Reduced blood vessel formation and tumor growth in alpha 5-integrin-negative teratocarcinomas and embryoid bodies

Embryonic stem (ES) cells-wild-type, heterozygous, or null for alpha5-integ rin-were injected ectopically into syngeneic mice to develop teratocarcinom as. alpha5-null-derived teratocarcinomas were significantly smaller than th e wild-type or alpha5 heterozygous tumors. Histological analysis revealed t he presence of tissues derived from all three germ layers, in all tumors. H owever, alpha5-null teratocarcinomas displayed less undifferentiated tissue than did the controls, Decreased proliferation and increased apoptosis wer e observed in the undifferentiated areas of the alpha5-null teratocarcinoma s, The expression of extracellular matrix proteins, fibronectin and tenasci n-C, and the basement membrane components, laminin, entactin/nidogen, and c ollagen IV, was similar in the different tumors, although the deposition of these molecules was more disorganized in alpha5-null teratocarcinomas, The absence of alpha5-integrin in the various tissues of the alpha5-null tumor s was confirmed by immunohistochemistry. Many vessels, but not all, stained positively for alpha5-integrin, showing that they were host derived. Analy sis of the area occupied by vessels revealed, on average, an 8-fold decreas e in alpha5-null teratocarcinomas compared with control tumors. Staining fo r smooth muscle ol-actin showed that pericytes and smooth muscle cells were recruited around the vessels in all tumors, suggesting similar vessel diff erentiation. Deposition of EIIIA and EIIIB and fibronectin around the vesse ls was observed in all tumors. The fact that some, although few, alpha5-int egrin-negative vessels existed in alpha5-null tumors indicated that alpha5- /- ES cells could differentiate into endothelial cells. Endothelial cell di fferentiation and vessel formation were analyzed also in vitro. alpha5-null ES cells were differentiated into embryoid bodies, although they were dela yed in growth and attachment. Differentiation into endothelial cells was ac hieved, but the organization into a complex vasculature was delayed compare d with controls. We conclude that alpha5 beta1-integrin plays a significant role in vessel formation both in ES cell cultures and in teratocarcinomas. Reduced vascularization likely contributed to the reduced proliferation an d increased apoptosis observed in alpha5-null teratocarcinomas.