Zileuton, a 5-lipoxygenase inhibitor, attenuates mouse microglial cell-mediated rotenone toxicity in PC12 cells / 浙江大学学报·医学版

<p><b>OBJECTIVE</b>To examine the effect of a selective inhibitor of 5-lipoxygenase (5-LOX) zileuton on microglia-mediated rotenone neurotoxicity.</p><p><b>METHODS</b>The supernatant from different concentrations of rotenone-stimulated mouse microglia BV2 cells was used as the conditioned media (CM) for PC12 cells. The viability of PC12 cells was determined by MTT assay and lactate dehydrogenase (LDH) release. Cell death was observed by LDH release and double fluorescence staining with Hoechst/propidiumiodide (PI). The effect of zileuton on microglia-mediated rotenone toxicity was evaluated by the above methods.</p><p><b>RESULTS</b>Rotenone at 1-10 nmol/L was nontoxic to PC12 cells directly. However, the CM from BV2 cells that were treated with rotenone (1-10 nmol/L) resulted in toxicity of PC12 cells. The BV2 CM which stimulated with rotenone (1-10 nmol/L) induced morphological changes, reduced cell viability, and increased LDH release and cell necrosis in PC12 cells. Pretreatment of BV2 cells with the 5-LOX inhibitor zileuton (0.01-1 μmol/L) protected PC12 cells from the microglia-mediated rotenone toxicity.</p><p><b>CONCLUSION</b>The 5-LOX inhibitor zileuton effectively attenuates microglia-mediated rotenone toxicity in PC12 cells. These results suggest that 5-LOX pathway may be involved in neuronal death induced by microglial inflammation.</p>

5-lipoxygenase is involved in rotenone-induced injury in PC12 cells / 浙江大学学报·医学版

<p><b>OBJECTIVE</b>To determine whether 5-lipoxygenase (5-LOX) is involved in rotenone-induced injury in PC12 cells, which is a cell model of Parkinson disease.</p><p><b>METHODS</b>After rotenone treatment for various durations, cell viability was determined by colorimetric MTT reduction assay, and 5-LOX translocation was detected by immunocytochemistry. The effect of 5-LOX inhibitor zileuton was also investigated.</p><p><b>RESULT</b>Rotenone (0.3-30 μmol/L) induced PC12 cell injury, and zileuton (3-100 μmol/L) attenuated this injury. Rotenone also time-and concentration-dependently induced 5-LOX translocation into the nuclear envelope, and zileuton (1-30 μmo/L) significantly inhibited rotenone-induced 5-LOX translocation.</p><p><b>CONCLUSION</b>5-LOX is involved in rotenone-induced injury in PC12 cells, and 5-LOX inhibitor zileuton can reduce rotenone-induced 5-LOX activation and cell injury.</p>

Transfection with 5-lipoxygenase/green fluorescence protein for evaluating injury-induced 5-lipoxygenase translocation to the nuclear membrane in PC12 cells / 浙江大学学报·医学版

<p><b>OBJECTIVE</b>To evaluate the translocation of 5-lipoxygenase (5-LOX)) after injuries by transfection with green fluorescence protein (GFP)/5-LOX in PC12 cells.</p><p><b>METHODS</b>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.</p><p><b>RESULT</b>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.</p><p><b>CONCLUSION</b>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.</p>