Selective Clock-Gene Responses to Photoperiodic and Glucocorticoid Exposure in Human Umbilical Cord-Derived Mesenchymal Stem Cells

Abstract

Objective: Circadian rhythms regulate essential cellular functions, yet their presence and entrainment in human umbilical cord-derived mesenchymal stem cells (hUC-MSCs) remain poorly defined.Methods: hUC-MSCs were cultured under 12-hour light/12-hour dark (light-dark [LD]) cycles or constant darkness (DD) for four days. Cell proliferation, senescence, and gene expression were assessed by Alamar Blue assay, senescence-associated β-galactosidase staining, fluorescence microscopy, and reverse transcription-quantitative polymerase chain reaction. Circadian rhythmicity was analyzed using Cosinor software. Dexamethasone (Dex; 100–500 nM, 2 hr) was applied to evaluate glucocorticoid-induced entrainment.Results: LD cycles were associated with increased cell proliferation and BMAL1 protein levels compared with DD conditions. Only PER3 showed evidence of rhythmic expression under LD exposure (Cosinor p<0.05), whereas other core clock genes did not show evidence of rhythmicity. Light exposure was not associated with a detectable increase in cellular senescence or changes in the expression of stemness markers (OCT4, SOX2, NANOG, C-MYC). Short-term Dex treatment (100–500 nM, 2 hr) did not markedly impair cell proliferation or migration. Notably, treatment with 500 nM Dex was associated with rhythmic BMAL1 mRNA expression over the 24-hour observation period, whereas CLOCK expression did not show evidence of rhythmicity.Conclusion: hUC-MSCs show partial and selective clock-gene responses to photoperiodic and glucocorticoid exposure. Moderate LD exposure was associated with increased proliferation and BMAL1 expression, while Dex was associated with rhythmic BMAL1 expression and was relatively well tolerated in hUC-MSCs under the conditions tested. These findings provide preliminary insight into how circadian rhythmicity may influence mesenchymal stem cells culture conditions.