PROJECT SUMMARY/ABSTRACT Restrictive allograft syndrome (RAS) is a particularly aggressive form of chronic rejection post-lung transplantation, marked by an extremely poor prognosis (median survival of < 1 year) and little therapeutic options. Pathogenesis of RAS remains elusive although association with donor specific antibodies and antibody mediated rejection have been found in human studies. We have recently characterized a murine lung transplant model which recapitulates the histologic changes of RAS and establishes the requisite role of humoral immune response in its development. A unique feature noted in the murine RAS lung allograft was the presence of activated B cells, plasmablasts, and fully differentiated plasma cell (PCs). PCs were also identified in human RAS lungs suggesting that a rejecting lung serves as an inflammatory local niche for humoral immune responses. These antibody secreting cells (ASCs) localized along the bronchovascular bundles (BVBs) and sub-pleural space, lying in close association with expanding mesenchymal cells (MCs). The key role of specialized stromal cells, via paracrine factors of C-X-C Motif Chemokine Ligand 12 (CXCL12) and interleukin (IL)-6, in establishment of a stable survival niche for ASCs is well recognized across a variety of lymphoid and hematopoietic tissues. We have recently characterized the in situ niche of a subpopulation of lung-resident mesenchymal cells which expand and contribute to fibroproliferation in a RAS allograft. This Foxf1+/Gli1+/Scal1+/Col1+ mesenchymal stromal cell (MSC) population forms a three dimensional network along the bronchovascular bundle (BVB-MSCs). Our new preliminary data demonstrates that these cells are the high CXCL12/IL-6 expressing population, and lie in close apposition to ASCs in the rejecting lung allograft. A novel mechanism of upregulation of CXCL12 expression in BVB-MSCs by IL-6 transsignaling and downstream JAK/Stat activation, with recruited monocytes contributing to the soluble IL-6R, was identified. In this proposal we will investigate an innovative and novel hypothesis of paracrine signaling between this graft-resident mesenchymal stromal cell population and immune cells and the role of IL-6 transsignaling/CXCL12 axis in regulating the humoral inflammatory niche in a rejecting lung. The proposed experiments will utilize our ability to identify and conditionally target the specific BVB-MSC subpopulation, and the novel orthotropic whole lung transplant model of RAS to elucidate the spatial in vivo niche of APCs, its temporal regulation by CXCl12 expressing BVB-MSCs, and its functional significance in the pathogenesis of RAS (Aim 1). The role of IL-6 and IL-6 trans-signaling in cellular communication between resident MCs and infiltrating immune cells within this niche and the pathogenesis of RAS will be determined (Aim 2). The effect of specific drugs targeting humoral cell responses and IL-6 signaling pathway will be tested in the murine lung allograft ...