Fibronectin interaction with α4β1 integrin prevents apoptosis in B cell chronic lymphocytic leukemia:: correlation with Bcl-2 and Bax

B cell chronic lymphocytic leukemia (B-CLL) consists of the accumulation of malignant cells that apparently escape normal apoptotic regulation. We have studied the role of alpha 4 beta 1 integrin/fibronectin interaction in preventing apoptosis of these cells in vitro. B cells from 16 patients showed constant expression of alpha 4 beta 1 and little or no alpha 5 beta 1. B-CLL cells cultured on fibronectin or two previously described fibronectin recombinant fragments (H89 and H0) which contain the ligands for alpha 4 beta l, consistently showed higher viability than control cells cultured on poly-lysine. The H89 fragment, containing the high affinity ligand CS-1, was the most efficient substrate with mean cell viability values of 72, 60 and 35% at days 2, 5 and 8 of culture, respectively. For control cells these values were 40, 27 and 15%, respectively. Parallel cell cycle analysis confirmed these results. The anti-apoptotic effect required direct contact with immobilized substrata since it was not observed when using B-CLL conditioned media alone or when clustering alpha 4 beta 1 with specific mAbs in suspension. Quantitation of the apoptosis regulatory proteins Bcl-2 and Fax revealed that cells cultured on the H89 fragment showed high/moderate levels of Bcl-2 (with some interpatient variation) and low levels of Fax resulting in an elevated Bcl-2/Bax ratio. These results indicate that adhesion of B-CLL cells to fibronectin upregulate the Bcl-2/Bax ratio and this may contribute to the anti-apoptotic effect induced via alpha 4 beta 1 integrin.