Hypoxic preconditioning (HP) is a promising approach to enhance the therapeutic efficacy of mesenchymal stem cells (MSCs) by modulating their oxidative stress response, metabolic activity, and secretome composition. Conditioned media (CM) obtained from MSCs cultured under hypoxia contains bioactive molecules and extracellular vesicles (EVs) that support regenerative processes. However, the effects of varying oxygen levels on the redox status and physicochemical characteristics of MSC-derived CM remain incompletely understood. This study aimed to investigate how two physiologically relevant oxygen concentrations (1% and 5%) influence oxidative stress parameters and nanoparticle features in Wharton’s jelly-derived MSC (WJ-MSC)-conditioned media. Cells were cultured under 1% or 5% O₂ and subjected to serum starvation for 48 or 72 h. CM samples were analyzed for total oxidant status (TOS), total antioxidant status (TAS), and oxidative stress index (OSI). Nanoparticle size and zeta potential were evaluated using dynamic light scattering (DLS), and HIF-1α levels were quantified via ELISA. Results showed that CM from 1% O₂ cultures exhibited significantly higher oxidative stress, with elevated TOS and OSI values and reduced TAS levels, particularly after 72 h. Nanoparticle size was initially larger under 1% O₂ but decreased with time, whereas 5% O₂ supported more stable size profiles. Zeta potential measurements revealed more negative values under 5% O₂, indicating greater colloidal stability. HIF-1α expression markedly increased under 1% O₂, confirming hypoxia-induced cellular adaptation. In conclusion, this study demonstrates that graded hypoxia distinctly modulates oxidative stress and nanoparticle characteristics in MSC-derived CM. These findings provide a basis for optimizing hypoxic preconditioning protocols to improve the quality and therapeutic potential of acellular MSC-based therapies.

Human umbilical cord matrix Wharton’s jelly mesenchymal stem cells (WJ-MSCs) are commonly utilized in regenerative medicine due to their therapeutic benefits. However, the microenvironmental stress present in patients with hyperglycemia can significantly reduce mesenchymal stem cell (MSC) viability under high-glucose conditions in the body, ultimately reducing their therapeutic effectiveness. Enhancing the survival rate of MSCs following cell transplantation remains a crucial challenge. This study investigates whether Quantum Hyperlight (QHL) can counteract the detrimental effects of high glucose (HG), thereby improving MSC survival, proliferation, and mitochondrial function. We aimed to evaluate the effect of QHL on cellular viability, proliferation, and mitochondrial activity in WJ-MSCs exposed to HG. MSCs were cultured in a medium containing normal glucose (NG) (1 g/L) and HG (4.5 g/L). MSCs in the HG medium were exposed to QHL for 90 s or 180 s with an energy density of 2.4 Joules/cm ² /minute and an average power density of 40 mW/cm ² . Then, proliferating cell nuclear antigen (PCNA), MTT assays, and Mitotracker Green staining were performed to evaluate cell viability and proliferation. The viability of MSCs was significantly increased in the QHL-treated groups (84% in QHL-90 s and 86% in QHL-180 s) compared to the untreated HG group (65%, p < 0.001). PCNA expression in QHL-90 s and QHL-180 s groups showed significant increases ( p < 0.001) compared to the untreated HG group. MitoTracker staining intensity was significantly higher in the QHL-treated groups compared to the untreated HG group ( p < 0.001). The HG environment reduced viability, proliferation, and mitochondrial staining. In the context of the NG environment, MSCs exhibited notable differences. However, the viability, proliferation, and mitochondrial staining rates of MSCs were significantly higher in the HG conditions when treated with QHL compared to the group that did not receive QHL. This study introduces QHL as a novel approach to enhance the therapeutic potential of WJ-MSCs under HG conditions, demonstrating its ability to improve cellular viability, proliferation, and mitochondrial activity. This study highlights its potential as a pretreatment to improve clinical outcomes in regenerative medicine.