Genes with significantly altered expression in the striatum of Pink1−/− mice.

Gene expression was analyzed using NF-κB, PI3 kinase/Akt and cAMP/Ca2+ signaling PCR Arrays (SA Biosciences) as described in the Methods. Ct values (mean ± SD) for individual genes are indicated for wildtype (WT) and Pink1−/− mice. In addition, expression of each gene relative to the housekeeping genes (HKG) is indicated for WT and Pink1−/− mice and was used to calculate the fold change in gene expression (Pink1−/−/WT). All PCR arrays contain five HKG (β-glucuronidase, hypoxanthine guanine phosphoribosyl transferase, heat shock protein 90-alpha, glyceraldehyde-3-phosphate dehydrogenase, and β-actin), to which the expression of the genes of interests is normalized. None of the HKG was differentially expressed between WT and Pink1−/− mice. Data were evaluated and calculated with the ΔΔCt method using the RT2 Profiler PCR Array Data Analysis software and resources available online (http://sabiosciences.com/pcr/arrayanalysis.php). The p values were calculated based on a Student's t-test of the replicate 2−(AVG ΔCt) values for each gene in the WT and Pink1−/− groups. All genes with a pboth genotypes after running five arrays (five mice per genotype were analyzed). Occasionally, a well (gene) yielded no signal at all for reasons that are unrelated to actual lack of expression. For example, if a specific gene became detectable at PCR cycle number 25 in four mice but showed no expression in the fifth mouse of the same genotype, we concluded that this must be an experimental/technical error rather than actual lack of expression and omitted the corresponding data. However, we still used five data points for the other genotype if available. For a description of the function of the listed genes in innate immunity, MAPK signaling and/or their involvement and regulation in PD and models of PD, see the main text and references therein.本研究采用核因子κB（NF-κB）、磷脂酰肌醇3-激酶（PI3 kinase）/蛋白激酶B（Akt）及环腺苷酸/钙离子（cAMP/Ca²⁺）信号通路PCR芯片（PCR Arrays，SA Biosciences公司），按照方法部分所述流程进行基因表达分析。针对野生型（wildtype, WT）与Pink1基因敲除（Pink1⁻/⁻）小鼠的各目标基因，其循环阈值（Ct值）以均值±标准差（mean ± SD）形式标注。此外，分别给出两类小鼠中各基因相对于持家基因（housekeeping genes, HKG）的表达水平，并以此计算基因表达倍数变化（Pink1⁻/⁻/WT）。 所有PCR芯片均包含5种持家基因：β-葡萄糖醛酸苷酶（β-glucuronidase）、次黄嘌呤鸟嘌呤磷酸核糖转移酶（hypoxanthine guanine phosphoribosyl transferase）、热休克蛋白90α（heat shock protein 90-alpha）、甘油醛-3-磷酸脱氢酶（glyceraldehyde-3-phosphate dehydrogenase）及β-肌动蛋白（β-actin），目标基因的表达量均以这些持家基因为参照进行标准化。两类小鼠的持家基因表达水平无显著差异。 本研究采用RT² Profiler PCR芯片数据分析软件及配套在线资源（http://sabiosciences.com/pcr/arrayanalysis.php），通过双ΔCt（ΔΔCt）法对数据进行评估与计算。P值通过对野生型与Pink1⁻/⁻小鼠组中各基因的重复2⁻(平均ΔCt)值进行学生t检验（Student's t-test）计算得到。完成5次芯片实验（每个基因型分析5只小鼠）后，两类基因型中组间差异具有统计学意义的所有基因均被纳入分析。 偶尔会出现某一反应孔（对应基因）未检测到信号的情况，该现象与基因实际不表达无关。例如，若同基因型的4只小鼠在PCR循环数25时可检测到某特定基因的信号，但第5只小鼠未检出，则我们判定该情况属于实验或技术误差，而非基因实际不表达，并剔除该对应数据。但若另一基因型可获取5个有效数据点，则仍予以使用。 关于所列基因在固有免疫、丝裂原活化蛋白激酶（MAPK）信号通路中的功能，及其在帕金森病（PD）及帕金森病模型中的参与与调控机制，详见正文及参考文献。