In vitro characterization of Human Olfactory Ensheathing Cells

Introduction and Aim: With the promising results of transplanted OECs in regeneration of various animal models of injured spinal cord, OECs have emerged as prime candidates for cell-mediated treatment of Spinal Cord Injury (SCI). This study aimed to isolate human Olfactory Ensheathing Cells (hOECs) by enzymatic dissociation of olfactory mucosal tissue and characterize their glial cell properties. Materials and Methods: Primary cultures of hOECs are grown on poly-L-lysine coated coverslips in multi wells supplemented with DMEM/F10, antibiotics and growth factors at 370C and 5% CO2. On the 14 day in vitro the second passage of the primary cultures of hOECs are fixed in 4% paraformaldehyde and labelled with primary antibodies for GFAP (Glial Fibrillary Acidic Protein), S-100 and p75NTR (p75 neurotrophin receptor) and with Alexa 488 secondary antibodies. The coverslips are mounted on glass slides with DAPI mounting medium visualized under fluorescence microscope. Results: Intense diffused immunofluorescence is observed with S-100, while it is punctate with P75NTR. GFAP displayed faint immunofluorescence. Intense diffuse staining with S-100 mark the Schawn cells like glial cells while punctate staining of the p75 neurotrophin receptor or p75NTR might indicate its key role cell modulation of extracellular matrix regulation. Positive immunofluorescence with GFAP, a cytoskeletal protein may suggest its role to promote cellular plasticity, in this study towards Schwann-like cellular plasticity needed for nerve regeneration. Conclusion: In an attempt to establish peripheral nerve regeneration in Phase-I clinical trials on spinal cord injured patients we established the primary cell cultures of hOECs to study their functional properties in vitro and to derive optimal dose for transplantation.