Regulation Of Matrix Synthesis Research Articles

The primary cilium conducts chondrocyte mechanotransduction

In several cell types fluid-flow deflection of primary cilia initiates a mechanotransduction pathway via polycystin 1 and 2 (PC1/2). In articular cartilage the chondrocyte primary cilium extends into cartilage matrix. Mechanical signals including compression and fluid flow trigger mechanosensitive regulation of matrix synthesis underpinning tissue homeostasis. Here we tested the hypothesis that the cilium plays a key role in chondrocyte mechanotransduction that includes ATP release, ATP-induced calcium transients and the subsequent regulation of matrix synthesis. These studies used murine chondrocytes with a hypomorphic mutation of Tg737, (encoding IFT88) which abolishes cilia growth. 3D agarose culture allowed compressive loading of WT and Tg737 chondrocytes followed by quantification of ATP release with a luciferase assay, calcium transients by Fluo-4 imaging, and matrix synthesis by qPCR and biochemical assay. Additionally, expression of purinergic receptors (P2R) and polycystins was assessed by western blot and immunocytochemistry. Compression of WT chondrocytes increased calcium transients and matrix production. By contrast this mechanosensitive behaviour was completely abolished in Tg737 cells. However mechanosensitive ATP release was present in both WT and Tg737 cells suggesting that IFT88 and the cilium are required for purinergic reception. Indeed exogenous ATP elicited calcium transients in WT but not in Tg737 cells. P2R expression profiles showed no global differences but polycystin-1 expression was altered in ORPK. We conclude that IFT88 plays a critical role in ATP-induced calcium signalling and is therefore essential to chondrocyte mechanotransduction. Furthermore, this suggests that IFT88 and the cilium may be fundamentally important for purinergic-calcium signalling pathways.

C-28/I2 and T/C-28a2 chondrocytes as well as human primary articular chondrocytes express sex hormone and insulin receptors—Useful cells in study of cartilage metabolism

Sex hormones and insulin have been implicated in articular cartilage metabolism. To supplement previous findings on the regulation of matrix synthesis with 17β-estradiol and insulin and to find a possible model to study cartilage metabolism in vitro, we evaluated the expression of estrogen receptors α and β (ERα, ERβ), androgen receptor (AR) and insulin receptor (IR), in immortalized C-28/I2 and T/C-28a2 chondrocytes and in human primary articular cartilage cells. Chondrocytes were treated with increasing concentrations of 17β-estradiol, dihydrotestosterone or insulin and analyzed by means of RT-PCR and Western blotting. Both cell lines as well as human articular chondrocytes expressed ER α and β, AR and IR at mRNA and protein levels. In immortalized C-28/I2 chondrocytes, we showed that increasing concentrations of 17β-estradiol diminished the 95kDa band of IR. Since 17β-estradiol suppresses insulin-induced proline incorporation and type II collagen synthesis, as we have previously demonstrated, our findings give the first clue that 17β-estradiol may have negative effects on cartilage anabolism triggered by insulin during hormonal imbalance. Compared to chondrocytes cultured without hormones, immunostaining for ERα/β, AR and IR was decreased in both cell lines after incubation of cells with the receptor-specific hormones. It can be assumed that C-28/I2 and T/C-28a2 chondrocytes interact with the respective hormones. Our findings provide a reproducible model for investigating sex hormone and insulin receptors, which are present in low concentrations in articular chondrocytes, in the tissue-specific context of cartilage metabolism.

Regulation of Matrix Synthesis, Remodeling and Accumulation in Glomerulosclerosis

After injury the body normally undergoes a repair process, however when this event becomes deregulated the pathological condition of fibrosis occurs. There are many similarities with respect to the fundamental mechanisms that regulate sclerosis in different organ systems. In this review we utilize the pathological entity of glomerulosclerosis in the kidney to highlight some of the general paradigms whereby extracellular matrix (ECM) is deposited in greater quantities than it is degraded. Our review discusses how genetic and structural abnormalities of specific ECM components can result in fibrosis. In addition, we outline how some key growth factors, integrins and oxidative stress play a role in the development of glomerulosclerosis.

Cell Death and Apoptosis in Ostearthritic Cartilage

Osteoarthritis (OA) is the most common chronic joint disease in the elderly population, causing significant pain and disability. Because the cardinal feature of OA is a progressive loss of articular cartilage, a great portion of the research endeavour into the pathogenesis of OA has been focused on the regulation of matrix synthesis and degradation. The phenotypic stability and survival of the chondrocytes are essential for the maintenance of a proper cartilage matrix. This has lead to the long-standing assumption that cell death is a central feature in OA cartilage degeneration. The important role of apoptosis in OA has been demonstrated in in vitro and in vivo models. However, it should be noted that the relative contribution of apoptotic cell death in the pathogenesis of OA is still difficult to assess because of the chronic nature of the disease process. Therefore, the apoptosis of chondrocytes seems to be a potential target for therapeutic interventions in OA. The death receptor, mitochondrial and endoplasmic reticulum pathways are the major cellular pathways of apoptosis. Of all these elements involved in the apoptosis of chondrocytes, caspase inhibition has been studied with the most detail. Other molecules with the capacity to modulate mitochondria function, phosphatase (PP-1A/B) activity and pro-apoptosis stimuli (NO, prostaglandins, cytokines, ROS) could be excellent targets to block apoptosis of chondrocytes. Finally, the regulation of the natural inhibitors of apoptosis (c-FLIP, BAR, ARC and HC-gp39) could complement the other strategies to reduce cartilage degradation.

Pharmacological Approaches to Prevent Abdominal Aortic Aneurysm Enlargement and Rupture

Current efforts to limit the mortality from abdominal aortic aneurysm (AAA) are dependent on detection and elective repair. Even by conservative estimates, there are more than 300,000 undetected AAAs in the United States, most of which are small and would not require immediate intervention. Current practice following detection of a small AAA includes education, risk factor management, and serial observation. This approach, based on the statistical probability of death from rupture compared to the morbidity and mortality of repair, can be unsettling to patients and lead to a decline in perceived quality of life. While the pathophysiology of AAA is not completely understood, observations from human tissues and animal studies have identified a number of potential targets for inhibiting aneurysm expansion. It is clear that the prominent inflammatory response identified in aneurysm tissue has a role in promoting aortic expansion. This inflammatory response is thought to account for increased expression of proteolytic enzymes. Recent work has suggested a unifying hypothesis centered on the MAP kinase family affecting both the regulation of matrix synthesis and the expression of proteolytic enzymes. The tetracycline antibiotics and antihypertensive medications that affect the angiotensin-converting enzyme system can inhibit proteolysis. There are adequate preliminary data to support a large prospective randomized trial of doxycycline to prevent aneurysm expansion.

Receptors for immune complexes activate gene expression and synthesis of matrix proteins in cultured rat and human mesangial cells: role of TGF-beta.

Most human glomerulonephritis are induced by the deposition and/or formation of immune complexes in the glomerular region. Recently, it has been demonstrated that cultured glomerular mesangial cells (MC) express Fc receptors for IgA and IgG (Fc-alpha and Fc-gamma receptors). In this work, we studied whether the interaction of IgA and IgG complexes with MC induces accumulation of mesangial matrix, the histologic hallmark of progressive glomerular diseases. The exposure of MC to IgA and IgG complexes increased extracellular matrix components, such as fibronectin (FN) and collagens, at both the mRNA and protein levels in a time- and dose-dependent manner. Monomeric IgA or F(ab')2 fragments did not increase FN production, indicating that a constant region of IgA and cross-linking of Fc-alpha receptors are required. We also explored the role of TGF-beta, a profibrogenic cytokine, in the regulation of matrix synthesis. Both IgA and IgG complexes caused in MC an augmentation in TGF-beta1 mRNA and TGF-beta activity and the conversion of latent TGF-beta to the biologically active form. The coincubation of cells with complexes and a neutralizing Ab to TGF-beta significantly reduced the FN synthesis. These results indicate that the Fc receptor occupancy of MC increases the production of extracellular matrix proteins. The autocrine TGF-beta synthesis appears to be largely responsible for this effect. These findings could have implications for a better understanding of the glomerulosclerosis process in immune complex nephritis.