Induced Pluripotency of Human Prostatic Epithelial Cells

Induced pluripotent stem (iPS) cells are a valuable resource for discovery of epigenetic changes critical to cell type-specific differentiation. Although iPS cells have been generated from other terminally differentiated cells, the reprogramming of normal adult human basal prostatic epithelial (E-PZ) cells to a pluripotent state has not been reported. Here, we attempted to reprogram E-PZ cells by forced expression of Oct4, Sox2, c-Myc, and Klf4 using lentiviral vectors and obtained embryonic stem cell (ESC)-like colonies at a frequency of 0.01%. These E-PZ-iPS-like cells with normal karyotype gained expression of pluripotent genes typical of iPS cells (Tra-1-81, SSEA-3, Nanog, Sox2, and Oct4) and lost gene expression characteristic of basal prostatic epithelial cells (CK5, CK14, and p63). E-PZ-iPS-like cells demonstrated pluripotency by differentiating into ectodermal, mesodermal, and endodermal cells in vitro, although lack of teratoma formation in vivo and incomplete demethylation of pluripotency genes suggested only partial reprogramming. Importantly, E-PZ-iPS-like cells re-expressed basal epithelial cell markers (CD44, p63, MAO-A) in response to prostate-specific medium in spheroid culture. Androgen induced expression of androgen receptor (AR), and co-culture with rat urogenital sinus further induced expression of prostate-specific antigen (PSA), a hallmark of secretory cells, suggesting that E-PZ-iPS-like cells have the capacity to differentiate into prostatic basal and secretory epithelial cells. Finally, when injected into mice, E-PZ-iPS-like cells expressed basal epithelial cell markers including CD44 and p63. When co-injected with rat urogenital mesenchyme, E-PZ-iPS-like cells expressed AR and expression of p63 and CD44 was repressed. DNA methylation profiling identified epigenetic changes in key pathways and genes involved in prostatic differentiation as E-PZ-iPS-like cells converted to differentiated AR- and PSA-expressing cells. Our results suggest that iPS-like cells derived from prostatic epithelial cells are pluripotent and capable of prostatic differentiation; therefore, provide a novel model for investigating epigenetic changes involved in prostate cell lineage specification.

诱导多能干细胞（induced pluripotent stem cells, iPS cells）是探索细胞类型特异性分化关键表观遗传变化的宝贵研究资源。尽管已有研究从其他终末分化细胞中成功诱导生成iPS细胞，但目前尚未见将正常成人前列腺基底上皮（E-PZ）细胞重编程至多能状态的相关报道。本研究尝试通过慢病毒载体（lentiviral vectors）强制表达Oct4、Sox2、c-Myc及Klf4，对E-PZ细胞进行重编程，最终以0.01%的诱导效率获得了胚胎干细胞（embryonic stem cell, ESC）样菌落。这些核型正常的E-PZ来源iPS样细胞，获得了典型iPS细胞的多能基因表达特征（Tra-1-81、SSEA-3、Nanog、Sox2及Oct4），同时丧失了前列腺基底上皮细胞的特征性基因表达（CK5、CK14及p63）。体外实验中，E-PZ-iPS样细胞可分化为外胚层、中胚层及内胚层细胞，展现出多向分化潜能；但体内未形成畸胎瘤（teratoma），且多能基因的去甲基化不完全，提示其重编程仅为部分完成。重要的是，在球体培养体系中，E-PZ-iPS样细胞可响应前列腺特异性培养基，重新表达基底上皮细胞标志物（CD44、p63、MAO-A）。雄激素可诱导雄激素受体（androgen receptor, AR）的表达，而与大鼠尿生殖窦共培养则进一步诱导了分泌细胞标志性蛋白——前列腺特异性抗原（prostate-specific antigen, PSA）的表达，表明E-PZ-iPS样细胞具备分化为前列腺基底上皮细胞及分泌型上皮细胞的能力。最后，将E-PZ-iPS样细胞注射至小鼠体内时，其可表达包括CD44及p63在内的基底上皮细胞标志物；当与大鼠尿生殖间质共移植时，E-PZ-iPS样细胞则表达AR，且p63与CD44的表达受到抑制。DNA甲基化谱分析显示，当E-PZ-iPS样细胞转化为表达AR及PSA的分化细胞时，其关键通路及参与前列腺分化的基因发生了表观遗传改变。本研究结果表明，源自前列腺上皮细胞的iPS样细胞具备多能性及前列腺定向分化能力，可为探索前列腺细胞谱系特化过程中的表观遗传调控机制提供全新的研究模型。