Porcine immortalised adipose-derived stem cells

Cell Source and Immortalisation

Following enzymatic digestion of fresh porcine abdominal adipose tissue, cells were cultured on treated cell culture plastic for 2 passages to isolate and expand the ADSC population. ADSCs were immortalised by incubation with Lentivirus containing the SV40 large T antigen for 24 hours. Immortalised cells were selected by treatment of cultures with neomycin for 7 days. Lentiviral vectors are integrative so the expression of the SV40 large T antigen is constitutive. The SV40 large T antigen suppresses the expression of the cell cycle regulator P53 locking cells in the cell cycle. piADSCs test positive for the expression of the SV40 large T antigen and negative for residual Lentivirus. Following immortalisation piADSCs retain their capacity for adipogenic differentiation.

Cell Characterisation

DB_002 cells demonstrate rapid proliferation under standard culture conditions, with growth curve analysis confirming high expansion potential over a 4-day culture period (Figure 1).

Adipogenic Differentiation Characterisation

piADSCs retain adipogenic lineage potential following immortalisation. Differentiation cultures accumulate intracellular lipid droplets following adipogenic induction, visualised by Oil Red O staining (Figure 2).

Microcarrier Suspension Adaptation

piADSCs readily attach to and expand on microcarriers while maintaining high viability under dynamic suspension conditions, supporting scale-up and bioprocess development applications (Figure 3)

‍

‍

‍