New in vivo model to analyse the expression of angiogenic genes in the borders of a cleft lip.

Defects in the fusion of facial buds can result from an anomaly in tissue development or apoptosis, or both. Our working hypothesis was that anomalies in the development of tissues could be caused by a genetic angiogenic defect. Our main objective was to design a reproducible experimental model to study the expression of angiogenic genes in the borders of cleft lips with or without cleft palate. We therefore prospectively studied seven non-syndromic patients, three with a cleft lip (2 right, 1 left), and four with a cleft lip and palate (1 bilateral, 2 right, 1 left), with no CGH (comparative genomic hybridisation) array, who had primary operations to repair their clefts. We also used four controls (cultured fibroblasts from healthy skin samples). The mean (range) age at operation was 44 (13-77) days. We studied the lateral and medial borders histologically and did qPCR (quantitative real-time polymerase chain reaction) analysis for gene expression with 22 genes of interest (and two housekeeping genes) involved in cleft lip and angiogenesis. The qPCR analysis found significant (p<0.05) overexpression of eight genes in the medial border and seven in the lateral border, and underexpression of nine genes in the medial, and ten in the lateral border. The difference in expression between the two borders was not significant. This preliminary study has enabled us to develop a new method to analyse the expression of angiogenic genes in the borders of cleft lips.