Mediation of LA-4 lung epithelial cell injury by alloreactive donor T cells.

Abstract

6575 Background: Acute noninfectious lung injury is associated with significant mortality in patients receiving allogeneic hematopoietic stem cell transplantation (allo-HSCT). While alloantigen-reactive donor T cells migrating into the lung seem critical, the role of T cell-mediated lung epithelial cell apoptosis is still unclear. Methods: Mouse LA-4 lung epithelial cells (H2-Dd) were analyzed for their surface marker expression by FACS. For cytotoxic T lymphocyte (CTL) chromium release assays, murine alloreactive C57BL/6 effector T cells (H2-Db) were purified following stimulation using allogeneic A/He mouse splenocytes (H2-Dd) and then plated against syngeneic EL-4 (H2-Db) or allogeneic LA-4 (H2-Dd) target cells. Results: Unstimulated LA-4 cells were H2-Kk - (MHCI), H2-Dd ++ (MHCI), I-Ak - (MHCII), TNFRI (+), TNFR II (+), Fas +, FasL -, CD86 -, CD80 (+), ICAM-1 -. Stimulation with lipopolysaccharide but not IFN gamma resulted in strong upregulation of H2-Dd and Fas, and neither stimulus changed any of the other markers tested. While LA-4 cells, consistent with their lack of MHCII expression, were not able to induce a significant proliferative T cell response, it seemed likely, that constitutive and inducible MHCI expression as well as TNRI/TNFRII and/or Fas expression rendered LA-4 cells as targets of alloreactive CD8+ T cell toxicity. While alloantigen specific lysis of LA-4 cells through C57BL/6 effector CD8+ T cells occurred in direct relation to the effector:target cell ratio applied in the CTL assay, no lysis was seen using syngeneic EL-4 target controls. To determine, whether perforin/granzyme, TNF or FasL:Fas were involved in T cell mediated LA-4 cell killing, we next used either effector T cells from perforin-deficient (pfp-/-) donor mice or targeted TNF or FasL using neutralizing antibodies (Ab). Anti-TNF Ab’s decreased cytotoxicity by 25%, whereas anti- FasL Ab’s only showed a minor and the use of pfp-/- T cells showed no effect. Conclusions: Our data demonstrate, that LA-4 lung epithelial cells are prone to alloreactive T cell injury involving TNF as a major cytotoxic mediator, and strengthen the concept of the lung as a target organ of GVHD.