Reactive arthritis is characterized by the presence of inflammation remote from the site of infection, associated with a number of infectious organism, such as Yersinia ssp. Attachment and entry of Yersinia ssp. into the nonphagocytic cells of the intestinal epithelium is primarily mediated by invasin, a ligand for beta1 integrins. The reactive arthritis, which occurs even in the absence of T-cells, includes hyperplasia of synovial lining cells, excess breakdown of ECM proteins and cellular exudates within the periarticular cavity beads coated with the wild type invasin, but not an adhesion deficient invasin (D911A). Preliminary experiments show that rabbit synovial fibroblasts (RSF) readily phagocytose, resulting in a 10-30 fold increase collagenase gene expression. To determine the mechanism of MMPs induction after ligation of beta 1 integrins by invasin signaling molecules involved in reorganizing the cytoskeleton that are induced as an early response to ligation of beta 1 integrins by invasin; in particular, the mechanism by which invasin-triggered endocytosis signals will compound with that triggered invasin-mediated RSF spreading. The molecules associating with the integins ligated by invasin in the two different situations, the immediate downstream signaling molecules and the mechanism of activation of NfkappaB, which is upstream from IL-1 induced by phagocytosis will be identified. One plausible hypothesis for the different signaling pathways is that the invasin-triggered endocytosis involves signal transduction machinery that is activated in the endosome compartment following phagocytosis and invasin-mediated receptor internalization. To test this model phosphorylation of the adapter protein-2, which is involved in clathrin assembly, will be studied. The rate of small GTPases of the Rho family will be studied by transfecting dominant negative and constitutively active constructs of these genes into RSF to see how they influence signal transduction. To compare signaling pathways elucidated in vitro with effects in vivo, a mouse model of Yersinia invasin-induced reactive arthritis will be established using purified invasin protein or Yersinia organisms.
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