The Effects of Alpha-Linolenic Acid on the Secretory Activity of Astrocytes and β Amyloid-Associated Neurodegeneration in Differentiated SH-SY5Y Cells: Alpha-Linolenic Acid Protects the SH-SY5Y cells against β Amyloid Toxicity

Figure 8

The CM and ALA-CM pretreatment inhibits Amyloid β- (Aβ_1-42-) induced depolarization of the mitochondrial membrane in differentiated SH-SY5Y cells. Representative fluorescence microscopy images of 5,5,6,6-tetrachloro-1,1,3,3 tetraethylbenzimi-dazoylcarbocyanine iodide (JC-1) staining (a) and the ratio of fluorescence intensity of J-aggregates to the fluorescence intensity of monomers (b, c) was used to measure mitochondrial membrane potential (ΔΨm) of differentiated SH-SY5Y cells. Results showed that Aβ_1-42 treatment decreased the ΔΨm of differentiated SH-SY5Y cells. The CM and ALA-CM pretreatment reversed the effect of Aβ_1-42 compared with group. The Insulin Degrading Enzyme (IDE) treatment of CM and ALA-CM reduced this effect. On the day 6th, the SH-SY5Y cells (differentiated) were pretreated for 1 h with CM or ALA-CM before the addition of 5 μM Aβ_1-42 for the next 24 h. The SH-SY5Y cells were also exposed to cotreatment of CM and ALA-CM with IDE to check whether insulin and IGF-I presence in CM and ALA-CM was responsible for the neuroprotective effect. Carbonyl cyanide 3-chlorophenylhydrazone (CCCP) was used as a mitochondrial membrane potential disruptor. Insulin was used as a positive control. Next, cells were subjected to JC-1 staining (see Materials and Methods section). Fluorescence of JC-1 was measured by a fluorescence microscope and microplate reader. One-way ANOVA followed by Tukey’s multiple comparisons test at the 0.05 level was used to determine differences between the treated cell and untreated control cells. Results are presented as (). The ratio of fluorescence intensity of J-aggregates (shown as red signals) to the fluorescence intensity of monomers (shown as green signals) was calculated according to the formula described in the Materials and Methods section. Statistical differences between the treated cells and untreated control cells are indicated by asterisks ( for ; for ; for ; ^# versus the control group). Scale bar is 50 μm.

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