In vitro injury model for oligodendrocytes: development, injury, and recovery.
In this study we investigated the effects of severe hypothermia (cryoinjury) on oligodendrocyte (OL) cell marker expression and morphological features. We used a chemically defined cell culture medium, glial development medium (GDM), which favored the optimal expression of the OL phenotype in CG4 cells. Experiments using CG4 cells cultured in 2% serum or in GDM were conducted in parallel. After severe hypothermia, cells were reanimated at 37 degrees C and 4.5% CO(2) and cultured in either GDM or in medium supplemented with 2% serum. In either medium, around 70% of the total number of cells detached within 2 to 4 hours following reanimation. Oligodendroglial markers such as A2B5, O4, Tf, ferritin, tubulin, and MBP were examined by double and triple immunofluorescence. All of these markers except MBP re-appeared at different times during the recovery period for up to 48 hours. Glial fibrillary acidic protein (GFAP) and heat shock protein 60 (HSP-60) were used as injury markers. The presence of serum induced HSP-60 expression, while GDM did not. All CG4 cells expressed HSP-60 in response to hypothermia independently of the cell culture medium used. Cryoinjury induced a spectrum of morphological changes in CG4 cells. The expression of OL specific markers was also influenced by hypothermia. Moreover both, serum and cryoinjury induced the expression of HSP-60 that colocalized with OL and myelin markers. The expression of GFAP by injured cells but not by normal cells corroborated the state of injury of CG4 cells.
Mental Retardation Research Center, Neuropsychiatric Institute, Department of Neurobiology UCLA School of Medicine, 760 Westwood Plaza, Room 78-177 NPI, Los Angeles, CA 90095-1759, USA. aespinosa@mednet.ucla.edu
A E
AE
De Los Monteros
P M
PM
Zhao
J
J
De Vellis
D000818:Animals; D015415:Biomarkers / Q000032:analysis; D002478:Cells, Cultured; D018834:Chaperonin 60 / Q000378:metabolism; D003470:Culture Media; D004195:Disease Models, Animal; D005615:Freezing; D005904:Glial Fibrillary Acidic Protein / Q000378:metabolism; D007035:Hypothermia / Q000503:physiopathology*; D009836:Oligodendroglia / Q000502:physiology*
D015415:Biomarkers; D018834:Chaperonin 60; D003470:Culture Media; D005904:Glial Fibrillary Acidic Protein