Proficiency of Carboxymethyl cellulose as a Cryoprotectant. Clinical and Histological Evaluation of Cryopreserved Heterogenous Mesenchymal Stem Cell-Exosomal Hydrogel on Critical Size Skin Wounds in Dogs

Abstract

Background: Fresh stem cell exosomes are usually obtained and reused in the same individual. It cannot be kept viable for a long period of time regardless of the lengthy preparation time. Freezing is typically used to preserve the viability of perishable materials and increase their lifetime. Regrettably, normal freezing of biomaterials leads to cell damage. Therefore, a cryoprotectant can save the cells from the conventional cryodamage. Sodium carboxymethylcellulose (NA-CMC) is a powdery substance that is used to manufacture bio-safe hydrofilm gels because of its high viscosity, cytocompatibility, and non-allergenic nature. Materials and Methods: Sterile CMC hydrogel was prepared, part of which was loaded with exosomal solution derived from MSCs. The gel was kept at −20°C for preservation. Two bilateral full-thickness circular skin wounds of 2-cm diameter were created on the back of experimental dogs. The wounds were at least 2.5 cm apart. Treatment started 24 hours after wound creation. Group I received CMC gel solely, whereas group II received frozen CMC exosomal gel. The gel was applied 4 times, a single application per day with 1-day interval. Results: Clinically, the frozen exosomal gel significantly promoted wound healing with no scaring. Histologically, enhanced dermal fibroblasts and organized collagen deposition were seen in the treated group. Conclusion: CMC proved to be an efficient cryoprotectant and a suitable vehicle for exosomes. Deep freezing was proven to conserve the viability, extended the preservation, and facilitated the usage of exosomal gel. This technique of preserved cell-free therapy is inexpensive, time-saving, and proficient and seems suitable for treating cutaneous wounds.

Keywords

• Critical-size defect; Skin wound; Heterogeneous; Frozen; Exosome; Carboxymethyl cellulose