The Relationship Between Autophagy-Lysosome Pathway And Neurodegenera-tive Disease In Acidic Conditions

ObjectiveTo explore the relationship between autophagy-lysosome pathway and neurodegenerative disease in acidic conditions.MethodsNHE-1knock-down cells were established by transfecting pAd-miR-NHE1interfering adenovirus plasmid into SH-SY5Y cells. Intracellular acidic cell model were established by using suitable acid medium to culture NHE-1knock-down cells. The most suitable pH of the medium is5.8. So in this research work, there were3groups including normal group, intracellular acidification group and the negative control group (NC group). The shape of autophagy body, changes of mitochondrial formation and number of lysosome in the three groups were observed by transmission electro microscopy. The expression of autophagy marker-LC3Ⅱ and LC3I was detected by western blot.Results1、In normal group and NC group, normal mitochondrial formation and normal number of lysosomes were observed under the transmission electron microscopy. After the NHE-1knock-down cells were cultured in the medium with pH5.8, the detected intracellular pH of cells was6.5. Compare with normal cells(pHi=7.27) and NC cells(pHi=7.24), the intracellular pH decreased significantly(P<0.01). During half an hour to4hours’culture in acidic medium, swelling mitochondria and increasing number of lysosomes and autophagy vesicles were observed, autophagy bodies appeared. After8hours, apoptotic body appeared, the number of mitochondria decreased, there was no autophagy body and only a bit lysosomes leaved.2、Western blot shows, there are low LC-3Ⅱ expression in normal group and NC group, which were0.82±0.15and0.94±0.18. In the acidic environment for half an hour, the expression of LC-3Ⅱ was1.63±0.24, the difference compared with the normal group and NC group has significance(P<0.01), then reached2.01±0.35after4hours,the difference compared with the normal group and NC group has significance (P<0.01), after8hours, it was0.80±0.12, and the difference compared with the normal group and NC group has significance (P<0.01). In this acidic environment for half an hour, LC3Ⅱ/LC3I was1.04±0.21and the difference compared with the normal group and NC group has significance (P<0.01), then reached1.15±0.24after4hours and the difference compared with the normal group and NC group has significance (P<0.01), after8hours, it was0.57±0.15, and the difference compared with the normal group and NC group has significance(P<0.01).ConclusionsIntracellular acidification can activate the autophagy through the autophagy-lysosome pathway. In the later stage of autophagy, apoptotic bodies appeared and cell apoptosis started. The common character of neurodegeneration diseases is intracellular acidification which provided us that autophagy-lysosome pathway may speed up the apoptosis of neurons in neurodegeneration diseases.