PROPOLIS-FUNCTIONALIZED GELATIN HYDROGELS FOR THREE-DIMENSIONAL CULTURE OF WHARTON’S JELLY-DERIVED MESENCHYMAL STEM CELLS

Propolis is a natural product rich in phenolics that has been widely investigated for  its antioxidant, anti-inflammatory, and other biological properties (Martinotti et al., 2025), and its incorporation into biomaterials poses a promising strategy for developing functional systems with potential biomedical applications (Lesmana et al., 2022). Therefore, this study aimed to develop gelatin-based hydrogels functionalized with microcapsules containing Brazilian Organic Propolis (MP) and Red Propolis (RP) and to evaluate their cytocompatibility with Wharton’s jelly-derived mesenchymal stem cells (MSC-WJ). Initially, MP and RP were evaluated for cytotoxicity using the WST-1 assay at different propolis concentrations (25-400 µg/mL). Based on these results, we selected the highest concentration that maintained cell viability above 70% for subsequent experiments. Gelatin-based hydrogels containing propolis were prepared at a final gelatin concentration of 16.7% (w/v). Briefly, porcine gelatin was exposed to UV light for 60 min and then mixed with α-MEM supplemented with 10% FBS and 1% PSA. The mixture was heated at 50 °C for 20 min, after which propolis was added and the formulation was continuously stirred for 15 min at 37 °C. Cell morphology and distribution were investigated by confocal microscopy after 7 days of culture using phalloidin/DAPI staining. In addition, MSC-WJ spheroids were integrated into gelatin-based hydrogels containing MP or RP, and cell migration from the spheroids into the hydrogels was evaluated. The WST-1 assay showed a concentration-dependent response, with lower cell viability at the highest propolis concentrations, particularly at 200 and 400 µg/mL, while at concentrations ranging from 25 to 100 µg/mL, cell viability was improved, with values approaching or exceeding those of the control depending on the propolis type. Confocal microscopy revealed marked differences in cell distribution among the evaluated formulations after 7 days of culture, with a higher cell density observed in the control and RP formulations and a markedly lower number of cells in the MP formulation. Moreover, cells successfully migrated from the spheroids into gelatin-based hydrogels containing both MP and RP, demonstrating the potential of these systems for three-dimensional cell culture. The total phenolic content and antioxidant activity of the propolis-functionalized hydrogels are being evaluated using the Folin-Ciocalteu and DPPH assays, respectively. Overall, the preliminary findings demonstrate that the biological response of MSC-WJ to propolis-functionalized gelatin hydrogels depends on the propolis type and concentration, while the successful migration of spheroid-derived cells suggests their potential as bioactive platforms for three-dimensional cell culture and future biomedical applications.

Acknowledgment: CAPES-COFECUB (Finance code 001).