| 1. |
- Abbate, F, et al.
(författare)
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Prolonged force increase following a high-frequency burst is not due to a sustained elevation of [Ca2+]i
- 2002
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Ingår i: American journal of physiology. Cell physiology. - : American Physiological Society. - 0363-6143 .- 1522-1563. ; 283:1, s. C42-C47
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Tidskriftsartikel (refereegranskat)abstract
- A brief high-frequency burst of action potentials results in a sustained force increase in skeletal muscle. The present study investigates whether this force potentiation is the result of a sustained increase of the free myoplasmic [Ca2+] ([Ca2+]i). Single fibers from mouse flexor brevis muscles were stimulated with three impulses at 150 Hz (triplet) at the start of a 350-ms tetanus or in the middle of a 700-ms tetanus; the stimulation frequency of the rest of the tetanus ranged from 20 to 60 Hz. After the triplet, force was significantly ( P < 0.05) increased between 17 and 20% when the triplet was given at the start of the tetanus and between 5 and 18% when the triplet was given in the middle ( n = 7). However, during this potentiation, [Ca2+]iwas not consistently increased. Hence, the increased force following a high-frequency burst is likely due to changes in the myofibrillar properties.
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| 2. |
- Abdelmoez, AM, et al.
(författare)
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Comparative profiling of skeletal muscle models reveals heterogeneity of transcriptome and metabolism
- 2020
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Ingår i: American journal of physiology. Cell physiology. - : American Physiological Society. - 1522-1563 .- 0363-6143. ; 318:3, s. C615-C626
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Tidskriftsartikel (refereegranskat)abstract
- Rat L6, mouse C2C12, and primary human skeletal muscle cells (HSMCs) are commonly used to study biological processes in skeletal muscle, and experimental data on these models are abundant. However, consistently matched experimental data are scarce, and comparisons between the different cell types and adult tissue are problematic. We hypothesized that metabolic differences between these cellular models may be reflected at the mRNA level. Publicly available data sets were used to profile mRNA levels in myotubes and skeletal muscle tissues. L6, C2C12, and HSMC myotubes were assessed for proliferation, glucose uptake, glycogen synthesis, mitochondrial activity, and substrate oxidation, as well as the response to in vitro contraction. Transcriptomic profiling revealed that mRNA of genes coding for actin and myosin was enriched in C2C12, whereas L6 myotubes had the highest levels of genes encoding glucose transporters and the five complexes of the mitochondrial electron transport chain. Consistently, insulin-stimulated glucose uptake and oxidative capacity were greatest in L6 myotubes. Insulin-induced glycogen synthesis was highest in HSMCs, but C2C12 myotubes had higher baseline glucose oxidation. All models responded to electrical pulse stimulation-induced glucose uptake and gene expression but in a slightly different manner. Our analysis reveals a great degree of heterogeneity in the transcriptomic and metabolic profiles of L6, C2C12, or primary human myotubes. Based on these distinct signatures, we provide recommendations for the appropriate use of these models depending on scientific hypotheses and biological relevance.
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| 3. |
- Al-Khalili, L, et al.
(författare)
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MEF2 activation in differentiated primary human skeletal muscle cultures requires coordinated involvement of parallel pathways
- 2004
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Ingår i: American journal of physiology. Cell physiology. - : American Physiological Society. - 0363-6143 .- 1522-1563. ; 286:6, s. C1410-C1416
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Tidskriftsartikel (refereegranskat)abstract
- The myocyte enhancer factor (MEF)2 transcription factor is important for development of differentiated skeletal muscle. We investigated the regulation of MEF2 DNA binding in differentiated primary human skeletal muscle cells and isolated rat skeletal muscle after exposure to various stimuli. MEF2 DNA binding activity in nonstimulated (basal) muscle cultures was almost undetectable. Exposure of cells for 20 min to 120 nM insulin, 0.1 and 1.0 mM hydrogen peroxide, osmotic stress (400 mM mannitol), or 1.0 mM 5-aminoimidazole-4-carboxamide-1-β- d-ribofuranoside (AICAR) led to a profound increase in MEF2 DNA binding. To study signaling pathways mediating MEF2 activity, we preincubated human skeletal muscle cell cultures or isolated rat epitrochlearis muscles with inhibitors of p38 mitogen-activated protein kinase (MAPK) (10 μM SB-203580), MEK1 (50 μM PD-98059), PKC (1 and 10 μM GF109203X), phosphatidylinositol (PI) 3-kinase (10 μM LY-294002), or AMP-activated protein kinase (AMPK; 20 μM compound C). All stimuli resulted primarily in activation of MEF2D DNA binding. Exposure of cells to osmotic or oxidative stress increased MEF2 DNA binding via pathways that were completely blocked by MAPK inhibitors and partially blocked by inhibitors of PKC, PI 3-kinase, and AMPK. In epitrochlearis muscle, MAPK inhibitors blocked contraction but not AICAR-mediated MEF2 DNA binding. Thus activation of MEF2 in skeletal muscle is regulated via parallel intracellular signaling pathways in response to insulin, cellular stress, or activation of AMPK.
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| 4. |
- Al-Khalili, L, et al.
(författare)
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Proteasome inhibition in skeletal muscle cells unmasks metabolic derangements in type 2 diabetes
- 2014
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Ingår i: American journal of physiology. Cell physiology. - : American Physiological Society. - 1522-1563 .- 0363-6143. ; 307:9, s. C774-C787
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Tidskriftsartikel (refereegranskat)abstract
- Two-dimensional difference gel electrophoresis (2-D DIGE)-based proteome analysis has revealed intrinsic insulin resistance in myotubes derived from type 2 diabetic patients. Using 2-D DIGE-based proteome analysis, we identified a subset of insulin-resistant proteins involved in protein turnover in skeletal muscle of type 2 diabetic patients, suggesting aberrant regulation of the protein homeostasis maintenance system underlying metabolic disease. We then validated the role of the ubiquitin-proteasome system (UPS) in myotubes to investigate whether impaired proteasome function may lead to metabolic arrest or insulin resistance. Myotubes derived from muscle biopsies obtained from people with normal glucose tolerance (NGT) or type 2 diabetes were exposed to the proteasome inhibitor bortezomib (BZ; Velcade) without or with insulin. BZ exposure increased protein carbonylation and lactate production yet impaired protein synthesis and UPS function in myotubes from type 2 diabetic patients, marking the existence of an insulin-resistant signature that was retained in cultured myotubes. In conclusion, BZ treatment further exacerbates insulin resistance and unmasks intrinsic features of metabolic disease in myotubes derived from type 2 diabetic patients. Our results highlight the existence of a confounding inherent abnormality in cellular protein dynamics in metabolic disease, which is uncovered through concurrent inhibition of the proteasome system.
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| 5. |
- Albinsson, Sebastian, et al.
(författare)
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Differential dependence of stretch and shear stress signaling on caveolin-1 in the vascular wall
- 2008
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Ingår i: American Journal of Physiology: Cell Physiology. - : American Physiological Society. - 1522-1563 .- 0363-6143. ; 294, s. 271-279
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Tidskriftsartikel (refereegranskat)abstract
- The role of caveolae in stretch- vs. flow-induced vascular responses was investigated using caveolin-1 deficient (KO) mice. Portal veins were stretched longitudinally for 5 min (acute) or 72 h (organ culture). Basal ERK1/2 and Akt phosphorylation were increased in organ-cultured KO veins, as were protein synthesis and vessel wall cross-section. Stretch stimulated acute phosphorylation of ERK1/2 and long-term phosphorylation of focal adhesion kinase (FAK) and cofilin, but did not affect Akt phosphorylation. Protein synthesis, and particularly synthesis of smooth muscle differentiation markers, was increased by stretch. These effects did not differ in portal veins from KO and control mice, which also showed the same contractile response to membrane depolarization and inhibition by the Rho kinase inhibitor Y-27632. KO carotid arteries had increased wall cross-section and responded to pressurization (120 mmHg) for 1 h with increased ERK1/2 but not Akt phosphorylation, similar to control arteries. Shear stress by flow for 15 min, on the other hand, increased phosphorylation of Akt in carotids from control but not KO mice. In conclusion, caveolin-1 contributes to a low basal ERK1/2 and Akt activity and is required for Akt-dependent signals in response to shear stress (flow), but is not essential for trophic effects of stretch (pressure) in the vascular wall. Key words: Hypertrophy, vasoconstriction, vascular smooth muscle, endothelium, nitric oxide.
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| 6. |
- Albinsson, Sebastian, et al.
(författare)
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INTEGRATION OF SIGNAL PATHWAYS FOR STRETCH-DEPENDENT GROWTH AND DIFFERENTIATION IN VASCULAR SMOOTH MUSCLE.
- 2007
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Ingår i: American Journal of Physiology: Cell Physiology. - : American Physiological Society. - 1522-1563 .- 0363-6143. ; 293:May 16, s. 772-782
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Tidskriftsartikel (refereegranskat)abstract
- Vascular smooth muscle phenotype is regulated by environmental factors, such as mechanical forces, which exert effects on signaling to differentiation and growth. We used the mouse portal vein in organ culture to investigate stretch-dependent activation of Akt, extracellular regulated protein kinase (ERK) and focal adhesion kinase (FAK), which have been suggested to be involved in the regulation of stretch-dependent protein synthesis. The role of actin polymerization in these signaling events was examined using the actin stabilizing agent jasplakinolide. Stretch caused a biphasic activation of FAK at 5-15 minutes and 24-72 hours, which may reflect first a direct phosphorylation of preexisting focal adhesions followed by a rearrangement of focal adhesions to accommodate for the increased mechanical load. Phosphorylation of ERK was increased by acute stretch but then decreased, and Akt did not have a distinct peak in stretch-induced phosphorylation. Inhibition of ERK, phosphatidylinositol 3-kinase (PI3K) or mammalian target of rapamycin (mTOR) reduced global but not contractile protein synthesis with maintained stretch sensitivity. Stabilization of actin filaments with jasplakinolide, in unstretched portal veins, resulted in increased ERK phosphorylation and global protein synthesis as well as synthesis of contractile proteins. In contrast, stretch during culture with jasplakinolide did not affect FAK phosphorylation or contractility. Therefore, remodeling of smooth muscle cells to adapt to stretch requires a dynamic cytoskeleton. Key words: actin polymerization, MAP kinase, PI3 kinase, focal adhesion kinase, protein synthesis.
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| 7. |
- Aperia, Anita Chatarina, et al.
(författare)
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Na+, K+-ATPase, a new class of plasma membrane receptors
- 2016
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Ingår i: American Journal of Physiology - Cell Physiology. - : American Physiological Society. - 0363-6143 .- 1522-1563. ; 310:7, s. C491-C495
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Tidskriftsartikel (refereegranskat)abstract
- The Na(+), K(+)-ATPase (NKA) differs from most other ion transporters not only in its capacity to maintain a steep electrochemical gradient across the plasma membrane but also as a receptor for a family of cardiotonic steroids, to which ouabain belongs. Studies from many groups, performed during the last fifteen years, have demonstrated that ouabain, a member of the cardiotonic steroid family, can activate a network of signaling molecules and that NKA will also serve as a signal transducer that can provide a feed back loop between NKA and the mitochondria. This brief review summarizes the current knowledge and controversies with regard to the understanding of NKA signaling.
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| 8. |
- Belusa, R, et al.
(författare)
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Changes in Na(+)-K(+)-ATPase activity influence cell attachment to fibronectin
- 2002
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Ingår i: American journal of physiology. Cell physiology. - : American Physiological Society. - 0363-6143 .- 1522-1563. ; 282:2, s. C302-C309
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Tidskriftsartikel (refereegranskat)abstract
- Most vital cellular functions are dependent on a fine-tuned regulation of intracellular ion homeostasis. Here we have demonstrated, using COS cells that were untransfected or transfected with wild-type rat ouabain-resistant Na+-K+-ATPase, that partial inhibition of Na+-K+-ATPase has a dramatic influence on cell attachment to fibronectin. Ouabain dose-dependently decreased attachment in untransfected cells and in cells expressing wild-type Na+-K+-ATPase, but not in cells expressing ouabain-insensitive Na+-K+-ATPase, whereas inhibition of Na+-K+-ATPase by lowering extracellular K+concentration decreased attachment in all three cell types. Thirty percent inhibition of Na+-K+-ATPase significantly attenuated attachment. Na+-K+-ATPase inhibition caused a sustained increase in the intracellular Ca2+concentration that obscured Ca2+transients observed in untreated cells during attachment. Inhibitors of Ca2+transporters significantly decreased attachment, but inhibition of Na+/H+exchanger did not. Ouabain reduced focal adhesion kinase autophosphorylation but had no effect on cell surface integrin expression. These results suggest that the level of Na+-K+-ATPase activity strongly influences cell attachment, possibly by an effect on intracellular Ca2+.
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| 9. |
- Benziane, B, et al.
(författare)
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AMP-activated protein kinase activator A-769662 is an inhibitor of the Na(+)-K(+)-ATPase
- 2009
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Ingår i: American journal of physiology. Cell physiology. - : American Physiological Society. - 1522-1563 .- 0363-6143. ; 297:6, s. C1554-C1566
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Tidskriftsartikel (refereegranskat)abstract
- Muscle contraction and metabolic stress are potent activators of AMP-activated protein kinase (AMPK). AMPK restores energy balance by activating processes that produce energy while inhibiting those that consume energy. The role of AMPK in the regulation of active ion transport is unclear. Our aim was to determine the effect of the AMPK activator A-769662 on Na+-K+-ATPase function in skeletal muscle cells. Short-term incubation of differentiated rat L6 myotubes with 100 μM A-769662 increased AMPK and acetyl-CoA carboxylase (ACC) phosphorylation in parallel with decreased Na+-K+-ATPase α1-subunit abundance at the plasma membrane and ouabain-sensitive86Rb+uptake. Notably, the effect of A-769662 on Na+-K+-ATPase was similar in muscle cells that do not express AMPK α1- and α2-catalytic subunits. A-769662 directly inhibits the α1-isoform of the Na+-K+-ATPase, purified from rat and human kidney cells in vitro with IC5057 μM and 220 μM, respectively. Inhibition of the Na+-K+-ATPase by 100 μM ouabain decreases sodium pump activity and cell surface abundance, similar to the effect of A-769662, without affecting AMPK and ACC phosphorylation. In conclusion, the AMPK activator A-769662 inhibits Na+-K+-ATPase activity and decreases the sodium pump cell surface abundance in L6 skeletal muscle cells. The effect of A-769662 on sodium pump is due to direct inhibition of the Na+-K+-ATPase activity, rather than AMPK activation. This AMPK-independent effect on Na+-K+-ATPase calls into question the use of A-769662 as a specific AMPK activator for metabolic studies.
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| 10. |
- Bergdahl, Andreas, et al.
(författare)
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Plasticity of TRPC expression in arterial smooth muscle: correlation with store-operated Ca2+ entry.
- 2005
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Ingår i: American Journal of Physiology: Cell Physiology. - : American Physiological Society. - 1522-1563 .- 0363-6143. ; 288:4, s. 872-880
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Tidskriftsartikel (refereegranskat)abstract
- Loss of the smooth muscle contractile phenotype is critical in atherosclerosis and in restenosis after angioplasty, but its early signals are incompletely understood. In this study, we have explored the role of transient receptor potential canonical (TRPC) proteins, which have been suggested to mediate store-operated Ca2+ entry (SOCE). Contractility of rat cerebral arteries in organ culture is preserved for several days, whereas SOCE is increased. In correlation with this increase is that nifedipine-insensitive whole cell current, activated by depletion of intracellular Ca2+ stores, was increased by 50% in cells isolated from arteries cultured for 3 days. TRPC1 and TRPC6 mRNA were more than fivefold increased in cells isolated after organ culture, whereas TRPC3 was decreased. Immunofluorescent staining and/or Western blotting of arteries and isolated cells showed upregulation of TRPC1 and TRPC6 proteins during organ culture. In intact arteries, TRPC4 expression correlated with the amount of endothelium present. Ca2+ addition after store depletion caused a contraction in cultured, but not in freshly dissected, arteries. A polyclonal TRPC1 antibody directed against an extracellular epitope inhibited this contraction by approximately 50%. To investigate the basis of the TRPC upregulation and assess its possible clinical significance, segments of human internal mammary artery were organ cultured for 24 h and then exposed to balloon dilatation in vitro, followed by further culturing for up to 48 h. After dilatation, TRPC1 and TRPC6 mRNA were progressively increased compared with undilated control segments. The results of this study indicate that vascular injury enhances plasticity in TRPC expression, that TRPC expression correlates with cellular Ca2+ handling, and that TRPC1 is a subunit of upregulated store-operated Ca2+ channels.
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