The present study for the first time in the isolated and sporadic VSD patients found that (a) there are five significant variations within the CITED2 gene promoter region; (b) four of these five variations (4778G, 4255C, 4078C or 4933A) caused cellular functional changes with a significant decrease in the CITED2 promoter activity by influencing TFBSs; and (c) the functional changes by these variations may affect a group of downstream genes and pathways and eventually cause VSD. Here, CITED2 is linked to ventricular septal defect.