Hepatocyte conditioned medium sustains long term culture of CD34+ derived adult human hemangioblasts and induces their differentiation to endothelial cells

Liver neo-angiogenesis plays a fundamental role in physiological and pathological processes such as regeneration, cirrhosis, autoimmune hepatitis, and alcoholic liver disease. How liver parenchymal cells influence angiogenesis is largely unknown. We studied the influence of soluble factors released by hepatocytes on hematopoietic and endothelial cell differentiation. Human CD34+ cells cultured for several weeks in a hepatocyte-conditioned medium gradually decrease the expression of CD34 and CD133 markers (i.e. after 4 weeks from 85% and 69%, respectively, to 6% and 3%, respectively), whereas expression of CD144 and CD14 cell markers increased (from 2% and 8%, respectively, to 54% and 55%, respectively). The cells' capacity to form hematopoietic colonies in methylcellulose declined with time, whereas they acquired endothelial morphology, expressed endothelial markers, and incorporated into newly forming vascular structures both in vitro and in vivo. Cultured single CD34+ cells formed colonies expressing both hematopoietic (CD45+) and endothelial (CD44+) markers, suggesting they constitute a bona fide hemangioblast population. Conclusion: This system allowed subsequent stages of differentiation of hematopoietic cells to endothelial cells to be defined, underlining the strict interrelationship between endothelial and hematopoietic cells in a hepatocyte environment.