2,685 research outputs found
Inhibition of FasL sustains phagocytic cells and delays myogenesis in regenerating muscles fibers
No full textJ Leukoc Biol. 2001 Mar;69(3):482-9.
Inhibition of fasL sustains phagocytic cells and delays myogenesis in regenerating muscle fibers.
Sandri M, Sandri C, Brun B, Giurisato E, Cantini M, Rossini K, Destro C, Arslan P, Carraro U.
Source
Department of Biomedical Sciences, University of Padova, Italy. [email protected]
Abstract
Macrophage-muscle cell interactions are complex, and the majority is unknown. The persistence of inflammatory cells in skeletal muscle could be critical for myofiber viability. In the present paper, we show that FasL plays a role in the resolution of muscle inflammation. We analyzed inflamed muscles of normal mice treated from day 3 to day 8 with a FasL inhibitor (Fas-Ig) or with control Ig. Treated muscles were collected at 3, 5, and 10 days. The treatment with recombinant Fas-Ig protein induced a severe persistence of inflammatory cells at 5 days (115,000+/-27,838 vs. 41,661+/-6848, p<0.01) and 10 days from injury (145,500+/-40,850 vs. 5000+/-1000, p<0.001). Myofiber regeneration was highly impaired (37+/-14 vs. 252+/-28, p<0.01). Apoptosis of phagocytic cells was absent during Fas-Ig treatment (0.9+/-0.6 vs. 1300+/-150, p<0.0001), but apoptotic, mononucleated cells appeared at day 10, 2 days after the suspension of Fas-Ig administration. The time course of FasL expression during muscle inflammation, at mRNA and protein level, reveals a peak during myoblast proliferation. The peak of FasL expression coincides with the peak of apoptosis of phagocytic cells. In situ hybridization shows the co-expression of FasL and MyoD mRNA in mononucleated cells, i.e., myoblasts. Experiments on the myoblast cell culture confirmed the expression of FasL in myoblasts. The findings shown here indicate one of the pathways to control myoblast-macrophage interaction and might be relevant for the control of inflammatory cells in muscle tissue. Perhaps altering FasL expression with recombinant proteins could ameliorate inflammation in degenerative myopathies and up-regulate muscle regeneration.
PMID:
11261797
[PubMed - indexed for MEDLINE
Caspase 3 expression correlates with skeletal muscle apoptosis in Duchenne and facio-scapulo human muscular dystrophy. A potential target for pharmacological treatment?
No full textJ Neuropathol Exp Neurol. 2001 Mar;60(3):302-12.
Caspase 3 expression correlates with skeletal muscle apoptosis in Duchenne and facioscapulo human muscular dystrophy. A potential target for pharmacological treatment?
Sandri M, El Meslemani AH, Sandri C, Schjerling P, Vissing K, Andersen JL, Rossini K, Carraro U, Angelini C.
Source
Department of Biomedical Sciences, Institute of Experimental and Laboratory Medicine, University of Padua, Italy.
Abstract
Apoptosis was detected in different muscular diseases, including severe dystrophin deficiency, but apoptotic mechanisms are not completely described in adult skeletal muscle. Studying patients affected by Duchenne muscular dystrophy (DMD) and by facio-scapulo-humeral dystrophy (FSHD) we showed an increase of apoptotic myonuclei, bax, and bcl-2-positive myofibers. Positive correlation was detected between apoptotic nuclei and bax expression (p < 0.01). Expression of caspases was analyzed by RNase protection. Caspase transcript was not detected in normal skeletal muscles. DMD muscles expressed caspase 8, 3, 5, 2, 7 and Granzyme B mRNAs. Low levels of caspase 6, 3, and Granzyme B transcripts were detected in FSHD patients. Tissue levels of caspase 3 protein significantly correlated with apoptotic myonuclei (p < 0.05) and with bax expression (p < 0.01). In all DMD cases the activity of caspase 3 was increased, while the FSHD samples were heterogeneous. These data indicate that human skeletal muscle fibers. during the dystrophic process, modulate the expression of caspases and that caspase 3 is involved in myofiber cell death. opening new perspective in the pharmacological treatments of muscular dystrophies, such as the use of caspase inhibitors.
PMID:
11245214
[PubMed - indexed for MEDLINE
High-resolution sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunochemical identification of the 2X and embryonic myosin heavy chains in complex mixtures of isomyosins
No full textElectrophoresis. 1995 Jan;16(1):101-4.
High-resolution sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunochemical identification of the 2X and embryonic myosin heavy chains in complex mixtures of isomyosins.
Rossini K, Rizzi C, Sandri M, Bruson A, Carraro U.
Source
Department of Biomedical Sciences, University of Padova, Italy.
Abstract
In mammals myosin heavy chains (MHC) are polypeptides with a molecular mass of about 200 kDa whose isoforms can be identified by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and immunochemistry. Electrophoretic analysis is the only method for quantitating MHC profiles in single myofibers and/or cryostat sections of biopsied muscle. We present a method for SDS-PAGE of adult rat skeletal muscle which resolves MHC into four bands: 1, 2B, 2X, and 2A from the faster to the slower migrating band. Furthermore, embryonic MHC can be also resolved in a complex mixture of isomyosins, e.g. developing or regenerating muscles. The method does not involve preparation of gradient gels or electrophoresis at low temperature. Improved reproducibility is obtained by: (i) modification of the sample buffer; (ii) use of 7% polyacrylamide in the separating gel; (iii) control of pH of running buffer by recirculation or change of the buffer during the run; and (iv) a 24 h run. The procedure is compatible with Coomassie Brilliant Blue, silver and immunoblot staining. Resolution is sufficient to permit transblotting of separated MHC after SDS-PAGE. The different isoforms are easily identified with monoclonal antibodies. The technique provides an improved method to separate MHC and quantitate MHC2X and MHCemb in complex mixtures of MHC from a few cryostat sections of normal and diseased muscle.
PMID:
7737081
[PubMed - indexed for MEDLINE
A new two-step precipitation method removes free-SDS and Thiol reagents from diluted solutions, and then allows recovery and quantitation of proteins
No full textBiochem Biophys Res Commun. 1994 Apr 29;200(2):916-24.
A new two-step precipitation method removes free-SDS and thiol reagents from diluted solutions, and then allows recovery and quantitation of proteins.
Carraro U, Doria D, Rizzi C, Sandri M.
Source
Department of Experimental Biomedical Sciences, University of Padua, Italy.
Abstract
Several polypeptides are present in a few copies per cell (i.e., membrane receptors or transcription factors), and therefore their concentration and quantitation from highly diluted solution after detergent solubilization is often an essential and difficult step in their purification by gel electrophoresis. A solution (optimized to Tris-glycine, Tris-borate, and Na-phosphate buffers) to all of these problems is here described, detailing a two-step procedure which takes advantage of the lower solubility of free potassium dodecyl sulfate (KDS) vs micellar KDS even at neutral pH. In the first step, more than 90% of free DS precipitates out from protein solution, the method being poorly sensitive to dodecyl sulfate/KCl ratio from almost no SDS to 10%. Indeed to obtain a residual 0.01% of SDS in the solution no adjustment of KCl concentration is needed from 0.1 to 2.3% SDS. When the supernatant is added with ice-cold TCA and K+, protein solubility is severely affected. Excellent protein recovery, at least 90%, is obtained with hydrophilic proteins. On the other hand, hydrophobic or low-ionic-strength-insoluble proteins are partially lost in step 1 precipitate, so that protein yield decreases, still remaining to about 80%. Furthermore our procedure simplifies determination of protein contents of diluted mercaptoethanol-SDS-solubilized proteins: since thiol reagents are discarded with the final supernatant, proteins can be quantitated in the final pellet by standard colorimetric methods.
PMID:
8179627
[PubMed - indexed for MEDLINE
Apoptosis, DNA damage and ubiquitin expression in normal and mdx muscle fibers after exercise.
No full textFEBS Lett. 1995 Oct 16;373(3):291-5.
Apoptosis, DNA damage and ubiquitin expression in normal and mdx muscle fibers after exercise.
Sandri M, Carraro U, Podhorska-Okolov M, Rizzi C, Arslan P, Monti D, Franceschi C.
Source
Department of Biomedical Sciences, University of Padova, Italy.
Abstract
The current view indicates that after eccentric exercise myofibers are mechanically damaged and therefore an inflammatory and necrotic process occurs. In the present paper we examine the possibility that apoptosis plays a role in normal and dystrophin-deficient muscles after running. We analysed for apoptosis normal and dystrophin-deficient mouse muscles after a night of spontaneous wheel-running followed by two days of rest. Terminal deoxynucleotidyl transferase-mediated end-labeling of DNA in nuclei in tissue sections and gel electrophoresis of extracted DNA showed the presence of fragmented DNA. Furthermore, ubiquitin, a protein whose appearance is related to apoptosis, increased in muscles of both dystrophic and normal runner mice. The present findings which confirm that DNA damage is absent in muscles of sedentary mice but present in muscles of runner mice offer a new hypothesis on early events of muscle damage.
PMID:
7589485
[PubMed - indexed for MEDLINE
Reciprocità e Free Riding: Un'Analisi Evolutiva
No full textL'analisi evolutiva sviluppata in questo lavoro getta luce sulla stabilità evolutiva di popolazioni motivazionalmente eterogenee composte da giocatori auto-interessati e giocatori guidati da reciprocità, nell'ambito di un contesto di interazione avente la struttura del dilemma del prigioniero
Effects of beta 1-integrin antisense phosphorothioate-modified oligonucleotide on myoblast behaviour in vitro.
No full textCell Biochem Funct. 1995 Jun;13(2):99-104.
Effects of beta 1-integrin antisense phosphorothioate-modified oligonucleotide on myoblast behaviour in vitro.
Carraro U, Bruson A, Catani C, Dalla Libera L, Massimino ML, Rizzi C, Rossini K, Sandri M, Cantini M.
Source
University of Padova, CNR Unit for Muscle Biology and Physiopathology, Department of Biomedical Sciences, Italy.
Abstract
Myoblasts gene-engineered in vitro and then injected in vivo are safe, efficient options for gene therapy. While isolation of satellite cells is routinely achieved, their proliferation potential in vitro remains a limiting factor for cell transplantation under clinical conditions. We have studied the role of reversible inhibition of gene expression by antisense oligonucleotides on the proliferation of the myogenic cells. Addition of antisense oligonucleotides to myoblast cultures has been used to inhibit specifically the expression of the beta 1-integrin subunit gene. Here we show that the effects of multiple pulses of a phosphorothioate oligodeoxinucleotide antisense on the attachment to substrata and on the proliferation of myoblasts are dose-dependent. The addition of antisense to rat myoblasts caused rounding up of the cells and most of the cells became detached after several days in culture. A single pulse did not show any consistent effect, while in the presence of continuously administered antisense, the relative numbers of myoblasts in the treated muscle culture increased. We have no evidence of inhibition of myoblast fusion under these conditions. On the other hand, [3H]-TdR incorporation, total DNA and total number of cells decreased in antisense-treated cultures thus demonstrating an inhibitory effect of the phosphorothioate oligonucleotides on DNA synthesis. These side-effects could be overcome by substituting the phosphorothioate by unmodified oligonucleotides, so decreasing the half-life of the antisense, but also its toxicity. The overall results suggest a potential role of integrin antisense strategy in modulating the potential of myoblasts to proliferate.
PMID:
7538914
[PubMed - indexed for MEDLINE
Football Mining with R
No full textThis chapter presents a data mining process for investigating the relationship between the outcome of a football match (win, lose or draw) and a set of variables describing the actions of each team, using the R environment and selected R packages for statistical computing. The analyses were implemented with parallel computing when possible. Our goals were to identify, from hundreds of covariates, those that most strongly affect the probability of winning a match and to construct a small number of composite indicators based on the most predictive variables. These two tasks were carried out using the Random Forest machine learning algorithm and Principal Component Analysis, respectively. Variable selection was performed using the novel approach developed by Sandri and Zuccolotto in 2008. Finally, we compared the results of several different classification models and algorithms (Random Forest, Classification Neural Network, K-Nearest Neighbor, Naïve Bayes classifier, and Multinomial Logit regression), assessing both their performance and the insightfulness of their results
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