Bioactive human platelet lysate gel for enhanced proliferation of human umbilical cord tissue derived mesenchymal stem cells.

Mesenchymal Stem cells (MSCs) have a wide range of therapeutic applications due to their self-renewal and multi-lineage differentiation ability; large-scale production of MSCs is possible only with a highly efficient medium, which facilitates increased proliferation of MSCs within a short period. Recently, Human Platelet Lysate (hPL) has emerged as a promising substitute for fetal bovine serum (FBS) for cell expansion. The goal of this study is to optimize a stable gel formulation for the 3D expansion of MSCs using hPL as a matrix material for the improved proliferation of Human Umbilical Cord Tissue derived MSCs (hUCT-MSCs) in comparison to FBS and hPL-supplemented media in 2D culture. To assess the potential benefits of the hPL gel system, in promoting cell proliferation capacity, hUCT-MSCs were cultured on hPL gel coated-dish supplemented with hPL CM, and in FBS CM. Among the varying concentrations, 20% hPL gel was optimized to have more functional stability and shorter gelation time. SEM analysis and gel degradation study at different concentrations revealed the structural integrity and morphology of the gel. Microscopic images and histological staining by H&E were conducted to understand the multi-layered proliferation of hUCT-MSCs in hPL Gel. Flow cytometry analysis reported the expression of positive markers for human umbilical cord MSCs, namely CD 90⁺ and CD 105⁺, in hPL Gel and hPL Complete Medium (CM) similar to that in FBS. The CCK8 Assay carried out for each culture system, generated OD values respective to cell viability and proliferation. OD values of 1.65 nm, 1.27 nm, and 0.92 nm on average were observed for hPL Gel, hPL CM, and FBS control, respectively. Cells in hPL gel showed a 50% higher proliferation rate of viable cells compared to other culture media. AO/EtBr staining with FBS CM, hPL CM, and hPL gel revealed an increase in viable cells and a decrease in early apoptotic and necrotic cells in hPL Gel. In conclusion, the results of this study highlight the potential of hPL-based gels as superior matrices for multi-layered and enhanced proliferation of hUCT-MSCs.