Encapsulation of Human Limbus-derived Stromal/Mesenchymal Stem Cells for Storage and Transportation at Room Temperature

Abstract

Purpose : Cell-based therapy for corneal scarring using human limbus-derived mesenchymal/stromal stem cells (hLMSCs) is a promising alternative to conventional corneal transplantation. However, reliable methods of storing and transporting these cells for prolonged periods of time and over long distances remains challenging. This study aimed to test a novel method of storing and transporting hLSMCs at room temperature by encapsulating them.

Methods : The cell suspension of hLMSCs was mixed in equal volumes with sodium alginate solution, at a final density of 2.5x10⁶/mL. Encapsulated hLSMCs with complete media were kept in transit at room temperature (RT) or 4°C for 3-5 days, considering it to be the likely maximum duration of transporting cells from bench-to-bedside. Cells without encapsulation were also transported at RT as controls. A specialized container pre-conditioned to maintain temperatures of ≤30°C was used for transportation. Post-storage, hLMSCs were released from encapsulation, their viability was assessed, and they were placed in culture. After 48 and 96-hours in culture, hLSMCs were quantified for their proliferation, gene-expression and phenotypic assessment.

Results : Overall 5 vials were transported in 3 batches of 3-5 days duration. The container under transit maintained an average temperature of 18.6±1.8°C, where the average atmospheric temperature was 31.4±1.2°C. Encapsulated hLSMCs under transit at RT were recovered with a high viability of 82.5±0.9% after a 3-day storage and 76.9±1.9% over a 5-day storage as compared to 4°C that showed 65.2±1.2% and 64.5±0.8% respectively (p=0.01). Cells under transit at RT had better proliferation rates of 74.3±2.9% and 67.7±9.8% than cells stored at 4°C (54.8±9.04% and 52.4±8.1%) after 3 and 5 days of storage, respectively (p<0.001). Non-encapsulated cells at RT had no viability after 3-5 days. Cells after transit maintained their characteristic phenotype, showing the expression of CD105⁺, CD45^-, CD73⁺, VIM⁺, COL-III⁺, HLA-DR^-(mesenchymal) and P63-α⁺, ABCG2⁺, PAX-6⁺ (stem cell) markers.

Conclusions : Alginate encapsulation is a promising method of cell storage and transportation, offering high cell viability over prolonged durations, in transit and at room temperatures. This provides the opportunity of expanding the scope of cell-based therapy for corneal blindness to remote centers without cell-cultivation cGMP facilities.

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.