Isolation and characterization of endothelial cells from intramuscular hemangioma.

BACKGROUND: Intramuscular hemangiomas (IMHs) are benign vascular tumors of deep soft tissue characterized by endothelial cell (EC) proliferation. The purpose of this study was to isolate ECs from IMH, characterize their angiogenic phenotype and functional characteristics, and search for a possible signaling pathway related to IMH development.

METHODS: EC Isolation from IMH was performed by digestion, filtration, washing, incubation, and purification in sequence. Tie2 expression was compared between ECs from IMH and controls using reverse transcriptase polymerase chain reaction (RT-PCR). Cell invasion and proliferation assays were used to analyze functional responses of ECs to angiopoietin 1 (Ang1) and vascular endothelial growth factor (VEGF). Expression of downstream targets was analyzed using Western blot analysis.

RESULTS: Isolated ECs showed typical cobblestone appearance under light microscopy and formed capillary-like tubular structures using Matrigel tube-forming assay. RT-PCR of isolated ECs from six patients showed increased expression of Tie2 and VEGF receptor 1 (VEGFR1) compared with control ECs. Tie2 activation by Ang1 compared with VEGFR1 by VEGF resulted in increased EC migration and proliferation. Western blot analysis showed increased Tie2 expression in hemangioma samples compared with normal ECs. Phosphorylated Akt and phosphorylated forkhead box O1 (FOXO1) expression was observed in hemangioma samples only.

CONCLUSION: EC isolation from IMH could be a useful tool for further research. These results suggest that increased Tie2 expression, via Akt-FOXO1 pathway activation, may play an important role in IMH pathogenesis.