Effect of Cryopreservation on Different Passages of Porcine Dorsal Root Ganglion Cell Culture

Abstract

The cell culture obtained from dorsal root ganglia (DRG) is a valuable model used in biology or medical research. However, the effect of cryopreservation on the properties of DRG-derived cell culture of different passages remains unclear up to date. The objective of the study is to assess the effect of cryopreservation with various concentrations of cryoprotectant dimethyl sulfoxide (DMSO) on the viability and morphological features of porcine neonatal DRG cell culture of different passages. Cell suspension was obtained from DRG of neonatal piglets and cultured in an α-MEM nutritional medium supplemented with 10% fetal calf serum (0, 1st, and 2nd passages). Cells of different passages were cryopreserved at a cooling rate of 0.5°C/min to – 20°C (step 1) and 1°C/min to – 80°C (step 2) followed by immersion into liquid nitrogen. Cryoprotective solutions based on α-MEM nutrient medium and 25% fetal calf serum (FCS) containing 5%, 7.5%, and 10% DMSO were used. It was established that the primary culture (passage 0) consisted of four cell morphological types: large rounded cell bodies of sensory neurons (SN) and three types of non-neuronal cells, namely, polygonal cells with pronounced elongated processes (type 1), spindle-shaped cells (type 2), and multipolar flattened fibroblastoid cells (type 3). As the number of passages increases, an elimination of SN from the culture, a decrease in the relative number of 1st and 2nd cell types, and expansion of 3rd cell type were observed. DRG cell culture had sufficiently high resistance to cryopreservation as cell viability was in the range from 83% to 90% using different concentrations of DMSO. The cells of passage 1 were more resistant to cryopreservation in comparison with primary culture cells (passage 0). The best result was achieved by freezing the culture of passage 1 in the cryoprotective medium with 7.5% DMSO, where 90.6% of viable cells were observed after thawing.