Gene expression profiles of human cervical primary cells from ecto- and endocervix

The cellular origin of cervical cancers remains unclear. Revealing molecular details of transformation in this tissue has been hampered by the lack of culture systems, resembling the in vivo cervical architecture. Here we established a long-term in vitro 3D cervical organoid model derived from stem cells of human or mouse cervical tissue which recapitulates the in vivo stratified ectocervical and columnar endocervical epithelium. Stratified and columnar cervical epithelia arise from two discrete unipotent stem cell populations of the endocervix. Unique stem cell signatures reveal a dependency on intrinsic Notch and Wnt microenvironmental signals. The genetic signatures of KRT5+ stratified vs KRT7+ columnar cervical cells establish discrete groups of cervical cancer of the squamous and adenocarcinoma types, respectively. Cervical tissue morphology is guided by the interplay of two discrete unipotent cervical stem cell populations and the spatio-temporal distribution of signals from the stroma. Primary cells were isolated from healthy tissue of surgery specimen taken from ecto and/or endocervix of patients undergoing hysterectomy or cononization, cultured in 2D, transferred to Matrigel and cultured for 3d or 2weeks. Total RNA was extracted from samples and hybridized on Agilent-8x60k Human custom arrays as single-color hybridizations.

宫颈癌的细胞起源至今仍不明确。由于缺乏能够模拟体内宫颈组织结构的体外培养体系，解析该组织癌变过程的分子细节一直受到阻碍。本研究构建了源自人类或小鼠宫颈组织干细胞的长期体外三维宫颈类器官（cervical organoid）模型，该模型可复现体内复层外宫颈上皮与柱状内宫颈上皮的组织结构。复层宫颈上皮与柱状宫颈上皮均起源于内宫颈中的两种独立单能干细胞群。独特的干细胞基因特征表明，其存活依赖于内源性Notch与Wnt微环境信号通路。KRT5+（角蛋白5阳性）复层宫颈细胞与KRT7+（角蛋白7阳性）柱状宫颈细胞的基因特征，可分别对应鳞状细胞癌与腺癌两种亚型的宫颈癌离散分类群。宫颈组织的形态由两种独立的单能宫颈干细胞群的相互作用，以及间质信号的时空分布共同调控。研究人员从接受子宫切除术（hysterectomy）或宫颈锥切术（conization）的患者的外宫颈和/或内宫颈手术标本的健康组织中分离原代细胞，先以二维培养体系培养，随后转移至基质胶（Matrigel）中分别培养3天或2周。从上述样本中提取总RNA，以单色杂交方式在Agilent-8x60k人类定制基因芯片（Agilent-8x60k Human custom arrays）上完成杂交实验。