[Transfection with 5-lipoxygenase/green fluorescence protein for evaluating injury-induced 5-lipoxygenase translocation to the nuclear membrane in PC12 cells].
To evaluate the translocation of 5-lipoxygenase (5-LOX)) after injuries by transfection with green fluorescence protein (GFP)/5-LOX in PC12 cells.



PC12 cells were stably transfected with pEGFP-C2/5-LOX (GFP/5-LOX) or pEGFP-C2 vectors (control). After treatment with oxygen-glucose deprivation (OGD), H(2)O(2) or NMDA, GFP/5-LOX localization in the cells was observed under a fluorescence microscope. Wild-type 5-LOX was determined by immunostaining after the treatment.



In the GFP/5-LOX-transfected cells, GFP/5-LOX was primarily localized in the nucleus; while in the GFP-transfected cells, GFP was localized in both the cytoplasm and nucleus. After OGD and H(2)O(2) treatments, GFP/5-LOX was translocated to the nuclear membrane in 50.6 % and 57.7% cells respectively. However, after NMDA treatment or in GFP-transfected cells, no translocation was observed. Wild-type 5-LOX was distributed in the nuclei and cytoplasm, and all the 3 treatments induced 5-LOX translocation to the nuclear membrane.



In the PC12 cells stably transfected with GFP/5-LOX, GFP/5-LOX is primarily distributed in the nuclei; the OGD-, H(2)O(2)- and NMDA-induced 5-LOX translocation exhibits different properties.
Yuan
Yu-Mei
YM
Department of Pharmacology, College of Medicine, Zhejiang University, Hangzhou 310058, China.
Li
Cheng-Tan
CT
Ma
Zhen-Qiu
ZQ
Peng
Fang
F
Zhang
Wei-Ping
WP
Shen
Yue-Liang
YL
Wei
Er-Qing
EQ
D000818:Animals; D001094:Arachidonate 5-Lipoxygenase / Q000235:genetics / Q000378:metabolism*; D002467:Cell Nucleus / Q000378:metabolism; D005947:Glucose / Q000494:pharmacology; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism*; D006861:Hydrogen Peroxide / Q000494:pharmacology; D008856:Microscopy, Fluorescence; D016202:N-Methylaspartate / Q000494:pharmacology; D009685:Nuclear Envelope / Q000378:metabolism*; D016716:PC12 Cells; D021381:Protein Transport / Q000187:drug effects; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014162:Transfection
D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D016202:N-Methylaspartate; D006861:Hydrogen Peroxide; D001094:Arachidonate 5-Lipoxygenase; D005947:Glucose