Modulating aging-associated phenotypes in keratinocytes using antisense oligonucleotides targeting glycogenes.

Age-related keratinocyte dysfunction has been associated with altered glycosylation and dysregulated expression of several glycogenes, including St6gal1, St3gal2, and Man1a. Modulation of these aging-associated glycogenes may therefore represent a potential approach for investigating cellular aging phenotypes. In this study, an in vitro aging-like keratinocyte model exhibiting increased expression of aging-associated glycogenes was established. Antisense oligonucleotides (ASOs) targeting the upregulated glycogenes were designed and evaluated using different oligonucleotide chemistries and delivery systems. Target gene expression, lectin-based glycan profiles, Lgr6 expression, and wound closure were assessed following ASO treatment. Among the ASOs tested, phosphorodiamidate morpholino (PMO)-linked Man1a ASOs demonstrated greater suppression of Man1a mRNA than ASOs targeting St6gal1 or St3gal2. Delivery of PMO-Man1a ASOs using milk-derived nanovesicles (NV) was associated with the greatest reduction in Man1a mRNA levels compared with commercial transfection reagents. Although no significant changes were detected in ConA-reactive mannose-rich glycans, treatment was associated with an increased proportion of Lgr6+ cells and accelerated wound closure in aging-like keratinocytes. Overall, this study provides proof-of-concept evidence that NV-mediated delivery of glycogene-targeting ASOs can modulate selected aging-associated phenotypes in keratinocytes.