Department of Urology, Institute of Basic Medical Sciences, Graduate School of Comprehensive Human Sciences, University of Tsukuba, Ibaraki, Japan.
Cancer Sci. 2011 Jan;102(1):267-74. doi: 10.1111/j.1349-7006.2010.01750.x. Epub 2010 Oct 26.
Apoptotic protease activating factor-1 (APAF-1) is a key regulator gene of apoptosis, located downstream from p53. Loss of APAF-1 expression is associated with chemorefractory malignant melanoma and neuronal cell differentiation. In order to make clear the function of APAF-1 in the carcinogenesis of germ cell tumors, we evaluated the expression levels of APAF-1 and several apoptosis and differentiation markers by immunohistochemistry in formalin-fixed paraffin-embedded samples from 43 cases of testicular germ cell tumor (TGCT) and six specimens of normal testis tissue. Expression of cleaved caspase-3, Oct-3/4, and Ki-67 were also examined by immunohistochemistry to evaluate apoptotic reactivity, tumor differentiation, and proliferation activity, respectively. APAF-1 was downregulated in two TGCT cell lines by siRNA transfection, and subsequent expression of the Ki-67 and Oct-3/4 genes and differentiation markers of three embryonic germ layers including keratin16 (KRT16) for ectoderm, vimentin (VIM) for mesoderm and GATA4 for endoderm were then tested. No significant relationship was found between APAF-1 expression and apoptotic activity in TGCTs. Expression of APAF-1, Oct-3/4, and Ki-67 was significantly higher in seminomas than in non-seminomas. In TGCTs, higher APAF-1 expression was correlated with higher proliferation (high Ki-67) and a lower degree of differentiation (high Oct-3/4). Interestingly, the expression of APAF-1 gradually decreased in accordance with tumor differentiation (seminoma and embryonal carcinoma > teratoma). Downregulation of APAF-1 in TGCT cell lines resulted in a decrease of Ki-67 and Oct-3/4 and an increase of VIM and KRT16 gene expression. These data show that higher expression of APAF-1 is related to an undifferentiated state in the TGCT pathway.
凋亡蛋白酶激活因子-1（APAF-1）是凋亡的关键调节基因，位于 p53 下游。APAF-1 表达缺失与化学耐药性恶性黑色素瘤和神经元细胞分化有关。为了明确 APAF-1 在生殖细胞肿瘤发生中的作用，我们通过免疫组织化学方法检测了 43 例睾丸生殖细胞肿瘤（TGCT）和 6 例正常睾丸组织的福尔马林固定石蜡包埋样本中 APAF-1 及几种凋亡和分化标志物的表达水平。还通过免疫组织化学检测了裂解的 caspase-3、Oct-3/4 和 Ki-67 的表达，分别评估凋亡反应性、肿瘤分化和增殖活性。通过 siRNA 转染下调了两个 TGCT 细胞系中的 APAF-1 表达，随后检测了 Ki-67 和 Oct-3/4 基因以及三个胚层的分化标志物的表达，包括外胚层的角蛋白 16（KRT16）、中胚层的波形蛋白（VIM）和内胚层的 GATA4。在 TGCT 中，APAF-1 表达与凋亡活性之间无显著相关性。在精原细胞瘤中，APAF-1、Oct-3/4 和 Ki-67 的表达明显高于非精原细胞瘤。在 TGCT 中，APAF-1 表达越高，增殖（高 Ki-67）越高，分化程度越低（高 Oct-3/4）。有趣的是，APAF-1 的表达随着肿瘤分化程度的降低（精原细胞瘤和胚胎癌>畸胎瘤）逐渐降低。在 TGCT 细胞系中下调 APAF-1 导致 Ki-67 和 Oct-3/4 减少，波形蛋白和 KRT16 基因表达增加。这些数据表明，APAF-1 的高表达与 TGCT 途径中的未分化状态有关。
