HUMAN EOSINOPHILS PREFERENTIALLY SURVIVE ON TISSUE FIBRONECTIN COMPARED WITH PLASMA FIBRONECTIN

Background Eosinophil-derived inflammatory mediators including cytokines are considered to be important in the pathogenesis of allergic inflammation. Fibronectin (Fn) has been shown to be a physiological trigger of autocrine cytokine production by human eosinophils. Fn is encoded by a single gene, but alternate splicing of the primary RNA transcript results in polypeptide diversity in a cell type-specific fashion. Thus, tissue Fn contains approximately 50% more of the CS-1 cell binding region recognized by the integrin alpha 4 beta 1 compared with plasma Fn.

Objective Since eosinophils are predominantly tissue-dwelling cells we compared the effect of tissue and plasma Fn on eosinophil survival in culture.

Methods The viability and cytokine generation of eosinophils (>99.9% pure) cultured for up to 4 days in 96 well plates coated with tissue Fn, plasma Fn or BSA was compared.

Results There was a significant difference in the ability of tissue Fn to support eosinophil survival compared with plasma Fn (P < 0.01). Optimal survival with tissue Fn was seen at 25 mu g/well (70% +/- 2.0% viability at 3 days vs 7% +/- 2.2% viability on BSA). Significant (P < 0.001) cell viability on tissue Fn was observed for up to 4 days in culture (54% +/- 6.0%) compared with BSA coated wells. Addition of autologous mononuclear cells (final concentration 0.5%, 1% or 2%) resulted in plasma Fn-dependent eosinophil survival comparable to that of 99.9% pure eosinophils adherent to tissue Fn. Tissue Fn-dependent survival was significantly inhibited by anti-interleukin-3, anti-granulocyte macrophage colony stimulating factor (GM-CSF) and anti-IL-5 monoclonal antibodies. Picogram quantities of these three cytokines were detected in supernatants from eosinophils cultured for 3 days on tissue Fn using specific enzyme-linked immunosorbent assays (ELISAs). Eosinophil survival on tissue Fn was significantly inhibited by anti-pr and alpha 4 beta 1 monoclonal antibody (MoAb) and also by a MoAb specific for the CS-1 motif in the IIICS region of Fn.

Conclusion These observations show preferential survival of eosinophils cultured on tissue Fn as a result of alpha 4 beta 1-dependent interaction with the CS-1 region of tissue Fn triggering autocrine cytokine synthesis and release, thereby promoting their survival and persistence within the tissues.