Central Role of SREBP-2 in the Pathogenesis of Osteoarthritis

Background: Recent studies have implied that osteoarthritis (OA) is a metabolic disease linked to deregulation of genes involved in lipid metabolism and cholesterol efflux. Sterol Regulatory Element Binding Proteins (SREBPs) are transcription factors regulating lipid metabolism with so far no association with OA. Our aim was to test the hypothesis that SREBP-2, a gene that plays a key role in cholesterol homeostasis, is crucially involved in OA pathogenesis and to identify possible mechanisms of action. Methodology/Principal Findings: We performed a genetic association analysis using a cohort of 1,410 Greek OA patients and healthy controls and found significant association between single nucleotide polymorphism (SNP) 1784G>C in SREBP-2 gene and OA development. Moreover, the above SNP was functionally active, as normal chondrocytes’ transfection with SREBP-2-G/C plasmid resulted in interleukin-1β and metalloproteinase-13 (MMP-13) upregulation. We also evaluated SREBP-2, its target gene 3-hydroxy-3-methylglutaryl-coenzymeA reductase (HMGCR), phospho-phosphoinositide3-kinase (PI3K), phospho-Akt, integrin-alphaV (ITGAV) and transforming growth factor-$\beta$ (TGF-$\beta$) mRNA and protein expression levels in osteoarthritic and normal chondrocytes and found that they were all significantly elevated in OA chondrocytes. To test whether TGF-$\beta$ alone can induce SREBP-2, we treated normal chondrocytes with TGF-$\beta$ and found significant upregulation of SREBP-2, HMGCR, phospho-PI3K and MMP-13. We also showed that TGF-$\beta$ activated aggrecan (ACAN) in chondrocytes only through Smad3, which interacts with SREBP-2. Finally, we examined the effect of an integrin inhibitor, cyclo-RGDFV peptide, on osteoarthritic chondrocytes, and found that it resulted in significant upregulation of ACAN and downregulation of SREBP-2, HMGCR, phospho-PI3K and MMP-13 expression levels. Conclusions/Significance: We demonstrated, for the first time, the association of SREBP-2 with OA pathogenesis and provided evidence on the molecular mechanism involved. We suggest that TGF-$\beta$ induces SREBP-2 pathway activation through ITGAV and PI3K playing a key role in OA and that integrin blockage may be a potential molecular target for OA treatment

Experimental evidence of Migfilin as a new therapeutic target of hepatocellular carcinoma metastasis

Migfilin is a novel cell-matrix adhesion protein known to interact with Vasodilator Stimulated Phosphoprotein (VASP) and be localized both at cell-matrix and cell-cell adhesions. To date there is nothing known about its role in hepatocellular carcinoma (HCC). As matrix is important in metastasis, we aimed to investigate the Migfilin's role in HCC metastasis using two human HCC cell lines that differ in their metastatic potential; non-invasive Alexander cells and the highly invasive HepG2 cells. We silenced Migfilin by siRNA and studied its effect on signaling and metastasis-related cellular properties. We show that Migfilin's expression is elevated in HepG2 cells and its silencing leads to upregulation of actin reorganization-related proteins, namely phosphor-VASP (Ser157 and Ser239), Fascin-1 and Rho-kinase-1, promoting actin polymerization and inhibiting cell invasion. Phosphor-Akt (Ser473) is decreased contributing to the upregulation of free and phosphor-beta-catenin (Ser33/37Thr41) and inducing proliferation. Migfilin elimination upregulates Extracellular Signal-regulated kinase, which increases cell adhesion in HepG2 and reduces invasiveness. This is the first study to reveal that Migfilin inhibition can halt HCC metastasis in vitro, providing the molecular mechanism involved and presenting Migfilin as potential therapeutic target against HCC metastasis. (C) 2015 Elsevier Inc. All rights reserved

Elimination of Ras Suppressor-1 from Hepatocellular Carcinoma Cells Hinders their In Vitro Metastatic Properties

Background/Aim: Extracellular matrix (ECM) is of great significance for homeostasis in the liver. In fact, one of the stags leading to hepatocellular carcinoma (HCC) includes accumulation of excess ECM. Ras Suppressor-1 (RSU-1) is localized in the cell-ECM adhesions but its role in HCC is unexplored. Materials and Methods: We investigated the expression and role of RSU-1 in two HCC cell lines that differ in aggressiveness; non-invasive Alexander cells and highly invasive HepG2 cells. Results: Our results showed that RSU-1 expression is elevated in HepG2 cells both at the mRNA and protein level, while its silencing leads to increased cell proliferation in both cell lines. Interestingly, RSU-1 depletion from highly invasive HepG2 cells reduces cell adhesion and invasion. Conclusion: This is the first study to provide in vitro evidence for the involvement of RSU-1 in HCC cell invasive behavior

Lipid metabolism and osteoarthritis: Lessons from atherosclerosis

Osteoarthritis (OA) is an age-related degenerative disease comprising the main reason of handicap in the Western world. Interestingly, to date, there are neither available biomarkers for early diagnosis of the disease nor any effective therapy other than symptomatic treatment and joint replacement surgery. OA has long been associated with obesity, mainly due to mechanical overload exerted on the joints. Recent studies however, point to the direction that OA is a metabolic disease, as it also involves non-weight bearing joints. In fact, altered lipid metabolism may be the underlying cause. First, adipokines have been shown to be key regulators of OA pathogenesis. Second, epidemiological studies have shown serum cholesterol to be a risk factor for OA development. Third, lipid deposition in the joint is observed at the early stages of OA before the occurrence of histological changes. Fourth, proteomic analyses have shown an important connection between OA and lipid metabolism. Finally, recent gene expression studies reveal a deregulation of cholesterol influx and efflux and in the expression of lipid metabolism-related genes. Interestingly, lipids and lipid metabolism are known to be implicated in the development and progression of another age-related degenerative disease, atherosclerosis (ATH). Thus, although it is tempting to speculate that the osteoarthritic chondrocyte has been transformed to foam cell, it has not been proven yet. However, this may be an intriguing theory linking ATH and OA, which may open new avenues to novel therapeutic interventions for OA taking advantage of previous knowledge from ATH. (C) 2010 Published by Elsevier Ltd

Proteomics of osteoarthritic chondrocytes and cartilage

Osteoarthritis (OA) is characterized by irreversible destruction of the articular cartilage. OA affects more than 100 million individuals worldwide and has a major impact on patients' quality of life. The lack of effective therapy that prevents, inhibits or reverses the progress of OA often leaves only the option of surgical interventions. Thus, identification of the factors that contribute to OA pathogenesis is necessary for better understanding of OA pathobiology and discovery of effective therapies. Recent proteomic studies have been conducted to identify pathological mediators and biomarkers of OA, which have pinpointed novel pathways involved in cartilage degeneration. This article summarizes the recent findings, compares major techniques used in OA proteomics and discusses key proteins in OA and their potential use as therapeutic targets

Actin cytoskeleton dynamics linked to synovial fibroblast activation as a novel pathogenic principle in TNF‐driven arthritis

Rheumatoid arthritis is a chronic inflammatory disorder whose origin of defect has been the subject of extensive research during the past few decades. While a number of immune and non‐immune cell types participate in the development of chronic destructive inflammation in the arthritic joint, synovial fibroblasts have emerged as key effector cells capable of modulating both joint destruction and propagation of inflammation. Ample evidence of aberrant changes in the morphology and biochemical behaviour of rheumatoid arthritis synovial fibroblasts have established the tissue evading and “transformed” character of this cell type. We have recently demonstrated that actin cytoskeletal rearrangements determine the pathogenic activation of synovial fibroblasts in modelled TNF‐mediated arthritis, a finding correlating with similar gene expression changes which we observed in human rheumatoid arthritis synovial fibroblasts. Here, we show that pharmacological inhibition of actin cytoskeleton dynamics alters potential pathogenic properties of the arthritogenic synovial fibroblast, such as proliferation, migration and resistance to apoptosis, indicating novel opportunities for therapeutic intervention in arthritis. Recent advances in this field of research are reviewed and discussed

Mitogen-inducible gene-2 (MIG2) and migfilin expression is reduced in samples of human breast cancer

Background: Cell adhesion proteins that connect each cell to neighboring cells and the extracellular matrix play a fundamental role in metastasis. Mitogen-inducible gene-2 (MIG2), is a cell-matrix adhesion protein, which through migfilin, interacts with filamin-A, being linked to actin cytoskeleton. Aim: Recent studies have implicated both MIG2 and migfilin in cancer, but little is known regarding their expression in breast cancer. In this study, we investigated this topic. Materials and Methods: mRNA and protein expression was examined in 30 breast cancer samples and compared to that of normal adjacent tissue using real time-polymerase chain reaction (PCR) and western blotting. Results: Our results showed that expression of MIG2 and migfilin was significantly reduced in the majority of the breast cancer tissues compared to normal tissues regardless of metastatic status and disease stage. Conclusion: Both MIG2 and migfilin are down-regulated in breast cancer

Depletion of Ras Suppressor-1 (RSU-1) promotes cell invasion of breast cancer cells through a compensatory upregulation of a truncated isoform

Extracellular matrix (ECM)-adhesion proteins and actin cytoskeleton are pivotal in cancer cell invasion. Ras Suppressor-1 (RSU-1), a cell-ECM adhesion protein that interacts with PINCH-1, thus being connected to Integrin Linked Kinase (ILK), alpha-parvin (PARVA), and actin cytoskeleton, is up-regulated in metastatic breast cancer (BC) samples. Apart from the originally-identified gene (RSU-1L), an alternatively-spliced isoform (RSU-1-X1) has been reported. We used non-invasive MCF-7 cells, expressing only RSU-1L, and highly invasive MDA-MB-231-LM2 expressing both isoforms and generated stable shRNA-transduced cells lacking RSU-1L, while the truncated RSU-1-X1 isoform was depleted by siRNA-mediated silencing. RSU-1L depletion in MCF-7 cells resulted in complete abrogation of tumor spheroid invasion in three-dimensional collagen gels, whereas it promoted MDA-MB-231-LM2 invasion, through a compensatory upregulation of RSU-1-X1. When RSU-1-X1 was also eliminated, RSU-1L-depletion-induced migration and invasion were drastically reduced being accompanied by reduced urokinase plasminogen activator expression. Protein expression analysis in 23 human BC samples corroborated our findings showing RSU-1L to be upregulated and RSU-1-X1 downregulated in metastatic samples. We demonstrate for the first time, that both RSU-1 isoforms promote invasion in vitro while RSU-1L elimination induces RSU-1-X1 upregulation to compensate for the loss. Hence, we propose that both isoforms should be blocked to effectively eliminate metastasis. © 2019, The Author(s)