A switch in Notch gene expression parallels stem cell to endothelial transition in infantile hemangioma

Abstract

Background Infantile hemangioma (IH) is the most common benign tumor of infancy, yet its pathogenesis is poorly understood. Notch family members are known to play a role in vascular development during embryogenesis and postnatal tumor angiogenesis, yet the role of Notch signaling in the pathogenesis of IH has not been investigated. This study aims to survey Notch expression in IH. Materials & Methods RNA from resected hemangioma tissue and hemangioma-derived stem cells (HemSCs) and endothelial cells (HemECs) were used for gene expression analyses by real-time PCR. Results were confirmed with immunofluorescence (IF) for protein expression in tissue. Results Real-time PCR showed that Notch family gene expression in IH is distinct from placenta and skin. Notch3 is expressed in HemSCs, but not in HemECs, indicating Notch3 is downregulated as HemSCs differentiate into HemECs. Moreover, expression of endothelial-associated Notch proteins, Notch1, -4, and Jagged-1 are increased in involuting hemangiomas and HemECs, suggesting that as hemangioma progresses towards involution, it acquires more differentiated endothelium. A subset of cells stained double positive for Notch3 and CD31, pointing to a potential intermediate between the HemSC cellular differentiation into HemEC. Conclusion HemSCs have distinct Notch expression patterns from differentiated HemECs and from normal human endothelial cells. Notch3 is expressed in HemSCs while Notch1, Notch4, and Jagged-1 have higher expression levels in HemECs. Notch3 was localized to the interstitial cells outside of the nascent vascular channels in proliferating IH tissue sections, but became more apparent in the perivascular cells in involuting IH. In summary, the pattern of Notch gene expression mirrors the progression from immature cells to endothelial-lined vascular channels (i.e., endothelial differentiation) that characterizes the growth and involution of IH.

Wnt3a Expression is Associated with MMP-9 Expression in Primary Tumor and Metastatic Site in Recurrent or Stage IV Colorectal Cancer

“…There is evidence that the blockade of VEGF in tumors results in a rapid decrease of Dll4 expression in tumor ECs [64]. Interestingly, although HemECs had higher VEGF-A levels and increased activation of VEGFR-2 compared with normal ECs [40], the Dll4 levels in HemECs were lower than those found in normal ECs [44]. These data argue against the idea that VEGF interacts with Notch signaling in IH.…”

“…These characteristics likely result from the genetic instability of HemECs as somatic missense mutations in the kinase insert of the VEGFR-2 gene have been found in IHs [41]. In addition, imbalances in gene expression have been reported in mesenchymal compartments compared to normal tissues, suggesting a possible reciprocal interaction between HemECs and the surrounding cells [42][43][44]. Alternatively, HemECs may originate from progenitor/stem-like cells, which are known to display robust proliferative and clonogenic capabilities and to express high levels of VEGF-A [15,21].…”

NOTCH3 regulates stem-to–mural cell differentiation in infantile hemangioma

“…HemSCs are pluripotent and form GLUT1 + endothelial cells (ECs) in a xenograft mouse model of IH (3). Significant research has focused on the differentiation of HemSCs into hemangioma endothelial cells (HemECs) (8,9). There is evidence that HemSCs can also differentiate into α smooth muscle actin + (αSMA + ) mural cells in culture and in a xenograft mouse model…”

Infantile Hemangiomas: A Disease Model in the Study of Vascular Development, Aberrant Vasculogenesis and Angiogenesis

“…Moreover, the Notch expression profile of proliferating hemangiomas is different than that of hemangiomas undergoing involution, with involuting hemangiomas demonstrating increased expression of Notch4 and Jagged-1 when compared with proliferating hemangiomas. HemSCs and HemECs also demonstrate completely different Notch expression profiles, with HemSCs expressing higher levels of Notch2 and Notch3, but lower levels of Notch1, Notch4, Jagged-1, and Dll-4 compared to HemECs (Wu et al, 2010). These data suggest a possible role for Notch signaling in the maintanence of HemSC pluripotency and differentiation into HemEC phenotype.…”