Multiomic analysis reveals conservation of cancer-associated fibroblast phenotypes across species and tissue of origin

1. Download : Download high-res image (241KB)
2. Download : Download full-size image

     

Mechanical stretch induces myelin protein loss in oligodendrocytes by activating Erk1/2 in a calcium-dependent manner

Myelin loss in the brain is a common occurrence in traumatic brain injury (TBI) that results from impact-induced acceleration forces to the head. Fast and abrupt head motions, either resulting from violent blows and/or jolts, cause rapid stretching of the brain tissue, and the long axons within the white matter tracts are especially vulnerable to such mechanical strain. Recent studies have shown that mechanotransduction plays an important role in regulating oligodendrocyte progenitors cell differentiation into oligodendrocytes. However, little is known about the impact of mechanical strain on mature oligodendrocytes and the stability of their associated myelin sheaths. We used an in vitro cellular stretch device to address these questions, as well as characterize a mechanotransduction mechanism that mediates oligodendrocyte responses. Mechanical stretch caused a transient and reversible myelin protein loss in oligodendrocytes. Cell death was not observed. Myelin protein loss was accompanied by an increase in intracellular Ca²⁺ and Erk1/2 activation. Chelating Ca²⁺ or inhibiting Erk1/2 activation was sufficient to block the stretch-induced loss of myelin protein. Further biochemical analyses revealed that the stretch-induced myelin protein loss was mediated by the release of Ca²⁺ from the endoplasmic reticulum (ER) and subsequent Ca²⁺-dependent activation of Erk1/2. Altogether, our findings characterize an Erk1/2-dependent mechanotransduction mechanism in mature oligodendrocytes that de-stabilizes the myelination program.     

Matrix-dependent adhesion mediates network responses to physiological stimulation of the osteocyte cell process

Osteocytes are bone cells that form cellular networks that sense mechanical loads distributed throughout the bone tissue. Interstitial fluid flow in the lacunar canalicular system produces focal strains at localized attachment sites around the osteocyte cell process. These regions of periodic attachment between the osteocyte cell membrane and its canalicular wall are sites where pN-level fluid-flow induced forces are generated in vivo. In this study, we show that focally applied forces of this magnitude using a newly developed Stokesian fluid stimulus probe initiate rapid and transient intercellular electrical signals in vitro. Our experiments demonstrate both direct gap junction coupling and extracellular purinergic P2 receptor signaling between MLO-Y4 cells in a connected bone cell network. Intercellular signaling was initiated by pN-level forces applied at integrin attachment sites along both appositional and distal unapposed cell processes, but not initiated at their cell bodies with equivalent forces. Electrical coupling was evident in 58% of all cell pairs tested with appositional connections; coupling strength increased with the increasing number of junctional connections. Apyrase, a nucleotide-degrading enzyme, suppressed and abolished force-induced effector responses, indicating a contribution from ATP released by the stimulated cell. This work extends the understanding of how osteocytes modulate their microenvironment in response to mechanical signals and highlights mechanisms of intercellular relay of mechanoresponsive signals in the bone network.     

Stochastic nanoroughness modulates neuron–astrocyte interactions and function via mechanosensing cation channels

Extracellular soluble signals are known to play a critical role in maintaining neuronal function and homeostasis in the CNS. However, the CNS is also composed of extracellular matrix macromolecules and glia support cells, and the contribution of the physical attributes of these components in maintenance and regulation of neuronal function is not well understood. Because these components possess well-defined topography, we theorize a role for topography in neuronal development and we demonstrate that survival and function of hippocampal neurons and differentiation of telencephalic neural stem cells is modulated by nanoroughness. At roughnesses corresponding to that of healthy astrocytes, hippocampal neurons dissociated and survived independent from astrocytes and showed superior functional traits (increased polarity and calcium flux). Furthermore, telencephalic neural stem cells differentiated into neurons even under exogenous signals that favor astrocytic differentiation. The decoupling of neurons from astrocytes seemed to be triggered by changes to astrocyte apical-surface topography in response to nanoroughness. Blocking signaling through mechanosensing cation channels using GsMTx4 negated the ability of neurons to sense the nanoroughness and promoted decoupling of neurons from astrocytes, thus providing direct evidence for the role of nanotopography in neuron–astrocyte interactions. We extrapolate the role of topography to neurodegenerative conditions and show that regions of amyloid plaque buildup in brain tissue of Alzheimer’s patients are accompanied by detrimental changes in tissue roughness. These findings suggest a role for astrocyte and ECM-induced topographical changes in neuronal pathologies and provide new insights for developing therapeutic targets and engineering of neural biomaterials.Cellular homeostasis in the brain tissue is believed to be regulated primarily by a complex spatiotemporal signaling environment involving soluble neurotrophic factors (1, 2). These factors, including neurotrophins such as brain-derived neurotrophic factor, the TGF-β family including bone morphogenetic proteins (BMPs), and the IL-6 superfamily including ciliary neurotrophic factor (CNTF), regulate survival, steer progenitor fate decision, and critically affect the development of the nervous system as well as the homeostasis of the adult CNS (3–6). However, developmental processes such as axon pathfinding, synapse formation, nervous system patterning, neuronal plasticity, and degeneration fail to be explained solely on the basis of soluble factors. There is increasing evidence that physical variables such as the stiffness of a cellular environment influence cell development (7–12). However, the cells of the brain tissue reside in a soft environment that is rich in polysaccharides (13, 14). In the context of neuronal development and neurophysiology, astrocytes have an established role in maintaining neuronal function. They form a vast network that provides the physical and biochemical matrix over which neurons thrive and function (15, 16). The plasticity found in the brain can be attributed in part to the morphological changes that occur in astrocyte processes that can not only alter the geometry of the neuronal environment but also induce dynamic changes in astrocyte–neuron interactions affecting neurotransmission, signal gradients, and the relationship between synapses (15). Interestingly, the changes to the physical aspects of a neuronal environment can originate from changes to morphology of support cells such as astrocytes and also changes to ECM structure and properties. Cells and ECM polysaccharides play an important role in growth, differentiation, and migration of neural precursors, as well as in repair and plasticity in the central nervous system (17, 18). However, in addition to a biological function, cells and macromolecules provide a physically defined environment (19, 20), and we postulate a significant role for topography in neural development. Studies to date have focused on the effects of microscale topography, deterministic roughness, and substrate chemistry on neurite outgrowth and neuronal function (7, 21–23). However, the influence of ECM-like nanotopography on neuronal development and fate is a realm that has not been investigated thus far. Therefore, in this work we specifically focus on the impact of stochastic nanoroughness as would be provided by neighboring cells and ECM molecules on neuronal cell interactions, function, and differentiation.     

Effects of shear stress on endothelial cells: go with the flow

Haemodynamic forces influence the functional properties of vascular endothelium. Endothelial cells (ECs) have a variety of receptors, which sense flow and transmit mechanical signals through mechanosensitive signalling pathways to recipient molecules that lead to phenotypic and functional changes. Arterial architecture varies greatly exhibiting bifurcations, branch points and curved regions, which are exposed to various flow patterns. Clinical studies showed that atherosclerotic plaques develop preferentially at arterial branches and curvatures, that is in the regions exposed to disturbed flow and shear stress. In the atheroprone regions, the endothelium has a proinflammatory phenotype associated with low nitric oxide production, reduced barrier function and increased proadhesive, procoagulant and proproliferative properties. Atheroresistant regions are exposed to laminar flow and high shear stress that induce prosurvival antioxidant signals and maintain the quiescent phenotype in ECs. Indeed, various flow patterns contribute to phenotypic and functional heterogeneity of arterial endothelium whose response to proatherogenic stimuli is differentiated. This may explain the preferential development of endothelial dysfunction in arterial sites with disturbed flow.     

P2Y2 receptors and GRK2 are involved in oscillatory fluid flow induced ERK1/2 responses in chondrocytes

Mechanical loading is an important factor regulating cartilage metabolism maintained by chondrocytes. However, some of its underlying mechanisms remain poorly understood. In this study, we employed a chondrogenic cell line ATDC5 to investigate roles of P2Y₂ and GRK2 in chondrocyte mechanotransduction. We first confirmed the expression of chondrocyte markers in differentiated ATDC5 cells. We then exposed both differentiated and undifferentiated ATDC5 cells to oscillatory fluid flow, and found that differentiated ATDC5 cells responded to oscillatory fluid flow by increasing COX‐2 and aggrecan expressions. More importantly, fluid flow induced ERK1/2 response in differentiated cells was increased more than 10 times compared to those in undifferentiated cells. Furthermore, we found that P2Y₂ mRNA and protein levels in differentiated ATDC5 cells were significantly higher than those in undifferentiated cells. In contrast, GRK2 protein levels in differentiated cells were significantly lower than those in undifferentiated cells. Finally, overexpressions of P2Y₂ and GRK2 in differentiated ATDC5 cells result in a 34% increase and a 21% decrease of the ERK1/2 phosphorylation, respectively, in response to oscillatory fluid flow, suggesting important roles of P2Y₂ and GRK2 in chondrocyte mechanotransduction. © 2010 Orthopaedic Research Society Published by Wiley Periodicals, Inc. J Orthop Res 29:828–833