Yong Huang1, Yuting Wang1, Lixin Lin1, Peng Wang1, Lei Jiang1, Jian Liu1, Xueming Wang2. 1. Department of Plastic and Aesthetic Surgery, Yantai Yuhuangding Hospital, China. 2. Department of Plastic and Aesthetic Surgery, Yantai Yuhuangding Hospital, China. Electronic address: wangy_yaw@163.com.
Abstract
AIM: Keloid is a benign dermal tumor with excessive hyperplasia and deposition of collagen. As a common tumor suppressor gene, miR-133a-3p has not been studied in keloid. This study will delve into the specific mechanism of miR-133a-3p in keloid. METHODS: Normal skin fibroblasts and keloid fibroblasts (KFs) were first isolated from patients' normal skin and keloid, and cells were identified by morphological observation and immunofluorescence. The expressions of miR-133a-3p and extracellular matrix (ECM)-associated markers (Collagen I, III and α smooth muscle activin) were detected by Quantitative Real Time-Polymerase Chain Reaction (qRT-PCR). Cell viability and apoptosis of KFs were examined by Cell Counting Kit-8 assay, flow cytometry, and Caspase-3 colorimetry. TargetScan predicted target gene for miR-133a-3p was verified by luciferase assay, qRT-PCR and Western Blot (WB). WB was used to study protein expression of TGFBR1, phosphorylated -Smad2 (p-Smad2) and Smad2. Finally, a series of rescue experiments were performed to verify the intervention of target genes on miR-133a-3p. RESULTS: MiR-133a-3p was lowly expressed in keloid tissue and KFs. Overexpression of miR-133a-3p inhibited the expression of ECM-associated markers, reduced KFs viability, and promoted apoptosis. It was verified that interference regulator 5 (IRF5) is miR-133a-3p target gene. The rescue experiments showed that IRF5 reversed the effect of miR-133a-3p mimic on inhibiting fibrosis, and reversed the effects on promoting apoptosis and reducing cell proliferation. CONCLUSION: Overexpressed miR-133a-3p inhibits fibrosis by down-regulating IRF5 and thus inhibiting the TGF-β/Smad2 pathway. And it also promotes KFs apoptosis and reduces proliferation.
AIM: Keloid is a benign dermal tumor with excessive hyperplasia and deposition of collagen. As a common tumor suppressor gene, miR-133a-3p has not been studied in keloid. This study will delve into the specific mechanism of miR-133a-3p in keloid. METHODS: Normal skin fibroblasts and keloid fibroblasts (KFs) were first isolated from patients' normal skin and keloid, and cells were identified by morphological observation and immunofluorescence. The expressions of miR-133a-3p and extracellular matrix (ECM)-associated markers (Collagen I, III and α smooth muscle activin) were detected by Quantitative Real Time-Polymerase Chain Reaction (qRT-PCR). Cell viability and apoptosis of KFs were examined by Cell Counting Kit-8 assay, flow cytometry, and Caspase-3 colorimetry. TargetScan predicted target gene for miR-133a-3p was verified by luciferase assay, qRT-PCR and Western Blot (WB). WB was used to study protein expression of TGFBR1, phosphorylated -Smad2 (p-Smad2) and Smad2. Finally, a series of rescue experiments were performed to verify the intervention of target genes on miR-133a-3p. RESULTS: MiR-133a-3p was lowly expressed in keloid tissue and KFs. Overexpression of miR-133a-3p inhibited the expression of ECM-associated markers, reduced KFs viability, and promoted apoptosis. It was verified that interference regulator 5 (IRF5) is miR-133a-3p target gene. The rescue experiments showed that IRF5 reversed the effect of miR-133a-3p mimic on inhibiting fibrosis, and reversed the effects on promoting apoptosis and reducing cell proliferation. CONCLUSION: Overexpressed miR-133a-3p inhibits fibrosis by down-regulating IRF5 and thus inhibiting the TGF-β/Smad2 pathway. And it also promotes KFs apoptosis and reduces proliferation.