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Mechanical loading of articular cartilage stimulates the metabolism of resident chondrocytes and induces the synthesis of molecules to maintain the integrity of the cartilage. Mechanical signals modulate biochemical activity and changes in cell behavior through mechanotransduction. Compression of cartilage results in complex changes within the tissue including matrix and cell deformation, hydrostatic and osmotic pressure, fluid flow, altered matrix water content, ion concentration and fixed charge density. These changes are detected by mechanoreceptors on the cell surface, which include mechanosensitive ion channels and integrins that on activation initiate intracellular signalling cascades leading to tissue remodelling. Excessive mechanical loading also influences chondrocyte metabolism but unlike physiological stimulation leads to a quantitative imbalance between anabolic and catabolic activity resulting in depletion of matrix components. In this article we focus on the role of mechanical signalling in the maintenance of articular cartilage, and discuss how alterations in normal signalling can lead to pathology.

Summary Objective Classical neuronal signalling molecules such as substance P and glutamate have been identified in cartilage and have roles in regulation of chondrocyte function. This study looks at expression and activity of the ionotropic glutamate NMDA (N-methyl-D-aspartic acid) receptor (NMDAR) in human osteoarthritic (OA) chondrocytes. Method Chondrocytes were obtained from human knee joint arthroplasty specimens. NMDAR subunits and PSD-95 (postsynaptic density protein 95) expression were analysed by reverse transcription-polymerase chain reaction and Western blotting. Activity of NMDAR was assayed by radioactive calcium 45 uptake and changes in membrane potential in the presence and absence of NMDA and NMDAR antagonists and blockade of cell membrane ion channels. Results NMDAR 1, 2A, 2B and PSD-95 were detected in human OA chondrocytes whereas NR2B was absent from normal chondrocytes. NMDA induced calcium flux into OA chondrocytes and cell membrane depolarisation. These responses were blocked by NMDAR antagonists, removal of extracellular calcium, inhibition of nNOS (neuronal nitric oxide synthase) activity and uncoupling of NMDAR from PSD-95. Blockade of sodium channels by tetrodotoxin resulted in NMDA-induced membrane hyperpolarisation which was, in turn inhibited by apamin, a blocker of SK channels. NMDA-induced changes in cell membrane potential were not affected by l-type and stretch activated calcium channel inhibitors. Conclusions Human OA and normal articular chondrocytes differ in the expression of NMDAR subunits. In OA chondrocytes NMDAR signalling requires extracellular calcium, association with PSD-95, and nNOS activity. Downstream signalling results in activation of tetrodotoxin sensitive sodium channels and SK channels, a response that differs from that of normal chondrocytes suggesting altered activity of NMDAR in OA.

The Biological General Repository for Interaction Datasets (BioGRID: http://thebiogrid.org) is an open access database that houses genetic and protein interactions curated from the primary biomedical literature for all major model organism species and humans. As of September 2014, the BioGRID contains 749 912 interactions as drawn from 43 149 publications that represent 30 model organisms. This interaction count represents a 50% increase compared to our previous 2013 BioGRID update. BioGRID data are freely distributed through partner model organism databases and meta-databases and are directly downloadable in a variety of formats. In addition to general curation of the published literature for the major model species, BioGRID undertakes themed curation projects in areas of particular relevance for biomedical sciences, such as the ubiquitin-proteasome system and various human disease-associated interaction networks. BioGRID curation is coordinated through an Interaction Management System (IMS) that facilitates the compilation interaction records through structured evidence codes, phenotype ontologies, and gene annotation. The BioGRID architecture has been improved in order to support a broader range of interaction and post-translational modification types, to allow the representation of more complex multi-gene/protein interactions, to account for cellular phenotypes through structured ontologies, to expedite curation through semi-automated text-mining approaches, and to enhance curation quality control.

Increased levels of C-reactive protein (CRP) and heat-shock protein-70 (Hsp70) in plasma are known to be associated with an increased risk of cardiovascular disease. In this study we have investigated the effects of environmental air pollution particles (PM10) and ultrafine carbon black (ufCB) on the expression of CRP and Hsp70 in the lung epithelial cell line, A549. After treatment with PM10 or ufCB the cells were found to have increased expression of CRP and Hsp70 localized in both the cell cytoplasm and nucleus. Analysis of the cell supernatants revealed that CRP and Hsp70 were present, suggesting secretion of both proteins in response to the particulate treatment. To investigate if the expression of CRP and Hsp70 was the result of free radical production, cells were treated with ufCB in the presence of antioxidants (NAL and Trolox). This revealed that antioxidants reduced the amount of CRP and Hsp70 secreted from the cells. These findings suggest that CRP and Hsp70 may be secreted from the lung epithelium as a result of oxidative stress and have important effects on the inflammatory response associated with inhalation of particulate matter.

Monocyte chemoattractant protein-induced protein 1 (MCPIP1) encoded by the ZC3H12a gene (also known as Regnase-1) is involved in the regulation of degradation of mRNA of inflammatory modulators and for processing of pre-miRNA. These functions depend on the presence of the PIN domain. Moreover, MCPIP1 was described as a negative regulator of NF-κB and AP-1 signaling pathways although mechanisms underlying such activity remain unknown. We aimed at determining the role of MCPIP1 in adipogenesis. Here, we present evidence that Mcpip1 transcription is transiently activated during 3T3-L1 transition from pre- to adipocytes. However Mcpip1 protein expression is also strongly decreased at day one after induction of adipogenesis. Knockdown of Mcpip1 results in an upregulation of C/EBPβ and PPARγ mRNAs, whereas overexpression of MCPIP1 reduces the level of both transcription factors and impairs adipogenesis. MCPIP1-dependend modulation of C/EBPβ and PPARγ levels results in a modulation of the expression of downstream controlled genes. In addition, decreased C/EBPβ, but not PPARγ, depends on the activity of the MCPIP1 PIN domain, which is responsible for RNase properties of this protein. Together, these data confirm that MCPIP1 is a key regulator of adipogenesis.

The BioGRID database is an extensive repository of curated genetic and protein interactions for the budding yeast Saccharomyces cerevisiae, the fission yeast Schizosaccharomyces pombe, and the yeast Candida albicans SC5314, as well as for several other model organisms and humans. This protocol describes how to use the BioGRID website to query genetic or protein interactions for any gene of interest, how to visualize the associated interactions using an embedded interactive network viewer, and how to download data files for either selected interactions or the entire BioGRID interaction data set.

The Biological General Repository for Interaction Datasets (BioGRID) is a freely available public database that provides the biological and biomedical research communities with curated protein and genetic interaction data. Structured experimental evidence codes, an intuitive search interface, and visualization tools enable the discovery of individual gene, protein, or biological network function. BioGRID houses interaction data for the major model organism species—including yeast, nematode, fly, zebrafish, mouse, and human—with particular emphasis on the budding yeast Saccharomyces cerevisiae and the fission yeast Schizosaccharomyces pombe as pioneer eukaryotic models for network biology. BioGRID has achieved comprehensive curation coverage of the entire literature for these two major yeast models, which is actively maintained through monthly curation updates. As of September 2015, BioGRID houses approximately 335,400 biological interactions for budding yeast and approximately 67,800 interactions for fission yeast. BioGRID also supports an integrated posttranslational modification (PTM) viewer that incorporates more than 20,100 yeast phosphorylation sites curated through its sister database, the PhosphoGRID.

Integrins are a family of cell surface receptors which mediate cell–matrix and cell–cell adhesions. Among other functions they provide an important mechanical link between the cells external and intracellular environments while the adhesions that they form also have critical roles in cellular signal-transduction. Cell–matrix contacts occur at zones in the cell surface where adhesion receptors cluster and when activated the receptors bind to ligands in the extracellular matrix. The extracellular matrix surrounds the cells of tissues and forms the structural support of tissue which is particularly important in connective tissues. Cells attach to the extracellular matrix through specific cell-surface receptors and molecules including integrins and transmembrane proteoglycans. Integrins work alongside other proteins such as cadherins, immunoglobulin superfamily cell adhesion molecules, selectins, and syndecans to mediate cell–cell and cell–matrix interactions and communication. Activation of adhesion receptors triggers the formation of matrix contacts in which bound matrix components, adhesion receptors, and associated intracellular cytoskeletal and signaling molecules form large functional, localized multiprotein complexes. Cell–matrix contacts are important in a variety of different cell and tissue properties including embryonic development, inflammatory responses, wound healing, and adult tissue homeostasis. This review summarizes the roles and functions of integrins and extracellular matrix proteins in mechanotransduction.

C-reactive protein (CRP) is the prototypic human acute-phase protein and is found at increased levels in the blood during episodes of inflammation. CRP was generally thought to be produced only by hepatocytes; however, several studies have shown extrahepatic synthesis of CRP. A previous study showed that PM10 and ultrafine carbon black (ufCB) were able to induce CRP expression in A549 cells. This study aims to examine the factors that lead to the production of CRP in A549 cells. A549 human lung epithelial cells were treated with cytokines (interleukin 6, tumor necrosis factor alpha, interferon gamma, or interleukin 1beta) or carbon particles (CB and ufCB) for 18 h. It was found that CRP could be expressed within the cells and that CRP was secreted from the cells particularly with tumor necrosis factor alpha, CB and ufCB treatments. It was also found that this expression of CRP with CB and ufCB treatments was dependent on nuclear factor kappa B (NFkappaB). The expression of CRP in A549 cells may indicate an important role for CRP expression and secretion from lung epithelial cells in response to inflammatory stimuli.

Background This study has investigated the ability of tobacco smoke, and ingredients of tobacco smoke, to induce apoptosis in the airway epithelial cell line A549. Method A549 cells were treated with 80 μg/ml Tobacco smoke condensate (TSC), 10 mM Nicotine, 10 μM paraldehyde, 10 μM hydrogen peroxide, 1 μM Taxol® (Paclitaxel), 100%, 50% and 25% cigarette smoke extract (CSE). Following 4–48 h incubation apoptosis was measured morphologically following staining of cells with DAPI. TUNEL staining was also used to assess DNA damage after 24 and 48 h incubation. In addition, loss of mitochondrial cytochrome C and activation of Bax-α, early events in the apoptotic process, were measured after 4 h of incubation. Results Incubation of A549 cells with vehicle, Taxol, TSC, nicotine, paraldehyde, hydrogen peroxide and CSE caused a time-dependent detachment of the cells from the flask between 6 and 48 h. DAPI staining revealed that the cells remaining adhered to the flask appeared healthy whereas some of those that had detached appeared to be either apoptotic or indeterminate. Treatment with Taxol, TSC, nicotine, paraldehyde, hydrogen peroxide and CSE caused a significant increase in the number of apoptotic cells. Similarly, treatment with Taxol, TSC, nicotine, hydrogen peroxide and CSE caused a significant increase in the number of apoptotic cells among the cells that had detached from the culture plate. After 4 h of incubation, Taxol, TSC, hydrogen peroxide and CSE caused a significant reduction in mitochondrial cytochrome C and an increase in cytosolic cytochrome C. At the same time point, hydrogen peroxide and CSE significantly increased the concentration of Bax-α in the mitochondria. Conclusion Tobacco smoke initiates apoptosis in A549 airway epithelial cells as a result of mitochondrial damage and that this results in a cell detachment and full apoptosis. This effect appears to result from factors in tobacco smoke other than nicotine and may result from free radical activity. However, additional stable factors may also be involved since the free radical content of TSC is likely to be low.

Purpose: Classical neuronal signalling molecules such as substance P and glutamate have been identified in cartilage and are being shown to have roles in regulation of chondrocyte function. This study looks at the expression and function of the metabotropic glutamate NMDA receptor (NMDAR) in OA chondrocytes. Methods: Chondrocytes from articular cartilage of human knee joint arthroplasty specimens. NMDAR subunit expression at gene and protein levels was identified by RTPCR and western blotting. NMDAR function was assayed using radolabelled Ca45 uptake and changes in cell membrane potential in response to NMDA and mechanical stimulation (MS; electrophys. only) in the presence and absence of the NMDA antagonists MK801 (noncompetitive), Ifenprodil (NR2B selective) and APV (competitive). Results: NMDAR1, 2A, and 2B, but not NR2C, 2D and 3, were detected at varying levels in OA chondrocytes at both the protein and RNA levels. Assessment of receptor function showed that OA chondrocytes responded to NMDA; this response was decreased by specific antagonists. NMDA induced an increase in Ca45 uptake which was reduced to baseline by the antagonists. Both NMDA and mechanical stimulation induced a membrane depolarisation which was decreased or abolished by NMDAR antagonists. Conclusions: This study shows the presence of functional NMDA receptors composed of NR1, 2A and 2B subunits on human OA chondrocytes. It has been suggested that the NMDA receptor is a mechanosensitive receptor in neurones and the current investigation suggests similar involvement in chondrocyte mechanotransduction.