Background/Purpose: Systemic Sclerosis (SSc) is a multisystem autoimmune connective tissue disorder characterized by vascular injury and fibrosis of the skin and internal organs. Methyl-CpG-binding protein 2 (MeCP2) is a transcription activator or repressor depending on its interacting complexes. Increasing evidence shows that MeCP2 is closely involved in lung, liver, and kidney fibrosis. In this study, we aim to elucidate the role of MeCP2 in dermal fibroblasts from patients with SSc.

Methods: Dermal fibroblasts were isolated from healthy controls or patients with diffuse cutaneous SSc (dcSSc). MeCP2 expression was analyzed by qRT-PCR and western blotting. A scratch wound healing assay was used to evaluate fibroblast migration. MeCP2 overexpression was achieved by transfecting fibroblasts with MeCP2 DNA plasmids, while transfection with empty vectors was used as control. MeCP2 knockdown was done using MeCP2 siRNA. RNA isolated from dcSSc fibroblasts after 48hrs of MeCP2 knockdown was sequenced by Ilumina HiSeq2500. Gene pathway analysis was performed using GeneMANIA and by searching literatures and GeneCards database.

Results: MeCP2 expression was increased by 1.8-fold (p=0.02) in dcSSc fibroblasts (n=6) compared to normal fibroblasts (n=7) at the protein level. However, no change was observed at the mRNA level, suggesting that dysregulation of MeCP2 in dcSSc occurs at the post-transcriptional level. Overexpression of MeCP2 in normal fibroblasts inhibited cell migration (n=4, p=0.02) while MeCP2 knocked down dcSSc fibroblasts showed higher wound repair ability than controls (n=5, p<0.01). In addition, the mRNA levels of pro-fibrotic genes such as α-SMA and COL1A1 were significantly downregulated in MeCP2 overexpressing normal fibroblasts (n=6, p=0.03) while anti-fibrotic PPAR-γ was upregulated (n=6, p<0.01). Using RNA-seq we identified 51 differentially expressed genes after MeCP2 knockdown in dcSSc fibroblasts (n=5, ≥1.2 fold or ≤0.8 fold, adjusted p-value<0.05). 14 out of 51 differentially expressed genes appeared to be involved in fibrosis, among which 5 downregulated genes including ITGB1 and NID2 were shown to be related to COL1A1, and 4 genes, including ITGB1, PLAU, ADA, and TNFPIA1, were involved in cell migration/adhesion. None of the 14 screened genes were differentially expressed between normal and dcSSc fibroblasts, suggesting that MeCP2 might suppress fibrosis via balancing the expression of some of these candidate genes. Moreover, we further confirmed that 9 downregulated fibrosis-related genes identified by RNA-seq were indeed MeCP2-regulated genes, as they were upregulated when MeCP2 was overexpressed in normal fibroblasts.

Conclusion: MeCP2 exerts anti-fibrotic effects in dcSSc fibroblasts by reducing cell migration and myofibroblast differentiation. Whole transcriptome profiling reveals that MeCP2 is involved in cell adhesion and controls extracellular matrix (ECM)-related genes. Importantly, MeCP2 overexpression in dcSSc fibroblasts appears to be a defense mechanism to counteract the pro-fibrotic nature of the disease. MeCP2 might be a novel target to restore normal skin function in SSc.

« Back to 2017 ACR/ARHP Annual Meeting

ACR Meeting Abstracts - https://acrabstracts.org/abstract/elevated-mecp2-expression-in-diffuse-cutaneous-systemic-sclerosis-dermal-fibroblasts-is-associated-with-anti-fibrotic-effects/