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Mitochondrial Dysfunction and Morphological Abnormality in Keloid Fibroblasts

ABSTRACT: Objective: As an aberrant response to cutaneous wound healing, keloid scarring is a challenge to cure, and its clinical management remains unsatisfactory. The purpose of this study was to investigate whether mitochondrial function and morphology were impaired in keloid pathology. Approach: Keloid fibroblasts (KFs) and normal skin fibroblasts (NFs) were isolated and cultured from excised samples. A real-time extracellular flux analyzer was used to monitor the oxygen consumption rate for evaluating the mitochondrial respiratory function. Real-time PCR or Western blot was performed to detect the levels of mRNAs or proteins, respectively, including those associated with the tricarboxylic acid cycle, mitochondrial fission and fusion, and mitochondrial biogenesis. The reactive oxygen species (ROS) level and the mitochondrial mass were measured by flow cytometry. Mitochondrial ultrastructure was observed by transmission electron microscopy. Results: Compared with NFs, KFs showed reduced basal oxidative respiration (p < 0.05), maximal oxidative respiratory capacity, ATP production, and coupling efficiency (p < 0.01) but increased proton leakage (p < 0.01). ROS were decreased (p < 0.05) in KFs, whereas the mitochondrial mass was increased (p < 0.01). The mRNA and protein expressions related to mitochondrial dynamics (fission and fusion) and biogenesis were significantly upregulated in KFs (p < 0.05). Mitochondria in both KFs and keloid tissue were distended and swollen, along with displaying vacuolization and effacement of the cristae. Innovation: The possible involvement of mitochondrial abnormities in keloid pathology may promote promising therapeutic strategies for wound healing disorders. Conclusion: Collectively, our data suggested that both mitochondrial dysfunction and morphological impairment might play vital roles during keloid pathogenesis.

SUBMITTER: Li Q

PROVIDER: S-EPMC7585624 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundA keloid is a disease of excessive fibrosis that is characterized by the aberrant proliferation of fibroblasts. However, the molecular mechanisms of fibroblasts during the development of keloids remain unclear. This study aims to identify new molecular targets that promote the proliferation and migration of keloid fibroblasts, providing new ideas for the prevention and treatment of keloids.MethodsWe utilized bioinformatics tools to analyze data from keloid fibroblasts (KFs) available in the Gene Expression Omnibus (GEO) database to identify the key genes involved in keloid development. Homeobox C6 (HOXC6) emerged as a hub gene in KFs from the GEO database was verified in keloid tissue samples and KFs using reverse transcription-quantitative polymerase chain reaction, western blot (WB) and immunohistochemistry. Subsequently, the effects of downregulated HOXC6 expression on the cellular behaviors of KFs were examined by performing Cell Counting Kit-8, flow cytometry, transwell migration and WB assays. Meanwhile, we performed transcriptome sequencing and gene set enrichment analysis (GSEA) to further explore HOXC6-related mechanisms and validated the signaling pathways by performing a series of experiments.ResultsHOXC6 was the top-ranking hub gene of KFs in microarray datasets from GEO and was upregulated in keloid tissue samples and KFs. Downregulation of HOXC6 inhibited proliferation, migration and extracellular matrix (ECM) accumulation and promoted KF apoptosis. GSEA predicted that the hypoxia signaling pathway was associated with HOXC6 in KFs. Transcriptome sequencing suggested that the extracellular regulated protein kinase (ERK) pathway was one of the downstream pathways of HOXC6 in KFs. Our experiments confirmed that hypoxia-inducible factor-1α (HIF-1α) upregulates HOXC6, contributing to KFs proliferation, migration, apoptosis inhibition and collagen accumulation through the ERK signaling pathway.ConclusionsOur findings first revealed that HOXC6 acts as an oncogenic driver in the molecular mechanisms of fibroblasts in keloids. The HIF-1α/HOXC6/ERK axis promotes proliferation, migration and ECM production by KFs, contributing to the progression of keloids. Taken together, HOXC6 may serve as a promising novel therapeutic target and new focus for research designed to understand the pathogenesis of keloids.

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Mitochondrial Dysfunction and Morphological Abnormality in Keloid Fibroblasts

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