Nanoencapsulation of Human Adipose Mesenchymal Stem Cells: Experimental Factors Role to Successfully Preserve Viability and Functionality of Cells

Cell nanoencapsulation is a novel delivery system based on a self-assembly technique mediated by electrostatic interactions called Layer-by-Layer (LbL) deposition, without an increase in volume implant because of the nanometric thickness of its layers. LbL coats the entire surface of individual cells, providing mechanical resistance to cells against manipulation and storage conditions prior to implantation in the patient. In this work, single-cell nanocapsule formation using human adipose-derived mesenchymal stem cells (ADSC) given their potentiality in regenerative medicine was assessed by fluorescence microscopy and Zeta potential assays. Both methodologies were conclusive in showing layerby-layer nanocapsule formation of every single ADSC. Significant differences in terms of viability and cell functionality preservation were observed depending on the polycation used. Using a combination of fluorescence microscopy and fluorimetric assays, we found that cell survival after nanocapsulation was only efficient when chitosan was added to cells. These results were consistent with other cell types used in this study. Other polycations such as poly(allylamine hydrochloride) (PAH), poly(diallyldimethylammonium chloride) (PDADMAC) and poly-L-lysine (PLL) markedly decreased cell viability (22%, 11% and 15%, respectively). In addition, the use of potassium-enriched saline solutions, such as Hanks and Ringer’s solution, during the nanoencapsulation process on ADSCs was harmful on cell viability compared to standard media (36% vs 79%, respectively). The addition of a mixture of polyanions such as hyaluronic acid and chondroitin sulfate did not affect cell viability (79% and 81%). The combination of chitosan/hyaluronic acid and chondroitin sulfate was also effective in preserving the cell functionality of ADSCs, including the proliferation and differentiation of these cells as assessed by MTT assay and microscopy, respectively. Taken together, these results indicate that ADSCs can be successfully nanoencapsulated using a first layer of chitosan and a second layer of a combination of hyaluronic acid and chondroitin sulfate with a standard potassium concentration in the culture medium.