3}, mdx{sup Cv5}) to determine the usefulness of alternate strains as models for the visual effects of dystropin. Abnormal ERGs similar to those seen in DMD/BMS patients exist in the mdx{sup Cv3} strain of muscular dystrophy mice. Normal ERGs were found the mdx{sup Cv5} strain. The mutations in the mdx and mdx{sup Cv5} mice have been mapped to the 5{prime} end of the dmd gene, while the mutation in the mdx{sup Cv3} mouse is in the 3{prime} end. Thus, there are position effects of the gene defect on the ERG phenotype that are conserved in the mouse. Such genotype-phenotype correlations may reflect differential expression of shorter isoforms of dystrophin.

  15. A Multidisciplinary Evaluation of the Effectiveness of Cyclosporine A in Dystrophic Mdx Mice

    PubMed Central

    De Luca, Annamaria; Nico, Beatrice; Liantonio, Antonella; Paola Didonna, Maria; Fraysse, Bodvael; Pierno, Sabata; Burdi, Rosa; Mangieri, Domenica; Rolland, Jean-François; Camerino, Claudia; Zallone, Alberta; Confalonieri, Paolo; Andreetta, Francesca; Arnoldi, Elisa; Courdier-Fruh, Isabelle; Magyar, Josef P.; Frigeri, Antonio; Pisoni, Michela; Svelto, Maria; Conte-Camerino, Diana

    2005-01-01

    Chronic inflammation is a secondary reaction of Duchenne muscular dystrophy and may contribute to disease progression. To examine whether immunosuppressant therapies could benefit dystrophic patients, we analyzed the effects of cyclosporine A (CsA) on a dystrophic mouse model. Mdx mice were treated with 10 mg/kg of CsA for 4 to 8 weeks throughout a period of exercise on treadmill, a protocol that worsens the dystrophic condition. The CsA treatment fully prevented the 60% drop of forelimb strength induced by exercise. A significant amelioration (P < 0.05) was observed in histological profile of CsA-treated gastrocnemius muscle with reductions of nonmuscle area (20%), centronucleated fibers (12%), and degenerating area (50%) compared to untreated exercised mdx mice. Consequently, the percentage of normal fibers increased from 26 to 35% in CsA-treated mice. Decreases in creatine kinase and markers of fibrosis were also observed. By electrophysiological recordings ex vivo, we found that CsA counteracted the decrease in chloride conductance (gCl), a functional index of degeneration in diaphragm and extensor digitorum longus muscle fibers. However, electrophysiology and fura-2 calcium imaging did not show any amelioration of calcium homeostasis in extensor digitorum longus muscle fibers. No significant effect was observed on utrophin levels in diaphragm muscle. Our data show that the CsA treatment significantly normalized many functional, histological, and biochemical endpoints by acting on events that are independent or downstream of calcium homeostasis. The beneficial effect of CsA may involve different targets, reinforcing the usefulness of immunosuppressant drugs in muscular dystrophy. PMID:15681831

  16. Expression of human dystrophin following the transplantation of genetically modified mdx myoblasts.

    PubMed

    Moisset, P A; Gagnon, Y; Karpati, G; Tremblay, J P

    1998-10-01

    Transplantation of genetically modified autologous myoblasts has been proposed as a possible solution to avoid long-term use of immunosuppressive drugs. To determine the conditions to be used in this kind of approach for possible treatment of dystrophin deficiency, mdx myoblasts were infected at different multiplicities of infection (MOI or 0.01-1000) with an adenoviral vector containing a CMV promoter/enhancer driven 6.3 kb human dystrophin cDNA (minigene) and tested in vitro for transgene expression. In these cultures, dystrophin mRNA was found to be proportionate with increasing MOI. Primary myoblast cultures derived from transgenic mdx mice expressing beta-Gal under a muscle-specific promoter and showing high expression of the human mini-dystrophin transgene introduced by the adenoviral vector were grafted into anterior tibialis muscles of SCID mice. Ten and 24 days after transplantation, numerous muscle fibers expressing both human dystrophin and beta-Gal were detected throughout the mouse muscles by immunohistochemistry using an antibody specific for human dystrophin. The presence of the human mini-dystrophin mRNA was also detected by RT-PCR. These results demonstrate that three essential conditions in autologous myoblast transplantation can be achieved: (1) in vivo survival of at least some of the transduced myoblasts; (2) efficient fusion of these cells with the host muscle fibers; and (3) the high expression of the dystrophin transgene in situ. Furthermore, this article provides a novel RT-PCR-based technique to quantify the human dystrophin minigene expression in vitro and in vivo. PMID:9930339

  17. Video endoscopy: removal of retained sewing needles from the duodenum.

    PubMed

    Gajbhiye, Ashok S; Gajbhiye, Raj N; Tirupude, Bhupesh H; Bajaj, Prasang P; Gupta, Tarush H

    2013-06-01

    We report an interesting case of a 21-year-old unmarried girl who swallowed six sewing needles. Her complaints were pain in the epigastrium, associated with nausea and vomiting. On examination, there was mild tenderness in the epigastrium. X-ray of the abdomen and endoscopy confirmed the presence of six needles in the duodenum, with tips lodged in the duodenal wall. Psychiatric opinion was sought which was normal. Under video endoscope (Pentax 2.8, EG 27708) guidance with Captura biopsy forceps without spikes (Cook DBF-2.4-160-S), six sewing needles were removed successfully from the duodenum through the endoscope channel without any complications. However, a video endoscopic removal of the retained six needles from duodenum is probably being reported for the first time. PMID:24426531

  18. Enhancement of Muscle T Regulatory Cells and Improvement of Muscular Dystrophic Process in mdx Mice by Blockade of Extracellular ATP/P2X Axis.

    PubMed

    Gazzerro, Elisabetta; Baldassari, Simona; Assereto, Stefania; Fruscione, Floriana; Pistorio, Angela; Panicucci, Chiara; Volpi, Stefano; Perruzza, Lisa; Fiorillo, Chiara; Minetti, Carlo; Traggiai, Elisabetta; Grassi, Fabio; Bruno, Claudio

    2015-12-01

    Infiltration of immune cells and chronic inflammation substantially affect skeletal and cardiac muscle degeneration in Duchenne muscular dystrophy. In the immune system, extracellular adenosine triphosphate (ATP) released by dying cells is sensed as a danger associated molecular pattern through P2 purinergic receptors. Specifically, the P2X7 subtype has a prominent role in regulating immune system physiology and contributes to inflammasome activation also in muscle cells. Here, we show that in vivo blockade of the extracellular ATP/P2X purinergic signaling pathway by periodate-oxidized ATP delayed the progression of the dystrophic phenotype and dampened the local inflammatory response in mdx mice, a spontaneous mouse model of dystrophin deficiency. Reduced infiltration of leukocytes and macrophages and decreased expression of IL-6 were revealed in the muscles of periodate-oxidized ATP-treated mdx mice. Concomitantly, an increase in Foxp3(+) immunosuppressive regulatory T cells was observed and correlated with enhanced myofiber regeneration. Moreover, we detected reduced concentrations of profibrotic cytokines, including transforming growth factor-β and connective tissue growth factor, in muscles of periodate-oxidized ATP-treated mdx mice. The improvement of inflammatory features was associated with increased strength and reduced necrosis, thus suggesting that pharmacologic purinergic antagonism altering the adaptive immune component in the muscle infiltrates might represent a promising therapeutic approach in Duchenne muscular dystrophy. PMID:26465071

  19. Beneficial cilostazol therapeutic effects in mdx dystrophic skeletal muscle.

    PubMed

    Hermes, Túlio de Almeida; Macedo, Aline Barbosa; Fogaça, Aline Reis; Moraes, Luis Henrique Rapucci; de Faria, Felipe Meira; Kido, Larissa Akemi; Cagnon, Valéria Helena Alves; Minatel, Elaine

    2016-02-01

    This study evaluated the possible protective effects of cilostazol against myonecrosis in dystrophic diaphragm muscle in vivo, focusing on oxidative stress, the inflammatory response and angiogenesis. Young mdx mice, the experimental animal for Duchenne muscular dystrophy, received cilostazol for 14 days. A second group of mdx mice and a control group of C57BL/10 mice received a saline solution. In the mdx mice, cilostazol treatment was associated with reduced loss of muscle strength (-34.4%), decreased myonecrosis, reduced creatine kinase levels (-63.3%) and muscle fibres stained for immunoglobulin G in dystrophic diaphragm muscle (-81.1%), and a reduced inflammatory response, with a decreased inflammatory area (-22%), macrophage infiltration (-44.9%) and nuclear factor-κB (-24%) and tumour necrosis factor-α (-48%) content in dystrophic diaphragm muscle. Furthermore, cilostazol decreased oxidative stress and attenuated reactive oxygen species production (-74%) and lipid peroxidation (-17%) in dystrophic diaphragm muscle, and promoted the up-regulation of angiogenesis, increasing the number of microvessels (15%). In conclusion, the present results show that cilostazol has beneficial effects in dystrophic muscle. More research into the potential of cilostazol as a novel therapeutic agent for the treatment of dystrophinopathies is required. PMID:26639107

  20. Traumatic injuries to the duodenum: a report of 98 patients.

    PubMed Central

    Corley, R D; Norcross, W J; Shoemaker, W C

    1975-01-01

    Data of 98 patients who had sustained traumatic injuries to the duodenum during a recent 7-year period is reviewed. The overall mortality was 23.5%; that of the blunt injury group was 35%, that of the penetrating injury group was 20%. However, after the establishment of a trauma unit, the mortality for duodenal injuries fell from 32% to 12%. Death from duodenal wounds may be reduced by earlier hospitalization, earlier diagnosis and consequently earlier surgical repair. Vigorous treatment of shock is essential. A specialized trauma unit with personnel experienced in the management of shock and trauma problems provides a better environment to carry out the preoperative and postoperative care of the acutely injured patient. Adequate surgical treatment of the blunt injury and missile injury of the duodenum should consist of the following procedures: 1) repair of the duodenal wall utilizing conventional techniques; 2) internal decompression of the repair by afferent jejunostomy; 3) efferent jejunostomy for postoperative feeding; 4) temporary gastrostomy; and 5) external drainage of the repair. In certain selected instances, the simple stab wound of the duodenum may be treated by conventional repair without decompression, but a loop of jujunum should be sutured over the repair to prevent delayed disruption. The majority of patients with injuries to the duodenum have associated organs injured which also require considered surgical judgment and action. PMID:1119875

  1. Efficient adenovirus-mediated transfer of a human minidystrophin gene to skeletal muscle of mdx mice.

    PubMed

    Ragot, T; Vincent, N; Chafey, P; Vigne, E; Gilgenkrantz, H; Couton, D; Cartaud, J; Briand, P; Kaplan, J C; Perricaudet, M

    1993-02-18

    Duchenne progressive muscular dystrophy is a lethal and common X-linked genetic disease caused by the absence of dystrophin, a 427K protein encoded by a 14 kilobase transcript. Two approaches have been proposed to correct the dystrophin deficiency in muscle. The first, myoblast transfer therapy, uses cells from normal donors, whereas the second involves direct intramuscular injection of recombinant plasmids expressing dystrophin. Adenovirus is an efficient vector for in vivo expression of various foreign genes. It has recently been demonstrated that a recombinant adenovirus expressing the lac-Z reporter gene can infect stably many mouse tissues, particularly muscle and heart. We have tested the ability of a recombinant adenovirus, containing a 6.3 kilobase pair Becker-like dystrophin complementary DNA driven by the Rous sarcoma virus promoter to direct the expression of a 'minidystrophin' in infected 293 cells and C2 myoblasts, and in the mdx mouse, after intramuscular injection. We report here that in vivo, we have obtained a sarcolemmal immunostaining in up to 50% of fibres of the injected muscle. PMID:8437625

  2. 100-fold but not 50-fold dystrophin overexpression aggravates electrocardiographic defects in the mdx model of Duchenne muscular dystrophy

    PubMed Central

    Yue, Yongping; Wasala, Nalinda B; Bostick, Brian; Duan, Dongsheng

    2016-01-01

    Dystrophin gene replacement holds the promise of treating Duchenne muscular dystrophy. Supraphysiological expression is a concern for all gene therapy studies. In the case of Duchenne muscular dystrophy, Chamberlain and colleagues found that 50-fold overexpression did not cause deleterious side effect in skeletal muscle. To determine whether excessive dystrophin expression in the heart is safe, we studied two lines of transgenic mdx mice that selectively expressed a therapeutic minidystrophin gene in the heart at 50-fold and 100-fold of the normal levels. In the line with 50-fold overexpression, minidystrophin showed sarcolemmal localization and electrocardiogram abnormalities were corrected. However, in the line with 100-fold overexpression, we not only detected sarcolemmal minidystrophin expression but also observed accumulation of minidystrophin vesicles in the sarcoplasm. Excessive minidystrophin expression did not correct tachycardia, a characteristic feature of Duchenne muscular dystrophy. Importantly, several electrocardiogram parameters (QT interval, QRS duration and the cardiomyopathy index) became worse than that of mdx mice. Our data suggests that the mouse heart can tolerate 50-fold minidystrophin overexpression, but 100-fold overexpression leads to cardiac toxicity. PMID:27419194

  3. Proteomic Profiling of the Dystrophin-Deficient MDX Heart Reveals Drastically Altered Levels of Key Metabolic and Contractile Proteins

    PubMed Central

    Lewis, Caroline; Jockusch, Harald; Ohlendieck, Kay

    2010-01-01

    Although Duchenne muscular dystrophy is primarily classified as a neuromuscular disease, cardiac complications play an important role in the course of this X-linked inherited disorder. The pathobiochemical steps causing a progressive decline in the dystrophic heart are not well understood. We therefore carried out a fluorescence difference in-gel electrophoretic analysis of 9-month-old dystrophin-deficient versus age-matched normal heart, using the established MDX mouse model of muscular dystrophy-related cardiomyopathy. Out of 2,509 detectable protein spots, 79 2D-spots showed a drastic differential expression pattern, with the concentration of 3 proteins being increased, including nucleoside diphosphate kinase and lamin-A/C, and of 26 protein species being decreased, including ATP synthase, fatty acid binding-protein, isocitrate dehydrogenase, NADH dehydrogenase, porin, peroxiredoxin, adenylate kinase, tropomyosin, actin, and myosin light chains. Hence, the lack of cardiac dystrophin appears to trigger a generally perturbed protein expression pattern in the MDX heart, affecting especially energy metabolism and contractile proteins. PMID:20508850

  4. 100-fold but not 50-fold dystrophin overexpression aggravates electrocardiographic defects in the mdx model of Duchenne muscular dystrophy.

    PubMed

    Yue, Yongping; Wasala, Nalinda B; Bostick, Brian; Duan, Dongsheng

    2016-01-01

    Dystrophin gene replacement holds the promise of treating Duchenne muscular dystrophy. Supraphysiological expression is a concern for all gene therapy studies. In the case of Duchenne muscular dystrophy, Chamberlain and colleagues found that 50-fold overexpression did not cause deleterious side effect in skeletal muscle. To determine whether excessive dystrophin expression in the heart is safe, we studied two lines of transgenic mdx mice that selectively expressed a therapeutic minidystrophin gene in the heart at 50-fold and 100-fold of the normal levels. In the line with 50-fold overexpression, minidystrophin showed sarcolemmal localization and electrocardiogram abnormalities were corrected. However, in the line with 100-fold overexpression, we not only detected sarcolemmal minidystrophin expression but also observed accumulation of minidystrophin vesicles in the sarcoplasm. Excessive minidystrophin expression did not correct tachycardia, a characteristic feature of Duchenne muscular dystrophy. Importantly, several electrocardiogram parameters (QT interval, QRS duration and the cardiomyopathy index) became worse than that of mdx mice. Our data suggests that the mouse heart can tolerate 50-fold minidystrophin overexpression, but 100-fold overexpression leads to cardiac toxicity. PMID:27419194

  5. Peptide conjugation of 2'-O-methyl phosphorothioate antisense oligonucleotides enhances cardiac uptake and exon skipping in mdx mice.

    PubMed

    Jirka, Silvana M G; Heemskerk, Hans; Tanganyika-de Winter, Christa L; Muilwijk, Daan; Pang, Kar Him; de Visser, Peter C; Janson, Anneke; Karnaoukh, Tatyana G; Vermue, Rick; 't Hoen, Peter A C; van Deutekom, Judith C T; Aguilera, Begoña; Aartsma-Rus, Annemieke

    2014-02-01

    Antisense oligonucleotide (AON)-mediated exon skipping is a promising therapeutic approach for Duchenne muscular dystrophy that is currently being tested in various clinical trials. This approach is based on restoring the open reading frame of dystrophin transcripts resulting in shorter but partially functional dystrophin proteins as found in patients with Becker muscular dystrophy. After systemic administration, a large proportion of AONs ends up in the liver and kidneys. Therefore, enhancing AON uptake by skeletal and cardiac muscle would improve the AONs' therapeutic effect. For phosphorodiamidate morpholino oligomer, AONs use nonspecific positively charged cell penetrating peptides to enhance efficacy. However, this is challenging for negatively charged 2'-O-methyl phosphorothioate oligomer. Therefore, we screened a 7-mer phage display peptide library to identify muscle and heart homing peptides in vivo in the mdx mouse model and found a promising candidate peptide capable of binding muscle cells in vitro and in vivo. Upon systemic administration in dystrophic mdx mice, conjugation of a 2'-O-methyl phosphorothioate AON to this peptide indeed improved uptake in skeletal and cardiac muscle, and resulted in higher exon skipping levels with a significant difference in heart and diaphragm. Based on these results, peptide conjugation represents an interesting strategy to enhance the therapeutic effect of exon skipping with 2'-O-methyl phosphorothioate AONs for Duchenne muscular dystrophy. PMID:24320790

  6. Pedunculated islet-cell tumour of the duodenum.

    PubMed

    Britt, R P

    1966-05-01

    An unusual islet-cell tumour found at necropsy in a patient who had died from a myocardial infarction is described. Of particular interest were the pedunculated nature and large size of the tumour. The clinical features of the case are considered. Four islet-cell tumours in the duodenum have previously been reported and it seems probable that such tumours arise in heterotopic pancreas. PMID:4287114

  7. Hepatoid Adenocarcinoma of the Duodenum: An Unusual Location

    PubMed Central

    Ogbonna, Onyekachi Henry; Sakruti, Susmita; Sulieman, Maha; Ali, Ahmed; Shokrani, Babak; Oneal, Patricia

    2016-01-01

    Hepatoid adenocarcinoma (HAC) is a rare extrahepatic tumor distinguished by having both hepatoid and adenomatous features, which can make the diagnosis challenging. Although it mostly originates in the stomach, several other sites of origin have been reported. We report a case of HAC originating in the duodenum, a very unusual location. We also discuss an approach to the diagnosis of HAC using morphological and immunohistochemical features, and explore possible therapeutic options. PMID:27064217

  8. Bubble liposomes and ultrasound exposure improve localized morpholino oligomer delivery into the skeletal muscles of dystrophic mdx mice.

    PubMed

    Negishi, Yoichi; Ishii, Yuko; Shiono, Hitomi; Akiyama, Saki; Sekine, Shoko; Kojima, Takuo; Mayama, Sayaka; Kikuchi, Taiki; Hamano, Nobuhito; Endo-Takahashi, Yoko; Suzuki, Ryo; Maruyama, Kazuo; Aramaki, Yukihiko

    2014-03-01

    Duchenne muscular dystrophy (DMD) is a genetic disorder that is caused by mutations in the DMD gene that lead to an absence of functional protein. The mdx dystrophic mouse contains a nonsense mutation in exon 23 of the dystrophin gene; a phosphorodiamidate morpholino oligomer (PMO) designed to skip this mutated exon in the mRNA induces dystrophin expression. However, an efficient PMO delivery method is needed to improve treatment strategies for DMD. We previously developed polyethylene glycol (PEG)-modified liposomes (Bubble liposomes) that entrap ultrasound contrast gas and demonstrated that the combination of Bubble liposomes with ultrasound exposure is an effective gene delivery tool in vitro and in vivo. In this study, to evaluate the ability of Bubble liposomes as a PMO delivery tool, we tested the potency of the Bubble liposomes combined with ultrasound exposure to boost the delivery of PMO and increase the skipping of the mutated exon in the mdx mouse. The results indicated that the combination of Bubble liposomes and ultrasound exposure increased the uptake of the PMO targeting a nonsense mutation in exon 23 of the dystrophin gene and consequently increased the PMO-mediated exon-skipping efficiency compared with PMO injection alone, leading to significantly enhanced dystrophin expression. This increased efficiency indicated the potential of the combination of Bubble liposomes with ultrasound exposure to enhance PMO delivery for treating DMD. Thus, this ultrasound-mediated Bubble liposome technique may provide an effective, noninvasive, nonviral method for PMO therapy for DMD muscle as well as for other muscular dystrophies. PMID:24433046

  9. Wild-Type Mouse Models to Screen Antisense Oligonucleotides for Exon-Skipping Efficacy in Duchenne Muscular Dystrophy

    PubMed Central

    Cao, Limin; Han, Gang; Gu, Ben; Yin, HaiFang

    2014-01-01

    A readily available animal model is essential for rapidly identifying effective treatments for Duchenne muscular dystrophy (DMD), a devastating neuromuscular disorder caused by the lack of dystrophin protein, which results from frame-disrupting mutations in the DMD gene. Currently, the mdx mouse is the most commonly used model for antisense oligonucleotide (AO)-mediated exon skipping pre-clinical studies, with a mild phenotype. However, the accessibility of mdx mouse colonies particularly in developing countries can constrain research. Therefore in this study we explore the feasibility of using wild-type mice as models to establish exon-skipping efficiency of various DMD AO chemistries and their conjugates. Four different strains of wild-type mice and six different AO chemistries were investigated intramuscularly and the results indicated that the same exon-skipping efficiency was achieved for all tested AOs as that from mdx mice. Notably, levels of exon-skipping obtained in C57BL6 and C3H and mdx mice were most closely matched, followed by ICR and BALB/C mice. Systemic validation revealed that wild-type mice are less responsive to AO-mediated exon skipping than mdx mice. Our study provides evidence for the first time that wild-type mice can be appropriate models for assessing DMD AO exon-skipping efficiency with similar sensitivity to that of mdx mice and this finding can further accelerate the development of effective DMD AOs. PMID:25365558

  10. Interleukin-15 Administration Improves Diaphragm Muscle Pathology and Function in Dystrophic mdx Mice

    PubMed Central

    Harcourt, Leah J.; Holmes, Anna Greer; Gregorevic, Paul; Schertzer, Jonathan D.; Stupka, Nicole; Plant, David R.; Lynch, Gordon S.

    2005-01-01

    Interleukin (IL)-15, a cytokine expressed in skeletal muscle, has been shown to have muscle anabolic effects in vitro and to slow muscle wasting in rats with cancer cachexia. Whether IL-15 has therapeutic potential for diseases such as Duchenne muscular dystrophy (DMD) is unknown. We examined whether IL-15 administration could ameliorate the dystrophic pathology in the diaphragm muscle of the mdx mouse, an animal model for DMD. Four weeks of IL-15 treatment improved diaphragm strength, a highly significant finding because respiratory function is a mortality predictor in DMD. Enhanced diaphragm function was associated with increased muscle fiber cross-sectional area and decreased collagen infiltration. IL-15 administration was not associated with changes in T-cell populations or alterations in specific components of the ubiquitin proteasome pathway. To determine the effects of IL-15 on myofiber regeneration, muscles of IL-15-treated and untreated wild-type mice were injured myotoxically, and their functional recovery was assessed. IL-15 had a mild anabolic effect, increasing fiber cross-sectional area after 2 and 6 days but not after 10 days. Our findings demonstrate that IL-15 administration improves the pathophysiology of dystrophic muscle and highlight a possible therapeutic role for IL-15 in the treatment of neuromuscular disorders especially in which muscle wasting is indicated. PMID:15793293

  11. Sustained Dystrophin Expression Induced by Peptide-conjugated Morpholino Oligomers in the Muscles of mdx Mice

    PubMed Central

    Jearawiriyapaisarn, Natee; Moulton, Hong M; Buckley, Brian; Roberts, Jennifer; Sazani, Peter; Fucharoen, Suthat; Iversen, Patrick L; Kole, Ryszard

    2009-01-01

    Cell-penetrating peptides (CPPs), containing arginine (R), 6-aminohexanoic acid (X), and/or β-alanine (B) conjugated to phosphorodiamidate morpholino oligomers (PMOs), enhance their delivery in cell culture. In this study, the potency, functional biodistribution, and toxicity of these conjugates were evaluated in vivo, in EGFP-654 transgenic mice that ubiquitously express the aberrantly spliced EGFP-654 pre-mRNA reporter. Correct splicing and enhanced green fluorescence protein (EGFP) upregulation serve as a positive readout for peptide-PMO (PPMO) entry into cells and access to EGFP-654 pre-mRNA in the nucleus. Intraperitoneal injections of a series of PPMOs, A-N (12 mg/kg), administered once a day for four successive days resulted in splicing correction in numerous tissues. PPMO-B was highly potent in the heart, diaphragm, and quadriceps, which are key muscles in the treatment of Duchenne muscular dystrophy. We therefore investigated PPMO M23D-B, designed to force skipping of stop-codon containing dystrophin exon 23, in an mdx mouse model of the disease. Systemic delivery of M23D-B yielded persistent exon 23 skipping, yielding high and sustained dystrophin protein expression in body-wide muscles, including cardiac muscle, without detectable toxicity. The rescued dystrophin reduced serum creatinine kinase to near-wild-type levels, indicating improvement in muscle integrity. This is the first report of oligonucleotide-mediated exon skipping and dystrophin protein induction in the heart of treated animals. PMID:18545222

  12. Age-related alterations in cyclic nucleotide phosphodiesterase activity in dystrophic mouse leg muscle.

    PubMed

    Bloom, Timothy J

    2005-11-01

    Previous reports have described both increased and decreased cyclic nucleotide phosphodiesterase (PDE) activity in dystrophic muscle. Total PDE activity was measured in hind leg muscle from a mouse model of Duchenne muscular dystrophy (mdx) and a genetic control strain at 5, 8, 10, and 15 weeks of age. Total PDE activity declined in fractions isolated from mdx muscle over this time period, but was stable in fractions from control mice. Compared with age-matched controls, younger mdx muscle had higher cAMP and cGMP PDE activity. However, at 15 weeks, fractions from both strains had similar cGMP PDE activity and mdx fractions had lower cAMP PDE activity than controls. Particulate fractions from mdx muscle showed an age-related decline in sensitivity to the PDE4 inhibitor RO 20-1724. A similar loss of sensitivity to the PDE2 inhibitor erythro-9-(2-hydroxyl-3-nonyl)-adenine (EHNA) was seen in a particulate fraction from mdx muscle and to a lesser degree in control muscle. These results suggest that the earlier disagreement regarding altered cyclic nucleotide metabolism in dystrophic muscle may be due to changes with age in PDE activity of dystrophic tissue. The age-related decline in particulate PDE activity seen in dystrophic muscle appears to be isozyme-specific and not due to a generalized decrease in total PDE activity. PMID:16391714

  13. Erythropoietin reduces the expression of myostatin in mdx dystrophic mice.

    PubMed

    Feder, D; Rugollini, M; Santomauro, A; Oliveira, L P; Lioi, V P; Santos, R dos; Ferreira, L G; Nunes, M T; Carvalho, M H; Delgado, P O; Carvalho, A A S; Fonseca, F L A

    2014-11-01

    Erythropoietin (EPO) has been well characterized as a renal glycoprotein hormone regulating red blood cell production by inhibiting apoptosis of erythrocyte progenitors in hematopoietic tissues. EPO exerts regulatory effects in cardiac and skeletal muscles. Duchenne muscular dystrophy is a lethal degenerative disorder of skeletal and cardiac muscle. In this study, we tested the possible therapeutic beneficial effect of recombinant EPO (rhEPO) in dystrophic muscles in mdx mice. Total strength was measured using a force transducer coupled to a computer. Gene expression for myostatin, transforming growth factor-β1 (TGF-β1), and tumor necrosis factor-α (TNF-α) was determined by quantitative real time polymerase chain reaction. Myostatin expression was significantly decreased in quadriceps from mdx mice treated with rhEPO (rhEPO = 0.60 ± 0.11, control = 1.07 ± 0.11). On the other hand, rhEPO had no significant effect on the expression of TGF-β1 (rhEPO = 0.95 ± 0.14, control = 1.05 ± 0.16) and TNF-α (rhEPO = 0.73 ± 0.20, control = 1.01 ± 0.09). These results may help to clarify some of the direct actions of EPO on skeletal muscle. PMID:25296358

  14. Effect of the IL-1 Receptor Antagonist Kineret® on Disease Phenotype in mdx Mice

    PubMed Central

    Benny Klimek, Margaret E.; Sali, Arpana; Rayavarapu, Sree; Van der Meulen, Jack H.; Nagaraju, Kanneboyina

    2016-01-01

    Duchenne muscular dystrophy (DMD) is an X-linked muscle disease caused by mutations in the dystrophin gene. The pathology of DMD manifests in patients with progressive muscle weakness, loss of ambulation and ultimately death. One of the characteristics of DMD is muscle inflammation, and dystrophin-deficient skeletal muscles produce higher levels of the pro-inflammatory cytokine interleukin 1β (IL-1β) in response to toll like receptor (TLR) stimulation compared to controls; therefore, blocking the IL-1β pathway could improve the disease phenotype in mdx mice, a mouse model of DMD. Kineret® or IL-1Ra is a recombinant IL-1 receptor antagonist approved by the FDA for treating rheumatoid arthritis. To determine the efficacy of IL-1Ra in a DMD model, we administered subcutaneous injections of saline control or IL-1Ra (25 mg/kg/day) to mdx mice daily for 45 days beginning at 5 weeks of age. Functional and histological parameters were measured at the conclusion of the study. IL-1Ra only partially inhibited this signaling pathway in this study; however, there were still interesting observations to be noted. For example, although not significantly changed, splenocytes from the IL-1Ra-treated group secreted less IL-1β after LPS stimulation compared to control mice indicating a blunted response and incomplete inhibition of the pathway (37% decrease). In addition, normalized forelimb grip strength was significantly increased in IL-1Ra-treated mice. There were no changes in EDL muscle-specific force measurements, histological parameters, or motor coordination assessments in the dystrophic mice after IL-1Ra treatment. There was a significant 27% decrease in the movement time and total distance traveled by the IL-1Ra treated mice, correlating with previous studies examining effects of IL-1 on behavior. Our studies indicate partial blocking of IL-1β with IL-1Ra significantly altered only a few behavioral and strength related disease parameters; however, treatment with

  15. Effect of the IL-1 Receptor Antagonist Kineret® on Disease Phenotype in mdx Mice.

    PubMed

    Benny Klimek, Margaret E; Sali, Arpana; Rayavarapu, Sree; Van der Meulen, Jack H; Nagaraju, Kanneboyina

    2016-01-01

    Duchenne muscular dystrophy (DMD) is an X-linked muscle disease caused by mutations in the dystrophin gene. The pathology of DMD manifests in patients with progressive muscle weakness, loss of ambulation and ultimately death. One of the characteristics of DMD is muscle inflammation, and dystrophin-deficient skeletal muscles produce higher levels of the pro-inflammatory cytokine interleukin 1β (IL-1β) in response to toll like receptor (TLR) stimulation compared to controls; therefore, blocking the IL-1β pathway could improve the disease phenotype in mdx mice, a mouse model of DMD. Kineret® or IL-1Ra is a recombinant IL-1 receptor antagonist approved by the FDA for treating rheumatoid arthritis. To determine the efficacy of IL-1Ra in a DMD model, we administered subcutaneous injections of saline control or IL-1Ra (25 mg/kg/day) to mdx mice daily for 45 days beginning at 5 weeks of age. Functional and histological parameters were measured at the conclusion of the study. IL-1Ra only partially inhibited this signaling pathway in this study; however, there were still interesting observations to be noted. For example, although not significantly changed, splenocytes from the IL-1Ra-treated group secreted less IL-1β after LPS stimulation compared to control mice indicating a blunted response and incomplete inhibition of the pathway (37% decrease). In addition, normalized forelimb grip strength was significantly increased in IL-1Ra-treated mice. There were no changes in EDL muscle-specific force measurements, histological parameters, or motor coordination assessments in the dystrophic mice after IL-1Ra treatment. There was a significant 27% decrease in the movement time and total distance traveled by the IL-1Ra treated mice, correlating with previous studies examining effects of IL-1 on behavior. Our studies indicate partial blocking of IL-1β with IL-1Ra significantly altered only a few behavioral and strength related disease parameters; however, treatment with

  16. Evaluation of Tris[2-(Acryloyloxy)Ethyl]Isocyanurate Cross-Linked Polyethylenimine as Antisense Morpholino Oligomer Delivery Vehicle in Cell Culture and Dystrophic mdx Mice

    PubMed Central

    Wu, Bo; Tucker, Jay D.; Lu, Peijuan; Cloer, Caryn

    2014-01-01

    Abstract Hyperbranched poly(ester amine)s (PEAs) based on tris[2-(acryloyloxy)ethyl]isocyanurate (TAEI) cross-linked low-molecular-weight polyethylenimine (Mw: 0.8k/1.2k/2.0k) have been evaluated for delivering antisense phosphorodiamidate morpholino oligomer (PMO) in vitro and in vivo in the dystrophic mdx mouse. The results show that the PEAs constructed with polyethylenimine (PEI) 2.0k (C series) improved PMO delivery more efficiently than those constructed with PEI 0.8k (A series) or 1.2k (B series) in a GFP reporter-based C2C12 mouse myoblast culture system. The highest efficiency of exon-skipping in vitro with the PMO oligonucleotide targeting human dystrophin exon 50 was obtained when the PEA C12 [TAEI-PEI 2.0k (1:2)] was used. Nearly all of the PEAs improved dystrophin expression in mdx mice by local injection with a 2–4-fold increase when compared with PMO alone. Improved transfection efficiency and lower toxicity indicate the potential of the biodegradable PEA polymers as safe and efficient PMO delivery vectors for in vivo applications. PMID:24405395

  17. Evaluation of Tris[2-(acryloyloxy)ethyl]isocyanurate cross-linked polyethylenimine as antisense morpholino oligomer delivery vehicle in cell culture and dystrophic mdx mice.

    PubMed

    Wang, Mingxing; Wu, Bo; Tucker, Jay D; Lu, Peijuan; Cloer, Caryn; Lu, Qi Long

    2014-05-01

    Hyperbranched poly(ester amine)s (PEAs) based on tris[2-(acryloyloxy)ethyl]isocyanurate (TAEI) cross-linked low-molecular-weight polyethylenimine (Mw: 0.8k/1.2k/2.0k) have been evaluated for delivering antisense phosphorodiamidate morpholino oligomer (PMO) in vitro and in vivo in the dystrophic mdx mouse. The results show that the PEAs constructed with polyethylenimine (PEI) 2.0k (C series) improved PMO delivery more efficiently than those constructed with PEI 0.8k (A series) or 1.2k (B series) in a GFP reporter-based C2C12 mouse myoblast culture system. The highest efficiency of exon-skipping in vitro with the PMO oligonucleotide targeting human dystrophin exon 50 was obtained when the PEA C12 [TAEI-PEI 2.0k (1:2)] was used. Nearly all of the PEAs improved dystrophin expression in mdx mice by local injection with a 2-4-fold increase when compared with PMO alone. Improved transfection efficiency and lower toxicity indicate the potential of the biodegradable PEA polymers as safe and efficient PMO delivery vectors for in vivo applications. PMID:24405395

  18. Major alteration of the pathological phenotype in gamma irradiated mdx soleus muscles

    SciTech Connect

    Weller, B.; Karpati, G.; Lehnert, S.; Carpenter, S. )

    1991-07-01

    Two thousand rads of gamma irradiation delivered to the lower legs of ten day old normal and x-chromosome linked muscular dystrophy (mdx) mice caused significant inhibition of tibial bone and soleus muscle fiber growth. In the irradiated mdx solei, there was a major loss of muscle fibers, lack of central nucleation, and some endomysial fibrosis. These features were caused by a failure of regeneration of muscle fibers due to impaired proliferative capacity of satellite cells. Gamma irradiation transforms the late pathological phenotype of mdx muscles, so that in one major aspect (muscle fiber loss) they resemble muscles in Duchenne muscular dystrophy. However, extensive endomysial fibrosis which is another characteristic feature of Duchenne muscular dystrophy did not develop. This experimental model could be useful for the functional investigation of possible beneficial effects of therapeutic interventions in mdx dystrophy.

  19. Gastric emptying, small intestinal transit and fecal output in dystrophic (mdx) mice.

    PubMed

    Mulè, Flavia; Amato, Antonella; Serio, Rosa

    2010-01-01

    Duchenne muscular dystrophy (DMD), which results from deficiency in dystrophin, a sarcolemma protein of skeletal, cardiac and smooth muscle, is characterized by progressive striated muscle degeneration, but various gastrointestinal clinical manifestations have been observed. The aim was to evaluate the possible impact of the dystrophin loss on the gastrointestinal propulsion in mdx mice (animal model for DMD). The gastric emptying of a carboxymethyl cellulose/phenol red dye non-nutrient meal was not significantly different at 20 min from gavaging between wild-type and mdx mice. The intestinal transit and the fecal output were significantly decreased in mdx versus normal animals, although the length of the intestine was similar in both animals. The present results provide evidence for motor intestinal alterations in mdx mice in in vivo conditions. PMID:19784719

  20. Low Six4 and Six5 gene dosage improves dystrophic phenotype and prolongs life span of mdx mice.

    PubMed

    Yajima, Hiroshi; Kawakami, Kiyoshi

    2016-08-01

    Muscle regeneration is an important process for skeletal muscle growth and recovery. Repair of muscle damage is exquisitely programmed by cellular mechanisms inherent in myogenic stem cells, also known as muscle satellite cells. We demonstrated previously the involvement of homeobox transcription factors, SIX1, SIX4 and SIX5, in the coordinated proliferation and differentiation of isolated satellite cells in vitro. However, their roles in adult muscle regeneration in vivo remain elusive. To investigate SIX4 and SIX5 functions during muscle regeneration, we introduced knockout alleles of Six4 and Six5 into an animal model of Duchenne Muscular Dystrophy (DMD), mdx (Dmd(mdx) /Y) mice, characterized by frequent degeneration-regeneration cycles in muscles. A lower number of small myofibers, higher number of thick ones and lower serum creatine kinase and lactate dehydrogenase activities were noted in 50-week-old Six4(+/-) 5(+/-) Dmd(mdx) /Y mice than Dmd(mdx) /Y mice, indicating improvement of dystrophic phenotypes of Dmd(mdx) /Y mice. Higher proportions of cells positive for MYOD1 and MYOG (markers of regenerating myonuclei) and SIX1 (a marker of regenerating myoblasts and newly regenerated myofibers) in 12-week-old Six4(+/-) 5(+/-) Dmd(mdx) /Y mice suggested enhanced regeneration, compared with Dmd(mdx) /Y mice. Although grip strength was comparable in Six4(+/-) 5(+/-) Dmd(mdx) /Y and Dmd(mdx) /Y mice, treadmill exercise did not induce muscle weakness in Six4(+/-) 5(+/-) Dmd(mdx) /Y mice, suggesting higher regeneration capacity. In addition, Six4(+/-) 5(+/-) Dmd(mdx) /Y mice showed 33.8% extension of life span. The results indicated that low Six4 and Six5 gene dosage improved dystrophic phenotypes of Dmd(mdx) /Y mice by enhancing muscle regeneration, and suggested that SIX4 and SIX5 are potentially useful de novo targets in therapeutic applications against muscle disorders, including DMD. PMID:27224259

  1. Arginine butyrate per os protects mdx mice against cardiomyopathy, kyphosis and changes in axonal excitability.

    PubMed

    Vianello, Sara; Bouyon, Sophie; Benoit, Evelyne; Sebrié, Catherine; Boerio, Delphine; Herbin, Marc; Roulot, Morgane; Fromes, Yves; de la Porte, Sabine

    2014-11-01

    Duchenne muscular dystrophy (DMD) is a progressive neuromuscular disease caused by lack of dystrophin, a sub-sarcolemmal protein, which leads to dramatic muscle deterioration. We studied in mdx mice, the effects of oral administration of arginine butyrate (AB), a compound currently used for the treatment of sickle cell anemia in children, on cardiomyopathy, vertebral column deformation and electromyographic abnormalities. Monthly follow-up by echocardiography from the 8th month to the 14th month showed that AB treatment protected the mdx mice against drastic reduction (20-23%) of ejection fraction and fractional shortening, and also against the ≈20% ventricular dilatation and 25% cardiac hypertrophy observed in saline-treated mdx mice. The phenotypic improvement was corroborated by the decrease in serum CK level and by better fatigue resistance. Moreover, AB treatment protected against the progressive spinal deformity observed in mdx mice, another similarity with DMD patients. The value of the kyphosis index in AB-treated mice reached 94% of the value in C57BL/10 mice. Finally, axonal excitability parameters such as the membrane resting potential, the threshold and amplitude of the action potential, the absolute and relative refractory periods and the supernormal and subnormal periods, recorded from caudal and plantar muscles in response to excitability tests, that were modified in saline-treated mdx mice were not significantly changed, compared with wild-type animals, in AB-treated mdx mice. All of these results suggest that AB could be a potential treatment for DMD patients. PMID:25167832

  2. Successful endoscopic submucosal dissection for mucosal cancer of the duodenum.

    PubMed

    Shinoda, Masahiro; Makino, Atsushi; Wada, Masahiro; Kabeshima, Yasuo; Takahashi, Tsunehiro; Kawakubo, Hirofumi; Shito, Masaya; Sugiura, Hitoshi; Omori, Tai

    2010-01-01

    We report a case of mucosal duodenal cancer in a 62-year-old woman, which was successfully removed en bloc by endoscopic submucosal dissection (ESD). The patient underwent an upper gastrointestinal endoscopy at our hospital, which revealed an elevated flat mucosal lesion (type IIa) measuring 10 mm in diameter in the second portion of the duodenum. Histopathological examination of a biopsy specimen revealed features suggestive of a tubulovillous adenoma with severe atypia. As the findings suggested that the lesion had an adenocarcinoma component but was confined to the mucosal layer, we decided to carry out ESD and successfully removed the tumor in one piece. The resected tumor was 20 x 15 mm in size. Histopathological examination revealed that the lesion was a well-differentiated mucosal adenocarcinoma with no lymphovascular invasion. Mucosal duodenal cancer is extremely rare, and ESD of a lesion in the duodenum requires a high level of skill. To the best of our knowledge, this case is the first report of successful ESD carried out in a case of mucosal duodenal cancer. PMID:20078665

  3. Metastatic Prostate Cancer to the Duodenum: A Rare Case

    PubMed Central

    Kaswala, Dharmesh H.; Patel, Nitin; Jadallah, Sana; Wang, Weizheng

    2014-01-01

    Prostate cancer is the third most common cancer in man. About 1 in 6 males developed prostate cancer and 1 in 35 males die of this disease. Prostate cancer behavior ranges from microscopic tumors to aggressive cancer with metastatic potential. While metastasis to bone is relatively common, prostate cancer rarely metastasizes to the cecum, pituitary gland, small bowel, maxillary sinus and skin. Our case report presents a rare presentation of metastatic prostate cancer to the duodenum. Our search of the literature found only 2 cases of prostate metastases to duodenum published from 1966 to the present. To our knowledge this is the third case of metastatic prostate cancer presenting with duodenal metastasis. Although it is rare but in symptomatic patients small intestine metastasis should not be ignored with advanced prostate cancer. The case demonstrates a novel presentation of a common malignancy, and should raise awareness in clinicians and radiologists that prostate cancer can present with distant metastases in absence of any local lymphadenopathy. PMID:25161979

  4. Comparative study of myocytes from normal and mdx mice iPS cells.

    PubMed

    Chen, Fei; Cao, Jiqing; Liu, Qiang; Qin, Jie; Kong, Jie; Wang, Yanyun; Li, Yaqin; Geng, Jia; Li, Qiuling; Yang, Liqing; Xiang, Andy Peng; Zhang, Cheng

    2012-02-01

    Recently, induced pluripotent stem cells (iPS cells) have been derived from various techniques and show great potential for therapy of human diseases. Furthermore, the iPS technique can be used to provide cell models to explore pathological mechanisms of many human diseases in vitro, such as Duchenne muscular dystrophy (DMD), which is a severe recessive X-linked form of muscular dystrophy without effective treatment. In this study, we try to determine whether there are different characteristics of myocytes from mdx iPS cells and C57BL/10 iPS cells. Our results showed that both of mdx and C57BL/10 cells could be induced into iPS cells in vitro, whereas colony-forming ability of mdx iPS cells was much weaker than that of C57BL/10 iPS cells. Meanwhile, mdx iPS cells could be induced to differentiate into myocytes, whereas their differentiation efficiency was much lower than that of C57BL/10 iPS cells. And, the number of apoptotic cells in differentiated myocytes from mdx iPS cells was significantly higher than that from C57BL/10 iPS cells. More importantly, treatment of a pan-caspase inhibitor (Z-VAD) produced a significant decrease in apoptotic cells. This study might add some insight to the biology study of dystrophin gene. PMID:21976068

  5. Malformed mdx myofibers have normal cytoskeletal architecture yet altered EC coupling and stress-induced Ca2+ signaling

    PubMed Central

    Ward, Christopher W.

    2009-01-01

    Skeletal muscle function is dependent on its highly regular structure. In studies of dystrophic (dy/dy) mice, the proportion of malformed myofibers decreases after prolonged whole muscle stimulation, suggesting that the malformed myofibers are more prone to injury. The aim of this study was to assess morphology and to measure excitation-contraction (EC) coupling (Ca2+ transients) and susceptibility to osmotic stress (Ca2+ sparks) of enzymatically isolated muscle fibers of the extensor digitorum longus (EDL) and flexor digitorum brevis (FDB) muscles from young (2–3 mo) and old (8–9 mo) mdx and age-matched control mice (C57BL10). In young mdx EDL, 6% of the myofibers had visible malformations (i.e., interfiber splitting, branched ends, midfiber appendages). In contrast, 65% of myofibers in old mdx EDL contained visible malformations. In the mdx FDB, malformation occurred in only 5% of young myofibers and 11% of old myofibers. Age-matched control mice did not display the altered morphology of mdx muscles. The membrane-associated and cytoplasmic cytoskeletal structures appeared normal in the malformed mdx myofibers. In mdx FDBs with significantly branched ends, an assessment of global, electrically evoked Ca2+ signals (indo-1PE-AM) revealed an EC coupling deficit in myofibers with significant branching. Interestingly, peak amplitude of electrically evoked Ca2+ release in the branch of the bifurcated mdx myofiber was significantly decreased compared with the trunk of the same myofiber. No alteration in the basal myoplasmic Ca2+ concentration (i.e., indo ratio) was seen in malformed vs. normal mdx myofibers. Finally, osmotic stress induced the occurrence of Ca2+ sparks to a greater extent in the malformed portions of myofibers, which is consistent with deficits in EC coupling control. In summary, our data show that aging mdx myofibers develop morphological malformations. These malformations are not associated with gross disruptions in cytoskeletal or t

  6. Restoration of dystrophin-associated proteins in skeletal muscle of mdx mice transgenic for dystrophin gene.

    PubMed

    Matsumura, K; Lee, C C; Caskey, C T; Campbell, K P

    1993-04-12

    Duchenne muscular dystrophy (DMD) patients and mdx mice are characterized by the absence of dystrophin, a membrane cytoskeletal protein. Dystrophin is associated with a large oligomeric complex of sarcolemmal glycoproteins, including dystroglycan which provides a linkage to the extracellular matrix component, laminin. The finding that all of the dystrophin-associated proteins (DAPs) are drastically reduced in DMD and mdx skeletal muscle supports the primary function of dystrophin as an anchor of the sarcolemmal glycoprotein complex to the subsarcolemmal cytoskeleton. These findings indicate that the efficacy of dystrophin gene therapy will depend not only on replacing dystrophin but also on restoring all of the DAPs in the sarcolemma. Here we have investigated the status of the DAPs in the skeletal muscle of mdx mice transgenic for the dystrophin gene. Our results demonstrate that transfer of dystrophin gene restores all of the DAPs together with dystrophin, suggesting that dystrophin gene therapy should be effective in restoring the entire dystrophin-glycoprotein complex. PMID:8462697

  7. Aquapuncture Using Stem Cell Therapy to Treat Mdx Mice

    PubMed Central

    Esper, Greyson Vitor Zanatta; Pignatari, Graciela Conceição; Rodrigues, Marcio Nogueira; Bertagnon, Heloisa Godoi; Fernandes, Isabella Rodrigues; Nascimento, Nanci; Tabosa, Angela Maria Florencio; Beltrão-Braga, Patrícia Cristina Baleeiro; Miglino, Maria Angelica

    2015-01-01

    Duchenne muscular dystrophy (DMD) occurs due to genetic mutations that lead to absence or decrease of dystrophin protein generating progressive muscle degeneration. Cell therapy using mesenchymal stem cell (MSC) has been described as a treatment to DMD. In this work, MSC derived from deciduous teeth, called stem cells from human exfoliated deciduous teeth (SHED), were injected in acupoint as an alternative therapy to minimize muscle degeneration in twenty-two mdx mice. The treatment occurred three times with intervals of 21 days, and animals were analyzed four times: seven days prior treatment (T-7); 10 days after first treatment (T10); 10 days after second treatment (T31); and 10 days after third treatment (T52). Animals were evaluated by wire test for estimate strength and blood was collected to perform a creatinine phosphokinase analysis. After euthanasia, cranial tibial muscles were collected and submitted to histological and immunohistochemistry analyses. Treated groups presented improvement of strength and reduced creatinine phosphokinase levels. Also, a slight dystrophin increase was observed in tibial cranial muscle when aquapuncture was associated SHED. All therapies have minimized muscle degeneration, but the association of aquapuncture with SHED appears to have better effect, reducing muscle damage, suggesting a therapeutic value. PMID:26074983

  8. Wnt signaling pathway improves central inhibitory synaptic transmission in a mouse model of Duchenne muscular dystrophy.

    PubMed

    Fuenzalida, Marco; Espinoza, Claudia; Pérez, Miguel Ángel; Tapia-Rojas, Cheril; Cuitino, Loreto; Brandan, Enrique; Inestrosa, Nibaldo C

    2016-02-01

    The dystrophin-associated glycoprotein complex (DGC) that connects the cytoskeleton, plasma membrane and the extracellular matrix has been related to the maintenance and stabilization of channels and synaptic receptors, which are both essential for synaptogenesis and synaptic transmission. The dystrophin-deficient (mdx) mouse model of Duchenne muscular dystrophy (DMD) exhibits a significant reduction in hippocampal GABA efficacy, which may underlie the altered synaptic function and abnormal hippocampal long-term plasticity exhibited by mdx mice. Emerging studies have implicated Wnt signaling in the modulation of synaptic efficacy, neuronal plasticity and cognitive function. We report here that the activation of the non-canonical Wnt-5a pathway and Andrographolide, improves hippocampal mdx GABAergic efficacy by increasing the number of inhibitory synapses and GABA(A) receptors or GABA release. These results indicate that Wnt signaling modulates GABA synaptic efficacy and could be a promising novel target for DMD cognitive therapy. PMID:26626079

  9. Contractile properties of skinned muscle fibres from young and adult normal and dystrophic (mdx) mice.

    PubMed Central

    Williams, D A; Head, S I; Lynch, G S; Stephenson, D G

    1993-01-01

    1. Single muscle fibres were enzymatically isolated from the soleus and extensor digitorum longus (EDL) muscles of genetically dystrophic mdx and normal (C57BL/10) mice aged 3-6 or 17-23 weeks. 2. Fibres of both muscles were chemically skinned with the non-ionic detergent Triton X-100 (2% v/v). Ca(2+)- and Sr(2+)-activated contractile responses were recorded and comparisons were made between several contractile parameters of various fibre types of normal and dystrophic mice of similar age. 3. There were no significant differences in the following contractile parameters of skinned fibres of normal and mdx mice of the same age: sensitivity to activating Ca2+ (pCa50) or Sr2+ (pSr50) and differential sensitivity to the activating ions (pCa50-pSr50). However the maximum isometric tension (Po) and the frequency of myofibrillar force oscillations in EDL fast-twitch fibres of young mdx mice were significantly lower than those of soleus fast-twitch fibres of the same animals, or fast-twitch fibres (EDL or soleus) of normal mice. 4. Age-related differences were apparent in some contractile parameters of both normal and mdx mice. In particular the steepness of force-pCa and force-pSr curves increased with age in normal mice, yet decreased with age in fibres of mdx mice. 5. A fluorescent probe, ethidium bromide, which interchelates with DNA, was used with laser-scanning confocal microscopy to determine the distribution of myonuclei in fibres. Fibres isolated from either muscle type of normal animals displayed a characteristic peripheral spiral of myonuclei. Fibres from muscles of mdx mice displayed three major patterns of nuclear distribution; the normal peripheral spiral, long central strands of nuclei, and a mixture of these two patterns. 6. The contractile characteristics of mdx fibres were not markedly influenced by the nuclear distribution pattern in that there were no discernible differences in the major contractile parameters (the Hill coefficients nCa and nSr, which

  10. What do mouse models of muscular dystrophy tell us about the DAPC and its components?

    PubMed Central

    Whitmore, Charlotte; Morgan, Jennifer

    2014-01-01

    There are over 30 mouse models with mutations or inactivations in the dystrophin-associated protein complex. This complex is thought to play a crucial role in the functioning of muscle, as both a shock absorber and signalling centre, although its role in the pathogenesis of muscular dystrophy is not fully understood. The first mouse model of muscular dystrophy to be identified with a mutation in a component of the dystrophin-associated complex (dystrophin) was the mdx mouse in 1984. Here, we evaluate the key characteristics of the mdx in comparison with other mouse mutants with inactivations in DAPC components, along with key modifiers of the disease phenotype. By discussing the differences between the individual phenotypes, we show that the functioning of the DAPC and consequently its role in the pathogenesis is more complicated than perhaps currently appreciated. PMID:25270874

  11. Emergency pancreatoduodenectomy for complex injuries of the pancreas and duodenum

    PubMed Central

    Krige, Jake E; Nicol, Andrew J; Navsaria, Pradeep H

    2014-01-01

    Background This single-centre study evaluated the outcome of a pancreatoduodenectomy for Grade 5 injuries of the pancreas and duodenum. Methods Prospectively recorded data of patients who underwent a pancreatoduodenectomy for trauma at a Level I Trauma Centre during a 22-year period were analysed. Results Nineteen (17 men and 2 women, median age 28 years, range 14–53 years) out of 426 patients with pancreatic injuries underwent a pancreatoduodenectomy (gunshot n = 12, blunt trauma n = 6 and stab wound n = 1). Nine patients had associated inferior vena cava (IVC) or portal vein (PV) injuries. Five patients had initial damage control procedures and underwent a definitive operation at a median of 15 h (range 11–92) later. Twelve had a pylorus-preserving pancreatoduodenectomy (PPPD) and 7 a standard Whipple. Three patients with APACHE II scores of 15, 18, 18 died post-operatively of multi-organ failure. All 16 survivors had Dindo-Clavien grade I (n = 1), grade II (n = 7), grade IIIa (n = 2), grade IVa (n = 6) post-operative complications. Factors complicating surgery were shock on admission, number of associated injuries, coagulopathy, hypothermia, gross bowel oedema and traumatic pancreatitis. Conclusions A pancreatoduodenectomy is a life-saving procedure in a small cohort of stable patients with non-reconstructable pancreatic head injuries. Damage control before a pancreatoduodenectomy will salvage a proportion of the most severely injured patients who have multiple injuries. PMID:24841125

  12. Acquired constricting and restricting lesions of the descending duodenum.

    PubMed

    Carbo, Alberto I; Sangster, Guillermo P; Caraway, Jessica; Heldmann, Maureen G; Thomas, Jaiyeola; Takalkar, Amol

    2014-01-01

    The descending duodenum is a structure with distinct pathologic processes and anatomic relationships that requires a systematic approach to the differential diagnosis. Because of its tubular shape and fixed position in the retroperitoneum, both intrinsic duodenal and juxtaduodenal diseases are capable of producing luminal narrowing and obstruction. Duodenal lesions may be located in the mucosa or submucosa. Extraduodenal lesions may originate in adjacent structures--such as the pancreas, liver, gallbladder, colon, and lymph nodes--or from other retroperitoneal structures. Causes of duodenal obstruction include intraluminal masses, such as bezoars; duodenal inflammation, such as as peptic ulcers and Crohn disease; hematomas; and benign or malignant mucosal and intramural tumors. Pancreatic inflammation; tumors; and extrinsic compression caused by gallbladder processes, hepatic masses, retroperitoneal fluid collections, and tumors, including lymphoma, may produce duodenal obstruction. Abdominal radiography, barium studies, multidetector computed tomography, magnetic resonance imaging, and positron emission tomography may be used to depict and characterize duodenal strictures. Integration of imaging, clinical, laboratory, and endoscopic findings plays a major role in establishing a diagnosis of obstructive duodenal strictures. PMID:25208276

  13. Simultaneous Clear Cell Sarcomas of the Duodenum and Jejunum

    PubMed Central

    Cruise, Michael

    2016-01-01

    Clear cell sarcoma (CCS) is an uncommon tumor that usually presents as an extremity mass but can rarely manifest as a gastrointestinal tumor with a diverse spectrum of symptoms, most commonly related to a mass effect or ulceration. Herein we report a case in which two separate tumors, one in the duodenum and the other in the jejunum, present concurrently. The subject presented with symptomatic anemia and underwent imaging and endoscopic studies that culminated in the discovery of the two lesions. He subsequently underwent operative treatment with resection of both tumors and made an unremarkable recovery. The resection specimen consisted of two separate clear cell sarcomas with negative margins. Under microscopic evaluation, they demonstrated nested growths of epithelioid cells with scattered spindled cells infiltrating the enteric wall. The neoplastic cells were positive for S100 with scattered expression of Melan A. Florescence in situ hybridization revealed a translocation at the EWRS1 locus. He was disease-free for 30 months following the procedure; then he developed a rapidly progressing metastatic disease with subsequent death 4 months later. PMID:27375743

  14. Simple diagrammatic approach to delineate duodenum on a radiotherapy planning CT scan.

    PubMed

    Kataria, Tejinder; Gupta, Deepak; Basu, Trinanjan; Gupta, Shivani; Goyal, Shikha; Banerjee, Susovan; Abhishek, Ashu; Bisht, Shyam S; Narang, Kushal

    2016-01-01

    In recent years, there has been increasing application of intensity-modulated radiotherapy and stereotactic body radiotherapy for the treatment of abdominal malignancies (stomach, pancreas, liver, spinal metastases). This warrants accurate delineation of organs at risk, especially the duodenum. The tortuous and curvy anatomy of duodenum often indistinguishable from adjoining organs is a practical challenge. Radiation Therapy Oncology Group (RTOG) has already published upper abdominal normal structure contouring guidelines to ease the delineation process. This pictorial essay following the RTOG guideline elaborates the step-by-step identification of the different parts of duodenum in relation to the adjoining important structures. PMID:26647654

  15. Effect of injecting primary myoblasts versus putative muscle-derived stem cells on mass and force generation in mdx mice.

    PubMed

    Mueller, Gunhild M; O'Day, Terry; Watchko, Jon F; Ontell, Marcia

    2002-06-10

    It is well established that the injection of normal myoblasts or of muscle-derived stem cells (MDSCs) into the muscle of dystrophin-deficient mdx mice results in the incorporation of a number of donor myoblasts into the host muscle. However, the effect of the injected exogenous cells on mdx muscle mass and functional capacity has not been evaluated. This study evaluates the mass and functional capacity of the extensor digitorum longus (EDL) muscles of adult, male mdx mice that received intramuscular injections of primary myoblasts or of MDSCs (isolated by a preplating technique; Qu, Z., Balkir, L., van Deutekom, J.C., Robbins, P.D., Pruchnic, R., and Huard, J., J. Cell Biol. 1998;142:1257-1267) derived from normal mice. Evaluations were made 9 weeks after cell transplantation. Uninjected mdx EDL muscles have a mass 50% greater than that of age-matched C57BL/10J (normal) EDL muscles. Injections of either primary myoblasts or MDSCs have no effect on the mass of mdx EDL muscles. EDL muscles of mdx mice generate 43% more absolute twitch tension and 43% less specific tetanic tension then do EDL muscles of C57BL/10J mice. However, the absolute tetanic and specific twitch tension of mdx and C57BL/10J EDL muscles are similar. Injection of either primary myoblasts or MDSCs has no effect on the absolute or specific twitch and tetanic tensions of mdx muscle. Approximately 25% of the myofibers in mdx EDL muscles that received primary myoblasts react positively with antibody to dystrophin. There is no significant difference in the number of dystrophin-positive myofibers when MDSCs are injected. Regardless of the source of donor cells, dystrophin is limited to short distances (60-900 microm) along the length of the myofibers. This may, in part, explain the failure of cellular therapy to alter the contractile properties of murine dystrophic muscle. PMID:12067441

  16. Short-chain fatty acid sensing in rat duodenum

    PubMed Central

    Akiba, Yasutada; Inoue, Takuya; Kaji, Izumi; Higashiyama, Masaaki; Narimatsu, Kazuyuki; Iwamoto, Ken-ichi; Watanabe, Masahiko; Guth, Paul H; Engel, Eli; Kuwahara, Atsukazu; Kaunitz, Jonathan D

    2015-01-01

    Intraduodenal fatty acids (FA) and bacterial overgrowth, which generate short-chain FAs (SCFAs), have been implicated in the generation of functional dyspepsia symptoms. We studied the mechanisms by which luminal SCFA perfusion affects duodenal HCO3− secretion (DBS), a measure of mucosal neurohumoral activation. Free fatty acid receptor (FFAR) 1 (FFA1), which binds long-chain FA (LCFA), and SCFA receptors FFA2 and FFA3 were immunolocalised to duodenal enteroendocrine cells. FFA3 colocalised with glucagon-like peptide (GLP)-1, whereas FFA2 colocalised with 5-HT. Luminal perfusion of the SCFA acetate or propionate increased DBS, enhanced by dipeptidyl peptidase-IV (DPPIV) inhibition, at the same time as increasing GLP-2 portal blood concentrations. Acetate-induced DBS was partially inhibited by monocarboxylate/HCO3− exchanger inhibition without affecting GLP-2 release, implicating acetate absorption in the partial mediation of DBS. A selective FFA2 agonist dose-dependently increased DBS, unaffected by DPPIV inhibition or by cholecystokinin or 5-HT3 receptor antagonists, but was inhibited by atropine and a 5-HT4 antagonist. By contrast, a selective FFA1 agonist increased DBS accompanied by GLP-2 release, enhanced by DPPIV inhibition and inhibited by a GLP-2 receptor antagonist. Activation of FFA1 by LCFA and presumably FFA3 by SCFA increased DBS via GLP-2 release, whereas FFA2 activation stimulated DBS via muscarinic and 5-HT4 receptor activation. SCFA/HCO3− exchange also appears to be present in the duodenum. The presence of duodenal fatty acid sensing receptors that signal hormone release and possibly signal neural activation may be implicated in the pathogenesis of functional dyspepsia. PMID:25433076

  17. Primary squamous cell carcinoma of the duodenum effectively treated with TS-1: a case report.

    PubMed

    Fujita, Toshihiro; Nasu, Yuichiro; Hamamoto, Hitomi; Miyata, Fumiko; Oshige, Akihiko; Shigenobu, Shuho; Kanmura, Shuji; Numata, Masatsugu; Aozaki, Shinichiro; Ido, Akio

    2014-12-01

    An 89-year-old man was admitted to our hospital for thorough investigation of refractory diabetes mellitus, which revealed primary squamous cell carcinoma of the duodenum. After two courses of chemotherapy, follow-up esophagoduodenogastroscopy and duodenal biopsy showed no evidence of tumor. No findings were suggestive of recurrence of the primary lesion 19 months after starting chemotherapy. This case suggests that chemotherapy including TS-1 may be effective for treating unresectable primary squamous cell carcinoma of the duodenum. PMID:25482907

  18. Use of water ingestion to distinguish the gallbladder and duodenum on cholescintigrams

    SciTech Connect

    Keller, I.A.; Weissmann, H.S.; Kaplun, L.L.; Freeman, L.M.

    1984-09-01

    Cholescintigraphic diagnosis of acute cholecystitis requires accurate assessment of gallbladder nonvisualization. Confusion may occur when the gallbladder overlies the duodenal sweep or when labeled bile pools in the duodenum. Gallbladder activity could not be differentiated from duodenal activity in 21 patients. The oral ingestion of 225 ml of water permitted successful differentiation of the gallbladder from the duodenum. In 25 control subjects, it was demonstrated that that volume of water did not have a cholecystokinetic effect.

  19. Structural and Functional Alterations of Skeletal Muscle Microvasculature in Dystrophin-Deficient mdx Mice.

    PubMed

    Latroche, Claire; Matot, Béatrice; Martins-Bach, Aurea; Briand, David; Chazaud, Bénédicte; Wary, Claire; Carlier, Pierre G; Chrétien, Fabrice; Jouvion, Grégory

    2015-09-01

    Duchenne muscular dystrophy (DMD) is a progressive neuromuscular disease, caused by an absence of dystrophin, inevitably leading to death. Although muscle lesions are well characterized, blood vessel alterations that may have a major impact on muscle regeneration remain poorly understood. Our aim was to elucidate alterations of the vascular network organization, taking advantage of Flk1(GFP/+) crossed with mdx mice (model for human DMD where all blood vessels express green fluorescent protein) and functional repercussions using in vivo nuclear magnetic resonance, combining arterial spin-labeling imaging of perfusion, and (31)P-spectroscopy of phosphocreatine kinetics. For the first time, our study focused on old (12-month-old) mdx mice, displaying marked chronic muscle lesions, similar to the lesions observed in human DMD, in comparison to young-adult (3-month-old) mdx mice displaying only mild muscle lesions with no fibrosis. By using an original approach combining a specific animal model, state-of-the-art histology/morphometry techniques, and functional nuclear magnetic resonance, we demonstrated that the microvascular system is almost normal in young-adult in contrast to old mdx mice, displaying marked microvessel alterations, and the functional repercussions on muscle perfusion and bioenergetics after a hypoxic stress vary depending on stage of pathology. This original approach clarifies disease evolution and paves the way for setting up new diagnostic markers or therapeutic strategies. PMID:26193666

  20. Losartan decreases cardiac muscle fibrosis and improves cardiac function in dystrophin-deficient mdx mice.

    PubMed

    Spurney, Christopher F; Sali, Arpana; Guerron, Alfredo D; Iantorno, Micaela; Yu, Qing; Gordish-Dressman, Heather; Rayavarapu, Sree; van der Meulen, Jack; Hoffman, Eric P; Nagaraju, Kanneboyina

    2011-03-01

    Recent studies showed that chronic administration of losartan, an angiotensin II type I receptor antagonist, improved skeletal muscle function in dystrophin-deficient mdx mice. In this study, C57BL/10ScSn-Dmd(mdx)/J female mice were either untreated or treated with losartan (n = 15) in the drinking water at a dose of 600 mg/L over a 6-month period. Cardiac function was assessed via in vivo high frequency echocardiography and skeletal muscle function was assessed using grip strength testing, Digiscan monitoring, Rotarod timing, and in vitro force testing. Fibrosis was assessed using picrosirius red staining and Image J analysis. Gene expression was evaluated using real-time polymerized chain reaction (RT-PCR). Percentage shortening fraction was significantly decreased in untreated (26.9% ± 3.5%) mice compared to losartan-treated (32.2% ± 4.2%; P < .01) mice. Systolic blood pressure was significantly reduced in losartan-treated mice (56 ± 6 vs 69 ± 7 mm Hg; P < .0005). Percentage cardiac fibrosis was significantly reduced in losartan-treated hearts (P < .05) along with diaphragm (P < .01), extensor digitorum longus (P < .05), and gastrocnemius (P < .05) muscles compared to untreated mdx mice. There were no significant differences in skeletal muscle function between treated and untreated groups. Chronic treatment with losartan decreases cardiac and skeletal muscle fibrosis and improves cardiac systolic function in dystrophin-deficient mdx mice. PMID:21304057

  1. Bodywide skipping of exons 45-55 in dystrophic mdx52 mice by systemic antisense delivery.

    PubMed

    Aoki, Yoshitsugu; Yokota, Toshifumi; Nagata, Tetsuya; Nakamura, Akinori; Tanihata, Jun; Saito, Takashi; Duguez, Stephanie M R; Nagaraju, Kanneboyina; Hoffman, Eric P; Partridge, Terence; Takeda, Shin'ichi

    2012-08-21

    Duchenne muscular dystrophy (DMD), the commonest form of muscular dystrophy, is caused by lack of dystrophin. One of the most promising therapeutic approaches is antisense-mediated elimination of frame-disrupting mutations by exon skipping. However, this approach faces two major hurdles: limited applicability of each individual target exon and uncertain function and stability of each resulting truncated dystrophin. Skipping of exons 45-55 at the mutation hotspot of the DMD gene would address both issues. Theoretically it could rescue more than 60% of patients with deletion mutations. Moreover, spontaneous deletions of this specific region are associated with asymptomatic or exceptionally mild phenotypes. However, such multiple exon skipping of exons 45-55 has proved technically challenging. We have therefore designed antisense oligo (AO) morpholino mixtures to minimize self- or heteroduplex formation. These were tested as conjugates with cell-penetrating moieties (vivo-morpholinos). We have tested the feasibility of skipping exons 45-55 in H2K-mdx52 myotubes and in mdx52 mice, which lack exon 52. Encouragingly, with mixtures of 10 AOs, we demonstrated skipping of all 10 exons in vitro, in H2K-mdx52 myotubes and on intramuscular injection into mdx52 mice. Moreover, in mdx52 mice in vivo, systemic injections of 10 AOs induced extensive dystrophin expression at the subsarcolemma in skeletal muscles throughout the body, producing up to 15% of wild-type dystrophin protein levels, accompanied by improved muscle strength and histopathology without any detectable toxicity. This is a unique successful demonstration of effective rescue by exon 45-55 skipping in a dystrophin-deficient animal model. PMID:22869723

  2. Selective modulation through the glucocorticoid receptor ameliorates muscle pathology in mdx mice

    PubMed Central

    Huynh, Tony; Uaesoontrachoon, Kitipong; Quinn, James L; Tatem, Kathleen S; Heier, Christopher R; Van Der Meulen, Jack H; Yu, Qing; Harris, Mark; Nolan, Christopher J; Haegeman, Guy; Grounds, Miranda D; Nagaraju, Kanneboyina

    2014-01-01

    The over-expression of NF-κB signalling in both muscle and immune cells contribute to the pathology in dystrophic muscle. The anti-inflammatory properties of glucocorticoids, mediated predominantly through monomeric glucocorticoid receptor inhibition of transcription factors such as NF-κB (transrepression), are postulated to be an important mechanism for their beneficial effects in Duchenne muscular dystrophy. Chronic glucocorticoid therapy is associated with adverse effects on metabolism, growth, bone mineral density and the maintenance of muscle mass. These detrimental effects result from direct glucocorticoid receptor homodimer interactions with glucocorticoid response elements of the relevant genes. Compound A, a non-steroidal selective glucocorticoid receptor modulator, is capable of transrepression without transactivation. We confirm the in vitro NF-κB inhibitory activity of compound A in H-2Kb-tsA58 mdx myoblasts and myotubes, and demonstrate improvements in disease phenotype of dystrophin deficient mdx mice. Compound A treatment in mdx mice from 18 days of post-natal age to 8 weeks of age increased the absolute and normalized forelimb and hindlimb grip strength, attenuated cathepsin-B enzyme activity (a surrogate marker for inflammation) in forelimb and hindlimb muscles, decreased serum creatine kinase levels and reduced IL-6, CCL2, IFNγ, TNF and IL-12p70 cytokine levels in gastrocnemius (GA) muscles. Compared with compound A, treatment with prednisolone, a classical glucocorticoid, in both wild-type and mdx mice was associated with reduced body weight, reduced GA, tibialis anterior and extensor digitorum longus muscle mass and shorter tibial lengths. Prednisolone increased osteopontin (Spp1) gene expression and osteopontin protein levels in the GA muscles of mdx mice and had less favourable effects on the expression of Foxo1, Foxo3, Fbxo32, Trim63, Mstn and Igf1 in GA muscles, as well as hepatic Igf1 in wild-type mice. In conclusion, selective

  3. Nifedipine treatment reduces resting calcium concentration, oxidative and apoptotic gene expression, and improves muscle function in dystrophic mdx mice.

    PubMed

    Altamirano, Francisco; Valladares, Denisse; Henríquez-Olguín, Carlos; Casas, Mariana; López, Jose R; Allen, Paul D; Jaimovich, Enrique

    2013-01-01

    Duchenne Muscular Dystrophy (DMD) is a recessive X-linked genetic disease, caused by mutations in the gene encoding dystrophin. DMD is characterized in humans and in mdx mice by a severe and progressive destruction of muscle fibers, inflammation, oxidative/nitrosative stress, and cell death. In mdx muscle fibers, we have shown that basal ATP release is increased and that extracellular ATP stimulation is pro-apoptotic. In normal fibers, depolarization-induced ATP release is blocked by nifedipine, leading us to study the potential therapeutic effect of nifedipine in mdx muscles and its relation with extracellular ATP signaling. Acute exposure to nifedipine (10 µM) decreased [Ca(2+)]r, NF-κB activity and iNOS expression in mdx myotubes. In addition, 6-week-old mdx mice were treated with daily intraperitoneal injections of nifedipine, 1 mg/Kg for 1 week. This treatment lowered the [Ca(2+)]r measured in vivo in the mdx vastus lateralis. We demonstrated that extracellular ATP levels were higher in adult mdx flexor digitorum brevis (FDB) fibers and can be significantly reduced after 1 week of treatment with nifedipine. Interestingly, acute treatment of mdx FDB fibers with apyrase, an enzyme that completely degrades extracellular ATP to AMP, reduced [Ca(2+)]r to a similar extent as was seen in FDB fibers after 1-week of nifedipine treatment. Moreover, we demonstrated that nifedipine treatment reduced mRNA levels of pro-oxidative/nitrosative (iNOS and gp91(phox)/p47(phox) NOX2 subunits) and pro-apoptotic (Bax) genes in mdx diaphragm muscles and lowered serum creatine kinase (CK) levels. In addition, nifedipine treatment increased muscle strength assessed by the inverted grip-hanging test and exercise tolerance measured with forced swimming test in mdx mice. We hypothesize that nifedipine reduces basal ATP release, thereby decreasing purinergic receptor activation, which in turn reduces [Ca(2+)]r in mdx skeletal muscle cells. The results in this work open new perspectives

  4. Nifedipine Treatment Reduces Resting Calcium Concentration, Oxidative and Apoptotic Gene Expression, and Improves Muscle Function in Dystrophic mdx Mice

    PubMed Central

    Henríquez-Olguín, Carlos; Casas, Mariana; López, Jose R.; Allen, Paul D.; Jaimovich, Enrique

    2013-01-01

    Duchenne Muscular Dystrophy (DMD) is a recessive X-linked genetic disease, caused by mutations in the gene encoding dystrophin. DMD is characterized in humans and in mdx mice by a severe and progressive destruction of muscle fibers, inflammation, oxidative/nitrosative stress, and cell death. In mdx muscle fibers, we have shown that basal ATP release is increased and that extracellular ATP stimulation is pro-apoptotic. In normal fibers, depolarization-induced ATP release is blocked by nifedipine, leading us to study the potential therapeutic effect of nifedipine in mdx muscles and its relation with extracellular ATP signaling. Acute exposure to nifedipine (10 µM) decreased [Ca2+]r, NF-κB activity and iNOS expression in mdx myotubes. In addition, 6-week-old mdx mice were treated with daily intraperitoneal injections of nifedipine, 1 mg/Kg for 1 week. This treatment lowered the [Ca2+]r measured in vivo in the mdx vastus lateralis. We demonstrated that extracellular ATP levels were higher in adult mdx flexor digitorum brevis (FDB) fibers and can be significantly reduced after 1 week of treatment with nifedipine. Interestingly, acute treatment of mdx FDB fibers with apyrase, an enzyme that completely degrades extracellular ATP to AMP, reduced [Ca2+]r to a similar extent as was seen in FDB fibers after 1-week of nifedipine treatment. Moreover, we demonstrated that nifedipine treatment reduced mRNA levels of pro-oxidative/nitrosative (iNOS and gp91phox/p47phox NOX2 subunits) and pro-apoptotic (Bax) genes in mdx diaphragm muscles and lowered serum creatine kinase (CK) levels. In addition, nifedipine treatment increased muscle strength assessed by the inverted grip-hanging test and exercise tolerance measured with forced swimming test in mdx mice. We hypothesize that nifedipine reduces basal ATP release, thereby decreasing purinergic receptor activation, which in turn reduces [Ca2+]r in mdx skeletal muscle cells. The results in this work open new perspectives towards

  5. PKC Theta Ablation Improves Healing in a Mouse Model of Muscular Dystrophy

    PubMed Central

    Madaro, Luca; Pelle, Andrea; Nicoletti, Carmine; Crupi, Annunziata; Marrocco, Valeria; Bossi, Gianluca; Soddu, Silvia; Bouché, Marina

    2012-01-01

    Inflammation is a key pathological characteristic of dystrophic muscle lesion formation, limiting muscle regeneration and resulting in fibrotic and fatty tissue replacement of muscle, which exacerbates the wasting process in dystrophic muscles. Limiting immune response is thus one of the therapeutic options to improve healing, as well as to improve the efficacy of gene- or cell-mediated strategies to restore dystrophin expression. Protein kinase C θ (PKCθ) is a member of the PKCs family highly expressed in both immune cells and skeletal muscle; given its crucial role in adaptive, but also innate, immunity, it is being proposed as a valuable pharmacological target for immune disorders. In our study we asked whether targeting PKCθ could represent a valuable approach to efficiently prevent inflammatory response and disease progression in a mouse model of muscular dystrophy. We generated the bi-genetic mouse model mdx/θ−/−, where PKCθ expression is lacking in mdx mice, the mouse model of Duchenne muscular dystrophy. We found that muscle wasting in mdx/θ−/− mice was greatly prevented, while muscle regeneration, maintenance and performance was significantly improved, as compared to mdx mice. This phenotype was associated to reduction in inflammatory infiltrate, pro-inflammatory gene expression and pro-fibrotic markers activity, as compared to mdx mice. Moreover, BM transplantation experiments demonstrated that the phenotype observed was primarily dependent on lack of PKCθ expression in hematopoietic cells. These results demonstrate a hitherto unrecognized role of immune-cell intrinsic PKCθ activity in the development of DMD. Although the immune cell population(s) involved remain unidentified, our findings reveal that PKCθ can be proposed as a new pharmacological target to counteract the disease, as well as to improve the efficacy of gene- or cell- therapy approaches. PMID:22348094

  6. Increased frequency of activated T-cells in the Helicobacter pylori-infected antrum and duodenum.

    PubMed

    Strömberg, E; Lundgren, A; Edebo, A; Lundin, S; Svennerholm, A-M; Lindholm, C

    2003-05-25

    Helicobacter pylori colonize the human stomach and duodenum. The infection has been shown to induce a strong T-cell response in the stomach, whereas the response within the duodenum has been poorly characterized. Furthermore, it remains to be elucidated whether the T-cell response may contribute to ulcer formation in the host. In this study, the frequency of different T-cell subsets, their degree of activation and expression of co-stimulatory receptors in biopsies from the duodenum as well as the antrum were studied by immunohistochemistry and flow cytometry. It was also evaluated whether there are differences in the T-cell responses between duodenal ulcer patients and asymptomatic carriers that might explain why only 10-15% of the infected subjects develop duodenal ulcers. The frequencies of CD4+, CD8+ and CD45RO+, i.e. memory T-cells, were significantly increased in the antrum, and the number of CD25+ cells was considerably higher in both the antrum and duodenum of duodenal ulcer patients and asymptomatic carriers as compared to uninfected individuals. Interestingly, the levels of immunosuppressive CTLA-4+ cells were significantly higher in the duodenum of duodenal ulcer patients, as compared to the asymptomatic carriers. H. pylori cause activation of T-cells in the duodenum as well as in the stomach. Our observation of higher levels of CTLA-4+ cells in the duodenum of duodenal ulcer patients than in the asymptomatic carriers suggests that a suppressive T-cell response may be related to the development of duodenal ulcers. PMID:12738386

  7. Hexose enhances oligonucleotide delivery and exon skipping in dystrophin-deficient mdx mice

    PubMed Central

    Han, Gang; Gu, Ben; Cao, Limin; Gao, Xianjun; Wang, Qingsong; Seow, Yiqi; Zhang, Ning; Wood, Matthew J. A.; Yin, HaiFang

    2016-01-01

    Carbohydrate-based infusion solutions are widely used in the clinic. Here we show that co-administration of phosphorodiamidate morpholino oligomers (PMOs) with glucose enhances exon-skipping activity in Duchenne muscular dystrophy (DMD) mdx mice. We identify a glucose–fructose (GF) formulation that potentiates PMO activity, completely corrects aberrant Dmd transcripts, restores dystrophin levels in skeletal muscles and achieves functional rescue without detectable toxicity. This activity is attributed to enhancement of GF-mediated PMO uptake in the muscle. We demonstrate that PMO cellular uptake is energy dependent, and that ATP from GF metabolism contributes to enhanced cellular uptake of PMO in the muscle. Collectively, we show that GF potentiates PMO activity by replenishing cellular energy stores under energy-deficient conditions in mdx mice. Our findings provide mechanistic insight into hexose-mediated oligonucleotide delivery and have important implications for the development of DMD exon-skipping therapy. PMID:26964641

  8. Hexose enhances oligonucleotide delivery and exon skipping in dystrophin-deficient mdx mice.

    PubMed

    Han, Gang; Gu, Ben; Cao, Limin; Gao, Xianjun; Wang, Qingsong; Seow, Yiqi; Zhang, Ning; Wood, Matthew J A; Yin, HaiFang

    2016-01-01

    Carbohydrate-based infusion solutions are widely used in the clinic. Here we show that co-administration of phosphorodiamidate morpholino oligomers (PMOs) with glucose enhances exon-skipping activity in Duchenne muscular dystrophy (DMD) mdx mice. We identify a glucose-fructose (GF) formulation that potentiates PMO activity, completely corrects aberrant Dmd transcripts, restores dystrophin levels in skeletal muscles and achieves functional rescue without detectable toxicity. This activity is attributed to enhancement of GF-mediated PMO uptake in the muscle. We demonstrate that PMO cellular uptake is energy dependent, and that ATP from GF metabolism contributes to enhanced cellular uptake of PMO in the muscle. Collectively, we show that GF potentiates PMO activity by replenishing cellular energy stores under energy-deficient conditions in mdx mice. Our findings provide mechanistic insight into hexose-mediated oligonucleotide delivery and have important implications for the development of DMD exon-skipping therapy. PMID:26964641

  9. The first exon duplication mouse model of Duchenne muscular dystrophy: A tool for therapeutic development.

    PubMed

    Vulin, Adeline; Wein, Nicolas; Simmons, Tabatha R; Rutherford, Andrea M; Findlay, Andrew R; Yurkoski, Jacqueline A; Kaminoh, Yuuki; Flanigan, Kevin M

    2015-11-01

    Exon duplication mutations account for up to 11% of all cases of Duchenne muscular dystrophy (DMD), and a duplication of exon 2 is the most common duplication in patients. For use as a platform for testing of duplication-specific therapies, we developed a mouse model that carries a Dmd exon 2 duplication. By using homologous recombination we duplicated exon 2 within intron 2 at a location consistent with a human duplication hotspot. mRNA analysis confirms the inclusion of a duplicated exon 2 in mouse muscle. Dystrophin expression is essentially absent by immunofluorescent and immunoblot analysis, although some muscle specimens show very low-level trace dystrophin expression. Phenotypically, the mouse shows similarities to mdx, the standard laboratory model of DMD. In skeletal muscle, areas of necrosis and phagocytosis are seen at 3 weeks, with central nucleation prominent by four weeks, recapitulating the "crisis" period in mdx. Marked diaphragm fibrosis is noted by 6 months, and remains unchanged at 12 months. Our results show that the Dup2 mouse is both pathologically (in degree and distribution) and physiologically similar to mdx. As it recapitulates the most common single exon duplication found in DMD patients, this new model will be a useful tool to assess the potential of duplicated exon skipping. PMID:26365037

  10. The action of porcine glucagon on the motility of the canine duodenum and jejunum.

    PubMed Central

    Evans, D. F.; Foster, G. E.; Hardcastle, J. D.; Jonhson, F.; Wright, J. W.

    1982-01-01

    1 Intravenous bolus doses of porcine glucagon of 0.001-0.05 mg kg-1 caused intense stimulation of the duodenum and jejunum of the dog. 2 Intravenous infusion of porcine glucagon at 0.025-0.05 mg kg-1 h-1 caused similar stimulation. In both cases the stimulation was phasic in nature. 3 Stimulation of the duodenum and jejunum following glucagon was accompanied by a decrease in frequency of the intestinal basic electrical rhythm (BER). No change was seen in the intervals between successive periods of phase III motor activity. PMID:7093585

  11. Regression of follicular lymphoma of the duodenum following eradication of H. pylori infection.

    PubMed

    Hayashi, Hiroki; Onishi, Yutaka; Mitsuoka, Hiroshi; Ogura, Takeshi; Maeda, Mitsuo; Nishigami, Takashi; Harada, Masaru

    2013-01-01

    A 64-year-old woman was referred for an examination of the upper gastrointestinal (GI) tract. Endoscopy showed an elevated lesion in the duodenum with central depression and multiple white granules. Biopsy specimens revealed lymphoid follicles composed predominantly of centrocytes with scattered centroblasts. The tumor cells were positive for bcl-2. The patient was diagnosed with follicular lymphoma and underwent antibiotic therapy for Helicobacter pylori (H. pylori) infection. The regression of the lesion was obvious. After 5.5 years of follow-up, there has been no evidence of recurrence. This case suggests that H. pylori eradication therapy is effective for treating follicular lymphoma in the duodenum. PMID:24292749

  12. Laparoscopic Pancreaticoduodenectomy for the Management of Localized Crohn's Disease of the Duodenum.

    PubMed

    Xingjun, Guo; Feng, Zhu; Min, Wang; Renyi, Qin

    2016-08-01

    Crohn's disease of the duodenum is an uncommon condition. Our case was an extremely rare manifestation of Crohn's disease, who presented with obstruction of the pylorus and the first and the second parts of the duodenum. Because of the severity of the obstruction, he underwent laparoscopic pancreaticoduodenectomy. Postoperative pancreatic leakage and bowel fistula were not observed, and there was no morbidity during the follow-up period. There was also no disturbance in digestive function, postoperatively. This is the first case employing laparoscopic pancreaticoduodenectomy to cure benign lesions leading to duodenal obstruction. Minimally invasive laparoscopic pancreaticoduodenectomy technology shows a very big advantage in treating this rare benign Crohn's disease. PMID:27574357

  13. Characterization of neuromuscular synapse function abnormalities in multiple Duchenne muscular dystrophy mouse models.

    PubMed

    van der Pijl, Elizabeth M; van Putten, Maaike; Niks, Erik H; Verschuuren, Jan J G M; Aartsma-Rus, Annemieke; Plomp, Jaap J

    2016-06-01

    Duchenne muscular dystrophy (DMD) is an X-linked myopathy caused by dystrophin deficiency. Dystrophin is present intracellularly at the sarcolemma, connecting actin to the dystrophin-associated glycoprotein complex. Interestingly, it is enriched postsynaptically at the neuromuscular junction (NMJ), but its synaptic function is largely unknown. Utrophin, a dystrophin homologue, is also concentrated at the NMJ, and upregulated in DMD. It is possible that the absence of dystrophin at NMJs in DMD causes neuromuscular transmission defects that aggravate muscle weakness. We studied NMJ function in mdx mice (lacking dystrophin) and wild type mice. In addition, mdx/utrn(+/-) and mdx/utrn(-/-) mice (lacking utrophin) were used to investigate influences of utrophin levels. The three Duchenne mouse models showed muscle weakness when comparatively tested in vivo, with mdx/utrn(-/-) mice being weakest. Ex vivo muscle contraction and electrophysiological studies showed a reduced safety factor of neuromuscular transmission in all models. NMJs had ~ 40% smaller miniature endplate potential amplitudes compared with wild type, indicating postsynaptic sensitivity loss for the neurotransmitter acetylcholine. However, nerve stimulation-evoked endplate potential amplitudes were unchanged. Consequently, quantal content (i.e. the number of acetylcholine quanta released per nerve impulse) was considerably increased. Such a homeostatic compensatory increase in neurotransmitter release is also found at NMJs in myasthenia gravis, where autoantibodies reduce acetylcholine receptors. However, high-rate nerve stimulation induced exaggerated endplate potential rundown. Study of NMJ morphology showed that fragmentation of acetylcholine receptor clusters occurred in all models, being most severe in mdx/utrn(-/-) mice. Overall, we showed mild 'myasthenia-like' neuromuscular synaptic dysfunction in several Duchenne mouse models, which possibly affects muscle weakness and degeneration. PMID:27037492

  14. Peptide Nucleic Acid Promotes Systemic Dystrophin Expression and Functional Rescue in Dystrophin-deficient mdx Mice.

    PubMed

    Gao, Xianjun; Shen, Xiaoyong; Dong, Xue; Ran, Ning; Han, Gang; Cao, Limin; Gu, Ben; Yin, HaiFang

    2015-01-01

    Antisense oligonucleotide (AO)-mediated exon-skipping therapeutics shows great promise for Duchenne muscular dystrophy (DMD) patients. However, recent failure with drisapersen, an AO candidate drug in phase 3 trial, highlights the importance of exploring other effective AO chemistries for DMD. Previously, we demonstrated the appreciable biological activity of peptide nucleic acid (PNA) AOs in restoring dystrophin expression in dystrophin-deficient mdx mice intramuscularly. Here, we further explore the systemic potential and feasibility of PNA AOs in mediating exon skipping in mdx mice as a comprehensive systemic evaluation remains lacking. Systemic delivery of PNA AOs resulted in therapeutic level of dystrophin expression in body-wide peripheral muscles and improved dystrophic pathology in mdx mice without any detectable toxicity. Up to 40% of dystrophin restoration was achieved in gastrocnemius, to a less extent with other skeletal muscles, with no dystrophin in heart. Notably, comparable systemic activity was obtained between PNA AOs and phosphorodiamidate morpholino oligomer, a DMD AO chemistry in phase 3 clinical trial, under an identical dosing regimen. Overall, our data demonstrate that PNA is viable for DMD exon-skipping therapeutics with 20 mer showing the best combination of activity, solubility, and safety and further modifications to increase PNA aqueous solubility can enable longer, more effective therapeutics without the associated toxicity. PMID:26440599

  15. Rapid depletion of muscle progenitor cells in dystrophic mdx/utrophin−/− mice

    PubMed Central

    Lu, Aiping; Poddar, Minakshi; Tang, Ying; Proto, Jonathan D.; Sohn, Jihee; Mu, Xiaodong; Oyster, Nicholas; Wang, Bing; Huard, Johnny

    2014-01-01

    Duchenne muscular dystrophy (DMD) patients lack dystrophin from birth; however, muscle weakness becomes apparent only at 3–5 years of age, which happens to coincide with the depletion of the muscle progenitor cell (MPC) pools. Indeed, MPCs isolated from older DMD patients demonstrate impairments in myogenic potential. To determine whether the progression of muscular dystrophy is a consequence of the decline in functional MPCs, we investigated two animal models of DMD: (i) dystrophin-deficient mdx mice, the most commonly utilized model of DMD, which has a relatively mild dystrophic phenotype and (ii) dystrophin/utrophin double knock-out (dKO) mice, which display a similar histopathologic phenotype to DMD patients. In contrast to age-matched mdx mice, we observed that both the number and regeneration potential of dKO MPCs rapidly declines during disease progression. This occurred in MPCs at both early and late stages of myogenic commitment. In fact, early MPCs isolated from 6-week-old dKO mice have reductions in proliferation, resistance to oxidative stress and multilineage differentiation capacities compared with age-matched mdx MPCs. This effect may potentially be mediated by fibroblast growth factor overexpression and/or a reduction in telomerase activity. Our results demonstrate that the rapid disease progression in the dKO model is associated, at least in part, with MPC depletion. Therefore, alleviating MPC depletion could represent an approach to delay the onset of the histopathologies associated with DMD patients. PMID:24781208

  16. Peptide Nucleic Acid Promotes Systemic Dystrophin Expression and Functional Rescue in Dystrophin-deficient mdx Mice

    PubMed Central

    Gao, Xianjun; Shen, Xiaoyong; Dong, Xue; Ran, Ning; Han, Gang; Cao, Limin; Gu, Ben; Yin, HaiFang

    2015-01-01

    Antisense oligonucleotide (AO)-mediated exon-skipping therapeutics shows great promise for Duchenne muscular dystrophy (DMD) patients. However, recent failure with drisapersen, an AO candidate drug in phase 3 trial, highlights the importance of exploring other effective AO chemistries for DMD. Previously, we demonstrated the appreciable biological activity of peptide nucleic acid (PNA) AOs in restoring dystrophin expression in dystrophin-deficient mdx mice intramuscularly. Here, we further explore the systemic potential and feasibility of PNA AOs in mediating exon skipping in mdx mice as a comprehensive systemic evaluation remains lacking. Systemic delivery of PNA AOs resulted in therapeutic level of dystrophin expression in body-wide peripheral muscles and improved dystrophic pathology in mdx mice without any detectable toxicity. Up to 40% of dystrophin restoration was achieved in gastrocnemius, to a less extent with other skeletal muscles, with no dystrophin in heart. Notably, comparable systemic activity was obtained between PNA AOs and phosphorodiamidate morpholino oligomer, a DMD AO chemistry in phase 3 clinical trial, under an identical dosing regimen. Overall, our data demonstrate that PNA is viable for DMD exon-skipping therapeutics with 20 mer showing the best combination of activity, solubility, and safety and further modifications to increase PNA aqueous solubility can enable longer, more effective therapeutics without the associated toxicity. PMID:26440599

  17. Effect of Dystrophin Deficiency on Selected Intrinsic Laryngeal Muscles of the "mdx" Mouse

    ERIC Educational Resources Information Center

    Fry, Lisa T.; Stemple, Joseph C.; Andreatta, Richard D.; Harrison, Anne L.; Andrade, Francisco H.

    2010-01-01

    Background: Intrinsic laryngeal muscles (ILM) show biological differences from the broader class of skeletal muscles. Yet most research regarding ILM specialization has been completed on a few muscles, most notably the thyroarytenoid and posterior cricoarytenoid. Little information exists regarding the biology of other ILM. Early evidence suggests…

  18. Selective modulation through the glucocorticoid receptor ameliorates muscle pathology in mdx mice.

    PubMed

    Huynh, Tony; Uaesoontrachoon, Kitipong; Quinn, James L; Tatem, Kathleen S; Heier, Christopher R; Van Der Meulen, Jack H; Yu, Qing; Harris, Mark; Nolan, Christopher J; Haegeman, Guy; Grounds, Miranda D; Nagaraju, Kanneboyina

    2013-10-01

    The over-expression of NF-κB signalling in both muscle and immune cells contribute to the pathology in dystrophic muscle. The anti-inflammatory properties of glucocorticoids, mediated predominantly through monomeric glucocorticoid receptor inhibition of transcription factors such as NF-κB (transrepression), are postulated to be an important mechanism for their beneficial effects in Duchenne muscular dystrophy. Chronic glucocorticoid therapy is associated with adverse effects on metabolism, growth, bone mineral density and the maintenance of muscle mass. These detrimental effects result from direct glucocorticoid receptor homodimer interactions with glucocorticoid response elements of the relevant genes. Compound A, a non-steroidal selective glucocorticoid receptor modulator, is capable of transrepression without transactivation. We confirm the in vitro NF-κB inhibitory activity of compound A in H-2K(b) -tsA58 mdx myoblasts and myotubes, and demonstrate improvements in disease phenotype of dystrophin deficient mdx mice. Compound A treatment in mdx mice from 18 days of post-natal age to 8 weeks of age increased the absolute and normalized forelimb and hindlimb grip strength, attenuated cathepsin-B enzyme activity (a surrogate marker for inflammation) in forelimb and hindlimb muscles, decreased serum creatine kinase levels and reduced IL-6, CCL2, IFNγ, TNF and IL-12p70 cytokine levels in gastrocnemius (GA) muscles. Compared with compound A, treatment with prednisolone, a classical glucocorticoid, in both wild-type and mdx mice was associated with reduced body weight, reduced GA, tibialis anterior and extensor digitorum longus muscle mass and shorter tibial lengths. Prednisolone increased osteopontin (Spp1) gene expression and osteopontin protein levels in the GA muscles of mdx mice and had less favourable effects on the expression of Foxo1, Foxo3, Fbxo32, Trim63, Mstn and Igf1 in GA muscles, as well as hepatic Igf1 in wild-type mice. In conclusion, selective

  19. In vivo gene editing in dystrophic mouse muscle and muscle stem cells#

    PubMed Central

    Cheng, Jason K.W.; Chew, Wei Leong; Widrick, Jeffrey J.; Yan, Winston X.; Maesner, Claire; Wu, Elizabeth Y.; Xiao, Ru; Ran, F. Ann; Cong, Le; Zhang, Feng; Vandenberghe, Luk H.; Church, George M.; Wagers, Amy J.

    2016-01-01

    Frame-disrupting mutations in the DMD gene, encoding dystrophin, compromise myofiber integrity and drive muscle deterioration in Duchenne muscular dystrophy (DMD). Removing one or more exons from the mutated transcript can produce an in-frame mRNA and a truncated but still functional protein. In this study, we develop and test a direct gene editing approach to induce exon deletion and recover dystrophin expression in the mdx mouse model of DMD. Delivery by adeno-associated virus (AAV) of clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 endonucleases coupled with paired guide RNAs flanking the mutated Dmd exon23 resulted in excision of intervening DNA and restored Dystrophin reading frame in myofibers, cardiomyocytes and muscle stem cells following local or systemic delivery. AAV-Dmd CRISPR-treatment partially recovered muscle functional deficiencies and generated a pool of endogenously corrected myogenic precursors in mdx mouse muscle. PMID:26721686

  20. In vivo gene editing in dystrophic mouse muscle and muscle stem cells.

    PubMed

    Tabebordbar, Mohammadsharif; Zhu, Kexian; Cheng, Jason K W; Chew, Wei Leong; Widrick, Jeffrey J; Yan, Winston X; Maesner, Claire; Wu, Elizabeth Y; Xiao, Ru; Ran, F Ann; Cong, Le; Zhang, Feng; Vandenberghe, Luk H; Church, George M; Wagers, Amy J

    2016-01-22

    Frame-disrupting mutations in the DMD gene, encoding dystrophin, compromise myofiber integrity and drive muscle deterioration in Duchenne muscular dystrophy (DMD). Removing one or more exons from the mutated transcript can produce an in-frame mRNA and a truncated, but still functional, protein. In this study, we developed and tested a direct gene-editing approach to induce exon deletion and recover dystrophin expression in the mdx mouse model of DMD. Delivery by adeno-associated virus (AAV) of clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 endonucleases coupled with paired guide RNAs flanking the mutated Dmd exon23 resulted in excision of intervening DNA and restored the Dmd reading frame in myofibers, cardiomyocytes, and muscle stem cells after local or systemic delivery. AAV-Dmd CRISPR treatment partially recovered muscle functional deficiencies and generated a pool of endogenously corrected myogenic precursors in mdx mouse muscle. PMID:26721686

  1. Differential ability of Ptf1a and Ptf1a-VP16 to convert stomach, duodenum and liver to pancreas

    PubMed Central

    Jarikji, Zeina H.; Vanamala, Sandeep; Beck, Caroline W.; Wright, Chris V.E.; Leach, Steven D.; Horb, Marko E.

    2014-01-01

    Summary Determining the functional attributes of pancreatic transcription factors is essential to understand how the pancreas is specified distinct from other endodermal organs, such as liver, stomach and duodenum, and to direct the differentiation of other cell types into pancreas. Previously, we demonstrated that Pdx1-VP16 was sufficient to convert liver to pancreas. In this paper we characterize the functional ability of another pancreatic transcription factor, Ptf1a, in promoting ectopic pancreatic fates at early stages throughout the endoderm and later in during organogenesis. Using the transthyretin promoter to drive expression in the early liver region/bud of transgenic Xenopus tadpoles, we find that Ptf1a-VP16 is able to convert liver to pancreas. Overexpression of the unmodified Ptf1a on the other hand, has no effect in liver, but is able to convert stomach and duodenum to pancreas. When overexpressed at earlier embryonic stages throughout the endoderm, Ptf1a activity is similarly limited, whereas Ptf1a-VP16 has increased activity. Interestingly, in all instances we find that Ptf1a-VP16 is only capable of promoting acinar cell fates, whereas Ptf1a promotes both acinar and endocrine fates. Lastly, we demonstrate that, similar to mouse and zebrafish, Xenopus Ptf1a is essential for the initial specification of both endocrine and exocrine cells during normal pancreas development. PMID:17320068

  2. Fibroadipogenic progenitors mediate the ability of HDAC inhibitors to promote regeneration in dystrophic muscles of young, but not old Mdx mice

    PubMed Central

    Mozzetta, Chiara; Consalvi, Silvia; Saccone, Valentina; Tierney, Matthew; Diamantini, Adamo; Mitchell, Kathryn J; Marazzi, Giovanna; Borsellino, Giovanna; Battistini, Luca; Sassoon, David; Sacco, Alessandra; Puri, Pier Lorenzo

    2013-01-01

    HDAC inhibitors (HDACi) exert beneficial effects in mdx mice, by promoting endogenous regeneration; however, the cellular determinants of HDACi activity on dystrophic muscles have not been determined. We show that fibroadipogenic progenitors (FAP) influence the regeneration potential of satellite cells during disease progression in mdx mice and mediate HDACi ability to selectively promote regeneration at early stages of disease. FAPs from young mdx mice promote, while FAPs from old mdx mice repress, satellite cell-mediated formation of myotubes. In young mdx mice HDACi inhibited FAP adipogenic potential, while enhancing their ability to promote differentiation of adjacent satellite cells, through upregulation of the soluble factor follistatin. By contrast, FAPs from old mdx mice were resistant to HDACi-mediated inhibition of adipogenesis and constitutively repressed satellite cell-mediated formation of myotubes. We show that transplantation of FAPs from regenerating young muscles restored HDACi ability to increase myofibre size in old mdx mice. These results reveal that FAPs are key cellular determinants of disease progression in mdx mice and mediate a previously unappreciated stage-specific beneficial effect of HDACi in dystrophic muscles. PMID:23505062

  3. Fulminant phlegmonitis of the esophagus, stomach, and duodenum due to Bacillus thuringiensis.

    PubMed

    Matsumoto, Hisatake; Ogura, Hiroshi; Seki, Masafumi; Ohnishi, Mitsuo; Shimazu, Takeshi

    2015-03-28

    We report a case of phlegmonitis of the esophagus, stomach, and duodenum in patient in an immunocompromised state. Culture of gastric juice and blood yielded Bacillus thuringiensis. This case showed that even low-virulence bacilli can cause lethal gastrointestinal phlegmonous gastritis in conditions of immunodeficiency. PMID:25834344

  4. Different mechanisms of actions of genistein, quercetin on spontaneous contractions of rabbit duodenum.

    PubMed

    Santos-Fagundes, Diego; Grasa, Laura; Gonzalo, Sergio; Valero, Marta Sofía; Castro, Marta; Arruebo, María Pilar; Plaza, Miguel Ángel; Divina-Murillo, María

    2015-07-01

    Flavonoids are known to relax precontracted intestinal smooth muscle and delay intestinal transit or intestinal peristalsis. The aim of this study was to determine the effects of genistein and quercetin on spontaneous contractions of rabbit duodenum in vitro in an organ bath. Genistein and quercetin (0.1-10µM) reduced the amplitude of spontaneous contractions in the longitudinal and circular smooth muscle of rabbit duodenum, but they did not modify the frequency. Bay K8644 (L-type Ca2+ channel activator), apamin, charybdotoxin, and tetraetylammonium (K+ channel blockers) reverted the inhibition of amplitude of spontaneous contractions induced by genistein in longitudinal and circular smooth muscle. H-89 (protein kinase A inhibitor) antagonized the reduction of the amplitude of spontaneous contractions induced by quercetin in longitudinal and circular smooth muscle of duodenum, while 2,5-dideoxiadenosine (adenylyl cyclase inhibitor) reverted only the reduction of the amplitude in circular smooth muscle. In conclusion, genistein and quercetin reduce the spontaneous contractions in the duodenum by different mechanisms of actions. The effect of genistein would be mediated by Ca2+ and K+ channels, while the effect of quercetin would be mediated by cAMP and protein kinase A. PMID:26140633

  5. Adenocarcinoma of the third and fourth portions of the duodenum: The capsule endoscopy value

    PubMed Central

    Paquissi, Feliciano Chanana; Lima, Ana Henriqueta Filipe Bunga Pimentel; Lopes, Maria de Fátima do Nascimento Vieira; Diaz, Francisco Viamontes

    2015-01-01

    Primary adenocarcinoma of the small intestine occurs in over 50% of cases in the duodenum. However, its location in the third and fourth duodenal portions occurs rarely and is a diagnostic challenge. The aim of this work is to report an adenocarcinoma of the third and fourth duodenal portions, emphasizing its diagnostic difficulty and the value of video capsule endoscopy. A man, 40 years old, with no medical history, with abdominal discomfort and progressive fatigue, presented four months ago with one episode of moderate melena. The physical examination was normal, except for mucosal pallor. Blood tests were consistent with microcytic, hypochromic iron deficiency anemia with 7.8 g/dL hemoglobin. The upper and lower endoscopy were normal. Additional work-up with video capsule endoscopy showed a polypoid lesion involving the third and fourth portions of the duodenum. Biopsy showed a moderately differentiated adenocarcinoma. Abdominal computed tomography showed a wall thickening from the third duodenal portion to the proximal jejunum, without distant metastasis. The patient underwent segmental resection (distal duodenum and proximal jejunum) with duodenojejunostomy. The surgical specimen histology confirmed the biopsy diagnosis, with transmural infiltration, without nodal involvement. Conclusion: Adenocarcinoma of the third and fourth portions of the duodenum is difficult to diagnose and capsule endoscopy is of great value. PMID:26309371

  6. cDNA cloning and mRNA expression of canine pancreatic and duodenum homeobox 1 (Pdx-1).

    PubMed

    Takemitsu, Hiroshi; Yamamoto, Ichiro; Lee, Peter; Ohta, Taizo; Mori, Nobuko; Arai, Toshiro

    2012-10-01

    Pancreatic and duodenal homeobox 1 (Pdx-1) is a critical insulin transcription factor expressed by pancreatic β-cells, and is crucial in the early stage of pancreas development. Unfortunately, nothing concerning Pdx-1 in canine has been elucidated yet. In this study, full length canine Pdx-1 cDNA was cloned and it was 1498 bp in length, consisting of a 99 bp 5'-untranslated region (UTR), a 849 bp coding region, and a 550 bp 3'-UTR region. A deduced 282 amino acid sequence of canine PDX-1 displayed high overall sequence identity with human, bovine, and mouse PDX-1. qRT-PCR analysis revealed that a high level of Pdx1 mRNA expression is exists in the duodenum and pancreas of canines. In addition, functional canine insulin promoter-luciferase reporter constructs with various canine insulin promoter region fragments revealed that our Pdx-1 isolated cDNA sequence encodes for a functionally active PDX-1 protein. Significant promoter activity was observed within the -583 bp 5'-upstream region of canine insulin gene with Chinese hamster ovary cells. In addition, Pdx-1 appears to have a synergistic effect with beta cell transactivator 2 (BETA2) and V-maf avian musculoaponeurotic fibrosarcoma oncogene homolog A (MafA), which also have important roles in the activation of the insulin gene promoter. Our results confirm that similar to humans, Pdx1 plays an important role in expression of insulin gene in canines. PMID:22172402

  7. Linking cytoarchitecture to metabolism: sarcolemma-associated plectin affects glucose uptake by destabilizing microtubule networks in mdx myofibers

    PubMed Central

    2013-01-01

    Background Duchenne muscular dystrophy (DMD) is one of the most frequent forms of muscular disorders. It is caused by the absence of dystrophin, a core component of the sarcolemma-associated junctional complex that links the cytoskeleton to the extracellular matrix. We showed previously that plectin 1f (P1f), one of the major muscle-expressed isoforms of the cytoskeletal linker protein plectin, accumulates at the sarcolemma of DMD patients as well as of mdx mice, a widely studied animal model for DMD. Based on plectin’s dual role as structural protein and scaffolding platform for signaling molecules, we speculated that the dystrophic phenotype observed after loss of dystrophin was caused, at least to some extent, by excess plectin. Thus, we hypothesized that elimination of plectin expression in mdx skeletal muscle, while probably resulting in an overall more severe phenotype, may lead to a partial phenotype rescue. In particular, we wanted to assess whether excess sarcolemmal plectin contributes to the dysregulation of sugar metabolism in mdx myofibers. Methods We generated plectin/dystrophin double deficient (dKO) mice by breeding mdx with conditional striated muscle-restricted plectin knockout (cKO) mice. The phenotype of these mice was comparatively analyzed with that of mdx, cKO, and wild-type mice, focusing on structural integrity and dysregulation of glucose metabolism. Results We show that the accumulation of plectin at the sarcolemma of mdx muscle fibers hardly compensated for their loss of structural integrity. Instead, it led to an additional metabolic deficit by impairing glucose uptake. While dKO mice suffered from an overall more severe form of muscular dystrophy compared to mdx or plectin-deficient mice, sarcolemmal integrity as well as glucose uptake of their myofibers were restored to normal levels upon ablation of plectin. Furthermore, microtubule (MT) networks in intact dKO myofibers, including subsarcolemmal areas, were found to be more robust

  8. Treatment with a Nitric Oxide-Donating NSAID Alleviates Functional Muscle Ischemia in the Mouse Model of Duchenne Muscular Dystrophy

    PubMed Central

    Thomas, Gail D.; Ye, Jianfeng; De Nardi, Claudio; Monopoli, Angela; Ongini, Ennio; Victor, Ronald G.

    2012-01-01

    In patients with Duchenne muscular dystrophy (DMD) and the standard mdx mouse model of DMD, dystrophin deficiency causes loss of neuronal nitric oxide synthase (nNOSμ) from the sarcolemma, producing functional ischemia when the muscles are exercised. We asked if functional muscle ischemia would be eliminated and normal blood flow regulation restored by treatment with an exogenous nitric oxide (NO)-donating drug. Beginning at 8 weeks of age, mdx mice were fed a standard diet supplemented with 1% soybean oil alone or in combination with a low (15 mg/kg) or high (45 mg/kg) dose of HCT 1026, a NO-donating nonsteroidal anti-inflammatory agent which has previously been shown to slow disease progression in the mdx model. After 1 month of treatment, vasoconstrictor responses to intra-arterial norepinephrine (NE) were compared in resting and contracting hindlimbs. In untreated mdx mice, the usual effect of muscle contraction to attenuate NE-mediated vasoconstriction was impaired, resulting in functional ischemia: NE evoked similar decreases in femoral blood flow velocity and femoral vascular conductance (FVC) in the contracting compared to resting hindlimbs (ΔFVC contraction/ΔFVC rest = 0.88±0.03). NE-induced functional ischemia was unaffected by low dose HCT 1026 (ΔFVC ratio = 0.92±0.04; P>0.05 vs untreated), but was alleviated by the high dose of the drug (ΔFVC ratio = 0.22±0.03; P<0.05 vs untreated or low dose). The beneficial effect of high dose HCT 1026 was maintained with treatment up to 3 months. The effect of the NO-donating drug HCT 1026 to normalize blood flow regulation in contracting mdx mouse hindlimb muscles suggests a putative novel treatment for DMD. Further translational research is warranted. PMID:23139842

  9. A note on the duodenum of the West Indian manatee (Trichechus manatus), with emphasis on the duodenal glands.

    PubMed

    Reynolds, J E; Krause, W J

    1982-01-01

    The anatomy of the duodenum of the West Indian manatee has both macroscopic and microscopic features that are unusual when compared to other mammalian forms. Macroscopically, the voluminous duodenal ampulla and the paired duodenal diverticula are distinctive. The general microscopic structure of the manatee duodenum is not unusual, but the duodenal glands secrete an acid mucin (sialomucin). The cells of these glands appear to be intermediate between classical serous and mucous cell types. These cells also contain granules with regions of high peripheral electron density. The overall structure of the manatee duodenum most strongly resembles that of the dugong, another member of the order Sirenia. PMID:7148376

  10. Perforated duodenum--an unusual etiology of Fournier's disease: a case report.

    PubMed

    Lee, Yung-Chin; Yang, Wen-Horng; Wu, Wen-Jeng; Chou, Yii-Her; Huang, Chun-Hsiung

    2003-12-01

    Fournier's disease, a form of necrotizing fasciitis, is a rapidly progressing subcutaneous infection of the male genitalia. We report a case of Fournier's disease with the unusual etiology of a perforated duodenum. This patient suffered from progressive right scrotal swelling after percutaneous transhepatic cholangeal drainage. Scrotal exploration revealed a large abscess with muscle necrosis that had spread up to the right retroperitoneal space. Radiologic studies and second abdominal exploration documented the origin as a perforated duodenum. The pus distribution in this case suggested that the infection process differed from that in previous reports. In future cases of Fournier's disease involving previous abdominal events, we recommend that abdominal origins be carefully surveyed before scrotal exploration. PMID:14719562

  11. Helicobacter spp. in the saliva, stomach, duodenum and faeces of colony dogs.

    PubMed

    Ekman, E; Fredriksson, M; Trowald-Wigh, G

    2013-01-01

    The role of Helicobacter spp. infection in canine gastrointestinal disease is unclear and routes of transmission are of epidemiological and zoonotic importance. The aim of this study was to identify Helicobacter spp. in the saliva, stomach, duodenum and faeces of dogs using a multiplex PCR, and to evaluate any attendant histopathological changes. Helicobacter canis was the most common species detected in saliva and faeces and no correlation between the presence of Helicobacter spp. and histopathological changes in either the stomach or duodenum was observed. All dogs examined were co-infected with up to four species of the organism. This is the first time these bacteria have been studied at species level at multiple sites within the canine alimentary tract. PMID:22683393

  12. Stomach and duodenum ulcer: comparing the efficiency of three laser therapeutic techniques

    NASA Astrophysics Data System (ADS)

    Myslovich, L. V.

    2001-04-01

    An investigation was made of how effective various therapeutic techniques are in treating stomach and duodenum ulcers. The investigation was made on 105 patients (70 patients were affected by duodenum ulcer and 25 patients suffered from stomach ulcer). Three different complex laser therapeutic techniques were compared with each other and with a generally accepted drug treatment. It was found that the most pronounced therapeutic effect was observed in patients administered a complex laser technique that included drug therapy, the intravenous laser irradiation of blood, and the focal-segmental laser therapy. This complex laser therapy enabled ulcer scarring within 8 to 15 days after the beginning of the treatment (with the average scarring term of 9 days).

  13. Endoscopic Resection of a Pedunculated Brunner's Gland Hamartoma of the Duodenum.

    PubMed

    Iwamuro, Masaya; Tanaka, Takehiro; Ando, Satoko; Gotoda, Tatsuhiro; Kanzaki, Hiromitsu; Kawano, Seiji; Kawahara, Yoshiro; Okada, Hiroyuki

    2016-01-01

    A 68-year-old Japanese woman presented with a solitary pedunculated polyp in the duodenum. Endoscopic ultrasonography showed multiple cystic structures in the polyp. The polyp was successfully resected by endoscopic snare polypectomy and pathologically diagnosed as Brunner's gland hamartoma. Because hamartomatous components were not identified in the stalk of the polyp, we speculate that the stalk developed from traction of the normal duodenal mucosa. When a solitary, pedunculated polyp with cystic structure within the submucosa is found in the duodenum, Brunner's gland hamartoma should be considered in the differential diagnosis, despite the rarity of the disease. This case underscores the usefulness of endoscopic ultrasonography for the diagnosis of duodenal subepithelial tumors. PMID:27579190

  14. Endoscopic Resection of a Pedunculated Brunner's Gland Hamartoma of the Duodenum

    PubMed Central

    Tanaka, Takehiro; Ando, Satoko; Gotoda, Tatsuhiro; Kanzaki, Hiromitsu; Kawano, Seiji; Kawahara, Yoshiro; Okada, Hiroyuki

    2016-01-01

    A 68-year-old Japanese woman presented with a solitary pedunculated polyp in the duodenum. Endoscopic ultrasonography showed multiple cystic structures in the polyp. The polyp was successfully resected by endoscopic snare polypectomy and pathologically diagnosed as Brunner's gland hamartoma. Because hamartomatous components were not identified in the stalk of the polyp, we speculate that the stalk developed from traction of the normal duodenal mucosa. When a solitary, pedunculated polyp with cystic structure within the submucosa is found in the duodenum, Brunner's gland hamartoma should be considered in the differential diagnosis, despite the rarity of the disease. This case underscores the usefulness of endoscopic ultrasonography for the diagnosis of duodenal subepithelial tumors. PMID:27579190

  15. Ventilation during air breathing and in response to hypercapnia in 5 and 16 month-old mdx and C57 mice

    PubMed Central

    Gayraud, Jérome; Matécki, Stefan; Hnia, Karim; Mornet, Dominique; Préfaut, Christian; Mercier, Jacques; Michel, Alain; Ramonatxo, Michèle

    2007-01-01

    Previous studies have shown a blunted ventilatory response to hypercapnia in mdx mice older than 7 months. We test the hypothesis that in the mdx mice ventilatory response changes with age, concomitantly with the increased functional impairment of the respiratory muscles. We thus studied the ventilatory response to CO2 in 5 and 16 month-old mdx and C57BL10 mice (n = 8 for each group). Respiratory rate (RR), tidal volume (VT), and minute ventilation (VE) were measured, using whole-body plethysmography, during air breathing and in response to hypercapnia (3, 5 and 8% CO2). The ventilatory protocol was completed by histological analysis of the diaphragm and intercostals muscles. During air breathing, the 16 month-old mdx mice showed higher RR and, during hypercapnia (at 8% CO2 breathing), significantly lower RR (226 ± 26 vs. 270 ± 21 breaths/min) and VE (1.81 ± 0.35 vs. 3.96 ± 0.59 ml min−1 g−1)(P < 0.001) in comparison to C57BL10 controls. On the other hand, 5 month-old C57BL10 and mdx mice did not present any difference in their ventilatory response to air breathing and to hypercapnia. In conclusion, this study shows similar ventilation during air breathing and in response to hypercapnia in the 5 month-old mdx and control mice, in spite of significant pathological structural changes in the respiratory muscles of the mdx mice. However in the 16 month-old mdx mice we observed altered ventilation under air and blunted ventilation response to hypercapnia compared to age-matched control mice. Ventilatory response to hypercapnia thus changes with age in mdx mice, in line with the increased histological damage of their respiratory muscles. PMID:17431804

  16. Absorption-Enhancing Effect of Nitric Oxide on the Absorption of Hydrophobic Drugs in Rat Duodenum.

    PubMed

    Kishimoto, Hisanao; Miyazaki, Kaori; Takizawa, Yusuke; Shirasaka, Yoshiyuki; Inoue, Katsuhisa

    2016-02-01

    Nitric oxide (NO), an endogenous gas that plays a versatile role in the physiological system, has the ability to increase the intestinal absorption of water-soluble compounds through the paracellular route. However, it remains unclear whether NO can enhance the absorption of hydrophobic drugs through the transcellular route. In this study, we examined the absorption-enhancing effect of NO on intestinal permeability of hydrophobic drugs in rat intestine. The pretreatment of rat gastrointestinal sacs with NOC7, a NO-releasing reagent, significantly increased the permeation of griseofulvin from mucosa to serosa in the sacs prepared from the duodenum, but not in those prepared from the other regions such as jejunum, ileum, and colon. The absorption-enhancing effect of NOC7 on the duodenal permeation varied depending on the hydrophobicity of the drugs used. Furthermore, NOC7 treatment was found to be apparently ineffective on the griseofulvin permeation in the duodenum pretreated with dithiothreitol (DTT) that was used as a mucus remover, even though the permeation was increased by pretreatment with DTT alone. These results suggest that NO increases the absorption of hydrophobic drugs through the transcellular route in the duodenum by modulating the mucus layer function. PMID:26458075

  17. Extramedullary plasmocytoma associated with a massive deposit of amyloid in the duodenum.

    PubMed

    Carneiro, Fabiana Pirani; Sobreira, Maria Nazareth Machado; Maia, Lívia Bravo; Sartorelli, Alesso Cervantes; Franceschi, Luiz Eduardo de Almeida Prado; Brandão, Mauro Brito; Calaça, Bárbara Wosnjuk; Lustosa, Fernando Silva; Lopes, João Vieira

    2009-07-28

    We report a rare case of extramedullary plasmocytoma associated with a massive deposit of amyloid in the duodenum. A 72-year-old Japanese man was admitted to our hospital presenting with a 3-mo history of epigastric pain, vomiting and weight loss. On computed tomography (CT) a wall thickening of the fourth part of the duodenum was observed. Multiple biopsies obtained from the lesion showed infiltration of plasma cells and lymphocytes, but they were not conclusive. The patient underwent resection of the lesion and, on histopathological examination, the lesion consisted of a dense and diffuse infiltrate of plasma cells and a few admixed lymphocytes with reactive follicles extending to the muscular propria. An extensive deposition of amyloid was also observed. Immunohistochemical stains revealed that a few plasmacytoid cells showed lambda light chain staining, though most were kappa light chain positive. These cells also were positive for CD138 and CD56 but negative for CD20 and CD79. The findings were consistent with extramedullary plasmocytoma associated with a massive deposit of amyloid in duodenum. A subsequent workup for multiple myeloma was completely negative. The patient showed no signs of local recurrence or dissemination of the disease after 12 mo follow-up. Because of the association of plasmocytoma and amyloidosis, the patient must be followed up because of the possible systemic involvement of the neoplasm and amyloidosis in future. PMID:19630116

  18. Effects of irradiating adult mdx mice before full-length dystrophin cDNA transfer on host anti-dystrophin immunity.

    PubMed

    Eghtesad, S; Zheng, H; Nakai, H; Epperly, M W; Clemens, P R

    2010-09-01

    Duchenne muscular dystrophy is a fatal, genetic disorder in which dystrophin-deficient muscle progressively degenerates, for which dystrophin gene transfer could provide effective treatment. The host immune response to dystrophin, however, is an obstacle to therapeutic gene expression. Understanding the dystrophin-induced host immune response will facilitate the discovery of strategies to prolong expression of recombinant dystrophin in dystrophic muscle. Using whole-body irradiation of the dystrophic mdx mouse before gene transfer, we temporally removed the immune system; a 600 rad dose removed peripheral immune cells, which were restored by self-reconstitution, and a 900 rad dose removed central and peripheral immune cells, which were restored by adoptive transfer of bone marrow from a syngeneic, dystrophin-normal donor. The anti-dystrophin humoral response was delayed and dystrophin expression was partially preserved in irradiated, vector-treated mice. Nonirradiated, vector-treated control mice lost muscle dystrophin expression completely, had an earlier anti-dystrophin humoral response and demonstrated muscle fibers focally surrounded with T cells. We conclude that dystrophin gene transfer induced anti-dystrophin humoral immunity and cell-mediated responses that were significantly diminished and delayed by temporal removal of the host central or peripheral immune cells. Furthermore, manipulation of central immunity altered the pattern of regulatory T cells in muscle. PMID:20827278

  19. Long-Term Efficacy of Systemic Multiexon Skipping Targeting Dystrophin Exons 45–55 With a Cocktail of Vivo-Morpholinos in Mdx52 Mice

    PubMed Central

    Echigoya, Yusuke; Aoki, Yoshitsugu; Miskew, Bailey; Panesar, Dharminder; Touznik, Aleksander; Nagata, Tetsuya; Tanihata, Jun; Nakamura, Akinori; Nagaraju, Kanneboyina; Yokota, Toshifumi

    2015-01-01

    Antisense-mediated exon skipping, which can restore the reading frame, is a most promising therapeutic approach for Duchenne muscular dystrophy. Remaining challenges include the limited applicability to patients and unclear function of truncated dystrophin proteins. Multiexon skipping targeting exons 45–55 at the mutation hotspot of the dystrophin gene could overcome both of these challenges. Previously, we described the feasibility of exons 45–55 skipping with a cocktail of Vivo-Morpholinos in vivo; however, the long-term efficacy and safety of Vivo-Morpholinos remains to be determined. In this study, we examined the efficacy and toxicity of exons 45–55 skipping by intravenous injections of 6 mg/kg 10-Vivo-Morpholino cocktail (0.6 mg/kg each vPMO) every 2 weeks for 18 weeks to dystrophic exon-52 knockout (mdx52) mice. Systemic skipping of the entire exons 45–55 region was induced, and the Western blot analysis exhibited the restoration of 5–27% of normal levels of dystrophin protein in skeletal muscles, accompanied by improvements in histopathology and muscle strength. No obvious immune response and renal and hepatic toxicity were detected at the end-point of the treatment. We demonstrate our new regimen with the 10-Vivo-Morpholino cocktail is effective and safe for long-term repeated systemic administration in the dystrophic mouse model. PMID:25647512

  20. Long-term efficacy of systemic multiexon skipping targeting dystrophin exons 45-55 with a cocktail of vivo-morpholinos in mdx52 mice.

    PubMed

    Echigoya, Yusuke; Aoki, Yoshitsugu; Miskew, Bailey; Panesar, Dharminder; Touznik, Aleksander; Nagata, Tetsuya; Tanihata, Jun; Nakamura, Akinori; Nagaraju, Kanneboyina; Yokota, Toshifumi

    2015-01-01

    Antisense-mediated exon skipping, which can restore the reading frame, is a most promising therapeutic approach for Duchenne muscular dystrophy. Remaining challenges include the limited applicability to patients and unclear function of truncated dystrophin proteins. Multiexon skipping targeting exons 45-55 at the mutation hotspot of the dystrophin gene could overcome both of these challenges. Previously, we described the feasibility of exons 45-55 skipping with a cocktail of Vivo-Morpholinos in vivo; however, the long-term efficacy and safety of Vivo-Morpholinos remains to be determined. In this study, we examined the efficacy and toxicity of exons 45-55 skipping by intravenous injections of 6 mg/kg 10-Vivo-Morpholino cocktail (0.6 mg/kg each vPMO) every 2 weeks for 18 weeks to dystrophic exon-52 knockout (mdx52) mice. Systemic skipping of the entire exons 45-55 region was induced, and the Western blot analysis exhibited the restoration of 5-27% of normal levels of dystrophin protein in skeletal muscles, accompanied by improvements in histopathology and muscle strength. No obvious immune response and renal and hepatic toxicity were detected at the end-point of the treatment. We demonstrate our new regimen with the 10-Vivo-Morpholino cocktail is effective and safe for long-term repeated systemic administration in the dystrophic mouse model. PMID:25647512

  1. Adeno-associated virus vector-mediated minidystrophin gene therapy improves dystrophic muscle contractile function in mdx mice.

    PubMed

    Watchko, Jon; O'Day, Terry; Wang, Bing; Zhou, Liqiao; Tang, Ying; Li, Juan; Xiao, Xiao

    2002-08-10

    Duchenne muscular dystrophy (DMD) is the most common disabling and lethal genetic muscle disorder, afflicting 1 of every 3500 males. Patients with DMD experience progressive muscle degeneration and weakness and succumb to respiratory or cardiac failure by their early twenties. No treatment is currently available for DMD. Mutations in the dystrophin gene result in lack of a functional dystrophin protein in striated muscle, which induces instability in the muscle cell membrane leading to persistent muscle injury after contraction. We have previously created novel minidystrophin genes and demonstrated that adeno-associated virus (AAV)-mediated intramuscular delivery of the minigenes effectively ameliorated mdx dystrophic histopathology and led to normal cell membrane integrity for more than 1 year. In this paper, we investigated whether AAV-minidystrophin could also improve mdx muscle contractile function. Two-month-old adult male mdx mice, with established muscular dystrophy, were given a single-dose injection of an AAV-minidystrophin vector in the tibialis anterior (TA) muscle of one leg, with the untreated contralateral leg used as a control. The treated TA muscle showed both (1) a significant increase in isometric force generation and (2) a significant increase in resistance to lengthening activation-induced muscle force decrements. We conclude that AAV-minidystrophin gene treatment is effective in improving mdx muscle contractile function. PMID:12215266

  2. Evolution of pathological changes in the gastrocnemius of the mdx mice correlate with utrophin and beta-dystroglycan expression.

    PubMed

    Roma, Josep; Munell, Francina; Fargas, Arnau; Roig, Manuel

    2004-11-01

    Utrophin can function in muscle as a substitute for dystrophin and its over-expression has been used successfully to ameliorate mdx muscle pathology. Despite of this fact, there are no detailed studies on the expression of endogenous skeletal muscle utrophin- and dystrophin-associated glycoproteins throughout the life span of mdx mice. We have monitored, sequentially, the expression of matrix metalloproteinase-9 (MMP-9), myosin heavy chain, utrophin and beta-dystroglycan, as well as the mRNA expression of utrophin and of structurally related proteins, in mdx and control mice. We found an inverse relationship between concentration of muscle utrophin and abundance of groups of degenerative-regenerative fibers and of MMP-9 expression. There was also temporal correlation between the decline of utrophin at 15 days of age and the onset of muscle necrosis. Conversely, reappearance of utrophin, with a peak around 2 months of age, was followed by a progressive decline of necrosis. A lineal correlation between utrophin and beta-dystroglycan levels, not seen in controls, indicates that improvement of mdx is due to utrophin binding to dystrophin-associated glycoproteins. Utrophin and other structurally related protein transcripts were not up-regulated, suggesting a post-transcriptional regulation for utrophin in skeletal muscle. PMID:15365724

  3. Fructose Promotes Uptake and Activity of Oligonucleotides With Different Chemistries in a Context-dependent Manner in mdx Mice.

    PubMed

    Cao, Limin; Han, Gang; Lin, Caorui; Gu, Ben; Gao, Xianjun; Moulton, Hong M; Seow, Yiqi; Yin, HaiFang

    2016-01-01

    Antisense oligonucleotide (AO)-mediated exon-skipping therapeutics shows great promise in correcting frame-disrupting mutations in the DMD gene for Duchenne muscular dystrophy. However, insufficient systemic delivery limits clinical adoption. Previously, we showed that a glucose/fructose mixture augmented AO delivery to muscle in mdx mice. Here, we evaluated if fructose alone could enhance the activities of AOs with different chemistries in mdx mice. The results demonstrated that fructose improved the potency of AOs tested with the greatest effect on phosphorodiamidate morpholino oligomer (PMO), resulted in a 4.25-fold increase in the number of dystrophin-positive fibres, compared to PMO in saline in mdx mice. Systemic injection of lissamine-labeled PMO with fructose at 25 mg/kg led to increased uptake and elevated dystrophin expression in peripheral muscles, compared to PMO in saline, suggesting that fructose potentiates PMO by enhancing uptake. Repeated intravenous administration of PMO in fructose at 50 mg/kg/week for 3 weeks and 50 mg/kg/month for 5 months restored up to 20% of wild-type dystrophin levels in skeletal muscles with improved functions without detectable toxicity, compared to untreated mdx controls. Collectively, we show that fructose can potentiate AOs of different chemistries in vivo although the effect diminished over repeated administration. PMID:27351681

  4. Myostatin/activin blocking combined with exercise reconditions skeletal muscle expression profile of mdx mice.

    PubMed

    Kainulainen, Heikki; Papaioannou, Konstantinos G; Silvennoinen, Mika; Autio, Reija; Saarela, Janne; Oliveira, Bernardo M; Nyqvist, Miro; Pasternack, Arja; 't Hoen, Peter A C; Kujala, Urho M; Ritvos, Olli; Hulmi, Juha J

    2015-01-01

    Duchenne muscular dystrophy is characterized by muscle wasting and decreased aerobic metabolism. Exercise and blocking of myostatin/activin signaling may independently or combined counteract muscle wasting and dystrophies. The effects of myostatin/activin blocking using soluble activin receptor-Fc (sActRIIB-Fc) administration and wheel running were tested alone or in combination for 7 weeks in dystrophic mdx mice. Expression microarray analysis revealed decreased aerobic metabolism in the gastrocnemius muscle of mdx mice compared to healthy mice. This was not due to reduced home-cage physical activity, and was further downregulated upon sActRIIB-Fc treatment in enlarged muscles. However, exercise activated pathways of aerobic metabolism and counteracted the negative effects of sActRIIB-Fc. Exercise and sActRIIB-Fc synergistically increased expression of major urinary protein, but exercise blocked sActRIIB-Fc induced phosphorylation of STAT5 in gastrocnemius muscle. In conclusion, exercise alone or in combination with myostatin/activin blocking corrects aerobic gene expression profiles of dystrophic muscle toward healthy wild type mice profiles. PMID:25304272

  5. Polyethylenimine-modified pluronics (PCMs) improve morpholino oligomer delivery in cell culture and dystrophic mdx mice.

    PubMed

    Wang, Mingxing; Wu, Bo; Lu, Peijuan; Cloer, Caryn; Tucker, Jay D; Lu, Qilong

    2013-01-01

    We investigated a series of small-sized polyethylenimine (PEI, 0.8/1.2 k)-conjugated pluronic copolymers (PCMs) for their potential to enhance delivery of an antisense phosphorodiamidate morpholino oligomer (PMO) in vitro and in dystrophic mdx mice. PCM polymers containing pluronics of molecular weight (Mw) ranging 2-6 k, with hydrophilic-lipophilic balance (HLB) 7-23, significantly enhanced PMO-induced exon-skipping in a green fluorescent protein (GFP) reporter-based myoblast culture system. Application of optimized formulations of PCMs with PMO targeted to dystrophin exon 23 demonstrated a significant increase in exon-skipping efficiency in dystrophic mdx mice. Consistent with our observations in vitro, optimization of molecular size and the HLB of pluronics are important factors for PCMs to achieve enhanced PMO delivery in vivo. Observed cytotoxicity of the PCMs was lower than Endo-porter and PEI 25 k. Tissue toxicity of PCMs in muscle was not clearly detected with the concentrations used, indicating the potential of the PCMs as effective and safe PMO carriers for treating diseases such as muscular dystrophy. PMID:23164938

  6. Short communication: Difructose anhydride III promotes calcium absorption from the duodenum in cattle.

    PubMed

    Teramura, M; Nakai, T; Itoh, M; Sato, T; Ohtani, M; Kawashima, C; Hanada, M

    2015-04-01

    Difructose anhydride (DFA) III promotes the intestinal absorption of calcium via a paracellular pathway in rats. In dairy cows, DFA III reaches the duodenum without being degraded by ruminal bacteria and hence could be used to control hypocalcemia. The aims of the present study were to investigate the percentage of DFA III that appears in the duodenum of cows and to determine the effect of DFA III on calcium absorption from duodenal fluid. The first experiment was performed in 3 ruminally and duodenally cannulated dry Holstein cows in a 3 × 3 Latin square design. Each experimental period lasted 7 d. On the first day, the cows were ruminally fed one of the following treatments: 0 (DFA0), 50 (DFA50), or 100 (DFA100) g/d of DFA III, using cobalt-EDTA as a liquid phase marker. Difructose anhydride III was detected in duodenal fluid 1 h after feeding, and its concentration peaked 4 h after feeding, in a dose-dependent manner. The percentages of DFA III that appeared in the duodenum after the DFA50 and DFA100 treatments were 69.1 ± 7.0% and 67.9 ± 5.6%, respectively. The second experiment used the everted duodenal sacs of cattle (n = 7 in each group). Sacs were incubated in artificial mucosal fluid containing 1 mM DFA III or no DFA III (control) for 60 min with 100% O2 in a water bath at 37 °C. After incubation, the calcium concentration of the artificial serosal fluid in the everted sacs was measured. Calcium absorption was higher in the DFA III-treated group than in the control group (803 ± 161 and 456 ± 74 nmol/cm of sac, respectively). The above results demonstrate that approximately 70% of administered DFA III reached the duodenum of cows intact. Moreover, similar to its effects on calcium absorption in rats, DFA III promoted calcium absorption via a paracellular pathway in the duodenum of cows. PMID:25648815

  7. Andrographolide attenuates skeletal muscle dystrophy in mdx mice and increases efficiency of cell therapy by reducing fibrosis

    PubMed Central

    2014-01-01

    Background Duchenne muscular dystrophy (DMD) is characterized by the absence of the cytoskeletal protein dystrophin, muscle wasting, increased transforming growth factor type beta (TGF-β) signaling, and fibrosis. At the present time, the only clinically validated treatments for DMD are glucocorticoids. These drugs prolong muscle strength and ambulation of patients for a short term only and have severe adverse effects. Andrographolide, a bicyclic diterpenoid lactone, has traditionally been used for the treatment of colds, fever, laryngitis, and other infections with no or minimal side effects. We determined whether andrographolide treatment of mdx mice, an animal model for DMD, affects muscle damage, physiology, fibrosis, and efficiency of cell therapy. Methods mdx mice were treated with andrographolide for three months and skeletal muscle histology, creatine kinase activity, and permeability of muscle fibers were evaluated. Fibrosis and TGF-β signaling were evaluated by indirect immunofluorescence and Western blot analyses. Muscle strength was determined in isolated skeletal muscles and by a running test. Efficiency of cell therapy was determined by grafting isolated skeletal muscle satellite cells onto the tibialis anterior of mdx mice. Results mdx mice treated with andrographolide exhibited less severe muscular dystrophy than untreated dystrophic mice. They performed better in an exercise endurance test and had improved muscle strength in isolated muscles, reduced skeletal muscle impairment, diminished fibrosis and a significant reduction in TGF-β signaling. Moreover, andrographolide treatment of mdx mice improved grafting efficiency upon intramuscular injection of dystrophin-positive satellite cells. Conclusions These results suggest that andrographolide could be used to improve quality of life in individuals with DMD. PMID:24655808

  8. The Effects of Experimental Sleep Apnea on Cardiac and Respiratory Functions in 6 and 18 Month Old Dystrophic (mdx) Mice.

    PubMed

    Chaudhari, Milind R; Fallavollita, James A; Farkas, Gaspar A

    2016-01-01

    Duchenne muscular dystrophy (DMD) is a fatal disease where over 90% of patients succumb to respiratory or cardiac failure. Sleep apnea and sleep disordered breathing (SDB) are noted in a plurality of DMD patients, and the resulting nocturnal episodic hypoxia (EH) cannot be ruled out as a contributing factor to cardiac and respiratory dysfunction. In this study, we investigated the impact of long-term episodic hypoxia, which mimics the cyclic hypoxia seen in sleep apnea, on cardiac and respiratory function in a murine model of DMD (mdx mice). Since the severity and prevalence of sleep apnea in DMD increases with age, we studied the impact of EH on young (6-month) and on older (18-month) mdx mice. Mice were either exposed for 12 weeks to EH (8 hours/day, 5 days/week) or to room air. We noted a significant increase in left ventricular (LV) dilatation (transthoracic echocardiography) on EH exposure in both age groups, but reduced LV contractility was seen only in 6-month old mice. With EH exposure, an increased fibrosis (hydroxyproline) was noted in both cardiac and diaphragm muscle in 18-month but not 6-month old mice. No significant change in relative diaphragm strength (in-vitro) was noted on EH exposure in 18-month old mice. In contrast, EH exposed 6-month old mice showed a significant increase in relative diaphragm strength. EH exposure did not result in any significant change in ventilatory parameters (barometric plethysmography) in awake 6-month old mdx mice. In contrast, 18-month old mdx mice showed considerable ventilatory dysfunction, consistent with reduced ventilatory reserve. Our findings highlight that sleep apnea impacts respiratory and cardiac function in muscular dystrophy, and that EH can have divergent effects on both systems. To our knowledge, this is the first comprehensive study to investigate the impact of EH on cardiac and respiratory function in mdx mice. PMID:26808526

  9. The Effects of Experimental Sleep Apnea on Cardiac and Respiratory Functions in 6 and 18 Month Old Dystrophic (mdx) Mice

    PubMed Central

    Fallavollita, James A.

    2016-01-01

    Duchenne muscular dystrophy (DMD) is a fatal disease where over 90% of patients succumb to respiratory or cardiac failure. Sleep apnea and sleep disordered breathing (SDB) are noted in a plurality of DMD patients, and the resulting nocturnal episodic hypoxia (EH) cannot be ruled out as a contributing factor to cardiac and respiratory dysfunction. In this study, we investigated the impact of long-term episodic hypoxia, which mimics the cyclic hypoxia seen in sleep apnea, on cardiac and respiratory function in a murine model of DMD (mdx mice). Since the severity and prevalence of sleep apnea in DMD increases with age, we studied the impact of EH on young (6-month) and on older (18-month) mdx mice. Mice were either exposed for 12 weeks to EH (8 hours/day, 5 days/week) or to room air. We noted a significant increase in left ventricular (LV) dilatation (transthoracic echocardiography) on EH exposure in both age groups, but reduced LV contractility was seen only in 6-month old mice. With EH exposure, an increased fibrosis (hydroxyproline) was noted in both cardiac and diaphragm muscle in 18-month but not 6-month old mice. No significant change in relative diaphragm strength (in-vitro) was noted on EH exposure in 18-month old mice. In contrast, EH exposed 6-month old mice showed a significant increase in relative diaphragm strength. EH exposure did not result in any significant change in ventilatory parameters (barometric plethysmography) in awake 6-month old mdx mice. In contrast, 18-month old mdx mice showed considerable ventilatory dysfunction, consistent with reduced ventilatory reserve. Our findings highlight that sleep apnea impacts respiratory and cardiac function in muscular dystrophy, and that EH can have divergent effects on both systems. To our knowledge, this is the first comprehensive study to investigate the impact of EH on cardiac and respiratory function in mdx mice. PMID:26808526

  10. [WEAK COMBINED MAGNETIC FIELDS ADJUSTED TO THE PARAMETRIC RESONANCE FOR Ca2+ INTENSIFY DYSTROPHIN SYNTHESIS IN MDX MICE SKELETAL MUSCLES AFTER CELL THERAPY].

    PubMed

    Sokolova, A V; Sokolov, G V; Mikhailov, V M

    2016-01-01

    The mdx mice are an X-linked myopathic mutants, an animal model for human Duchenne muscular dystrophy (DMD). Mdx mice muscles are characterized by high level of striated muscle fibers (SMF) death followed by regeneration. As a result most SMFs of mdx mice have centrally located nuclei. The possibility of using stem cells therapy for the correction of DMD is actively being studied. One of the approaches to the usage of bone marrow stem cells for cellular therapy of DMD is the replacement of bone marrow after irradiation by X-rays. This method however does not give significant increase of dystrophin synthesis in mdx mice muscles fibers. We have tried to affect the mice after bone marrow transplantation by weak combined magnetic fields adjusted to the parametric resonance for Ca2+(Ca(2+)-MF) based on the data that the weak combined magnetic fields influence on tissues regeneration. We observed a significant increase in the proportion of dystrophin-positive SMFs in group of mdx mice radiation chimera 5 Gy and 3 Gy which was additionally exposed in Ca(2+)-MF in comparison with the control mdx mice and the group of mdx mice radiation chimera 5 Gy and 3 Gy which was kept in terrestrial magnetic field 2 months after chimera preparation--up to 15.8 and 18.3%, respectively. Also, there was an accumulation of SMFs without central nuclei. These data indicate a significanly increased efficacy of cell therapy in the case of additional exposition in Ca(2+)-MF. Thus, the efficiency of bone marrow transplantation mdx mice after both in doses 3 and 5 Gy was considerably enhanced by additional exposition to Ca(2+)-MF. Apparently, such magnetic field can intensify functioning of donor's nuclei which had been incorporated into muscle fibers. PMID:27228662

  11. Effects of Mechanical Overloading on the Properties of Soleus Muscle Fibers, with or without Damage in MDX and Wild Type Mice

    NASA Astrophysics Data System (ADS)

    Terada, Masahiro; Kawano, Fuminori; Ohira, Takashi; Oke, Yoshihiko; Nakai, Naoya; Ohira, Yoshinobu

    2008-06-01

    Effects of mechanical overloading on the characteristics of regenerating or not-regenerating soleus muscle fibers were studied. The muscle fibers of mdx mice were characterized by the localization of myonuclei. Muscle damage was also induced in wild type (WT) mice by injection of cardiotoxin (CTX) into soleus muscle. Overloading was applied for 14 days to the left soleus muscle in mdx and intact and CTX-injected WT mice by removing the distal tendons of plantaris and gastrocnemius muscles. The contralateral muscle served as the normal control. These animals were then allowed ambulation recovery in the cage. Central myonuclei were noted in many fibers of mdx and CTX-injected mice with or without overloading. In general, the fibers with central nuclei were considered as regenerating fibers. The fibers with more central nuclei were increased in mdx mice, but the fibers with more peripheral nuclei were increased in CTX-injected WT mice by overloading. The muscle satellite cells, neuromuscular junctions (NMJ), and myonuclei were stained. Most of the properties, such as number of myonuclei and satellite cells, size of NMJ, and fiber length, were not influenced by mechanical overloading in all mice. Approximately 0.6% branched fibers were seen in the intact soleus of mdx mice, although these fibers were not detected in WT mice. However, the percentage of these fibers was increased by overloading especially in mdx mice (~50% vs. ~2.5% in WT). In CTX-injected WT mice, these fibers were ~15% with or without overloading. The fiber cross sectional area in normal WT, but not in mdx and CTX-injected WT mice, was increased by overloading (p<0.05). These results suggested that the functional overload induced muscle damage in mdx mice, but promoted the regeneration in CTX-injected WT mice.

  12. Intraluminal superior vena cava metastasis from adenosquamous carcinoma of the duodenum: A case report

    PubMed Central

    TAKAYOSHI, KOTOE; ARIYAMA, HIROSHI; TAMURA, SHINGO; YODA, SHUNSUKE; ARITA, TAKESHI; YAMAGUCHI, TOSHIHIRO; OZONO, KEIGO; YAMAMOTO, HIDETAKA; INADOMI, KYOKO; KUMAGAI, HOZUMI; TANAKA, MAMORU; OKUMURA, YUTA; SAGARA, KOSUKE; NIO, KENTA; NAKANO, MICHITAKA; ARITA, SHUJI; KUSABA, HITOSHI; ODASHIRO, KEITA; ODA, YOSHINAO; AKASHI, KOICHI; BABA, EISHI

    2016-01-01

    In 2013, a 76-year-old male with a cardiac pacemaker was diagnosed with adenosquamous carcinoma of the duodenum. Subsequently, a pancreatoduodenectomy and lymph node dissection were performed, and 12 cycles of adjuvant chemotherapy (modified FOLFOX6 regimen), which consisted of fluorouracil, leucovorin and oxaliplatin, were administered via a central venous catheter. At 5 months after the completion of adjuvant chemotherapy, the patient experienced the sudden onset of severe pain at the back right of the ear, edema of the right side of the face and right jugular vein dilatation. Computed tomography (CT) revealed filling defects in the superior vena cava (SVC) and right brachiocephalic vein, indicating catheter-induced venous thrombosis. Although the catheter was removed and anti-coagulation therapy, aspiration of the thrombosis and ballooning dilatation were performed immediately, the patient's symptoms were not ameliorated. Notably, histological examination following thrombus aspiration revealed metastatic cancer cells, and fluorodeoxyglucose-positron emission tomography/CT identified metabolically active nodules in the SVC at locations consistent with the initial duodenal tumors detected by CT and in the first thoracic vertebrae. The tumor thrombus rapidly increased in size and resulted in worsening dyspnea. Subsequently, radiotherapy was performed, followed by chemotherapy, which relieved the systemic symptoms and suppressed the tumor growth. Adenosquamous carcinoma of the duodenum is extremely rare, and to the best of our knowledge, intraluminal SVC metastasis as a result of adenosquamous carcinoma of the duodenum has not been reported previously. The placement of a cardiac pacemaker, central venous catheter and tumor cells possessing high metastatic potential are hypothesized to have contributed to this rare case of metastasis. PMID:26870254

  13. Kinetics of acute infection with Toxoplasma gondii and histopathological changes in the duodenum of rats.

    PubMed

    Trevizan, Aline Rosa; Vicentino-Vieira, Suellen Laís; da Silva Watanabe, Paulo; Góis, Marcelo Biondaro; de Melo, Gessilda de Alcântara Nogueira; Garcia, João Luiz; José de Almeida Araújo, Eduardo; Sant'Ana, Débora de Mello Gonçales

    2016-06-01

    Toxoplasma gondii crosses the intestinal barrier to spread into the body. We investigate the intestinal wall and epithelial cells of the duodenum of rats infected with T. gondii during different time points of acute infection. Male Wistar rats, 60 days of age, were assigned into groups that were orally inoculated with 5000 sporulated oocysts T. gondii for 6 h (G6), 12 h (G12), 24 h (G24), 48 h (G48), 72 h (G72), 7 days (G7d), and 10 days (G10d). The control group (CG) received saline. The rats were killed and the duodenum was processed to obtain histological sections stained with hematoxylin and eosin, Periodic Acid Schiff, and Alcian blue (pH 2.5 and 1.0). Morphometry was performed on the layers of the intestinal wall and enterocytes, and the number of goblet cells and intraepithelial lymphocytes was counted. The data were compared by ANOVA considering 5% as level of significance. The infection provoked an increase in the width of villi and crypts; decrease in enterocyte height; increase in the smaller-diameter and reduction in the larger-diameter of the enterocytes nuclei, increased number of goblet cells secreting neutral (G6, G12 and G7d) and acidic (G7d and G10d) mucus, and increase in the number of intraepithelial lymphocytes (G48). The infected groups showed atrophy of the submucosa and muscular layers and the total wall. Acute infection with T. gondii caused morphological changes in the intestinal wall and epithelial cells of the duodenum in rats. PMID:26993084

  14. Villous Adenocarcinoma of the Duodenum Invading the Ampulla of Vater: Case Report and Review of Literature

    PubMed Central

    Cardi, Francesco; Migliore, Marcello; Romeo, Gaetano

    1996-01-01

    We report a case of villous adenocarcinoma of duodenum arising from the ampulla of Vater with a review of the literature. Although preoperative endoscopic biopsies were performed, no malignancy was identified. Because of the pathological uncertainty we decided to perform a pylorus-preserving pancreatoduodenectomy. Microscopic examination demonstrated glandular dysplasia with aspects of villous adenoma and well differentiated adenocarcinoma. We conclude that both in malignant cases and in cases with uncertain diagnosis a pylorus-preserving pancreatoduodenectomy is the best surgical treatment because it results in better 5 year survival. PMID:9184865

  15. Primary Follicular Lymphoma of the Duodenum with Erosions as Atypical Macroscopic Features

    PubMed Central

    Takeuchi, Keiko; Iwamuro, Masaya; Imagawa, Atsushi; Kubota, Yoshitsugu; Miyatani, Katsuya; Takata, Katsuyoshi; Okada, Hiroyuki

    2012-01-01

    A 52-year-old Japanese woman who was eventually diagnosed with primary follicular lymphoma of the duodenum showed atypical endoscopic features, namely, erosions with peripheral whitish edematous mucosa. Initial biopsy specimens taken from the erosions revealed insufficient numbers of lymphoma cells for histological diagnosis. Subsequent biopsy specimens from the peripheral mucosa containing the whitish enlarged villi showed infiltration of the lymphoma cells forming lymphoid follicles, which led us to the appropriate diagnosis. This case indicates that endoscopists should take biopsy samples from the peripheral mucosa with whitish enlarged villi rather than erosions in the rare instances that erosions appear as the main macroscopic feature of intestinal follicular lymphoma. PMID:22690224

  16. Prevention of exercised induced cardiomyopathy following Pip-PMO treatment in dystrophic mdx mice

    PubMed Central

    Betts, Corinne A.; Saleh, Amer F.; Carr, Carolyn A.; Hammond, Suzan M.; Coenen-Stass, Anna M. L.; Godfrey, Caroline; McClorey, Graham; Varela, Miguel A.; Roberts, Thomas C.; Clarke, Kieran; Gait, Michael J.; Wood, Matthew J. A.

    2015-01-01

    Duchenne muscular dystrophy (DMD) is a fatal neuromuscular disorder caused by mutations in the Dmd gene. In addition to skeletal muscle wasting, DMD patients develop cardiomyopathy, which significantly contributes to mortality. Antisense oligonucleotides (AOs) are a promising DMD therapy, restoring functional dystrophin protein by exon skipping. However, a major limitation with current AOs is the absence of dystrophin correction in heart. Pip peptide-AOs demonstrate high activity in cardiac muscle. To determine their therapeutic value, dystrophic mdx mice were subject to forced exercise to model the DMD cardiac phenotype. Repeated peptide-AO treatments resulted in high levels of cardiac dystrophin protein, which prevented the exercised induced progression of cardiomyopathy, normalising heart size as well as stabilising other cardiac parameters. Treated mice also exhibited significantly reduced cardiac fibrosis and improved sarcolemmal integrity. This work demonstrates that high levels of cardiac dystrophin restored by Pip peptide-AOs prevents further deterioration of cardiomyopathy and pathology following exercise in dystrophic DMD mice. PMID:25758104

  17. Prevention of exercised induced cardiomyopathy following Pip-PMO treatment in dystrophic mdx mice.

    PubMed

    Betts, Corinne A; Saleh, Amer F; Carr, Carolyn A; Hammond, Suzan M; Coenen-Stass, Anna M L; Godfrey, Caroline; McClorey, Graham; Varela, Miguel A; Roberts, Thomas C; Clarke, Kieran; Gait, Michael J; Wood, Matthew J A

    2015-01-01

    Duchenne muscular dystrophy (DMD) is a fatal neuromuscular disorder caused by mutations in the Dmd gene. In addition to skeletal muscle wasting, DMD patients develop cardiomyopathy, which significantly contributes to mortality. Antisense oligonucleotides (AOs) are a promising DMD therapy, restoring functional dystrophin protein by exon skipping. However, a major limitation with current AOs is the absence of dystrophin correction in heart. Pip peptide-AOs demonstrate high activity in cardiac muscle. To determine their therapeutic value, dystrophic mdx mice were subject to forced exercise to model the DMD cardiac phenotype. Repeated peptide-AO treatments resulted in high levels of cardiac dystrophin protein, which prevented the exercised induced progression of cardiomyopathy, normalising heart size as well as stabilising other cardiac parameters. Treated mice also exhibited significantly reduced cardiac fibrosis and improved sarcolemmal integrity. This work demonstrates that high levels of cardiac dystrophin restored by Pip peptide-AOs prevents further deterioration of cardiomyopathy and pathology following exercise in dystrophic DMD mice. PMID:25758104

  18. Toward Integrated Molecular Diagnostic System (iMDx): Principles and Applications

    PubMed Central

    Park, Seung-min; Sabour, Andrew F.; Son, Jun Ho; Lee, Sang Hun

    2014-01-01

    Integrated molecular diagnostic systems (iMDx), which are automated, sensitive, specific, user-friendly, robust, rapid, easy-to-use, and portable, can revolutionize future medicine. This review will first focus on the components of sample extraction, preservation, and filtration necessary for all point-of-care devices to include for practical use. Subsequently, we will look for low-powered and precise methods for both sample amplification and signal transduction, going in-depth to the details behind their principles. The final field of total device integration and its application to the clinical field will also be addressed to discuss the practicality for future patient care. We envision that microfluidic systems hold the potential to breakthrough the number of problems brought into the field of medical diagnosis today. PMID:24759281

  19. Differential roles of MMP-9 in early and late stages of dystrophic muscles in a mouse model of Duchenne muscular dystrophy.

    PubMed

    Shiba, Naoko; Miyazaki, Daigo; Yoshizawa, Takahiro; Fukushima, Kazuhiro; Shiba, Yuji; Inaba, Yuji; Imamura, Michihiro; Takeda, Shin'ichi; Koike, Kenichi; Nakamura, Akinori

    2015-10-01

    Matrix metalloprotease (MMP)-9 is an endopeptidase associated with the pathogenesis of Duchenne muscular dystrophy (DMD). The precise function of MMP-9 in DMD has not been elucidated to date. We investigated the effect of genetic ablation of MMP-9 in the mdx mouse model (mdx/Mmp9(-/-)). At the early disease stage, the muscles of mdx/Mmp9(-/-) mice showed reduced necrosis and neutrophil invasion, accompanied by down-regulation of chemokine MIP-2. In addition, muscle regeneration was enhanced, which coincided with increased macrophage infiltration and upregulation of MCP-1, and resulted in increased muscle strength. The mdx/Mmp9(-/-) mice also displayed accelerated upregulation of osteopontin expression in skeletal muscle at the acute onset phase of dystrophy. However, at a later disease stage, the mice exhibited muscle growth impairment through altered expression of myogenic factors, and increased fibroadipose tissue. These results showed that MMP-9 might have multiple functions during disease progression. Therapy targeting MMP-9 may improve muscle pathology and function at the early disease stage, but continuous inhibition of this protein may result in the accumulation of fibroadipose tissues and reduced muscle strength at the late disease stage. PMID:26170062

  20. [Frequency dynamics of stomach and duodenum ulcer found in liquidators of consequence of Chernobyl accident (according to the data of long-term observation)].

    PubMed

    Gasanova, E V; Kovalenko, A N

    2004-01-01

    The authors have studied the frequency dynamics of stomach and duodenum ulcer occurrence revealed in liquidators of consequence of Chernobyl accident (1986-1987) after long-term medical observation with age and radiation absorbed dose variations. According to the findings, duodenum ulcer incidence prevails over stomach ulcer in all scale of age. The connection between the frequency dynamics of duodenum ulcer and age of the patients wasn't found. PMID:15724602

  1. Pathologic effects of fractionated fast neutrons or photons on the pancreas, pylorus and duodenum of dogs

    SciTech Connect

    Zook, B.C.; Bradley, E.W.; Casarett, G.W.; Rogers, C.C.

    1983-10-01

    Thirty-nine adult male Beagles received either fast neutron or photon irradiation to the right thorax to determine the relative biological effectiveness (RBE) of fast neutrons on normal pulmonary tissue. The right anterior abdomen was included in the field of radiation. Twenty-four dogs (six/group) received fast neutrons with an average energy of 15 MeV to total doses of 1000, 1500, 2250 or 3375 rad in four fractions per week for six weeks. Fifteen dogs received 3000, 4500, or 6750 rad of photons (five/group) in an identical fractionation pattern. All neutron irradiated dogs receiving 3375 and 2250 rad and one receiving 1500 rad developed clinical signs of pancreatic, hepatic and gastrointestinal disturbances. The liver enzymes of these dogs became elevated and they died or were euthanized in extremis 47-367 days after irradiation. Only one 6750 rad photon dog developed similar signs and died 708 days post-irradiation. Five neutron and 10 photon exposed dogs died of other causes. Neutron-induced lesions in the stomach and duodenum included hemorrhages, erosions, ulcerations and fibrosis. Ulcers perforated the GI tract of five dogs. Pancreatic lesions included degranulation and necrosis of acinar cells, fibrosis and atrophy. Islet cells were not obviously damaged. All lesions were associated with degenerative and occlusive vascular changes. The RBE of fast neutrons, assessed by clinical signs, gross and microscopic pathology, is approximately 3-4.5 for pancreas and about 4.5 for pylorus and duodenum.

  2. Bacterial overgrowth in the duodenum of dogs with exocrine pancreatic insufficiency.

    PubMed

    Williams, D A; Batt, R M; McLean, L

    1987-07-15

    Bacterial overgrowth (greater than 10(5) colony-forming units/ml duodenal juice) in the duodenum was demonstrated in 8 of 11 dogs with exocrine pancreatic insufficiency (EPI). In 4 of these 8 dogs, the overgrowth included large numbers (greater than 10(4) colony-forming units/ml) of obligate anaerobic bacteria and was associated with decreased activities of several brush border marker enzymes and, in 2 dogs, with partial villous atrophy in the jejunum. Changes in the jejunal mucosa of the remaining dogs (with either no overgrowth or overgrowth of aerobic bacteria alone) were characterized by increased activities of some brush border disaccharides and of lysosomal hydrolases. One dog was euthanatized without treatment, at the owner's request. The response of 4 of the remaining 10 dogs treated with enzyme replacement alone was poor or suboptimal, and all of these 4 dogs had bacterial overgrowth. One of these dogs had an excellent clinical response when also given oxytetracycline orally for 14 days, but the other 3 dogs did not improve further in response to the same treatment. It was concluded that bacterial overgrowth in the duodenum is common in dogs with EPI and that, when such overgrowth includes large numbers of obligate anaerobes, there may be associated biochemical and morphologic abnormalities in jejunal mucosa. Functional disturbances related to abnormal intestinal microflora may be responsible for the failure of some dogs with EPI to respond fully to oral pancreatic enzyme supplementation without antibiotic therapy. PMID:3610795

  3. Glutamine oxidation and utilization by rat and human oesophagus and duodenum.

    PubMed

    James, L A; Lunn, P G; Middleton, S; Elia, M

    1999-04-01

    The rates of utilization and oxidation of glutamine and glucose by oesophageal and duodenal tissues have been investigated in both rats and human subjects. In the rat, glutamine utilization by oesophageal tissue was 2-3-fold lower than that in the duodenum, and this substrate contributed less than 10% to the total oxidative metabolism of the tissue, even when glutamine was the only substrate provided. In contrast, rat duodenal tissue derived about 34% of the total CO2 production from glutamine-C, and this contribution was not suppressed by the addition of either glucose or a mixture of the other substrates. Rates of glucose utilization and oxidation by the duodenum were lower than those for glutamine, and were significantly (P < 0.001) suppressed by addition of glutamine. In both oesophageal and duodenal tissues, less than 10% of the glutamine-C utilized was fully oxidized, approximately 60-70% was converted to glutamate, and 30-40% to alanine. Results obtained using human biopsy tissue samples were similar to those observed in the rat. Glutamine oxidation contributed 34 (SD 4)% of the total CO2 production by the duodenal tissue, but only 8 (SD 4)% to oesophageal tissue oxidation. The findings suggest that glutamine is not an important or preferred fuel for oesophageal tissue, whereas it is for duodenal tissue. Thus, these tissues can be expected to respond differently to glutamine administration. PMID:10999020

  4. Identification of telocytes in the lamina propria of rat duodenum: transmission electron microscopy

    PubMed Central

    Carmona, I Cantarero; Bartolomé, M J Luesma; Escribano, C Junquera

    2011-01-01

    Abstract Recently the new term ‘telocytes’ has been proposed for cells formerly known as interstitial Cajal-like cells. In fact, telocytes are not really Cajal-like cells, they being different from all other interstitial cells by the presence of telopodes, which are cell-body prolongations, very thin, extremely long with a moniliform aspect. The identification of these cells is based on ultrastructural criteria. The presence of telocytes in others organs was previously documented. We reported for the first time, an ultrastructural study of telocytes in the lamina propria of rat duodenum. Our findings show that typical telocytes are present in the rat duodenum. Telocytes are located in the lamina propria, immediately below mucosal crypts. Telopodes frequently establish close spatial relationships with immune cells, blood vessels and nerve endings. On the basis of their distribution and morphology, we suggest that these cells may be involved in immune response and in our opinion, it may be possible that different locations of telocytes could be associated with different roles. PMID:21054782

  5. Massive Idiosyncratic Exon Skipping Corrects the Nonsense Mutation in Dystrophic Mouse Muscle and Produces Functional Revertant Fibers by Clonal Expansion

    PubMed Central

    Lu, Q.L.; Morris, G.E.; Wilton, S.D.; Ly, T.; Artem'yeva, O.V.; Strong, P.; Partridge, T.A.

    2000-01-01

    Conventionally, nonsense mutations within a gene preclude synthesis of a full-length functional protein. Obviation of such a blockage is seen in the mdx mouse, where despite a nonsense mutation in exon 23 of the dystrophin gene, occasional so-called revertant muscle fibers are seen to contain near-normal levels of its protein product. Here, we show that reversion of dystrophin expression in mdx mice muscle involves unprecedented massive loss of up to 30 exons. We detected several alternatively processed transcripts that could account for some of the revertant dystrophins and could not detect genomic deletion from the region commonly skipped in revertant dystrophin. This, together with exon skipping in two noncontiguous regions, favors aberrant splicing as the mechanism for the restoration of dystrophin, but is hard to reconcile with the clonal idiosyncrasy of revertant dystrophins. Revertant dystrophins retain functional domains and mediate plasmalemmal assembly of the dystrophin-associated glycoprotein complex. Physiological function of revertant fibers is demonstrated by the clonal growth of revertant clusters with age, suggesting that revertant dystrophin could be used as a guide to the construction of dystrophin expression vectors for individual gene therapy. The dystrophin gene in the mdx mouse provides a favored system for study of exon skipping associated with nonsense mutations. PMID:10704448

  6. Merkel cell carcinoma of unknown primary with lymph node and mesenteric metastasis involving the pancreas and duodenum

    PubMed Central

    Lim, Brian S.; Flannery, Christopher M.; Koh, Stephen S.; Yaghsezian, Harout

    2016-01-01

    Merkel cell carcinoma (MCC) of skin is a rare, aggressive cutaneous malignancy of neuroendocrine origin. MCC predominantly affects elderly Caucasians and has high predilection for sun exposed areas. Histologic exam and immunohistochemical profile is required to establish the diagnosis. It has high propensity for local recurrence and metastasis, and carries poor prognosis. However, metastasis to mesentery involving the duodenum is very uncommon and rarely reported in literature. We hereby describe a patient with lymph node and mesenteric metastasis invading duodenum and pancreas with unknown primary origin of MCC. PMID:27034815

  7. Increased Intraepithelial Vα24 Invariant NKT Cells in the Celiac Duodenum

    PubMed Central

    Montalvillo, Enrique; Bernardo, David; Martínez-Abad, Beatriz; Allegretti, Yessica; Fernández-Salazar, Luis; Calvo, Carmen; Chirdo, Fernando G.; Garrote, José A.; Arranz, Eduardo

    2015-01-01

    Celiac Disease (CD) is an interferon (IFN)γ-mediated duodenal hypersensitivity to wheat gluten occurring in genetically predisposed individuals. Gluten-free diet (GFD) leads to a complete remission of the disease. Vα24-restricted invariant NKT (iNKT) cells are important to maintain immune homeostasis in the gut mucosa because of their unique capacity to rapidly produce large quantities of both T-helper (Th)1 and Th2 cytokines upon stimulation. We studied the presence of these cells in the CD duodenum. Duodenal biopsies were obtained from 45 untreated-CD patients (uCD), 15 Gluten Free Diet-CD patients (GFD-CD), 44 non-inflamed non-CD controls (C-controls) and 15 inflamed non-CD controls (I-controls). Two populations from Spain and Argentina were recruited. Messenger RNA (mRNA) expression of Vα24-Jα18 (invariant TCRα chain of human iNKT cells), IFNγ and intracellular transcription factor Forkhead Box P3 (Foxp3), and flow cytometry intraepithelial lymphocyte (IEL) profile were determined. Both uCD and GFD-CD patients had higher Vα24-Jα18 mRNA levels than non-CD controls (I and C-controls). The expression of Vα24-Jα18 correlated with Marsh score for the severity of mucosal lesion and also with increased mRNA IFNγ levels. uCD and GFD-CD patients had decreased mRNA expression of FoxP3 but increased expression of Vα24-Jα18, which revealed a CD-like molecular profile. Increased numbers of iNKT cells were confirmed by flow cytometry within the intraepithelial lymphocyte compartment of uCD and GFD-CD patients and correlated with Vα24-Jα18 mRNA expression. In conclusion, we have found an increased number of iNKT cells in the duodenum from both uCD and GFD-CD patients, irrespective of the mucosal status. A CD-like molecular profile, defined by an increased mRNA expression of Vα24-Jα18 together with a decreased expression of FoxP3, may represent a pro-inflammatory signature of the CD duodenum. PMID:26529008

  8. Electrical Impedance Myography to Detect the Effects of Electrical Muscle Stimulation in Wild Type and Mdx Mice

    PubMed Central

    Li, Jia; Yim, Sung; Pacheck, Adam; Sanchez, Benjamin; Rutkove, Seward B.

    2016-01-01

    Objective Tools to better evaluate the impact of therapy on nerve and muscle disease are needed. Electrical impedance myography (EIM) is sensitive to neuromuscular disease progression as well as to therapeutic interventions including myostatin inhibition and antisense oligonucleotide-based treatments. Whether the technique identifies the impact of electrical muscle stimulation (EMS) is unknown. Methods Ten wild-type (wt) C57B6 mice and 10 dystrophin-deficient (mdx) mice underwent 2 weeks of 20 min/day EMS on left gastrocnemius and sham stimulation on the right gastrocnemius. Multifrequency EIM data and limb girth were obtained before and at the conclusion of the protocol. Muscle weight, in situ force measurements, and muscle fiber histology were also assessed at the conclusion of the study. Results At the time of sacrifice, muscle weight was greater on the EMS-treated side than on the sham-stimulated side (p = 0.018 for wt and p = 0.007 for mdx). Similarly, in wt animals, EIM parameters changed significantly compared to baseline (resistance (p = 0.009), reactance (p = 0.0003) and phase (p = 0.002); these changes were due in part to reductions in the EIM values on the EMS-treated side and elevations on the sham-simulated side. Mdx animals showed analogous but non-significant changes (p = 0.083, p = 0.064, and p = 0.57 for resistance, reactance and phase, respectively). Maximal isometric force trended higher on the stimulated side in wt animals only (p = 0.06). Myofiber sizes in wt animals were also larger on the stimulated side than on the sham-stimulated side (p = 0.034); no significant difference was found in the mdx mice (p = 0.79). Conclusion EIM is sensitive to stimulation-induced muscle alterations in wt animals; similar trends are also present in mdx mice. The mechanisms by which these EIM changes develop, however, remains uncertain. Possible explanations include longer-term trophic effects and shorter-term osmotic effects. PMID:26986564

  9. Organoaxial partial rotation of duodenum with midgut malrotation in an adult

    PubMed Central

    Amarakoon, Luckshika Udeshani; Rathnamali, Baj Gamage Anushka; Jayasundara, Jasin Arachchige Saman Bingumal; de Silva, Ajith

    2014-01-01

    Midgut malrotation includes a range of developmental abnormalities that occur during fetal intestinal rotation. Manifestations of intestinal malrotation are generally seen in the paediatric population and are uncommon in adults. Symptomatic patients may present with either acute abdominal pain due to midgut volvulus, or chronic abdominal pain due to proximal midgut partial obstruction in the presence of congenital bands. A limited number of paediatric cases of duodenal occlusion due to duodenal malrotation has been previously reported in the medical literature. We herein report the case of a 57-year-old woman who presented with duodenal obstruction due to organoaxial partial rotation of the distal duodenum associated with midgut malrotation. This is probably the first of such a case diagnosed in adulthood reported in the medical literature. Our patient underwent Roux-en-Y duodenojejunostomy and had symptomatic relief following the successful surgery. PMID:25630324

  10. The role of mast cell in tissue morphogenesis. Thymus, duodenum, and mammary gland as examples.

    PubMed

    Ribatti, Domenico; Crivellato, Enrico

    2016-02-01

    Mast cells (MCs) are strategically located at host/environment interfaces like skin, airways, and gastro-intestinal and uro-genital tracts. MCs also populate connective tissues in association with blood and lymphatic vessels and nerves. MCs are absent in avascular tissues, such as mineralized bone, cartilage, and cornea. MCs have various functions and different functional subsets of MCs are encountered in different tissues. However, we do not' know exactly what is the physiological function of MC. Most of these functions are not essential for life, as various MC-deficient strains of mice and rats seems to have normal life spans. In this review article, we have reported and discussed the literature data concerning the role of MCs in tissue morphogenesis, and in particular their role in the development of thymus, duodenum, and mammary gland. PMID:26615957

  11. Histological structure of duodenum in gilts receiving low doses of zearalenone and deoxynivalenol in feed.

    PubMed

    Lewczuk, Bogdan; Przybylska-Gornowicz, Barbara; Gajęcka, Magdalena; Targońska, Krystyna; Ziółkowska, Natalia; Prusik, Magdalena; Gajęcki, Maciej

    2016-01-01

    Deoxynivalenol (DON) and zearalenone (ZEN), produced by microfungi of the Fusarium family, are among the most commonly occurring mycotoxins. They are considered important factors affecting human and animal health as well as livestock productivity. The aim of this study was to determine the effect of low doses of these mycotoxins on the histological structure of the pig duodenum. The study was performed on 72 gilts, with initial weights of approximately 25kg, divided into 4 equal groups. Group I received per os ZEN (40μg/kg BW), group II-DON (12μg/kg BW), group III-ZEN (40μg/kg BW) and DON (12μg/kg BW), and group IV-vehicle. The pigs were killed after 1, 2, 3, 4, 5 and 6 weeks of the treatment, and the duodenum samples were prepared for histological investigations. The slides were digitalized and subjected to morphometrical analysis. The treatment with DON and ZEN did not change the architecture of the mucosa or the ratio between goblet and adsorptive cells in the epithelium. The administration of DON induced an increase in the number of lymphocytes in the mucosal epithelium. Both mycotoxins, administered alone or together, increased the quantity of lymphocytes, plasma cells and macrophages with black-brown granules in the lamina propria. The time-courses of changes in the number of defense system cells evoked by DON and ZEN were different. In conclusion, dietary exposure to low doses of Fusarium mycotoxins should be considered an important risk factor for subclinical inflammation in the small intestine. PMID:26679981

  12. A COMPARATIVE REVIEW OF THE FLOW OF NITROGEN FRACTIONS AT THE OMASAL CANAL AND DUODENUM OF DAIRY COWS

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The objective of this paper was to review and compare published data for the passage of N fractions to the omasal canal and duodenum of lactating dairy cows. Two data sets were created; one with data from 14 studies in which digesta was sampled from the omasal canal (57 means) and the other with dat...

  13. Duodenum has the greatest potential to absorb soluble non-ammonia nitrogen in the nonmesenteric gastrointestinal tissues of dairy cows*

    PubMed Central

    Xie, Ying-ming; Xu, Qing-biao; Wu, Yue-ming; Huang, Xin-bei; Liu, Jian-xin

    2015-01-01

    In cattle, dietary protein is gradually degraded into peptide-bound amino acids (PBAAs), free amino acids (FAAs), and ultimately into ammonia by the rumen microbes. Both PBAA and FAA are milk protein precursors, and the rumen and small intestines are the main sites where such precursors are produced and absorbed. This work was designed to investigate the expression of the peptide transporter PepT1 and the AA transporters ASCT2, y+LAT1, and ATB0,+, and the concentrations of PBAA, FAA, and soluble protein in the rumen, omasum, and duodenum of dairy cows. Tissues and digesta were collected from six healthy Chinese Holstein dairy cows immediately after the animals were slaughtered. The expression of transporters was analyzed by real-time quantitative polymerase chain reaction (PCR). The FAA concentration was assessed using an amino acid (AA) analyzer, PBAA concentration by quantification of AA before and after acid-hydrolysis by 6 mol/L HCl, and soluble protein concentration by quantification of the bicinchoninic acid content. The results showed that the relative abundance of mRNA of the transporters and the soluble non-ammonia nitrogen (SNAN) concentration of each fraction were greater in the duodenum than in the rumen or omasum. These results indicate that the duodenum is the predominant location within the nonmesenteric digestive tract for producing milk protein precursors. In addition, PBAA was the largest component of SNAN in the digesta from the rumen, omasum, and duodenum. In conclusion, the duodenum has the greatest concentrations of SNAN and PBAA, and the greatest potential for absorption of SNAN in the form of PBAA in the nonmesenteric gastrointestinal tissues of dairy cows. PMID:26055912

  14. Nuclear receptor and target gene mRNA abundance in duodenum and colon of dogs with chronic enteropathies.

    PubMed

    Greger, D L; Gropp, F; Morel, C; Sauter, S; Blum, J W

    2006-11-01

    Nuclear receptors (NR), such as constitutive androstane receptor (CAR), pregnane X receptor (PXR) and peroxisome proliferator-associated receptors alpha and gamma (PPARalpha, PPARgamma) are mediators of inflammation and may be involved in inflammatory bowel disease (IBD) and food responsive diarrhea (FRD) of dogs. The present study compared mRNA abundance of NR and NR target genes [multi drug-resistance gene-1 (MDR1), multiple drug-resistance-associated proteins (MRD2, MRD3), cytochrome P450 (CYP3A12), phenol-sulfating phenol sulfotransferase (SULT1A1) and glutathione-S-transferase (GST A3-3)] in biopsies obtained from duodenum and colon of dogs with IBD and FRD and healthy control dogs (CON; n=7 per group). Upon first presentation of dogs, mRNA levels of PPARalpha, PPARgamma, CAR, PXR and RXRalpha in duodenum as well as PPARgamma, CAR, PXR and RXRalpha in colon were not different among groups (P>0.10). Although mRNA abundance of PPARalpha in colon of dogs with FRD was similar in both IBD and CON (P>0.10), PPARalpha mRNA abundance was higher in IBD than CON (P<0.05). Levels of mRNA of MDR1 in duodenum were higher in FRD than IBD (P<0.05) or CON (P<0.001). Compared with CON, abundances of mRNA for MRP2, CYP3A12 and SULT1A1 were higher in both FRD and IBD than CON (P<0.05). Differences in mRNA levels of PPARalpha and MRP2 in colon and MDR1, MRP2, CYP3A12 and SULT1A1 in duodenum may be indicative for enteropathy in FRD and (or) IBD dogs relative to healthy dogs. More importantly, increased expression of MDR1 in FRD relative to IBD in duodenum may be a useful diagnostic marker to distinguish dogs with FRD from dogs with IBD. PMID:16446074

  15. Pathways of abnormal stress-induced Ca2+ influx into dystrophic mdx cardiomyocytes

    PubMed Central

    Fanchaouy, M.; Polakova, E.; Jung, C.; Ogrodnik, J.; Shirokova, N.; Niggli, E.

    2009-01-01

    In Duchenne muscular dystrophy, deficiency of the cytoskeletal protein dystrophin leads to well-described defects in skeletal muscle, but also to dilated cardiomyopathy, accounting for about 20% of the mortality. Mechanisms leading to cardiomyocyte cell death and cardiomyopathy are not well understood. One hypothesis suggests that the lack of dystrophin leads to membrane instability during mechanical stress and to activation of Ca2+ entry pathways. Using cardiomyocytes isolated from dystrophic mdx mice we dissected the contribution of various putative Ca2+ influx pathways with pharmacological tools. Cytosolic Ca2+ and Na+ signals as well as uptake of membrane impermeant compounds were monitored with fluorescent indicators using confocal microscopy and photometry. Membrane stress was applied as moderate osmotic challenges while membrane current was quantified using the whole-cell patch-clamp technique. Our findings suggest a major contribution of two primary Ca2+ influx pathways, stretch-activated membrane channels and short-lived microruptures. Furthermore, we found evidence for a secondary Ca2+ influx pathway, the Na+-Ca2+ exchange (NCX), which in cardiac muscle has a large transport capacity. After stress it contributes to Ca2+ entry in exchange for Na+ which had previously entered via primary stress-induced pathways, representing a previously not recognized mechanism contributing to subsequent cellular damage. This complexity needs to be considered when targeting abnormal Ca2+ influx as a treatment option for dystrophy. PMID:19604578

  16. Increased resting intracellular calcium modulates NF-κB-dependent inducible nitric-oxide synthase gene expression in dystrophic mdx skeletal myotubes.

    PubMed

    Altamirano, Francisco; López, Jose R; Henríquez, Carlos; Molinski, Tadeusz; Allen, Paul D; Jaimovich, Enrique

    2012-06-15

    Duchenne muscular dystrophy (DMD) is a genetic disorder caused by dystrophin mutations, characterized by chronic inflammation and severe muscle wasting. Dystrophic muscles exhibit activated immune cell infiltrates, up-regulated inflammatory gene expression, and increased NF-κB activity, but the contribution of the skeletal muscle cell to this process has been unclear. The aim of this work was to study the pathways that contribute to the increased resting calcium ([Ca(2+)](rest)) observed in mdx myotubes and its possible link with up-regulation of NF-κB and pro-inflammatory gene expression in dystrophic muscle cells. [Ca(2+)](rest) was higher in mdx than in WT myotubes (308 ± 6 versus 113 ± 2 nm, p < 0.001). In mdx myotubes, both the inhibition of Ca(2+) entry (low Ca(2+) solution, Ca(2+)-free solution, and Gd(3+)) and blockade of either ryanodine receptors or inositol 1,4,5-trisphosphate receptors reduced [Ca(2+)](rest). Basal activity of NF-κB was significantly up-regulated in mdx versus WT myotubes. There was an increased transcriptional activity and p65 nuclear localization, which could be reversed when [Ca(2+)](rest) was reduced. Levels of mRNA for TNFα, IL-1β, and IL-6 were similar in WT and mdx myotubes, whereas inducible nitric-oxide synthase (iNOS) expression was increased 5-fold. Reducing [Ca(2+)](rest) using different strategies reduced iNOS gene expression presumably as a result of decreased activation of NF-κB. We propose that NF-κB, modulated by increased [Ca(2+)](rest), is constitutively active in mdx myotubes, and this mechanism can account for iNOS overexpression and the increase in reactive nitrogen species that promote damage in dystrophic skeletal muscle cells. PMID:22549782

  17. Increased Resting Intracellular Calcium Modulates NF-κB-dependent Inducible Nitric-oxide Synthase Gene Expression in Dystrophic mdx Skeletal Myotubes*

    PubMed Central

    Altamirano, Francisco; López, Jose R.; Henríquez, Carlos; Molinski, Tadeusz; Allen, Paul D.; Jaimovich, Enrique

    2012-01-01

    Duchenne muscular dystrophy (DMD) is a genetic disorder caused by dystrophin mutations, characterized by chronic inflammation and severe muscle wasting. Dystrophic muscles exhibit activated immune cell infiltrates, up-regulated inflammatory gene expression, and increased NF-κB activity, but the contribution of the skeletal muscle cell to this process has been unclear. The aim of this work was to study the pathways that contribute to the increased resting calcium ([Ca2+]rest) observed in mdx myotubes and its possible link with up-regulation of NF-κB and pro-inflammatory gene expression in dystrophic muscle cells. [Ca2+]rest was higher in mdx than in WT myotubes (308 ± 6 versus 113 ± 2 nm, p < 0.001). In mdx myotubes, both the inhibition of Ca2+ entry (low Ca2+ solution, Ca2+-free solution, and Gd3+) and blockade of either ryanodine receptors or inositol 1,4,5-trisphosphate receptors reduced [Ca2+]rest. Basal activity of NF-κB was significantly up-regulated in mdx versus WT myotubes. There was an increased transcriptional activity and p65 nuclear localization, which could be reversed when [Ca2+]rest was reduced. Levels of mRNA for TNFα, IL-1β, and IL-6 were similar in WT and mdx myotubes, whereas inducible nitric-oxide synthase (iNOS) expression was increased 5-fold. Reducing [Ca2+]rest using different strategies reduced iNOS gene expression presumably as a result of decreased activation of NF-κB. We propose that NF-κB, modulated by increased [Ca2+]rest, is constitutively active in mdx myotubes, and this mechanism can account for iNOS overexpression and the increase in reactive nitrogen species that promote damage in dystrophic skeletal muscle cells. PMID:22549782

  18. Loss of nNOS inhibits compensatory muscle hypertrophy and exacerbates inflammation and eccentric contraction-induced damage in mdx mice

    PubMed Central

    Froehner, Stanley C.; Reed, Sarah M.; Anderson, Kendra N.; Huang, Paul L.; Percival, Justin M.

    2015-01-01

    Approaches targeting nitric oxide (NO) signaling show promise as therapies for Duchenne and Becker muscular dystrophies. However, the mechanisms by which NO benefits dystrophin-deficient muscle remain unclear, but may involve nNOSβ, a newly discovered enzymatic source of NO in skeletal muscle. Here we investigate the impact of dystrophin deficiency on nNOSβ and use mdx mice engineered to lack nNOSμ and nNOSβ to discern how the loss of nNOS impacts dystrophic skeletal muscle pathology. In mdx muscle, nNOSβ was mislocalized and its association with the Golgi complex was reduced. nNOS depletion from mdx mice prevented compensatory skeletal muscle cell hypertrophy, decreased myofiber central nucleation and increased focal macrophage cell infiltration, indicating exacerbated dystrophic muscle damage. Reductions in muscle integrity in nNOS-null mdx mice were accompanied by decreases in specific force and increased susceptibility to eccentric contraction-induced muscle damage compared with mdx controls. Unexpectedly, muscle fatigue was unaffected by nNOS depletion, revealing a novel latent compensatory mechanism for the loss of nNOS in mdx mice. Together with previous studies, these data suggest that localization of both nNOSμ and nNOSβ is disrupted by dystrophin deficiency. They also indicate that nNOS has a more complex role as a modifier of dystrophic pathology and broader therapeutic potential than previously recognized. Importantly, these findings also suggest nNOSβ as a new drug target and provide a new conceptual framework for understanding nNOS signaling and the benefits of NO therapies in dystrophinopathies. PMID:25214536

  19. Clinical map document based on XML (cMDX): document architecture with mapping feature for reporting and analysing prostate cancer in radical prostatectomy specimens

    PubMed Central

    2010-01-01

    Background The pathology report of radical prostatectomy specimens plays an important role in clinical decisions and the prognostic evaluation in Prostate Cancer (PCa). The anatomical schema is a helpful tool to document PCa extension for clinical and research purposes. To achieve electronic documentation and analysis, an appropriate documentation model for anatomical schemas is needed. For this purpose we developed cMDX. Methods The document architecture of cMDX was designed according to Open Packaging Conventions by separating the whole data into template data and patient data. Analogue custom XML elements were considered to harmonize the graphical representation (e.g. tumour extension) with the textual data (e.g. histological patterns). The graphical documentation was based on the four-layer visualization model that forms the interaction between different custom XML elements. Sensible personal data were encrypted with a 256-bit cryptographic algorithm to avoid misuse. In order to assess the clinical value, we retrospectively analysed the tumour extension in 255 patients after radical prostatectomy. Results The pathology report with cMDX can represent pathological findings of the prostate in schematic styles. Such reports can be integrated into the hospital information system. "cMDX" documents can be converted into different data formats like text, graphics and PDF. Supplementary tools like cMDX Editor and an analyser tool were implemented. The graphical analysis of 255 prostatectomy specimens showed that PCa were mostly localized in the peripheral zone (Mean: 73% ± 25). 54% of PCa showed a multifocal growth pattern. Conclusions cMDX can be used for routine histopathological reporting of radical prostatectomy specimens and provide data for scientific analysis. PMID:21078179

  20. Protein-DNA array-based identification of transcription factor activities differentially regulated in skeletal muscle of normal and dystrophin-deficient mdx mice.

    PubMed

    Dogra, Charu; Srivastava, Daya Shankar; Kumar, Ashok

    2008-05-01

    Inactivation of dystrophin gene is the primary cause of Duchenne muscular dystrophy (DMD) in humans and mdx mice. However, the underpinning mechanisms, which govern the pathogenesis of dystrophin-deficient skeletal muscle, remain poorly understood. We have previously reported activation of mitogen-activated protein kinases (MAPK), nuclear factor-kappa B (NF-kappaB), and phosphatidyl-inositol 3-kinase/Akt (PI3K/Akt) signaling pathways in diaphragm muscle of mdx mice. In this study, using a protein-DNA array-based approach, we have investigated the activation of 345 transcription factors in diaphragm muscle of 6-week old normal and dystrophin-deficient mdx mice. Our data demonstrate increased activation of a number nuclear transcription factors including AP1, HFH-3, PPARalpha, c.myb BP, ETF, Fra-1/JUN, kBF-A, N-rasBP, lactoferrin BP, Myb(2), EBP40_45, EKLF(1), p53(2), TFEB, Myc-Max; c-Rel; E2, ISRE; NF-kB; Stat1 p84/p91, Antioxidant RE, EVI-1, Stat3, AP3, p53, Stat4, AP4, HFH-1, FAST-1, Pax-5, and Beta-RE in the diaphragm muscle of mdx mice compared to corresponding normal mice. The level of activation for p53 was highest among all the transcription factors studied. Furthermore, higher activation of p53 in diaphragm muscle of mdx mice was associated with its increased phosphorylation and nuclear translocation. Collectively, our data suggest that the primary deficiency of dystrophin leads to the aberrant activation of nuclear transcription factors which might further contribute to muscle pathogenesis in mdx mice. PMID:18278580

  1. Identification of novel, therapy-responsive protein biomarkers in a mouse model of Duchenne muscular dystrophy by aptamer-based serum proteomics

    PubMed Central

    Coenen-Stass, Anna M. L.; McClorey, Graham; Manzano, Raquel; Betts, Corinne A.; Blain, Alison; Saleh, Amer F.; Gait, Michael J.; Lochmüller, Hanns; Wood, Matthew J. A.; Roberts, Thomas C.

    2015-01-01

    There is currently an urgent need for biomarkers that can be used to monitor the efficacy of experimental therapies for Duchenne Muscular Dystrophy (DMD) in clinical trials. Identification of novel protein biomarkers has been limited due to the massive complexity of the serum proteome and the presence of a small number of very highly abundant proteins. Here we have utilised an aptamer-based proteomics approach to profile 1,129 proteins in the serum of wild-type and mdx (dystrophin deficient) mice. The serum levels of 96 proteins were found to be significantly altered (P < 0.001, q < 0.01) in mdx mice. Additionally, systemic treatment with a peptide-antisense oligonucleotide conjugate designed to induce Dmd exon skipping and recover dystrophin protein expression caused many of the differentially abundant serum proteins to be restored towards wild-type levels. Results for five leading candidate protein biomarkers (Pgam1, Tnni3, Camk2b, Cycs and Adamts5) were validated by ELISA in the mouse samples. Furthermore, ADAMTS5 was found to be significantly elevated in human DMD patient serum. This study has identified multiple novel, therapy-responsive protein biomarkers in the serum of the mdx mouse with potential utility in DMD patients. PMID:26594036

  2. Early right ventricular fibrosis and reduction in biventricular cardiac reserve in the dystrophin-deficient mdx heart.

    PubMed

    Meyers, Tatyana A; Townsend, DeWayne

    2015-02-15

    Duchenne muscular dystrophy (DMD) is a progressive disease of striated muscle deterioration. Respiratory and cardiac muscle dysfunction are particularly clinically relevant because they result in the leading causes of death in DMD patients. Despite the clinical and physiological significance of these systems, little has been done to understand the cardiorespiratory interaction in DMD. We show here that prior to the onset of global cardiac dysfunction, dystrophin-deficient mdx mice have increased cardiac fibrosis with the right ventricle being particularly affected. Using a novel biventricular cardiac catheterization technique coupled with cardiac stress testing, we demonstrate that both the right and left ventricles have significant reductions in both systolic and diastolic function in response to dobutamine. Unstimulated cardiac function is relatively normal except for a significant reduction in the ventricular pressure transient duration compared with controls. These biventricular analyses also reveal the absence of a dobutamine-induced increase in isovolumic relaxation in the right ventricle of control hearts. Simultaneous assessment of biventricular pressure demonstrates a dobutamine-dependent enhancement of coupling between the ventricles in control mice, which is absent in mdx mice. Furthermore, studies probing the passive-extension properties of the left ventricle demonstrate that the mdx heart is significantly more compliant compared with age-matched C57BL/10 hearts, which have an age-dependent stiffening that is completely absent from dystrophic hearts. These new results indicate that right ventricular fibrosis is an early indicator of the development of dystrophic cardiomyopathy, suggesting a mechanism by which respiratory insufficiency may accelerate the development of heart failure in DMD. PMID:25485898

  3. Toll-like receptor 4 ablation in mdx mice reveals innate immunity as a therapeutic target in Duchenne muscular dystrophy

    PubMed Central

    Giordano, Christian; Mojumdar, Kamalika; Liang, Feng; Lemaire, Christian; Li, Tong; Richardson, John; Divangahi, Maziar; Qureshi, Salman; Petrof, Basil J.

    2015-01-01

    Toll-like receptor 4 (TLR4) recognizes specific structural motifs associated with microbial pathogens and also responds to certain endogenous host molecules associated with tissue damage. In Duchenne muscular dystrophy (DMD), inflammation plays an important role in determining the ultimate fate of dystrophic muscle fibers. In this study, we used TLR4-deficient dystrophic mdx mice to assess the role of TLR4 in the pathogenesis of DMD. TLR4 expression was increased and showed enhanced activation following agonist stimulation in mdx diaphragm muscle. Genetic ablation of TLR4 led to significantly increased muscle force generation in dystrophic diaphragm muscle, which was associated with improved histopathology including decreased fibrosis, as well as reduced pro-inflammatory gene expression and macrophage infiltration. TLR4 ablation in mdx mice also altered the phenotype of muscle macrophages by inducing a shift toward a more anti-inflammatory (iNOSneg CD206pos) profile. In vitro experiments confirmed that lack of TLR4 is sufficient to influence macrophage activation status in response to classical polarizing stimuli such as IFN-gamma and IL-4. Finally, treatment of dystrophic mice with glycyrrhizin, an inhibitor of the endogenous TLR4 ligand, high mobility group box (HMGB1), also pointed to involvement of the HMGB1–TLR4 axis in promoting dystrophic diaphragm pathology. Taken together, our findings reveal TLR4 and the innate immune system as important players in the pathophysiology of DMD. Accordingly, targeting either TLR4 or its endogenous ligands may provide a new therapeutic strategy to slow disease progression. PMID:25552658

  4. Comparative proteomic profiling of dystroglycan-associated proteins in wild type, mdx, and Galgt2 transgenic mouse skeletal muscle.

    PubMed

    Yoon, Jung Hae; Johnson, Eric; Xu, Rui; Martin, Laura T; Martin, Paul T; Montanaro, Federica

    2012-09-01

    Dystroglycan is a major cell surface glycoprotein receptor for the extracellular matrix in skeletal muscle. Defects in dystroglycan glycosylation cause muscular dystrophy and alterations in dystroglycan glycosylation can impact extracellular matrix binding. Here we describe an immunoprecipitation technique that allows isolation of beta dystroglycan with members of the dystrophin-associated protein complex (DAPC) from detergent-solubilized skeletal muscle. Immunoprecipitation, coupled with shotgun proteomics, has allowed us to identify new dystroglycan-associated proteins and define changed associations that occur within the DAPC in dystrophic skeletal muscles. In addition, we describe changes that result from overexpression of Galgt2, a normally synaptic muscle glycosyltransferase that can modify alpha dystroglycan and inhibit the development of muscular dystrophy when it is overexpressed. These studies identify new dystroglycan-associated proteins that may participate in dystroglycan's roles, both positive and negative, in muscular dystrophy. PMID:22775139

  5. Dystrophin deficient cardiomyopathy in mouse: Expression of Nox4 and Lox are associated with fibrosis and altered functional parameters in the heart

    PubMed Central

    Spurney, Christopher F.; Knoblach, Susan; Pistilli, Emidio E.; Nagaraju, Kanneboyina; Martin, Gerard R.; Hoffman, Eric P.

    2008-01-01

    Duchenne muscular dystrophy (DMD; dystrophin-deficiency) causes dilated cardiomyopathy in the second decade of life in affected males. We studied the dystrophin-deficient mouse heart (mdx) using high frequency echocardiography, histomorphometry, and gene expression profiling. Heart dysfunction was prominent at 9-10 months of age and showed significantly increased LV internal diameter (end systole) and decreased posterior wall thickness. This cardiomyopathy was associated with a 30% decrease in shortening fraction. Histologically, there was a 10-fold increase in connective tissue volume (fibrosis). mRNA profiling with RT-PCR validation showed activation of key pro-fibrotic genes, including Nox4 and Lox. The Nox gene family expression differed in mdx heart and skeletal muscle, where Nox2 was specifically induced in skeletal muscle while Nox4 was specifically induced in heart. This is the first report of an altered profibrotic gene expression profile in cardiac tissue of dystrophic mice showing echocardiographic evidence of cardiomyopathy. PMID:18440230

  6. Treatment of gastric and duodenum ulcers by means of copper laser irradiation

    NASA Astrophysics Data System (ADS)

    Averbush, G. I.; Beliy, K. P.; Berezin, J. D.; Gidkov, B. N.; Solovjov, A. F.

    1996-04-01

    One of the leading problems of gastroenterology of our country is the task of ulcerative disease of the stomach treatment, duodenum treatment and diseases of large intestinal treatment. In spite of introduction of clinical practice the new anti-ulcerous preparations, the process of ulcerative disease of the stomach treatment remains a long process and makes in average about 2 - 4 weeks for acute ulcers and 2 - 3 months for chronic ulcers. In accordance with a world statistics an ulcerative disease of the stomach remains a very spread sickness (on the average 5 cases on 1 thousand persons), and even in the majority of countries at the last decade an increase of number of illness, stipulated by breach of ecology, stress, general worsening of human living conditions took place. Notice that the growth of exposure of morbidity in the area of alimentary tract to a certain extent is connected with introduction into diagnostics practice the endoscopical methods during a checkup of patients. Within Russia the statistics data give the following picture: (1) 500 - 700 persons per 100 thousand of inhabitants have an ulcerative disease and 15 - 20 persons from them have a steady resistance to an anti- ulcerative medicamental therapy. (2) At complex and electoral treatment of ulcerative disease patients with various preparations an epithelization have 60 - 94% of patients in the time of 20 - 48 days. (3) Insufficient efficiency of means of a conservative treatment results in that 50% of the sicks need urgent operations, sometimes at complicated for them conditions.

  7. Expression of nutrient transporters in duodenum, jejunum, and ileum of Eimeria maxima-infected broiler chickens.

    PubMed

    Fetterer, Raymond H; Miska, Katarzyna B; Jenkins, Mark C; Wong, Eric A

    2014-10-01

    The uptake of amino acids is mediated by active transporters located on the basolateral and brush border membranes of intestinal epithelial cells. The current study investigated the expression of amino acid transporters (AAT) and other genes in the intestine of chicks infected with Eimeria maxima. At 7-day postinfection (PI), tissue from each intestinal segment (duodenum, jejunum, and ileum) was taken from birds inoculated with 3 × 10(3) oocysts/bird and processed to recover RNA. Analysis of gene expression was performed using real-time reverse transcription polymerase chain reaction (qRT-PCR). Results were given as relative expression using β₂-microglobulin as an endogenous control. All the genes studied were expressed in three segments of the intestines, and expression of the genes was altered by infection with E. maxima. Even though the jejunum is considered the parasite's primary predilection site, there was no segment-related difference in expression of most of the genes studied. The antimicrobial peptide (LEAP2) was downregulated in all three segments of the intestine. The results also demonstrate that transporters associated with brush border membranes were downregulated while transporters associated with the basolateral membranes were upregulated and that E. maxima alters the expression of AAT and LEAP2 throughout the small intestine. PMID:25193050

  8. Common Bile Duct Stones Detected After Cholecystectomy:Advancement into the Duodenum via the Percutaneous Route

    SciTech Connect

    Ozcan, Nevzat Erdogan, Nuri; Baskol, Mevlut

    2003-04-15

    Purpose: To report our experience in the use of percutaneous extraction of common bile duct stones detected in the post-cholecystectomy period. Methods: Forty-two patients in whom endoscopic cannulation and/or sphincterotomy had failed or could not be done due to several reasons underwent balloon dilatation of the ampulla of Vater and subsequent advancement of the stones via the percutaneous transhepatic route or T-tube tract. Results: The procedure was successful in 42 cases. In three patients, stones were crushed in the common bile duct and pushed as fragments into the duodenum. In all cases transient adverse effects were observed. There were no major complications. All cases were checked with ultrasonography for 6 months after the procedure. Conclusion: Percutaneous extraction of common bile duct stones is an effective method of treatment with a high success rate,low complication rate and shorter hospital stay. It may serve as an alternative method in cases where endoscopic removal of stones fails.

  9. Treatment with human immunoglobulin G improves the early disease course in a mouse model of Duchenne muscular dystrophy.

    PubMed

    Zschüntzsch, Jana; Zhang, Yaxin; Klinker, Florian; Makosch, Gregor; Klinge, Lars; Malzahn, Dörthe; Brinkmeier, Heinrich; Liebetanz, David; Schmidt, Jens

    2016-01-01

    Duchenne muscular dystrophy (DMD) is a severe hereditary myopathy. Standard treatment by glucocorticosteroids is limited because of numerous side effects. The aim of this study was to test immunomodulation by human immunoglobulin G (IgG) as treatment in the experimental mouse model (mdx) of DMD. 2 g/kg human IgG compared to human albumin was injected intraperitoneally in mdx mice at the age of 3 and 7 weeks. Advanced voluntary wheel running parameters were recorded continuously. At the age of 11 weeks, animals were killed so that blood, diaphragm, and lower limb muscles could be removed for quantitative PCR, histological analysis and ex vivo muscle contraction tests. IgG compared to albumin significantly improved the voluntary running performance and reduced muscle fatigability in an ex vivo muscle contraction test. Upon IgG treatment, serum creatine kinase values were diminished and mRNA expression levels of relevant inflammatory markers were reduced in the diaphragm and limb muscles. Macrophage infiltration and myopathic damage were significantly ameliorated in the quadriceps muscle. Collectively, this study demonstrates that, in the early disease course of mdx mice, human IgG improves the running performance and diminishes myopathic damage and inflammation in the muscle. Therefore, IgG may be a promising approach for treatment of DMD. Two monthly intraperitoneal injections of human immunoglobulin G (IgG) improved the early 11-week disease phase of mdx mice. Voluntary running was improved and serum levels of creatine kinase were diminished. In the skeletal muscle, myopathic damage was ameliorated and key inflammatory markers such as mRNA expression of SPP1 and infiltration by macrophages were reduced. The study suggests that IgG could be explored as a potential treatment option for Duchenne muscular dystrophy and that pre-clinical long-term studies should be helpful. PMID:26230042

  10. Successful surgical management for duodenum obstruction in a 66 year-old woman previously undiagnosed intestinal malrotation.

    PubMed

    Motomura, Takashi; Takahashi, Ikuo; Noguchi, Shinichi; Ochi, Tomohiro; Kajiwara, Yuichiro; Mano, Yohei; Nakanishi, Ryota; Fujinaka, Yoshihiko; Nishida, Kojiro; Yamashita, Yo-ichi; Saeki, Hiroshi; Kawanaka, Hirofumi; Morita, Masaru; Ikeda, Tetsuo; Soejima, Yuji; Nishizaki, Takashi; Maehara, Yoshihiko

    2013-12-01

    Intestinal malrotation is a congenital abnormality and is rarely seen in the adulthood. Most adult cases would be classified to the non-rotation type with Ladd's band and Ladd procedure is the treatment of choice. A 66 year-old woman admitted to our hospital due to duodenum obstruction. Several tests revealed that she had intestinal malrotation previously undiagnosed. Operative findings showed the fusion of duodenum with jejunum by the incomplete Treitz ligament. There was no Ladd's band and the right colon was unfixed. Dissection of the fusion completely released her symptom and she discharged without any complication. This is the first report of untypically intestinal malrotation in the adulthood without Ladd's band. PMID:24693686

  11. Preparation and characterization of a novel pH-sensitive coated microsphere for duodenum-specific drug delivery.

    PubMed

    Zhou, Dan; Zhu, Xi; Wang, Yang; Jin, Yun; Xu, Xuefan; Fan, Tingting; Liu, Yan; Zhang, Zhirong; Huang, Yuan

    2012-05-01

    The aim of this study is to develop a duodenum-specific drug delivery system on the basis of a pH-sensitive coating and a mucoadhesive inner core for eradication of Helicobacter pylori (H. pylori) in the ulcer duodenum. Hydroxypropyl methylcellulose acetate maleate (HPMCAM) was used as the pH-sensitive material, which dissolves around pH 3.0. The mucoadhesive microspheres loaded with furazolidone (FZD-ad-MS) were prepared by the emulsification-solvent evaporation method using Carbopol 971NP as the mucoadhesive polymer. The prepared pH-sensitive coated mucoadhesive microspheres (AM-coated-MS) were characterized in regards to particle size, drug loading efficiency, morphological change, drug stability, drug release and in vitro anti-H. pylori activity. The particle size was 160.97 ± 47.24 μm and 336.44 ± 129.34 μm, and the drug content was 42.33 ± 3.43% and 10.96 ± 1.29% for FZD-ad-MS and AM-coated-MS, respectively. The morphological changes in different pH media were characterized by scanning electron microscopy (SEM). HPMCAM coating improved the stability of the FZD-ad-MS and these particles were expected to remain intact until their arrival in the duodenum. The drug release was extremely suppressed at pH 1.2 for AM-coated-MS, but increased at pH 4.0 after regeneration of FZD-ad-MS. In addition, FZD-ad-MS exhibited excellent anti-H. pylori activity in vitro. Thus, the HPMCAM-coated microspheres developed in this study hold great promise for use as a duodenum-specific drug delivery system for H. pylori clearance. PMID:22644851

  12. Mechanism for enhanced absorption of a solid dispersion formulation of LY2300559 using the artificial stomach duodenum model.

    PubMed

    Polster, Christopher S; Wu, Sy-Juen; Gueorguieva, Ivelina; Sperry, David C

    2015-04-01

    An artificial stomach duodenum (ASD) model has been used to demonstrate the performance difference between two formulations of LY2300559, a low-solubility acidic developmental drug. The two formulations investigated were a conventional high-shear wet granulation (HSWG) formulation and a solid dispersion formulation. A pharmacokinetic study in humans demonstrated the enhanced performance of the solid dispersion formulation relative to the HSWG formulation. The Cmax and AUC of the solid dispersion was 2.6 and 1.9 times greater, respectively, compared to the HSWG formulation. In the ASD, the solid dispersion formulation performance was characterized by three main phases: (1) rapid release in the stomach, creating a supersaturated concentration of drug, (2) precipitation in the stomach, and (3) rapid redissolution of the precipitate in the duodenum to concentration levels that are supersaturated relative to crystalline drug. A series of complementary experiments were employed to describe this performance behavior mechanistically. Imaging experiments with a pH indicating dye showed that local pH gradients from meglumine in the solid dispersion formulation were responsible for creating a high initial supersaturation concentration in the stomach. Upon dissipation of meglumine, the drug precipitated in the stomach as an amorphous solid. Because the precipitated drug is in an amorphous form, it can then rapidly redissolve as it transits to the more neutral environment of the duodenum. This unexpected sequence of physical state changes gives a mechanistic explanation for the enhanced in vivo performance of the solid dispersion formulation relative to the HSWG formulation. PMID:25723790

  13. Low intensity training of mdx mice reduces carbonylation and increases expression levels of proteins involved in energy metabolism and muscle contraction.

    PubMed

    Hyzewicz, Janek; Tanihata, Jun; Kuraoka, Mutsuki; Ito, Naoki; Miyagoe-Suzuki, Yuko; Takeda, Shin'ichi

    2015-05-01

    High intensity training induces muscle damage in dystrophin-deficient mdx mice, an animal model for Duchenne muscular dystrophy. However, low intensity training (LIT) rescues the mdx phenotype and even reduces the level of protein carbonylation, a marker of oxidative damage. Until now, beneficial effects of LIT were mainly assessed at the physiological level. We investigated the effects of LIT at the molecular level on 8-week-old wild-type and mdx muscle using 2D Western blot and protein-protein interaction analysis. We found that the fast isoforms of troponin T and myosin binding protein C as well as glycogen phosphorylase were overcarbonylated and downregulated in mdx muscle. Some of the mitochondrial enzymes of the citric acid cycle were overcarbonylated, whereas some proteins of the respiratory chain were downregulated. Of functional importance, ATP synthase was only partially assembled, as revealed by Blue Native PAGE analysis. LIT decreased the carbonylation level and increased the expression of fast isoforms of troponin T and of myosin binding protein C, and glycogen phosphorylase. In addition, it increased the expression of aconitate hydratase and NADH dehydrogenase, and fully restored the ATP synthase complex. Our study demonstrates that the benefits of LIT are associated with lowered oxidative damage as revealed by carbonylation and higher expression of proteins involved in energy metabolism and muscle contraction. Potentially, these results will help to design therapies for DMD based on exercise mimicking drugs. PMID:25660994

  14. Effective Dystrophin Restoration by a Novel Muscle-Homing Peptide–Morpholino Conjugate in Dystrophin-Deficient mdx Mice

    PubMed Central

    Gao, Xianjun; Zhao, Jingwen; Han, Gang; Zhang, Yajie; Dong, Xue; Cao, Limin; Wang, Qingsong; Moulton, Hong M; Yin, HaiFang

    2014-01-01

    Antisense oligonucleotide (AO)–mediated splice correction therapy for Duchenne muscular dystrophy has shown huge promise from recent phase 2b clinical trials, however high doses and costs are required and targeted delivery can lower both of these. We have previously demonstrated the feasibility of targeted delivery of AOs by conjugating a chimeric peptide, consisting of a muscle-specific peptide and a cell-penetrating peptide, to AOs in mdx mice. Although increased uptake in muscle was observed, the majority of peptide–AO conjugate was found in the liver. To search for more effective muscle-homing peptides, we carried out in vitro biopanning in myoblasts and identified a novel 12-mer peptide (M12) showing preferential binding to skeletal muscle compared to the liver. When conjugated to phosphorodiamidate morpholino oligomers, ~25% of normal level of dystrophin expression was achieved in body-wide skeletal muscles in mdx mice with significant recovery in grip strength, whereas <2% in corresponding tissues treated with either muscle-specific peptide–phosphorodiamidate morpholino oligomer or unmodified phosphorodiamidate morpholino oligomer under identical conditions. Our data provide evidences for the first time that a muscle-homing peptide alone can enhance AO delivery to muscle without appreciable toxicity at 75 mg/kg, suggesting M12-phosphorodiamidate morpholino oligomer can be an alternative option to current AOs. PMID:24732757

  15. Sulforaphane mitigates muscle fibrosis in mdx mice via Nrf2-mediated inhibition of TGF-β/Smad signaling.

    PubMed

    Sun, Chengcao; Li, Shujun; Li, Dejia

    2016-02-15

    Sulforaphane (SFN), an activator of NF-E2-related factor 2 (Nrf2), has been found to have an antifibrotic effect on liver and lung. However, its effects on dystrophic muscle fibrosis remain unknown. This work was undertaken to evaluate the effects of SFN-mediated activation of Nrf2 on dystrophic muscle fibrosis. Male mdx mice (age 3 mo) were treated with SFN by gavage (2 mg/kg body wt per day) for 3 mo. Experimental results demonstrated that SFN remarkably attenuated skeletal and cardiac muscle fibrosis as indicated by reduced Sirius Red staining and immunostaining of the extracellular matrix. Moreover, SFN significantly inhibited the transforming growth factor-β (TGF-β)/Smad signaling pathway and suppressed profibrogenic gene and protein expressions such as those of α-smooth muscle actin (α-SMA), fibronectin, collagen I, plasminogen activator inhibitor-1 (PAI-1), and tissue inhibitor metalloproteinase-1 (TIMP-1) in an Nrf2-dependent manner. Furthermore, SFN significantly decreased the expression of inflammatory cytokines CD45, TNF-α, and IL-6 in mdx mice. In conclusion, these results show that SFN can attenuate dystrophic muscle fibrosis by Nrf2-mediated inhibition of the TGF-β/Smad signaling pathway, which indicates that Nrf2 may represent a new target for dystrophic muscle fibrosis. PMID:26494449

  16. Partial restoration of cardiac function with ΔPDZ nNOS in aged mdx model of Duchenne cardiomyopathy

    PubMed Central

    Lai, Yi; Zhao, Junling; Yue, Yongping; Wasala, Nalinda B.; Duan, Dongsheng

    2014-01-01

    Transgenic gene deletion/over-expression studies have established the cardioprotective role of neuronal nitric oxide synthase (nNOS). However, it remains unclear whether nNOS-mediated heart protection can be translated to gene therapy. In this study, we generated an adeno-associated virus (AAV) nNOS vector and tested its therapeutic efficacy in the aged mdx model of Duchenne cardiomyopathy. A PDZ domain-deleted nNOS gene (ΔPDZ nNOS) was packaged into tyrosine mutant AAV-9 and delivered to the heart of ∼14-month-old female mdx mice, a phenotypic model of Duchenne cardiomyopathy. Seven months later, we observed robust nNOS expression in the myocardium. Supra-physiological ΔPDZ nNOS expression significantly reduced myocardial fibrosis, inflammation and apoptosis. Importantly, electrocardiography and left ventricular hemodynamics were significantly improved in treated mice. Additional studies revealed increased phosphorylation of phospholamban and p70S6K. Collectively, we have demonstrated the therapeutic efficacy of the AAV ΔPDZ nNOS vector in a symptomatic Duchenne cardiomyopathy model. Our results suggest that the cardioprotective role of ΔPDZ nNOS is likely through reduced apoptosis, enhanced phospholamban phosphorylation and improved Akt/mTOR/p70S6K signaling. Our study has opened the door to treat Duchenne cardiomyopathy with ΔPDZ nNOS gene transfer. PMID:24463882

  17. Metagenomics Reveals Dysbiosis and a Potentially Pathogenic N. flavescens Strain in Duodenum of Adult Celiac Patients

    PubMed Central

    D'Argenio, Valeria; Casaburi, Giorgio; Precone, Vincenza; Pagliuca, Chiara; Colicchio, Roberta; Sarnataro, Daniela; Discepolo, Valentina; Kim, Sangman M; Russo, Ilaria; Del Vecchio Blanco, Giovanna; Horner, David S; Chiara, Matteo; Pesole, Graziano; Salvatore, Paola; Monteleone, Giovanni; Ciacci, Carolina; Caporaso, Gregory J; Jabrì, Bana; Salvatore, Francesco; Sacchetti, Lucia

    2016-01-01

    OBJECTIVES: Celiac disease (CD)-associated duodenal dysbiosis has not yet been clearly defined, and the mechanisms by which CD-associated dysbiosis could concur to CD development or exacerbation are unknown. In this study, we analyzed the duodenal microbiome of CD patients. METHODS: The microbiome was evaluated in duodenal biopsy samples of 20 adult patients with active CD, 6 CD patients on a gluten-free diet, and 15 controls by DNA sequencing of 16S ribosomal RNA libraries. Bacterial species were cultured, isolated and identified by mass spectrometry. Isolated bacterial species were used to infect CaCo-2 cells, and to stimulate normal duodenal explants and cultured human and murine dendritic cells (DCs). Inflammatory markers and cytokines were evaluated by immunofluorescence and ELISA, respectively. RESULTS: Proteobacteria was the most abundant and Firmicutes and Actinobacteria the least abundant phyla in the microbiome profiles of active CD patients. Members of the Neisseria genus (Betaproteobacteria class) were significantly more abundant in active CD patients than in the other two groups (P=0.03). Neisseria flavescens (CD-Nf) was the most abundant Neisseria species in active CD duodenum. Whole-genome sequencing of CD-Nf and control-Nf showed genetic diversity of the iron acquisition systems and of some hemoglobin-related genes. CD-Nf was able to escape the lysosomal compartment in CaCo-2 cells and to induce an inflammatory response in DCs and in ex-vivo mucosal explants. CONCLUSIONS: Marked dysbiosis and an abundance of a peculiar CD-Nf strain characterize the duodenal microbiome in active CD patients thus suggesting that the CD-associated microbiota could contribute to the many inflammatory signals in this disorder. PMID:27045926

  18. Circadian gastric acidity in Helicobacter pylori positive ulcer patients with and without gastric metaplasia in the duodenum.

    PubMed Central

    Savarino, V; Mela, G S; Zentilin, P; Mele, M R; Lapertosa, L; Patetta, R; Dallorto, E; Vassallo, A; Mansi, C; Vigneri, S; Celle, G

    1996-01-01

    BACKGROUND: The presence of gastric metaplasia allows helicobacter pylori to colonise the duodenum and this condition is thought to be acquired as a response to acid hypersecretion. This functional disorder, however, is present only in a subgroup of duodenal ulcer patients and, in addition, surface gastric metaplasia has been frequently found in the proximal duodenum of normal subjects and patients with non-ulcer dyspepsia, who cannot be certainly considered as acid hypersecretors. AIMS: To clarify the role of acid in inducing gastric type epithelium in the duodenum. This study aimed at assessing whether the pattern of circadian gastric acidity differs between H pylori positive duodenal ulcer patients with and without duodenal gastric metaplasia. PATIENTS: Seventy one patients with duodenal ulcer confirmed by endoscopy and who were found to be positive for H pylori infection by histology on antrum biopsy specimens were enrolled into this study. METHODS: Gastric type epithelium in the duodenum was found in 49 of 71 ulcer patients (69%). Continuous 24 hour gastric pH metry was performed in 50 healthy subjects and in the two subgroups of duodenal ulcer patients with and without gastric metaplasia in the duodenum. Gastric acidity was calculated for 24 hours (1700-1659), night (2000-0759) and day-time (0800-1959). RESULTS: Ulcer patients without gastric metaplasia showed a significantly higher gastric acidity (p < 0.001) than controls for every time interval considered, while the ulcer subgroup with gastric metaplasia was more acid than healthy subjects (p < 0.001) during the whole 24 hour period and the daytime. There was no difference between the two subgroups of duodenal ulcer patients with and without gastric metaplasia during the various time segments analysed. CONCLUSION: The findings confirm that the circadian gastric acidity of duodenal ulcer patients is higher than that of controls. As there is no difference in gastric pH between duodenal ulcer patients with

  19. Effects of trimebutine maleate (TM-906) on the spontaneous contraction of isolated duodenum and ileum in both guinea pigs and rabbits.

    PubMed

    Takenaga, H; Magaribuchi, T; Tamaki, H

    1986-01-01

    Effects of trimebutine maleate (TM-906) on the spontaneous contraction of isolated duodenum and ileum were studied in both guinea pigs and rabbits. In the duodenum and ileum of both guinea pigs and rabbits, TM-906 (10(-6) g/ml, 10(-5) g/ml) produced a potentiation of the spontaneous contraction in preparations with low contractile activity (low tone or small contraction), while it caused an inhibition of the spontaneous contraction in preparations with high contractile activity (high tone or large contraction). The potentiation of spontaneous contraction by TM-906 was more pronounced in the ileum than in the duodenum of both guinea pigs and rabbits. When the spontaneous contraction of duodenum and ileum was decreased by atropine, the potentiation of spontaneous contraction by TM-906 was further augmented and was more pronounced in the ileum than in the duodenum. When the spontaneous contraction was remarkably potentiated by physostigmine or acetylcholine, TM-906 markedly inhibited the potentiated spontaneous contraction, and the potentiation by TM-906 seen in preparations with low contractile activity disappeared. From these results, it is concluded that TM-906 produces, depending on the contractile activity of the preparations, a potentiation or an inhibition of the spontaneous contraction of duodenum and ileum in both guinea pigs and rabbits and that the potentiation by TM-906 is more pronounced in the ileum than in the duodenum. It is suggested that the endogenous acetylcholine partly modifies the effects of TM-906, but that it does not relate to the more pronounced potentiation by TM-906 in the ileum than in the duodenum. PMID:3959348

  20. Overexpression of Galgt2 in skeletal muscle prevents injury resulting from eccentric contractions in both mdx and wild-type mice.

    PubMed

    Martin, Paul T; Xu, Rui; Rodino-Klapac, Louise R; Oglesbay, Elaine; Camboni, Marybeth; Montgomery, Chrystal L; Shontz, Kim; Chicoine, Louis G; Clark, K Reed; Sahenk, Zarife; Mendell, Jerry R; Janssen, Paul M L

    2009-03-01

    The cytotoxic T cell (CT) GalNAc transferase, or Galgt2, is a UDP-GalNAc:beta1,4-N-acetylgalactosaminyltransferase that is localized to the neuromuscular synapse in adult skeletal muscle, where it creates the synaptic CT carbohydrate antigen {GalNAcbeta1,4[NeuAc(orGc)alpha2, 3]Galbeta1,4GlcNAcbeta-}. Overexpression of Galgt2 in the skeletal muscles of transgenic mice inhibits the development of muscular dystrophy in mdx mice, a model for Duchenne muscular dystrophy. Here, we provide physiological evidence as to how Galgt2 may inhibit the development of muscle pathology in mdx animals. Both Galgt2 transgenic wild-type and mdx skeletal muscles showed a marked improvement in normalized isometric force during repetitive eccentric contractions relative to nontransgenic littermates, even using a paradigm where nontransgenic muscles had force reductions of 95% or more. Muscles from Galgt2 transgenic mice, however, showed a significant decrement in normalized specific force and in hindlimb and forelimb grip strength at some ages. Overexpression of Galgt2 in muscles of young adult mdx mice, where Galgt2 has no effect on muscle size, also caused a significant decrease in force drop during eccentric contractions and increased normalized specific force. A comparison of Galgt2 and microdystrophin overexpression using a therapeutically relevant intravascular gene delivery protocol showed Galgt2 was as effective as microdystrophin at preventing loss of force during eccentric contractions. These experiments provide a mechanism to explain why Galgt2 overexpression inhibits muscular dystrophy in mdx muscles. That overexpression also prevents loss of force in nondystrophic muscles suggests that Galgt2 is a therapeutic target with broad potential applications. PMID:19109526

  1. Clear cell sarcoma of tendons and aponeuroses (malignant melanoma of soft parts) in the duodenum: the first visceral case.

    PubMed

    Ekfors, T O; Kujari, H; Isomäki, M

    1993-03-01

    An ulcerated tumour was removed by a Whipple's operation from the descending part of the duodenum of a 38-year-old male. The tumour cells were mainly spindle-shaped, arranged in nests and had very prominent nucleoli. A few cells contained melanin and melanosomes. Immunoreactivity for S-100 protein and focally for HMB-45 was observed. These features are diagnostic for clear cell sarcoma of tendons and aponeuroses. Because no other primary tumour could be found and the search for similar cases from the literature was unsuccessful, we believe that this tumour is the first reported clear cell sarcoma in a visceral location. PMID:7684355

  2. Postruminal synthesis modifies the odd- and branched-chain fatty acid profile from the duodenum to milk.

    PubMed

    Vlaeminck, B; Gervais, R; Rahman, M M; Gadeyne, F; Gorniak, M; Doreau, M; Fievez, V

    2015-07-01

    Milk odd- and branched-chain fatty acids (OBCFA) have been suggested as potential biomarkers for rumen function. The potential of milk OBCFA as a biomarker depends on whether their profile reflects the profile observed in the duodenum. The objective of this study was to evaluate whether the OBCFA profile in duodenum samples is reflected in plasma and milk. For this, 2 dairy cattle experiments were used. In experiment 1, 4 Holstein cows fitted with rumen and proximal duodenum cannulas were used in a 4×4 Latin square design. The treatments consisted of 2 nitrogen levels (143 vs. 110g of crude protein/kg of dry matter for high and low N, respectively) combined with either 1 of the 2 energy sources (i.e., starch from barley, corn, and wheat or fiber from soybean hulls and dehydrated beet pulp). In experiment 2, 4 Holstein cows fitted with rumen and proximal duodenum cannulas were used in a 3×3 Latin square design, with the treatments consisting of 3 diets: (1) RNB-, a diet with a crude protein content of 122g/kg of dry matter, predicted to provide protein digested in the small intestine according to the requirement of the animals, but with a shortage of rumen degradable protein; (2) RNB- to which 6g/d of niacin was added through inclusion in the mineral and vitamin premix, and (3) RNB- to which urea was added to balance rumen degradable N supply resulting in a CP content of 156g/kg of dry matter. In both experiments, samples of duodenal digesta, plasma, and milk were collected and analyzed for fatty acids. Additionally, lipids in plasma samples were separated in lipid classes and analyzed for fatty acids. The OBCFA profile in milk was enriched in 15:0, iso-17:0, anteiso-17:0, and cis-9-17:1 as compared with duodenal samples, and milk secretions even exceeded duodenal flows, which suggests occurrence of postruminal synthesis, such as de novo synthesis, desaturation, and elongation. The postruminal modification of the OBCFA profile might hamper the application of OBCFA

  3. Detailed ordering of markers localizing to the Xq26-Xqter region of the human X chromosome by the use of an interspecific Mus spretus mouse cross

    SciTech Connect

    Avner, P.; Amar, L.; Arnaud, D.; Hanauer, A.; Cambrou, J.

    1987-03-01

    Five probes localizing to the Xq26-Xqter region of the human X chromosome have been genetically mapped on the mouse X chromosome using an interspecific cross involving Mus spretus to a contiguous region lying proximally to the Tabby (Ta) locus. Pedigree and recombinational analysis establish the marker order as being Hprt-FIX-c11-G6PD-St14-1. The size of this contiguous region is such that the X-linked muscular dystrophy (mdx) mouse mutation probably maps within this segment. This in turn suggests that it is highly improbable that the mouse mdx locus represents a model for Duchenne muscular dystrophy (DMD). It is, however, compatible with the idea that this mutation may correspond in man to Emery Dreifuss muscular dystrophy. The high frequency of restriction fragment length polymorphisms found in this interspecific system for all the human cross-reacting probes examined up until now, using only a limited number of restriction enzymes, suggests that the Mus spretus mapping system may be of great potential value for establishing the linkage relationships existing in man when conserved chromosomal regions are concerned and human/mouse cross-reacting probes are available or can be obtained.

  4. Dystrophin expression following the transplantation of normal muscle precursor cells protects mdx muscle from contraction-induced damage.

    PubMed

    Rousseau, Joel; Dumont, Nicolas; Lebel, Carl; Quenneville, Simon P; Côté, Claude H; Frenette, Jérome; Tremblay, Jacques P

    2010-01-01

    Duchenne muscular dystrophy (DMD) is the most frequent muscular dystrophy. Currently, there is no cure for the disease. The transplantation of muscle precursor cells (MPCs) is one of the possible treatments, because it can restore the expression of dystrophin in DMD muscles. In this study, we investigated the effects of myoblasts injected with cardiotoxin on the contractile properties and resistance to eccentric contractions of transplanted and nontransplanted muscles. We used the extensor digitorum longus (EDL) as a model for our study. We conclude that the sole presence of dystrophin in a high percentage of muscle fibers is not sufficient by itself to increase the absolute or the specific force in the EDL of transplanted mdx muscle. This lack of strength increase may be due to the extensive damage that was produced by the cardiotoxin, which was coinjected with the myoblasts. However, the dystrophin presence is sufficient to protect muscle from eccentric damage as indicated by the force drop results. PMID:20650035

  5. Fluid secretion in the duodenum and intestinal handling of water and electrolytes in Zollinger-Ellison syndrome.

    PubMed

    Rambaud, J C; Modigliani, R; Emonts, P; Matuchansky, C; Vidon, N; Besterman, H; Bernier, J J

    1978-12-01

    The slow marker perfusion technique was used in five patients with the Zollinger-Ellison syndrome in order to determine the basal and postcibal flow rates of fluids passing the duodenojejunal junction and distal ileum, and the composition of those fluids. Fecal water and electrolyte excretions were also measured. The 24-hr outputs at the ligament of Treitz were markedly increased, while fecal losses were normal or only slightly increased. Thus, the overall intestinal reabsorption of water was 96%. Fasting rates of fluid and electrolyte flow at the ligament of Treitz were also measured during a basal period, followed by a period of continuous gastric aspiration. Removal of gastric secretion had the following effects on the fluid passing through the duodenum: (1) dramatic decrease in flow rate; (2) an increase in osmolality, from hypotonicity to isotonicity; (3) rise of pH, from acid to alkaline values; (4) a decrease of PCO2, from high to normal values. No increase in fasting plasma levels of immunoreactive secretin and motilin was observed in Zollinger-Ellison syndrome, whereas normal subjects respond to acid in the duodenum by a marked rise in the circulating levels of these hormones. These facts suggest that, in Zollinger-Ellison syndrome: (1) the ability of the small bowel and colon to reabsorb water and electrolytes is normal: (2) duodenal dissipation of hydrogen ions is mainly due to intraluminal neutralization by bicarbonate; and (3) stimulation of water and electrolyte secretion by the pancreas is inadequate. PMID:32768

  6. A case of primary adenocarcinoma of the third portion of the duodenum resected by laparoscopic and endoscopic cooperating surgery

    PubMed Central

    Tamaki, Ichiro; Obama, Kazutaka; Matsuo, Koichi; Kami, Kazuhiro; Uemoto, Yusuke; Sato, Teruyuki; Ito, Tetsuo; Tamaki, Nobuyuki; Kubota, Keiko; Inoue, Hidenobu; Yamamoto, Eiji; Morimoto, Taisuke

    2015-01-01

    Introduction We report a case of primary adenocarcinoma in the third portion of the duodenum (D3) curatively resected by laparoscopic and endoscopic cooperating surgery (LECS). Presentation of case A 65-year-old woman had a routine visit to our hospital for a follow-up of rectal cancer resected curatively 2 years ago. A routine screening gastroduodenal endoscopy revealed an elevated lesion of 20 mm in diameter in the D3. The preoperative diagnosis was adenoma with high-grade dysplasia; however, suspicion about potential adenocarcinoma was undeniable. Curative resection was performed by LECS. Pathological examination revealed intramucosal adenocarcinoma arising from normal duodenal mucosa. The tumor was stage I (T1/N0/M0) in terms of the tumor, nodes, metastasis (TNM) classification. LECS for duodenal tumor has seldom been reported previously, and this is the first report of LECS for primary adenocarcinoma in the D3. The transverse mesocolon was removed from the head of pancreas to expose the duodenum, and the accessory right colic vein was cut; this was followed by the Kocher maneuver for mobilization of the lesion site. Discussion LECS enabled en bloc resection with adequate surgical margins and secure intra-abdominal suturing. Thorough mobilization of the mesocolon and pancreas head is essential for this procedure because it facilitates correct resection and suturing. Conclusion LECS is a feasible treatment option for duodenal neoplasms, including intramucosal adenocarcinoma, even though it exists in the D3. PMID:25723745

  7. The effect of Mn2+, Zn2+, Ba2+ and Ca2+ on spontaneous motility in sheep duodenum in vitro.

    PubMed

    Murillo, M D; Plaza, M A; Arruebo, M P

    1997-01-01

    1. The effects of several ions, Mn2+, Zn2+, Ba2+ and Ca2+, on spontaneous motility were investigated in longitudinal smooth muscle strips from sheep duodenum, in vitro. 2. Mn2+ (0.5-1.5 mM) and Zn2+ (0.5-5 mM) inhibited both the amplitude and frequency of motility in Krebs solution and in Ca(2+)-free medium. 3. Ba2+ (0.5-5 mM) evoked three types of contractile responses: (i) an increase in the frequency and a reduction of the amplitude of spontaneous contractions; (ii) a slight increase in muscle tone of the phasic contractions; and (iii) a rapid initial phasic contraction followed by slowly fading contraction. Ca2+ induced two kinds of responses in spontaneous motility: (i) a fast phasic contraction, followed by an increase in the amplitude and frequency of phasic contractions with no changes in its tone; and (ii) an increase in the amplitude of contractions. 4. The Ba(2+)-induced contractions were inhibited by EDTA, verapamil and diltiazem, but were not modified by sodium nitroprusside. The Ca(2+)-induced contractions were reduced by verapamil and diltiazem. 5. Our results show that Mn2+ and Zn2+ behave as inhibitors of sheep duodenum motility. In contrast, Ba2+ and Ca2+ stimulate motility. It is suggested that Ba2+ can penetrate the cells through voltage-dependent Ca2+ channels and behave as a partial substitute for Ca2+. PMID:9112079

  8. Observations on the submucous plexus and mucosal arteries of the dog's stomach and first part of the duodenum.

    PubMed Central

    Piasecki, C

    1975-01-01

    Arteriolar patterns of the submucous plexus were studied in all areas of the dog's stomach and in the first inch of the duodenum. There appeared to be no poverty of plexus, although in some cases the vessels were somewhat smaller in the pyloric part of the lesser curvature than elsewhere. Mucosal arteries arose from the plexus, and none appeared to have an extramural origin. In man, on the other hand, there is a poverty of the submucous plexus in the 'ulcer region', i.e. in the incisural region of the lesser curvature and in the first inch of the duodenum, associated in some cases with mucosal end arteries of extramural origin. The absence of these features in the dog, which does not suffer from spontaneous chronic ulceration, lends further support to the view that they play a role in the aetiology of the disease in man. Images Fig. 1 Fig. 2 Fig. 4A Fig. 4B Fig. 5 Fig. 6 Fig. 7 Fig. 8 PMID:1133082

  9. Altered ROS production, NF-κB activation and Interleukin-6 gene expression induced by electrical stimulation of in dystrophic mdx skeletal muscle cells

    PubMed Central

    Henríquez-Olguín, Carlos; Altamirano, Francisco; Valladares, Denisse; López, José R.; Allen, Paul D.; Jaimovich, Enrique

    2015-01-01

    Duchenne Muscular Dystrophy (DMD) is a fatal X-linked genetic disease, caused by mutations in the dystrophin gene, which cause functional loss of this protein. DMD pathology is associated with an increased production of reactive oxygen and nitrogen species (ROS and RNS). The aim of this work was to study the alterations in NF-κB activation and Interleukin-6 (IL-6) expression induced by membrane depolarization in dystrophic mdx myotubes. Membrane depolarization elicited by electrical stimulation (ES) increased p65 phosphorylation, NF-κB transcriptional activity and NF-κB-dependent IL-6 expression in wt myotubes, whereas in mdx myotubes it had the opposite effect. We have previously shown that depolarization-induced intracellular Ca2+ increases and ROS production are necessary for NF-κB activation and stimulation of gene expression in wt myotubes. Dystrophic myotubes showed a reduced amplitude and area under the curve of the Ca2+ transient elicited by ES. On the other hand, ES induced higher ROS production in mdx than wt myotubes, which were blocked by NOX2 inhibitors. Moreover, mRNA expression and protein levels of the NADPH oxidase subunits; p47phox and gp91phox were increased in mdx myotubes. Looking at ROS-dependence of NF-κB activation we found that in wt myotubes external administration of 50µM H2O2 increased NF-κB activity; after administration of 100 and 200 µM H2O2 there was no effect. In mdx myotubes there was a dose-dependent reduction in NF-κB activity in response to external administration of H2O2, with a significant effect of 100 µM and 200 µM, suggesting that ROS levels are critical for NF-κB activity. Prior blockage with NOX2 inhibitors blunted the effects of ES in both NF-κB activation and IL-6 expression. Finally, to ascertain whether stimulation of NF-κB and IL-6 gene expression by the inflammatory pathway is also impaired in mdx myotubes, we studied the effect of lipopolysaccharide (LPS) on both NF-κB activation and IL-6 expression

  10. Altered ROS production, NF-κB activation and interleukin-6 gene expression induced by electrical stimulation in dystrophic mdx skeletal muscle cells.

    PubMed

    Henríquez-Olguín, Carlos; Altamirano, Francisco; Valladares, Denisse; López, José R; Allen, Paul D; Jaimovich, Enrique

    2015-07-01

    Duchenne muscular dystrophy is a fatal X-linked genetic disease, caused by mutations in the dystrophin gene, which cause functional loss of this protein. This pathology is associated with an increased production of reactive oxygen (ROS) and nitrogen species. The aim of this work was to study the alterations in NF-κB activation and interleukin-6 (IL-6) expression induced by membrane depolarization in dystrophic mdx myotubes. Membrane depolarization elicited by electrical stimulation increased p65 phosphorylation, NF-κB transcriptional activity and NF-κB-dependent IL-6 expression in wt myotubes, whereas in mdx myotubes it had the opposite effect. We have previously shown that depolarization-induced intracellular Ca2+ increases and ROS production are necessary for NF-κB activation and stimulation of gene expression in wt myotubes. Dystrophic myotubes showed a reduced amplitude and area under the curve of the Ca2+ transient elicited by electrical stimulation. On the other hand, electrical stimuli induced higher ROS production in mdx than wt myotubes, which were blocked by NOX2 inhibitors. Moreover, mRNA expression and protein levels of the NADPH oxidase subunits: p47phox and gp91phox were increased in mdx myotubes. Looking at ROS-dependence of NF-κB activation we found that in wt myotubes external administration of 50 μM H2O2 increased NF-κB activity; after administration of 100 and 200 μM H2O2 there was no effect. In mdx myotubes there was a dose-dependent reduction in NF-κB activity in response to external administration of H2O2, with a significant effect of 100 μM and 200 μM, suggesting that ROS levels are critical for NF-κB activity. Prior blockage with NOX2 inhibitors blunted the effects of electrical stimuli in both NF-κB activation and IL-6 expression. Finally, to ascertain whether stimulation of NF-κB and IL-6 gene expression by the inflammatory pathway is also impaired in mdx myotubes, we studied the effect of lipopolysaccharide on both NF