VEGF is crucial for metaphyseal bone vascularization. In contrast, the angiogenic factors required for vascularization of epiphyseal cartilage are unknown, although this represents a developmentally and clinically important aspect of bone growth. The VEGF gene is alternatively transcribed into VEGF 120, VEGF 164, and VEGF 188 isoforms that differ in matrix association and receptor binding. Their role in bone development was studied in mice expressing single isoforms. Here we report that expression of only VEGF 164 or only VEGF 188 (in VEGF 188/188 mice) was sufficient for metaphyseal development. VEGF 188/188 mice, however, showed dwarfism, disrupted development of growth plates and secondary ossification centers, and knee joint dysplasia. This phenotype was at least partly due to impaired vascularization surrounding the epiphysis, resulting in ectopically increased hypoxia and massive chondrocyte apoptosis in the interior of the epiphyseal cartilage. In addition to the vascular defect, we provide in vitro evidence that the VEGF 188 isoform alone is also insufficient to regulate chondrocyte proliferation and survival responses to hypoxia. Consistent herewith, chondrocytes in or close to the hypoxic zone in VEGF 188/188 mice showed increased proliferation and decreased differentiation. These findings indicate that the insoluble VEGF 188 isoform is insufficient for establishing epiphyseal vascularization and regulating cartilage development during endochondral bone formation.