Project description:Accumulating evidence suggests that synovitis is associated with osteoarthritic process. Macrophages are predominant cell type in synovium and play principal role in development of synovitis. Our earlier study suggests that the interaction between cartilage fragments and macrophages exacerbates osteoarthritic process. However, molecular mechanisms by which cartilage fragments trigger cellular responses remain to be investigated. Therefore, the current study aims at analyzing molecular response of macrophages to cartilage fragments by RNA-seq
Project description:Joint diseases are often characterized by inflammatory processes resulting in pathological changes in joint tissues, including cartilage degradation and release of components to the synovial fluid. The complement system plays a central role in promoting the inflammation. Since several cartilage proteins are known to interact with complement, causing either activation or inhibition of the system, we aimed to investigate these interactions comprehensively. Bovine cartilage explants were cultured with interleukin-1alpha (IL-1a) to induce cartilage degradation, followed by incubation with human serum to allow interactions with complement. Label-free selected reaction monitoring (SRM) mass spectrometry (MS) was then used to specifically quantify complement proteins interacting with the cartilage explant. In parallel, the time-dependent degradation of cartilage was detected using tandem MS (MS/MS). Complement proteins resulting from activation of the classical and alternative pathway as well as the terminal pathway were detected on IL-1a stimulated cartilage at time points when clear alterations in the extracellular matrix composition had occurred. To confirm SRM results indicating complement activation, increased levels of the complement activation product C4d were detected by ELISA in serum after incubation with IL-1a stimulated cartilage. Further, typical activated (cleaved) C3 fragments were detected by western blotting of urea extracts of IL-1a stimulated cartilage. No complement activation was triggered by cartilage cultured in the absence of IL-1a. Components released from IL-1a stimulated cartilage during culture had an inhibitory effect on complement activation. These were released after a longer incubation period with IL-1a and may represent a feedback reaction to cartilage-triggered complement activation observed after a shorter incubation period.
Project description:We report high-throughput profiling of gene expression from murine primary macrophages. We profile mRNA in control and endotoxin stimulated macrophages, and examine the effect of AHR ligand (SGA360) under inflammatory status. This study provides a framework for understanding transcriptional changes caused by SGA360 during activated inflammatory signaling .
Project description:Synovial macrophages that are activated by cartilage fragments initiate synovitis, a condition that promotes hypertrophic changes in chondrocytes leading to cartilage degeneration in OA. In this study, we analyzed the molecular response of chondrocytes under condition of this type of stimulation to identify a molecular therapeutic target. Stimulated macrophages promoted hypertrophic changes in chondrocytes resulting in production of matrix-degrading enzymes of cartilage. Among the top-upregulated genes, FliI was found to be released from activated chondrocytes and exerted autocrine/paracrine effects on chondrocytes leading to an increase in expression of catabolic and hypertrophic factors. Silencing FliI in stimulated cells significantly reduced expression of catabolic and hypertrophic factors in cocultured chondrocytes. Our further results demonstrated that the FliI-TLR4-ERK1/2 axis is involved in the hypertrophic signaling of chondrocytes and catabolism of cartilage. Our findings provide a new insight into the pathogenesis of OA and identify a potentially new molecular target for diagnostics and therapeutics.
Project description:We used RNA-seq to profile the effect of a bacterially derived fatty on LPS-stimulated murine macrophages. From the transcriptional response, there was a significant decrease of inflammatory signals with lipid treatment, and pathway analysis of differentially expressed genes suggests PPAR alpha as potential anti-inflammatory mechanism.
Project description:The Toll-like receptors (TLRs) recognize different pathogen associated molecular patterns (PAMPs) and promote MyD88 dependent and independent responses. Previously, we have reported the discovery of the temporal changes in signaling cascades of macrophage proteome and secretome post-stimulation with different PAMPs. Here we present strategies to profile the secretome of TLR2-, TLR4, and TLR7- stimulated macrophages using whole pathogens were developed. Stable isotope labeling with amino acids in cell culture of macrophages was integrated with whole pathogen macrophage stimulation and subsequent targeted proteomics to quantify cytokines, chemokines, and transcription factors.
Project description:This SuperSeries is composed of the following subset Series: GSE5099: Expression Data from Macrophage Maturation and Polarization Experiment GSE35433: Genome-wide analysis of human macrophages stimulated with IL-4 (20ng/ml) (Illumina) GSE35434: Genome-wide analysis of human macrophages stimulated with IL-4 (10ng/ml) (Illumina) GSE35435: Genome-wide analysis of mouse macrophages stimulated with IL-4 (bone marrow macrophages) (Affymetrix) GSE35436: Genome-wide analysis of mouse macrophages stimulated with IL-4 (Biogel and thioglycollate macrophages) (Affymetrix) Refer to individual Series