Infantile hemangioma (IH) is the most common childhood benign vascular tumor. Recently, propranolol has been found to be an effective therapy for IH, but its mechanism of action is not yet understood. Hemangioma stem cells (HemSCs) have a mesenchymal morphology, robust proliferation, and multilineage differentiation (into adipocytes). Therefore, we hypothesized that propranolol could accelerate the transdifferentiation of HemSCs and prevent the growth of proliferating IH. In this study, the fibrofatty tissue of IH that received therapy with propranolol appeared much earlier than without the treatment.
We isolated HemSCs with CD133-tagged immunomagnetic beads, and then we used flow cytometry technology to analyze the HemSC phenotypes and determine whether propranolol induced HemSC death. The proliferation and adipogenesis abilities of propranolol-treated HemSCs were analyzed by 3-(4,5)-dimethylthiahiazo(-z-y1)-3,5-diphenytetrazoliumromide assay, Oil red O staining, and Western blotting.
We observed that the HemSC morphological traits gradually became spindle shaped, like fibroblasts, and the average extraction yield of HemSCs was about 0.25%. The HemSCs had high rates of expression for CD90 (98.8%) and CD105 (97.8%) but did not significantly express CD31 (0.7%). We also found a 100 μM concentration of propranolol cutoff point. Propranolol did not affect HemSC survival significantly at low concentrations (6.25, 12.5, 25.0, and 50.0 μM). However, propranolol resulted in a sharp and significant variation in cell morphology and survival rates at high concentrations (100, 200, and 400 μM). The results suggest that treatment with propranolol inhibited HemSC proliferation and induced cell death and apoptosis in a concentration-dependent manner. Oil droplets determined by Oil red O staining showed that propranolol increased the transdifferentiation rate of HemSCs into adipocytes. Furthermore, the expressions of phosphorylated AKT and peroxisome proliferator-activated receptor gamma (PPARγ) were increased with a 100 μM concentration of propranolol in HemSC culture.
Our study found that propranolol inhibited proliferation, induced apoptosis and necrosis, and promoted differentiation of HemSCs. Propranolol may upregulate PPARγ via PI3K pathways, thereby accelerating lipogenesis and enhancing IH HemSC adipogenesis.