DNA methylation of the CDC2L1 gene promoter region decreases the expression of the CDK11p58 protein and reduces apoptosis in keloid fibroblasts

Abstract

The excessive growth of fibroblasts in keloid is closely related to the status of gene methylation. The aim of this project was to study whether keloid development is related to DNA methylation in the CDC2L1 gene promoter region. DNA methylation of the promoter of this gene was analyzed by bisulfite sequencing and verified by DNA methylation-specific polymerase chain reaction. The results showed that the DNA methylation rate of CpG islands in the CDC2L1 gene promoter region was 50.0% (12/24) in patient keloid tissues and 0% (0/24) in normal skin-tissues from healthy controls. Patient keloid tissues with (n = 12) DNA methylation of the CDC2L1 gene promoter showed higher growth rates than those without (n = 12). Samples from keloid tissues with DNA methylation of the CDC2L1 gene promoter region had dramatically lower levels of CDK11p58 protein than samples from keloid tissues without DNA methylation of the CDC2L1 gene promoter region or healthy normal skin-tissues. In the fibroblasts with DNA methylation of the CDC2L1 gene promoter region from keloid tissues treated with DNA methyl-transferase inhibitor 5 aza 2′-deoxycytidine (5-aza-dC) for 48 h, CDK11p58 levels in the fibroblasts were significantly increased in a dose-dependent manner; the apoptotic rate of the fibroblasts was significantly higher in the treated group than in the non-treated group. This study revealed that DNA methylation exists in the CDC2L1 gene promoter region in keloid tissue fibroblasts. DNA methylation of the CDC2L1 gene promoter region dramatically inhibits the expression of CDK11p58 protein in keloid tissues. A specific demethylation drug, 5-aza-dC, suppressed DNA methylation of the promoter region, which increased the expression of CDK11p58. The elevated expression of CDK11p58 resulted in increased fibroblast apoptosis, thus restraining the development of keloids.