The Study Of The Molecular Mechanisms Of Propranolol Treatment For Infantial Hemangioma

Objective: Infantile hemangioma(IH) is the most common tumor of infancy. Oral propranolol(PRN) has recently been shown to be highly effective for infantile hemangiomas since 2008, and is currently recommended as the first-line treatment of IH. However, the therapeutic mechanisms still remain unclear.To explore the molecular mechanisms of propranolol for infantile hemangioma, we will study the effect of propranolol on the proliferation activity and apoptosis of hemangioma endothelial cells(Hem EC)and search the different genes expression of Hem EC after the treatment of propranolol by c DNA Microarray.Methods: 1.Culturing infantile hemangioma endothelial cells in vitro, and culturing human umbilical vein endothelial cells as a control group, with different concentrations of propranolol to deal two kinds of cells(0 umol/L, 25 umol/L, 50 umol/L, 75 umol/L, 100 umol/L, 125 umol/L), after 24, 48, 72 hours using MTT cell proliferation assay kit to check two kinds of cell proliferation activity. To deal Hem EC in the same way,then detect the cell apoptosis with flow cytometry.2.c DNA Microarray experiment: Hem EC total RNA of Propranolol treated group and control group was extracted by Trizol. Hybridization was performed in Agilent’s Sure Hyb Hybridization Chambers. Slides were scanned with the Agilent DNA microarray Scanner. The differentially expressed genes were screened after data analysis. Choose some gene differences, are verified by Real-time PCR.Results:1. Proliferation of Hem EC was promoted by 25μM propranolol with 24h-48h(P < 0.05);Proliferation of Hem EC was inhibited by 100-150 μM propranolol from 24 hours to 72 hours(P <0.05). The HUVEC proliferation was inhibited by 100-150 μM propranolol from 48 hours to 72 hours(P <0.05). when propranolol concentration≥100umol/L, Hem EC is more sensitive to propranolol when compared with HUVEC(P < 0.05).2. Apoptosis of Hem EC significantly increased with ≥100 umol/L propranolol in a dose-and time-dependent manner.(P < 0.05).3.Compared with control group, 128 genes in up-regulated and 58 genes down-regulated were screened by c DNA Microarray. The PCSK9, FABP3 gene expression significantly raised, respectively 3.22 times and 10.54 times.4.The expressions of PCSK9, FABP3 m RNA were verified by Real-time PCR, reaching 9.88 土 2.19 times and 21.9 土 8.18 times respectively compared with the control group.Conclusion:1. Low concentrations propranolol could promote proliferation of Hem EC; High concentrations Propranolol could inhibit the proliferation of Hem EC and HUVEC in a dose-and time-dependent manner; Hem EC is more sensitive to the propranolol compared with HUVEC.2.Propranolol is able to inhibit the proliferation of Hem EC and ind uce the apoptosis of Hem EC,resulting in inhibition of hemangioma.3. There are obviously differential genes in Hem EC after treated with propranolol by c DNA Microarray. These genes might be related to apoptosis, cell metabolism, cell cycle, growth regulation, signal transduction, steroid synthesis and lipid biosynthesis-related and so on.4. Propranolol inducing the apoptosis of Hem EC may be related to raise the expression of PCSK9 and FABP3 m RNA.