A peripheral neuroimmune link: glutamate agonists upregulate NMDA NR1 receptor mRNA and protein, vimentin, TNF-α, and RANTES in cultured human synoviocytes

Abstract

Human primary and clonal synovial cells were incubated with glutamate receptor agonists to assess their modulating influence on glutamate receptors N-methyl-d-aspartate (NMDA) NR1 and NR2 and inflammatory cytokines to determine potential for paracrine or autocrine (neurocrine) upregulation of glutamate receptors, as has been shown for bone and chondrocytes. Clonal SW982 synoviocytes constitutively express vimentin, smooth muscle actin (SMA), and NMDA NR1 and NR2. Coincubation (6 h) with glutamate agonists NMDA (5 microM), and the NMDA NR1 glycine site activator (+/-)1-aminocyclopentane-cis-1,3-dicarboxylic acid (5 muM), significantly increases cellular mRNA and protein levels of glutamate receptors, as well as increasing vimentin, SMA, tumor necrosis factor-alpha, and RANTES (regulated on activation, normal T-cell expressed and secreted), assessed qualitatively and quantitatively with nucleotide amplification, image analysis of immunocytochemical staining, fluorescein-activated cell sorting, Western blotting, and immunoassays. Human primary synovial cells harvested from patients with arthritic conditions also constitutively expressed NMDA NR1 with increases after agonist treatment. Glutamate receptor agonist-induced increases were blocked by the noncompetitive glutamate antagonist MK-801 (8 microg/ml) and NR1 blocking antibody. Coincubation with glutamate agonists and phorbol 12-myristate 13-acetate, a protein kinase C activator, significantly enhanced mean levels of TNF-alpha and RANTES in SW982 cell supernatants compared with incubation with either agent alone. Increases were diminished with protein kinase inhibitor and NR1 blocking antibody. The functional activation of glutamate receptors on human synoviocytes establishes a neurogenic cell signaling link between neurotransmitter glutamate released from nerve terminals and target cells in the joint capsule. The influence of glutamate on subsequent release of cellular proinflammatory mediators in non-neural tissue for activation of downstream immune events supports a peripheral neuroimmune link in arthritis.

Impact of Central and Peripheral TRPV1 and ROS Levels on Proinflammatory Mediators and Nociceptive Behavior

“…In the present study, the clonal type B synoviocytes were utilized as an in vitro model of knee joint inflammation since these synoviocytes produce inflammatory mediators when activated with another channel activator, glutamate [21]. Unstable hydroxyl radicals (HO•) were induced by TRPV1 agonist resiniferatoxin in cultured clonal SW982 synoviocytes.…”

Potential Role of Fibroblast-Like Synoviocytes in Joint Damage Induced by Brucella abortus Infection through Production and Induction of Matrix Metalloproteinases

“…The immortalized human FLS cell line SW982 was obtained from the ATCC (Rockville, MD). The fibroblast phenotype of SW982 cells has been demonstrated by their expression of vimentin, a fibroblast marker (30,51). This cell line has been successfully used to study the expressions of inflammatory cytokines and MMPs by FLS upon different stimuli (30,47,56).…”

“…The fibroblast phenotype of SW982 cells has been demonstrated by their expression of vimentin, a fibroblast marker (30,51). This cell line has been successfully used to study the expressions of inflammatory cytokines and MMPs by FLS upon different stimuli (30,47,56). The SW982 cell line was cultured as monolayers in a 5% CO 2 atmosphere at 37°C in a Dulbecco's modified Eagle medium F-12 nutrient mixture (Gibco, Grand Island, NY) supplemented with 2 mM L-glutamine, 10% heat-inactivated fetal bovine serum (FBS) (Gibco), 100 U/ml penicillin, and 100 g/ml streptomycin.…”

Anti-inflammatory effect of triterpene 3β, 6β, 16β-trihydroxylup-20(29)-ene obtained from Combretum leprosum Mart & Eich in mice

“…The activation of glutamate NMDA receptors causes the secretion of inflammatory mediators, i.e., TNF-a and IL-1b, that in turn can sensitize peripheral nerve endings and activate the immune response (Kane et al, 2005;McNearney et al, 2010;Sweeney and Firestein, 2004;Zelenka et al, 2005). Additionally, Feng et al (2003) found an increase in the concentration of glutamate and aspartate in the cerebrospinal fluid after i.p.…”