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Pigs of different ryanodine receptor 1 (RyR1) genotypes serve as a model for the evaluation of various non-invasive in vivo methods to measure muscle energymetabolism and body composition. The main focus is set on one hand on 31P and 13C nuclear magnetic resonance spectroscopy and on the other hand on nuclear magnetic resonance imaging and dual energy X-ray absorptiometry. In addition to a reference dissection and chemical analysis, the above mentioned methods are being extensively compared to other methods in the methodology part. An invasive muscle shot biopsy serves as an additional method in order to determine the muscle fiber composition of the longissimus dorsi muscle. Totally, 111 animals have been considered in the data analysis.13C- and 31P NMR spectroscopy are very appropriate methods to measure in vivo or post mortem, non-invasively and continuously changes in the concentration of glycogen and creatine or phosphocreatine (PCr), inorganic phosphate (Pi), adenosintriphosphate (ATP), and pH-value directly in relative or absolute amounts over an „unlimited“ time period. 31P NMR spectroscopy compared to 13C NMR spectroscopy has advantages in the sensitivity and timely resolution for measuring changes in the concentration within the components of muscle metabolism. While 31P NMR spectra in vivo yield sufficient results for measuring changes of PCr, ATP, Pi and pH value within a time interval < 1minute, it takes > 5 minutes to acquire reasonable 13C NMR spectra in order to measure changes in the concentration of glycogen and creatine.Progress in the techniques of body composition analysis is mainly based onelectronic and computer driven methods in order to provide non-invasive, fast and objective measurements. The selection of the appropriate method depends on the objective (research, performance testing, production, or diagnosis) and financial budget for each single study and clinical application. Technical details considering accuracy, precision, parameter selection and maximum/minimum size of an individual affect the decision as well as does a simple, robust, environment-safe, patient-comfortable and risk-free handling of the technique.The highest accuracy for whole body studies within the imaging methods providemagnetic resonance imaging (MRI) and computer tomography (CT) followed by dualenergy x-ray absorptiometry (DXA). DXA, however, outperforms MRI and CT due toits simple handling and easy data analysis for whole body or regional body composition studies. In addition, DXA is beside the Quantitative CT the only non-invasive method which provides direct bone mineral density measurements.According to the literature survey, the neutron activation analysis as a noninvasive method should be preferred for the calibration of body compositionmeasurement devices instead of chemical analysis or dissection. Chemical analysis and total dissection will remain standard reference methods as long as neutron activation facilities are available only on a very limited basis. However, the non-invasive (imaging) methods are less prone to error sources than are the diverse dissection or chemical analysis procedures.The main advantages of the non-invasive imaging and/or spectroscopic methodsin vivo are:1. the easy way to standardize the methods with a high repeatability (>75 %),2. the opportunity of analyzing separately large volumes of interest of the whole body, body tissues and/or body parts,3. the opportunity of performing continuous measurements over an “unlimited”period of time, and 4. the harm “free” animal/patient precautions with a high degree of hygienicsafety.An advantage for magnetic resonance, ultrasound and digital imaging is providedby the function without radiation, while especially the “spiral computer tomography” is characterized by a very high speed of analysis. The muscle metabolism of the defect allele carriers at the RyR1 locus Nn and nnshows already in vivo stress associated deviations in comparison to the “normal” genotype NN. After generating muscle stress by halothane, defect genotypes react with a decline in the glycogen, phosphocreatine, ATP, and pH level. Parallel inorganic phosphate and body temperature increase significantly. In the average show the homozygous defect allele carriers a stronger metabolic distress than do the heterozygous carriers. Differences among genotypes are higher post mortem compared to the metabolism in vivo. The muscle metabolic distress is connected with a muscle fiber hypertrophy in both defect allele genotypes. During growth, the homozygous defect allele genotype deposits more muscle mass (volume) and less fat than does the heterozygous genotype followed by the homozygous normal genotype.Already at a live body weight of about 10 kg, magnetic resonance imagingprovides evidence for a larger muscle volume in the homozygous defect genotype in comparison to the two other genotypes. Beside the studied RyR1 genotypes (and the RN—-allel especially present in Hampshire), a large number of genetic, morphologic and environmental factors influence muscle metabolism and body composition in Swine -- as described in the discussion part. The RN—-allel causes a reduced glycogen depletion in vivo (and post mortem). Corresponding, the Hampshire line (≥ 50 % Hampshire genes) is beside the US-Landrace and the synthetic line “Duroc x Hampshire x US-Landrace x Spotted x Yorkshire” less obese than are the lines Duroc, “Poland China x Landrace”, Spotted, and “Duroc x Minzhu”. Spotted and “Duroc x Minzhu” are the most obese lines. In addition comparing all lines, Spotted (all Nn) responses --unexpectedly -- most severely to (muscle) stress caused by halothane administration.
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We identified a novel T cell Ag in the South African clawed toad (Xenopus laevis) by a mAb designated 2B1. This Ag is present in relatively high levels on most thymocytes, approximately 65% of splenocytes, 55% of PBL, and 65% of intestinal lymphocytes, but is rarely seen on IgM+ B cells in any of these tissues. Lymphocytes bearing the 2B1 Ag proliferate in response to stimulation with Con A or PHA, whereas the 2B1- lymphocytes are reactive to LPS. Biochemical analysis indicates that this Ag is a differentially phosphorylated glycoprotein of 71 to 82 kDa. The protein core of 64 kDa bears both N- and O-linked carbohydrate side chains. The amino-terminal protein sequence of the 2B1 Ag shares significant homology with both the macrophage scavenger receptor type 1 motif and the mammalian CD5/CD6 family. The biochemical characteristics and cellular distribution of the 2B1 Ag suggest that it represents the CD5 homologue in X. laevis. While T cells constitutively express this highly conserved molecule, Xenopus B cells acquire the CD5 homologue only when they are stimulated in the presence of T cells
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Bovine rotavirus (BRV) V1005, like 34 further cell culture-adapted strains in a 6-year survey in Upper Bavaria, Germany, is not a P12 but a P5 P-type rotavirus. The conclusion is based on dot blot hybridization with P1-, P5-, and P11-specific cDNAs, encompassing the VP8* region of major sequence diversity, and on PCR using P1-, P5-, and P11-specific primer pairs derived from the VP5* region of VP4 (VP5* and VP8*, respectively, are the larger and smaller tryptic cleavage products of VP4). Sequencing of the hyperdivergent region of VP4 confirmed the close relatedness of BRV V1005 to BRV UK, the P5 prototype virus
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T-cell receptor (TCR) -chain (TCR) and ß-chain (TCRß) genes are well characterized in mammals, while only TCRß genes have been identified in other vertebrates. To identify avian TCR genes, we used monoclonal anti-CD3 antibodies to isolate chicken TCR for peptide sequence analysis. Degenerate oligonucleotide probes were then used to isolate a candidate TCR cDNA clone that hybridized with a 1.7-kb mRNA species present only in ß T cells and in tissues populated by these cells. Southern blot analysis revealed gene rearrangement in thymocytes and ß T-cell lines. The TCR cDNA candidate encoded an openreading frame of 275 amino acids, the predicted variable (V)-, joining (J)-, and constant (C)-region amino acid sequences of which shared 40%, 60%, and 25% homology with corresponding mammalian sequences. A single C gene and 25 V genes were identified by using region-specific probes. The V cDNA probe isolated from a Vß1+ cell line reacted with transcripts from one of five Vß2+ cell lines, suggesting shared use of V genes by Vß1+ and Vß2+ T cells and the existence of other V gene families. A genomic V sequence was flanked by classical recombination signal sequences but, unlike previously defined V genes, the leader and V region were encoded by a single exon. The data indicate evolutionary conservation of the basic TCR gene structure in birds and mammals
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Natural killer (NK) cell activity appears to be conserved throughout vertebrate development but NK cells have only been well characterized in mammals. Candidate NK cells have been identified in the chicken as cytoplasmic CD3+ and surface T cell receptor (TCR)/CD3- (TCRO) lymphocytes that often express CD8. The fact that the TCRO cells are abundant in the embryonic spleen before T cells enter this organ allowed us to cultivate the embryonic TCRO cells using growth factors derived from activated adult lymphocytes. These TCRO cells were cytotoxic for an NK target cell line. They expressed cell surface CD8, a putative interleukin-2 receptor, CD45 and a receptor for IgG, but did not express CD4, major histocompatibility complex class II or immunoglobulin. Biochemical analysis of the cytoplasmic CD3 antigen revealed two of the three CD3 , and homologues, and RNA transcripts for the third. The CD3 monoclonal antibody also precipitated a 32-kDa dimer that may represent a heterodimer of different CD3 constituents. TCR and gene transcripts were not detected in the TCRO cells. These results indicate that the avian TCRO cell is the mammalian NK cell homologue. The shared evolutionary features of T cells and NK cells in birds and mammals support the idea that they derive from a common progenitor
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