A Putative Role for VEGFR-1 (FLT-1) in B Cell Commitment and Differentiation.

VEGF and its receptors are expressed in the hematopoietic system. A role for FLT-1 in particular was described in monocyte-macrophage migration and lineage differentiation (Sawano A et al, 2001), megakaryocytes maturation (Casella I et al, 2003) and dendritic cell differentiation (Dikov M et al, 2005). Given that the expression of this receptor in the lymphoid lineage is not known, we to studied FLT-1 expression and a putative function in normal lymphoid progenitors. To address this question we induced in vitro CD34⁺ cells differentiation into the B cell lineage using a well established assay (on S17 stromal cells). With this approach, we observed that FLT-1 is expressed throughout B cell differentiation increasing along the differentiation process, and reaching its highest at the “immature B cell” stage. We also neutralized FLT-1 during B cell differentiation in vitro. Surprisingly, in the presence of the FLT-1 neutralizing antibody (6.12 monoclonal Ab, from ImClone systems), at the end of the assays (4 different experiments) a significantly higher number of CD19⁺ cells (mainly immature B cells) were detected. Analyzing some of the transcription factors known to be involved in the commitment and differentiation of lymphoid B cells, we observed that the expression of PU.1, Pax5 and E47 was up-regulated by FLT-1 neutralization. Next, given that FLT-1 function was mainly associated with cell migration, and since it is expressed in B cells that are ready to exit the bone marrow into secondary lymphoid organs, we reasoned that FLT-1 might have a role in B cells exit from the bone marrow. For this purpose, we treated mice with the FLT-1 neutralizing Ab for 3 days and analyzed B cells levels in bone marrow and peripheral blood. FLT-1 neutralization led to a significant decrease (p<0.05) in B cells in the bone marrow and peripheral blood. Taken together, our data supports a clear role for FLT-1 in B cell commitment. To understand if VEGF/PlGF signalling through FLT-1 promotes myeloid differentiation, suppresses B cell differentiation or simply regulates the quiescent state of hematopoietic stem cells, we differentiated in vitro CD34⁺/FLT-1⁻ cells and CD34⁺/FLT-1⁺ cells (10% of CD34+ cells) using the assay described above. Interestingly, CD34⁺/FLT-1⁻ differentiation in vitro largely promoted B cell differentiation, while CD34⁺/FLT-1⁺ cells originated mostly myeloid cell differentiation. We are currently exploiting the molecular basis whereby FLT-1 signalling may impair B cells commitment and possibly promotes myeloid differentiation.