Chronic lymphocytic leukemia (CLL) cells rapidly undergo apoptosis in vitro, suggesting that the in vivo microenvironment provides crucial antiapoptotic signals. Overexpression of the antiapoptotic proteins Bcl-2 and Mcl-1 is a hallmark of CLL, and their expression is further enhanced in the lymphoid tissues. However, the high levels of Mcl-1 found in peripheral blood samples, coupled with its short half-life, led us to hypothesize that it must be actively maintained in the peripheral circulation. Coculture of CLL cells with human vascular endothelial cells significantly enhanced tumor cell survival, an effect that was not observed with normal B cells. This was associated with elevated levels of the antiapoptotic proteins Bcl-2, Mcl-1, and Bcl-X L and marked increased expression of CD38 and CD49d, both of which are associated with clinically aggressive disease. Because CD38, CD49d, and some Bcl-2 family genes are transcriptional targets for NF-κB, we assessed NF-κB activation following coculture with endothelial cells. DNA binding of the NF-κB subunit Rel A was significantly increased and strongly correlated with changes in transcription of CD38, CD49d, BCL2, MCL1, and BCLXL, effects that were reversed by a peptide inhibitor of Rel A. These effects were not observed following coculture with nonendothelial cell lines. Therefore, CLL cells receive specific survival signals following interaction with endothelial cells mediated through the activation of NF-κB and the induction of downstream target genes. This type of interaction in the peripheral vasculature may explain the constitutive NF-κB activation and the overexpression of Bcl-2 family proteins commonly seen in this disease.
Summary
Interactions in the tumour microenvironment can promote chronic lymphocytic leukaemia (CLL) cell survival, proliferation and drug resistance. A detailed comparison of three co‐culture systems designed to mimic the CLL lymph node and vascular microenvironments were performed; two were mouse fibroblast cell lines transfected with human CD40LG or CD31 and the third was a human microvascular endothelial cell line, HMEC‐1. All three co‐culture systems markedly enhanced CLL cell survival and induced a consistent change in CLL cell phenotype, characterized by increased expression of CD38, CD69, CD44 and ITGA4 (CD49d); this phenotype was absent following co‐culture on untransfected mouse fibroblasts. In contrast to HMEC‐1 cells, the CD40LG and CD31‐expressing fibroblasts also induced ZAP70 expression and marked CLL cell proliferation as evidenced by carboxyfluorescein succinimidyl ester labelling and increased Ki‐67 expression. Taken together, our data show that co‐culture on different stroma induced a remarkably similar activation phenotype in CLL cells but only the CD40LG and CD31‐expressing fibroblasts increased ZAP70 expression and CLL cell proliferation, indicating that ZAP70 may play a critical role in this process. This comparative study reveals a number of striking similarities between the co‐culture systems tested but also highlights important differences that should be considered when selecting which system to use for in‐vitro investigations.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.