A. Immunolocalization of NOS1-3 isoforms at functional compartments and biochemical analysis of NOS 1-3 and related pathway molecules (Homer, mTor, others) analyzed before and after spaceflight, i.e. pre- and post-flight biopsies, of mixed/fast and slow-type human skeletal muscle.Previous Flight Experiments
B. To study altered expression patterns of the muscular NOS/NO system and related molecules (Capon, PIN, immediate early genes, Homer) following countermeasure after short-term exposure to microgravity.
C. To study altered expression patterns of the muscular NOS/NO system and related molecules (Capon, PIN, IEGs, Homer) following countermeasure after long-term exposure to microgravity.
M. Salanova, G. Schiffl, B. Püttmann, B.G. Schoser, D. Blottner, (2008), "Molecular biomarkers monitoring human skeletal muscle fibres and micro-vasculature following long-term bed rest with and without countermeasure", Journal of Anatomy, 212, pp. 306-318.
D. Blottner, B. Püttmann, M. Salanova, G. Schiffl, J. Rittweger, H.C. Gunga, D. Felsenberg, B.G. Schoser, (2006), "Skeletal muscle deconditioning, nitric oxide (NO) biomarker, and exercise countermeasure - Five years of bed rest studies", Journal of Gravitational Physiology, 13, 2, pp. 49-58.
D. Blottner, M. Salanova, B. Püttmann, G. Schiffl, D. Felsenberg, J. Rittweger, (2006), "Human skeletal muscle structure and function preserved by vibration muscle exercise following 55-days of bed rest", European Journal of Applied Physiology, 97, pp. 261-271.
A. Moukhina, B. Shenkman, D. Blottner, T. Nemirovskaya, Y. Lemesheva, B. Püttmann, I. Kozlovskaya, (2004), "Effects of support stimulation on human soleus fiber characteristics during exposure to "dry" immersion", Journal of Gravitational Physiology, 11, pp. P137-8.
J. Rudnick, B. Puttmann, P.A. Tesch, B. Alkner, B.G. Schoser, M. Salanova, K. Kirsch, H.C. Gunga, G. Schiffl, G. Lück, D. Blottner, (2004), "Differential expression of nitric oxide synthases (NOS 1-3) in human skeletal muscle following exercise countermeasure during 12 weeks of bed rest", FASEB Journal, 18, pp. 1228-1230.
M. Salanova, G. Schiffl, J. Rittweger, D. Felsenberg, D. Blottner, (2008), "Ryanodine receptor type-1 (RyR1) expression and protein S-nitrosylation pattern in human soleus myofibres following bed rest and exercise countermeasure", Histochemistry and cell biology, 130, 1, pp. 105-118.
M. Salanova, G. Schiffl, D. Blottner, (2009), "Atypical fast SERCA1a protein expression in slow myofibers and differential S-nitrosylation prevented by exercise during long term bed rest", Histochemistry and Cell Biology, 132, 4, pp. 383-394.
D. Blottner, N. Serradj, M. Salanova, C. Touma, R. Palme, M. Silva, J.M. Aerts, D. Berckmans, L. Vico, Y. Liu, A. Giuliani, F. Rustichelli, R. Cancedda, M. Jamon, (2009), "Morphological, physiological and behavioural evaluation of a Mice in Space housing system", Journal of Comparative Physiology B, 179, 4, doi: 10.1007/s00360-008-0330-4, pp. 519-533.
D.L. Belavý, O. Bock, H. Börst, G. Armbrecht, U. Gast, C. Degner, G. Beller, H. Soll, M. Salanova, H. Habazettl, M. Heer, A. de Haan, D.F. Stegeman, P. Cerretelli, D. Blottner, J. Rittweger, C. Gelfi, U. Kornak, D. Felsenberg, (2010), "The 2nd Berlin BedRest Study: protocol and implementation", Journal of Musculoskeletal and Neuronal Interactions - JMNI, 10, 3, pp. 207-219.
M. Moriggi, M. Vasso, C. Fania, D. Capitanio, G. Bonifacio, M. Salanova, D. Blottner, J. Rittweger, D. Felsenberg, P. Cerretelli, C. Gelfi, (2010), "Long-term bed rest with and without vibration exercise countermeasure: effects on human muscle protein dysregulation", Proteomics, 10, 21, doi: 10.1002/pmic.200900817, pp. 3756-3774.
M. Salanova, E. Bortoloso, G. Schiffl, M. Gutsmann, D.L. Belavy, D. Felsenberg, S. Furlan, P. Volpe, D. Blottner, (2011), "Expression and regulation of Homer in human skeletal muscle during neuromuscular junction adaptation to disuse and exercise", FASEB Journal, 25, 12, doi: 10.1096/fj.11-186049, pp. 4312-4325.
D. Sandonà, J.F. Desaphy, G.M. Camerino, E. Bianchini, S. Ciciliot, D. Danieli-Betto, G. Dobrowolny, S. Furlan, E. Germinario, K. Goto, M. Gutsmann, F. Kawano, N. Nakai, T. Ohira, Y. Ohno, A. Picard, M. Salanova, G. Schiffl, D. Blottner, A. Musarò, Y. Ohira, R. Betto, D. Conte, S. Schiaffino, (2012), "Adaptation of mouse skeletal muscle to long-term microgravity in the MDS mission", PLoS ONE, 7, 3, doi: 10.1371/journal.pone.0033232, pp. e33232.
M. Salanova, P. Volpe, D. Blottner, (2013), "Homer family regulation in skeletal muscle and neuromuscular adaptation", International Union of Biochemistry and Molecular Biology Life - IUBMB Life, 65, 9, doi: 10.1002/iub.1198, pp. 769-776.
M. Salanova, G. Schiffl, M. Gutsmann, D. Felsenberg, S. Furlan, P. Volpe, A. Clarke, D. Blottner, (2013), "Nitrosative stress in human skeletal muscle attenuated by exercise countermeasure after chronic disuse", Redox Biology, 1, doi: 10.1016/j.redox.2013.10.006, pp. 514-526.
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Figure 1: Confocal images showing activity-induced plasma membrane expression of nitric oxide synthase type-1 (NOS1, green immunofluorescence) at plasma membranes of crosssectioned biopsy samples of (A) inactively vs. (B) trained human m. soleus myofibers following extended (60 d) bed rest. (C) Without exercise (Control), NOS membrane expression is significantly decreased after bed rest. After bed rest and exercise countermeasure, NOS1 is highly preserved in both slow and fast myofibers (1 and 2). (D) Western immunoblot analysis showing altered NOS1 protein content in human skeletal muscle before (Pre) and after (Post) bed rest (exercise group). compare reference documents 3 and 5
Figure 2: Confocal microscopy showing similar distribution patterns of slow type-1 (dark) and fast type-2 (red) myofibers in (A) normal and (B) exercised human calf m. soleus following long-term (60d) bed rest. (C) Quantitative bar graph showing that the typical slow-to-fast fiber-shift (slow < fast type) of unloaded human m.soleus (Control) is prevented by exercise countermeasure (Exercise) during bed rest suggesting phenotype preservation of the soleus calf muscle. compare reference documents 3 and 5
Figure 3: Expression of Homer (green label) in the subsynaptic area of a human neuromuscular junction following exercise countermeasure. Homer proteins are regulated by motor nerve activity and help to control intramyocellular calcium balance. compare reference documents 11 and 13
Figure 4: Sketch showing nitrosative stress in human skeletal muscle via s-nitrosylated functional protein (SNO-proteins) analysis attenuated by exercise countermeasure following chronic disuse. Chronic disuse (60 days bed rest) results in excess levels of calcium-signaling SNO-proteins (RyR1, SERCA, PMCA) in leg skeletal muscle that can be normalized to baseline by exercise as countermeasure. compare reference document 14