Oxygen glucose deprivation model of cerebral stroke in PC-12 cells: glucose as a limiting factor.
Optimum time points for oxygen-glucose deprivation (OGD) and re-oxygenation have been identified to suggest the suitability of PC-12 cells as rapid and sensitive in vitro model of cerebral stroke. Further, the precise role of glucose as one of the limiting factors was ascertained. PC-12 cells were subjected to receive OGD of 1-8 h followed by re-oxygenation for 6 to 96 h in medium having glucose 0-10 mg/ml. Loss of cell viability was assessed using trypan blue dye exclusion and MTT assays. The significant (p < 0.05) reduction in percent viable cell count was started at 2 h of OGD (80.7 +/- 2.0) and continued in further OGD periods (3, 4, 5, 6, 7, and 8 h), i.e. 65.7 +/- 3.5, 59.7 +/- 4.6, 54.3 +/- 3.2, 44.7 +/- 2.9, 20.3 +/- 4.3, 5.7 +/- 2.0 of counted cells, respectively. Cells growing in glucose-free medium have shown a gradual (p < 0.001) decrease in cell viability throughout the re-oxygenation. Re-oxygenation of 24 h was found to be first statistically significant time point for all the glucose concentrations. Glucose concentration during re-oxygenation was found to be one of the key factors involved in the growth and proliferation in PC-12 cells. The OGD of 6 h followed by a re-oxygenation period of 24 h with 4-6 mg/ml glucose concentration could be recorded as optimum conditions under our experimental conditions.
Singh
G
G
Industrial Toxicology Research Centre, Lucknow, India.
Siddiqui
M A
MA
Khanna
V K
VK
Kashyap
M P
MP
Yadav
S
S
Gupta
Y K
YK
Pant
K K
KK
Pant
A B
AB
D000818:Animals; D002470:Cell Survival; D004195:Disease Models, Animal; D005947:Glucose / Q000378:metabolism*; D006801:Humans; D020925:Hypoxia-Ischemia, Brain / Q000378:metabolism* / Q000503:physiopathology; D010100:Oxygen / Q000378:metabolism*; D016716:PC12 Cells / Q000378:metabolism*; D051381:Rats; D020521:Stroke / Q000378:metabolism* / Q000503:physiopathology; D013997:Time Factors
D005947:Glucose; D010100:Oxygen