169 research outputs found
Differential response of skeletal muscles to mTORC1 signaling during atrophy and hypertrophy
No full textBACKGROUND: Skeletal muscle mass is determined by the balance between protein synthesis and degradation. Mammalian target of rapamycin complex 1 (mTORC1) is a master regulator of protein translation and has been implicated in the control of muscle mass. Inactivation of mTORC1 by skeletal muscle-specific deletion of its obligatory component raptor results in smaller muscles and a lethal dystrophy. Moreover, raptor-deficient muscles are less oxidative through changes in the expression PGC-1alpha, a critical determinant of mitochondrial biogenesis. These results suggest that activation of mTORC1 might be beneficial to skeletal muscle by providing resistance to muscle atrophy and increasing oxidative function. Here, we tested this hypothesis by deletion of the mTORC1 inhibitor tuberous sclerosis complex (TSC) in muscle fibers. METHOD: Skeletal muscles of mice with an acute or a permanent deletion of raptor or TSC1 were examined using histological, biochemical and molecular biological methods. Response of the muscles to changes in mechanical load and nerve input was investigated by challenging the mice by denervation or ablation of synergistic muscles. RESULTS: Genetic deletion or knockdown of raptor, causing inactivation of mTORC1, was sufficient to prevent muscle growth and enhance muscle atrophy. Conversely, short-term activation of mTORC1 by knockdown of TSC induced muscle fiber hypertrophy and atrophy-resistance upon denervation, in both fast tibialis anterior (TA) and slow soleus muscles. Surprisingly, however, sustained activation of mTORC1 by genetic deletion of Tsc1 caused muscle atrophy in all but soleus muscles. In contrast, oxidative capacity was increased in all muscles examined. Consistently, TSC1-deficient soleus muscle was atrophy-resistant whereas TA underwent normal atrophy upon denervation. Moreover, upon overloading, plantaris muscle did not display enhanced hypertrophy compared to controls. Biochemical analysis indicated that the atrophy respo of muscles was based on the suppressed phosphorylation of PKB/Akt via feedback inhibition by mTORC1 and subsequent increased expression of the E3 ubiquitin ligases MuRF1 and atrogin-1/MAFbx. In contrast, expression of both E3 ligases was not increased in soleus muscle suggesting the presence of compensatory mechanisms in this muscle. CONCLUSIONS: Our study shows that the mTORC1- and the PKB/Akt-FoxO pathways are tightly interconnected and differentially regulated depending on the muscle type. These results indicate that long-term activation of the mTORC1 signaling axis is not a therapeutic option to promote muscle growth because of its strong feedback induction of the E3 ubiquitin ligases involved in protein degradation
sj-docx-1-tan-10.1177_17562864231193530 – Supplemental material for Lipid profile with eslicarbazepine acetate and carbamazepine monotherapy in adult patients with newly diagnosed focal seizures: post hoc analysis of a phase III trial and open-label extension study
No full textSupplemental material, sj-docx-1-tan-10.1177_17562864231193530 for Lipid profile with eslicarbazepine acetate and carbamazepine monotherapy in adult patients with newly diagnosed focal seizures: post hoc analysis of a phase III trial and open-label extension study by Eugen Trinka, Rodrigo Rocamora, João Chaves, Mathias J. Koepp, Stephan Rüegg, Martin Holtkamp, Joana Moreira, Miguel M. Fonseca, Guillermo Castilla-Fernández and Fábio Ikedo in Therapeutic Advances in Neurological Disorders</p
Molecular mechanisms of simvastatin-induced myopathy and insulin resistance
No full textStatins or 3-hydroxy-3-methyl-glutaryl-coenzyme A reductase inhibitors are the most prescribed lipid-lowering drugs worldwide, used to treat hypercholesterolemia and efficient to reduce mortality and morbidity associated to cardiovascular diseases. They act primarily in the liver, where they inhibit the biosynthesis of cholesterol, alongside with other non-sterol intermediates, such as mevalonate, dolichol, farnesyl pyrophosphate and geranylgeranyl pyrophosphate, leading to the impairment of several cellular processes for example protein post-translational modifications and proliferation.
Statins have several beneficial effects on the cardiovascular system and are in general well tolerated. However, inhibition of cholesterol synthesis pathway can induce adverse effects, mainly towards the skeletal muscle. These adverse effects range from muscle pain to rhabdomyolysis in rare cases, which can ultimately lead to death. Moreover, recently, an increased occurrence of insulin resistance and new-onset diabetes have been reported in patients treated with statins.
Considering the huge proportion of people under statin therapy worldwide and the prevalence of cardiovascular diseases and type 2 diabetes, it is urgent to elucidate molecular mechanisms leading to myopathy and new-onset diabetes.
This thesis includes four papers, two that are published and two that are in preparation.
The first paper presents the effects of insulin on simvastatin-induced toxicity as well as on the impairments induced by simvastatin on the insulin receptor (IR) signaling in C2C12 myotubes. Simvastatin strongly reduced membrane integrity and depleted the intracellular ATP in C2C12 cells. Additionally, simvastatin induced endoplasmic reticulum (ER) stress. Insulin was potent to not only prevent, but also rescue partially and time-dependently simvastatin toxicity. Simvastatin significantly reduced Akt phosphorylation on the serine 473 residue, done by mTORC2, while inhibiting only by trend the phosphorylation on the threonine 308 residue (done by PDK1). In like manner, downstream effectors of Akt were affected, inducing a reduced mTORC1 activity, atrophy and apoptosis. Insulin prevented these effects in a dose-dependent fashion. These data demonstrate that impaired Akt activation is a consequence of impaired mTORC2 activity and that insulin can prevent deleterious effects of simvastatin on the insulin receptor transduction pathway.
Our second paper correlates potentiality of insulin to prevent cell death and maintaining insulin receptor signaling in simvastatin-treated C2C12 myotubes, to the reported new-onset diabetes and insulin resistance concomitant to statin therapy. We demonstrated the effects of simvastatin on glucose metabolism in mice treated orally with simvastatin and elucidated the mechanisms leading to insulin resistance using C2C12 myotubes. Simvastatin-treated mice had higher plasmatic glucose during ip glucose tolerance test (IGTT) and a reduced glucose uptake in skeletal muscle compared to water-treated mice. A reduced glucose uptake was also observed in C2C12 myotubes treated with the statin as well as an impaired expression and phosphorylation of the insulin receptor β chain. Akt (Ser473) phosphorylation was significantly decreased in treated myocytes, which was explained and demonstrated with a decreased mTOR phosphorylation. Cells displayed also an impaired phosphorylation of GSK3β, leading to a reduced glucose transporter 4 (GLUT4) translocation to the cell surface. These data provide the evidence that simvastatin can cause insulin resistance in mice, and highlights new potential molecular targets for the management of insulin resistance during statin therapy, with the identification of a defect of mTORC2 activity and of GLUT4 translocation to the cell membrane for glucose absorption.
The third paper integrates the knowledge we acquired with simvastatin-treated myotubes to perform a comparison between C2C12 myoblasts and myotubes, the precursor and mature muscle cells respectively, in order to evaluate how post-natal myogenesis was affected by simvastatin. We observed that myoblasts were more sensitive to toxic effects of simvastatin in comparison to their differentiated form. We identified that geranylgeraniol was strongly potent in rescuing simvastatin toxicity. We assessed mitochondrial respiration and superoxide generation and found out that only myoblasts were disturbed by simvastatin at this level, whereas the mitochondrial function was not affected in myotubes, probably due to a higher expression of superoxide dismutase 2 (SOD2). We then characterized proliferation, differentiation and fusion processes in simvastatin-treated myoblasts. Proliferation of myoblasts was strongly inhibited, as well as the expression of differentiation and fusion markers. Mevalonate could prevent these effects in co-treatment. Last, upon simvastatin treatment, both cell models underwent apoptosis, which was prevented by insulin. This study demonstrates differences in sensitivity between C2C12 myoblasts and differentiated myotubes treated with simvastatin and might represent a good start point in the understanding of why statin-treated patients experience muscle pain or weakness during exercise or muscular stress.
Our fourth paper evaluates the contribution of mTORC1 and mTORC2 in simvastatin-induced myopathy, and confirms for the first time that mTORC2 inhibition is the key event in statin-induced myotoxicity. We showed that mTORC1 inhibition was cytoprotective in C2C12 myotubes and did not recapitulate simvastatin myotoxicity and impaired insulin receptor signaling. Inhibiting mTORC2 by knocking down Rictor displayed a similar toxicity pattern to simvastatin treatment in control cells and led to a reduced Akt (Ser473) and downstream effectors phosphorylation, thus recapitulating simvastatin-induced impairments. The mechanisms leading to mTORC2 and subsequently Akt inactivation in myocytes treated with statins were unprenylation of cellular GTPases and induction of mitochondrial reactive oxygen species (ROS) production. These findings highlight the primary molecular events occurring with simvastatin therapy, giving future opportunities for solutions to better manage and prevent statin-induced myopathy
Dissertatio pneumatica de anima separata
No full textquam ... sub praesidio ... dn. Iohannis Lavateri ... examini subiicit Iohannes Henricus Ruegius, phil. cand. author & respondens ...Dedikation an Joh. Conrad Grebel, Joh. Heinrich Hirzel, Caspar Escher, Joh. Heinrich Rahn, Joh. Ulrich Escher, Joh. Heinrich Heidegger (2), Joh. Heinrich Zeller und Joh. Wilhelm Simler auf dem Titelbl. verso. Gedichte von Humbertus a Stavia und Andreas Rauch auf Bl. A4v.Diss. Hohe Schule Zürich, 166
mTORC2 controls neuron size and Purkinje cell morphology independent of mTORC1
No full textPrenatal brain development is mainly accomplished by extensive proliferation of neuronal precursor cells whereas postnatal brain growth in mammals is mainly mediated by the growth of those post-mitotic nerve cells. The neuron size and the branching pattern of the dendritic tree are highly controlled during development to enable the proper connectivity of neuronal circuits and the accurate electrical transmission in the adult which is a prerequisite for the brain to function normally. Aberrations in size, morphology or connectivity have been shown to be the cause for various brain disorders. Neuron size and dendrite development are controlled by intrinsic mechanisms, trophic factors and neuronal activity, processes that need the concerted action of a plethora of signaling molecules. A central integrator of various signaling cascades is the mammalian target of rapamycin (mTOR) and as such it contributes to brain development and function and is thus also implicated in the pathophysiology of psychiatric disorders.
mTOR is a serine threonine protein kinase that is highly conserved from yeast to humans and has been found to be part of at least two multi-protein complexes mTORC1 and mTORC2. The formation of mTORC1 is dependent on the protein raptor whereas mTORC2 assembly relies on the protein rictor. In recent years a complex picture about the function of mTORC1 has emerged by use of rapamycin, an immunosuppressive drug that acutely inhibits mTORC1 formation and activity and has attributed mTORC1 a major role in the regulation of cell size and proliferation. However, because the activity of mTORC2 is only depleted upon long term application of rapamycin, research advancement on its function was thus far impeded. Due to the early embryonic lethality of raptor or rictor knockout in mammals conditional knockout models were constructed. Whereas tissue specific knockout of raptor led to characteristic alterations, knockout of rictor in several organs such as skeletal muscle and adipose tissue provided none or only a weak phenotype. Several cell culture studies assigned mTORC2 a role in cytoskeletal modifications but in vivo confirmation is still lacking. The current knowledge about mTORC2 is restricted to the downstream targets Akt/PKB (proteinkinase B) and PKC (protein kinase C) which belong to the AGC kinase family. Those kinases are reported to influence cell morphology, growth and survival and are also essential regulators of brain development and function. PKCs are involved in synaptic plasticity and neurotransmitter release and, hence, also in the pathophysiological mechanisms of psychiatric disorders especially in schizophrenia and bipolar disorder. Concordantly, several psychiatric agents have been shown to alter PKC signaling. This emphasizes the urge to analyze the role of mTORC2 in the central nervous system.
In this dissertation the role of mTORC2 was analyzed in the central nervous system and in specific sub-populations of neurons by deletion of rictor. I discovered, that in contrast to all other organs analyzed so far, rictor knockout in the brain reveals a pronounced phenotype. The brain-size of those mice shows an enormous reduction to almost half of that of control mice which is caused mainly by the reduction of neuron size. The reduced cell size is observed in neurons derived from different brain areas in vitro and in vivo but is most prominent in Purkinje cells of the cerebellum, the cell type with highest rictor expression. In addition, dendrite morphology is majorly disrupted and the formation of dendritic spines is affected which correlates with a decreased neuronal activity. The Purkinje cell phenotype can also be reproduced in a Purkinje cell specific knockout of rictor and thus demonstrates that the effect of rictor deletion in neurons is cell autonomous. Moreover, Purkinje cell axonal path-finding is affected which correlates with the decrease in phosphorylation of the neuron specific PKC target protein GAP-43, a known regulator for axon growth and path-finding. Molecular analysis reveals that rictor is essential for the activity of all conventional PKC isoforms and the novel PKCε in vivo and in vitro in neurons which influences the function of downstream targets important for cytoskeleton modifications such as GAP-43, MARCKs and neurofascin. In addition, rictor controls the phosphorylation of Akt but does not alter mTORC1 signaling towards its downstream effectors. In summary it becomes clear that rictor is important in the development and maturation of neurons and controls their size and neuron structure which influences the entire brain function and affects the behavior of the mice. Thus, these data encompass a new role of rictor in CNS disorders
Evaluating the contributions of One Health initiatives to social sustainability
No full textOne Health is an approach that integrates perspectives from human, animal and environmental health to address health challenges. As the idea of One Health is grounded in achieving sustainable outcomes, an important aspect is the contribution of One Health to social sustainability. In this chapter we ask, what social sustainability is, what the indicators of social sustainability related to One Health are, and, through what measures we can evaluate the contributions of One Health to social sustainability, in terms of its operations, its supporting infrastructures and outcomes. We adopt a wider conceptualization of social sustainability and propose an approach based on basic needs, capabilities and emancipation, environmental justice, solidarity and social cohesion. First, we identify indicators used in literature to capture social sustainability in human, animal and environmental health and propose ways to integrate them into a framework for the evaluation of One Health initiatives. Second, we formulate questions that can be used to evaluate the social sustainability of One Health initiatives. Third, we discuss the viability of operationalising the indicators, the trade-Offs that might arise and identify how they can be minimised. We then discuss methodological issues and highlight the importance of transdisciplinary deliberative approaches for adapting the framework to specific contexts
mTOR complex 2 - akt signaling is physically and functionally at mam
No full textThe target of rapamycin (TOR) is a conserved protein kinase and a
central controller of growth. TOR can be part of two structurally and
functionally distinct complexes, termed TOR complex 1 and TOR
complex 2. Mammalian TOR complex 2 (mTORC2) is composed of
mTOR, Rictor, Sin1 and mLST8. Both mTORC1 and mTORC2 are activated
by growth factors. The mechanism via which growth factors
regulate mTORC2 has been elusive until recently. mTORC2 binds ribosomes
in a growth factor stimulated manner and this association is
required for mTORC2 activity.
mTOR complex 2 functions include control of spatial cell growth
and metabolism and thus, mTORC2 deregulation has been linked
to various disorders including cancer and diabetes. mTORC2 phosphorylates
and thereby activates the AGC kinase family member Akt
(PKB). Akt has many different targets and functions, not all of which
depend on mTORC2 mediated Akt phosphorylation.
In order to gain a better understanding of mTORC2 function, we
asked where mTORC2 signaling is localized. A number of studies
localized mTORC2, functionally or physically, either to the endoplasmic
reticulum (ER) or to mitochondria.We investigated whether these
seemingly unrelated observations concerning mTORC2 localization,
might be the consequence of mTORC2 signaling at MAM. MAM
or mitochondria-associated ER membrane is a quasi-synaptic subdomain
between the ER and mitochondria. MAM plays a crucial role in
the regulation of mitochondrial metabolism and cell survival by gating
both the calcium flux and phospholipid trafficking between the
ER and mitochondria.
First, we analyzed mTORC2 subcellular localization. mTORC2 is
localized to the ER adjacent to mitochondria and mTORC2 can be
biochemically isolated from MAM structures. mTOR complex 2 interacts
with the IP3R-Grp75-VDAC1 complex, a tether that connects ER
and mitochondria at MAM. Insulin stimulates mTORC2 localization
to MAM and mTORC2 interaction with the IP3R-Grp75-VDAC1 complex.
MAM localization of mTORC2 depends on mTORC2-ribosome
interaction.
Next we investigated the function of mTORC2 at MAM. Rictor
(mTORC2) knockout causes a decrease in MAM formation. Growth
factors stimulate MAM formation via mTORC2 and the Akt substrate
PACS2, a MAM resident protein. As expected for MAM deficient
cells, mTORC2 disruption changes the calcium flux from the ER to
mitochondria at MAM. Furthermore, we observe a reduction of Akt
mediated phosphorylation of the MAM calcium channel IP3R upon
Rictor knockout. Thus, mTORC2 signaling at MAM controls MAM mediated
calcium release via the Akt targets PACS2 and IP3R.
Since MAM disruption and calcium signaling both affect mitochondrial
metabolism, we proceeded by analyzing the mitochondrial physiology
of mTORC2 deficient cells. Rictor knockout cells exhibit a disruption
of VDAC1-HK2 binding, caused by a lack of Akt mediated
phosphorylation of HK2 at MAM. This, together with the defect in
MAM, induces an increase in basal respiration, mitochondrial inner
membrane potential, and ATP production in the mTORC2 deficient
cells, culminating in apoptosis. Thus, mTORC2 at MAM appears to
control several aspects of mitochondrial physiology.
These findings emphasize the role of MAM as a signaling hub that
controls cell physiology. By identifying the integral role of mTORC2
at the core of this platform, our results provide new insights on
the mechanisms that regulate growth and metabolism. These observations
may offer new therapeutic strategies against mTORC2 and
MAM driven diseases such as diabetes, Alzheimer’s and cancer
Das wertbasierte Geschäftsmodell – Ein aktualisierter Strukturierungsansatz
No full textDieser Beitrag stellt den wertbasierten Geschäftsmodellansatz als Herangehensweise zur strukturierten Beschreibung und Konzeption von Geschäftsmodellen vor. Dieser Ansatz ist eine Weiterentwicklung des Geschäftsmodellansatzes von Bieger, Rüegg-Stürm und von Rohr (2002), die dem aktuellen Stand der Forschung zu Geschäftsmodellen Rechnung trägt. Insbesondere werden neben der Orientierung an der Wertschaffung neu auch die Dimensionen Innovation und Wertverteilung eingeführt.</p
Notch2 signaling in development and cancer
No full textNotch signaling via cell to cell interaction is a key mechanism in regulating proliferation, cell fate decisions and survival of various cell types in both invertebrates and vertebrates. Among the four Notch receptors (Notch1-4) described in mammals, most studies focused on Notch1, while Notch2 functions are poorly understood. In humans, Notch2 mutations are associated with a variety of developmental disorders and tumor formation in several tissues. In the liver, Notch2 deletion was shown to cause Alagille syndrome (AGS), which is characterized by impaired intrahepatic bile duct (IHBD) development. In glioblastoma multiforme (GBM), the most aggressive form of CNS tumors, Notch2 is amplified and high Notch2 levels correlate with poor prognosis. However, the exact role of Notch2 in these human pathologies is not established. In order to investigate the function of Notch2 in AGS and GBM at the level of cells, tissues and organs, I generated transgenic mice that allow for tissue-specific expression of activated Notch2 (Notch2ICD), which mimics ligand-induced activation of Notch2 signaling.
To address the function of Notch2 signaling in AGS and IHBD development, Notch2ICD expression was induced in hepatoblasts. Hepatoblasts are the bipotential progenitors in the liver that give rise to either hepatocytes or biliary epithelial cells (BECs) that undergo tubulogenesis to form IHBDs. I observed that ectopic Notch2ICD expression in hepatoblasts induces biliary epithelial cell (BEC) differentiation, tubulogenesis of IHBDs, and BEC survival. These findings shed light on the role for Notch2 in AGS, since they provide an explanation why AGS patients with Notch2 mutations suffer from impaired IHBD development.
It is believed that GBM originates from glioma stem cells (GSCs) which can derive from developmentally stalled neural stem cells (NSCs). Thus, I addressed whether Notch2 plays a role in regulating NSC proliferation and differentiation, possibly predisposing NSCs to become GSCs and eventually GBMs. Therefore, I generated mice that ectopically express activated Notch2 in NSCs and compared the induced molecular alterations to those in GSCs from GBM cell lines and primary GBM biopsies. I show that key features of GSCs, such as increased proliferation and astrocytic lineage commitment, are induced by ectopic Notch2 signaling in NSCs. Aberrant Notch2 expression may therefore predispose NSCs to become GSCs that give rise to GBMs. Moreover, Notch2 signaling enhanced survival of GBM cells, possibly explaining the increased aggressiveness of GBMs with high Notch2 levels. Therefore, blockade of Notch2 signaling may interfere with GBM cell survival, and the formation and proliferation of GSCs and thus be of therapeutic benefit for the treatment of GBMs, for which no cure is available yet
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