Labeling of olfactory ensheathing glial cells with fluorescent tracers for neurotransplantation.
Development of cell-based treatment strategies for repair of the injured nervous system requires cell tracing techniques to follow the fate of transplanted cells and their interaction with the host tissue. The present study investigates the efficacy of fluorescent cell tracers Fast Blue, PKH26, DiO and CMFDA for long-term labeling of olfactory ensheathing glial cells (OEC) in culture and following transplantation into the rat spinal cord. All tested dyes produced very efficient initial labeling of p75-positive OEC in culture. The number of Fast Blue-positive cells remained largely unchanged during the first 4 weeks but only about 21% of the cells retained tracer 6 weeks after labeling. In contrast, the number of cells labeled with PKH26 and DiO was reduced to 51-55% after 2 weeks in culture and reached 8-12% after 4-6 weeks. CMFDA had completely disappeared from the cells 2 weeks after labeling. AlamarBlue assay showed that among four tested tracers only CMFDA reduced proliferation rate of the OEC. After transplantation into spinal cord, Fast Blue-labeled OEC survived for at least 8 weeks but demonstrated very limited migration from the injection sites. Additional immunostaining with glial and neuronal markers revealed signs of dye leakage from the transplanted cells resulted in weak labeling of microglia and spinal neurons. The results show that Fast Blue is an efficient cell marker for cultured OEC. However, transfer of the dye from the transplanted cells to the host tissue should be considered and correctly interpreted.
Pettersson
Jonas
J
Department of Integrative Medical Biology, Section of Anatomy, Umeå University, SE-901 87 Umeå, Sweden.
Lobov
Sergei
S
Novikova
Liudmila N
LN
D000578:Amidines; D000818:Animals; D016380:Brain Tissue Transplantation / Q000379:methods*; D002452:Cell Count; D018929:Cell Culture Techniques; D002465:Cell Movement; D002478:Cells, Cultured; D005260:Female; D005452:Fluoresceins; D005456:Fluorescent Dyes; D017628:Microglia / Q000502:physiology; D009457:Neuroglia / Q000502:physiology / Q000637:transplantation*; D009474:Neurons / Q000502:physiology; D009830:Olfactory Bulb; D009930:Organic Chemicals; D051381:Rats; D017207:Rats, Sprague-Dawley; D013116:Spinal Cord / Q000502:physiology / Q000601:surgery*; D013997:Time Factors
D000578:Amidines; D005452:Fluoresceins; D005456:Fluorescent Dyes; D009930:Organic Chemicals; C070080:PKH 26; C031455:diamidino compound 253-50; C069306:5-chloromethylfluorescein