神経変性疾患の細胞モデルを用いた、発症機序と予防法についての研究

我们一直在使用我们的帕金森病诱导PC12细胞模型研究α -突触核蛋白引起细胞死亡的机制。我们通过免疫细胞化学和免疫印迹实验分别证实了强力霉素不存在和不存在时α -突触核蛋白的诱导和抑制作用。我们在A53T突变细胞系中观察到细胞质中含有α -突触核蛋白和泛素。在细胞毒性评估中，我们采用台盼蓝排除法和DAPI核染色，发现神经元样分化和诱导A53T α -突触核蛋白表达后凋亡细胞死亡增加。我们在这里观察到的细胞死亡依赖于α -突触核蛋白的表达。我们提出阐明NF-kappaB通路在α -突触核蛋白突变引起的细胞死亡中的作用。我们发现A53T细胞系胞质包涵体IkappaB免疫染色阳性。我们尝试了三种NF-kappaB抑制剂，包括PDTC、DDTC和sn50，发现在抑制剂存在的情况下，A53T α -突触核蛋白引起的细胞死亡的衰减。我们的研究结果表明，NF-kappaB通路可能在突变型α -突触核蛋白引起的细胞死亡中发挥作用。此外，NF-kappaB通路可能是治疗α -突触核蛋白引起的神经细胞死亡的潜在靶点。

We have been studying the mechanisms underlying cell death caused by alpha-synuclein using our inducible PC12 cell model of Parkinson's disease. We confirmed induction and suppression of alpha-synuclein in the absence and presence of doxycycline, respectively, with immunocytochemistry and western blotting experiments. We observed cytoplasmic inclusions stained for alpha-synuclein and ubiquitin in A53T mutant cell lines. In cell toxicity evaluation, we employed the trypan-blue exclusion assay and DAPI nuclear staining, finding increased apoptotic cell death after neuron-like differentiation and induction of A53T alpha-synuclein expression. Cell death we observed here was in an alpha-synuclein expression dependent manner.We have proposed to elucidate the role for the NF-kappaB pathway in cell death caused by mutant alpha-synuclein. We found that cytoplasmic inclusions in A53T cell lines were positive for IkappaB immunostaining. We tried three NF-kappaB inhibitors including PDTC,DDTC, and SN5O, finding attenuation of A53T alpha-synuclein-caused cell death in the presence of the inhibitors.Our findings suggest that the NF-kappaB pathway may play a role in mutant alpha-synuclein-caused cell death. In addition, the NF-kappaB pathway may be a potential target to treat neural cell death caused by alpha-synuclein.