Vascular endothelial growth factor B (VEGF-B), a member of the VEGF/PDGF family, is highly expressed in many tissues with two differentially spliced transcripts generating two secreted isoforms, VEGF-B₁₆₇ and VEGF-B₁₈₆. In this work, we have investigated the expression of VEGF-B in tissues and cell lines using techniques that can distinguish the two isoforms. The results showed that the VEGF-B₁₆₇ isoform was predominantly expressed in most tissues, accounting for more than 80% of the total VEGF-B transcripts. The VEGF-B₁₈₆ isoform was expressed at lower levels and only in a limited number of tissues. Moreover, the VEGF-B₁₈₆ isoform was up-regulated in mouse and human tumor cell lines and primary tumors compared with their corresponding normal tissues. Taken together, our data suggest a fme genetic control of the expression of the two isoforms of VEGF-B, implying tissue- and cell-specific roles of the two VEGF-B isoforms.