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.

Purpose: Wharton's jelly-derived mesenchymal stem cells (WJ-MSCs) are self-regenerative and able to differentiate into multipotent stem cells. There may be different sources of mesenchymal stem cells (MSCs) involved in the repair mechanism of damaged tissues in the organism. WJ-MSCs may differentiate into osteocytes, chondrocytes, adipocytes, and myocyte cells. Furthermore, MSCs show neuroprotective effects on neurons. Today, many MSC neuroregenerative treatments have been shown to be effective. Studies have shown that MSCs are more involved in paracrine effects due to their neuroprotective effect in multiple diseases such as multiple sclerosis, acute spinal cord injury and encephalomyelitis. The main aim of this study was to investigate the neuronal markers of stem cells after incubation with β-mercaptoethanol (BME). Materials and methods: In our study, WJ-MSCs were thawed in a water bath at 37°C and cultivated in cell culture dishes. When the cell occupancy rate reached 60-70%, they were treated with 2 mM BME. At the first and third hours, MSCs were removed from the dishes, and flow cytometry and immunostaining revealed that BME's nestin, neuron filament light (NF-L), SOX1, SOX2, doublecortin (DCX), glial fibrillary acidic protein (GFAP), Ki67, and CD44 were evaluated. Results: Immunocytochemically, nestin and NF-L values ​​of MSCs exposed to BME increased at the first hour. In the flow cytometric evaluation, it was observed that nestin was high in the first hour. Conclusion: One of our aims in this study was to reduce the possible toxic side effects of BME for MSCs by exposing the BME used in previous studies at the minimum dose for neuronal differentiation. In our study, we showed first-hour changes similar to the neuronal differentiation obtained with pre- and post-induction in other studies.

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.

Shaping the ear cartilage and preserving the shape are important and quite difficult. The aim of this study was to demonstrate the effectiveness of the Wharton's jelly–derived stem cell–assisted electromechanical reshaping method in a rabbit ear cartilage defect model and to compare it with surgical reshaping. For the purpose of 25 × 4-mm cartilage defect reconstruction, 48 rabbit ears were divided into 2 main groups according to the shaping method, and these main groups were divided into 3 subgroups according to stem cell injection: control, sham, and stem cell. A rabbit ear cartilage defect was created, and rib cartilage was collected for reconstruction. Although electromechanical reshaping was performed in accordance with the rabbit ear geometry angle, surgical scoring and suturing were performed in the classical method. Stem cells were applied in the first week, and the grafts were removed in the first month. Analyses included angular change, weight change, and histopathology. In this study, electromechanical reshaping was histopathologically similar to surgical reshaping and is more effective in preserving the shape. Cartilage thickness and weight were higher in stem cell groups. Electromechanical reshaping is emerging as an effective and standardized method to maintain cartilage stability and geometry and offers a viable alternative to classic surgical techniques. In addition, stem cell application gave physical strength to cartilage. It is a method that allows us to obtain more stable and more durable cartilages that maintain given shape with the combination of Wharton jelly–assisted electromechanical reshaping method.

Background: Critical-sized peripheral nerve defects remain a major reconstructive challenge. Although autografts are considered the gold standard, their use is limited by donor-site morbidity and graft availability. Acellular nerve allografts (ANAs) offer a promising alternative but are often hampered by inadequate revascularization and limited functional outcomes in large defects. This study investigates whether combining mesenchymal stem cell-derived secretome therapy with surgical angiogenesis can enhance the regenerative capacity of ANAs. Methods: Forty male Wistar rats were randomly assigned to 5 groups (n=8): group 1 (reversed autograft), group 2 (ANA), group 3 (ANA+secretome), group 4 (ANA+surgical angiogenesis), and group 5 (ANA+secretome+surgical angiogenesis). A 10 mm sciatic nerve defect was created in all animals. Functional recovery was evaluated using the Sciatic Functional Index (SFI) at 6 and 9 weeks. Electrophysiological, histologic, immunohistochemical (CD34), and muscle weight/volume assessments were performed at 9 weeks. Results: Group 5 demonstrated the highest functional recovery among ANA-based interventions, achieving outcomes comparable to autografts. Groups 4 and 5 showed significantly improved axonal regeneration compared with ANA alone. CD34 immunostaining revealed enhanced vascularization in groups 4 and 5, with levels similar to the autograft group. No significant differences were observed in electrophysiological or muscle mass parameters. Conclusions: The combination of secretome therapy and surgical angiogenesis synergistically enhances the regenerative performance of acellular nerve allografts. This strategy may provide a clinically translatable alternative to autografts, particularly in challenging repair scenarios involving avascular or scarred recipient beds.