Subcellular Fractionation and Metaproteogenomic Identification and Validation of Key Differentially Expressed Molecular Targets for Keloid Disease.

Keloid disease (KD) is a common connective tissue disorder of unknown aetiopathogenesis with ill-defined treatment. Keloid scars present as exophytic fibroproliferative reticular lesions post-cutaneous injury, remain benign yet behave locally aggressive and expansive. To date, there is limited understanding, and validation of biomarkers identified through combined proteomic and genomic evaluation of KD. Therefore, the aim here was to identify putative-causative candidates in KD, by performing a comprehensive proteomics analysis of subcellular fractions as well as the whole cell, coupled with transcriptomics data analysis of normal compared with KD fibroblasts. We then applied novel integrative bioinformatics analyses to demonstrate that NF-kappa-Beta-p65 (RELA) from the cytosolic fraction and Calpain-2 (CAPN2) from the whole cell lysate were significantly up-regulated in KD and associated with alterations in relevant key signalling pathways including apoptosis. Our findings were further confirmed by showing upregulation of both RELA and CAPN2 in KD using flow cytometry and immunohistochemistry. Moreover, functional evaluation using real-time cell analysis and flow cytometry, demonstrated that both omeprazole and dexamethasone inhibited the growth of KD fibroblasts by enhancing the rate of apoptosis. In conclusion, to our knowledge previously unreported, subcellular fractionation and metaproteogenomic analyses have identified two novel biomarkers of relevance to keloid diagnostics and therapeutics.

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