Evidence for keratinocyte stem cells in vitro:: Long term engraftment and persistence of transgene expression from retrovirus-transduced keratinocytes
Evidence for keratinocyte stem cells in vitro:: Long term engraftment and persistence of transgene expression from retrovirus-transduced keratinocytes
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DOI：
10.1073/pnas.95.8.4356
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发表时间：
1998-04-14
影响因子：
11.1
通讯作者：
Taichman, LB
中科院分区：
文献类型：
--
作者：
Kolodka, TM;Garlick, JA;Taichman, LB
表皮被干细胞群更新，所述干细胞群在体内通过缓慢更新、标记保留、在表皮中的位置和β（1）整合素的富集而被定义，在体外通过克隆生长、延长连续传代和快速粘附于细胞外基质而被定义。本研究的目的是确定具有体外长期生长潜力的克隆形成细胞是否在体内持续存在并产生完全分化的表皮。通过用编码lacZ基因的逆转录病毒转导，在培养物中对人角质形成细胞进行遗传标记，并移植到无胸腺小鼠。移植前培养物的分析表明，21.1-27.8%的具有>30代能力的克隆形成细胞被成功地转导。在体内，β-半乳糖苷酶（β-gal）阳性细胞参与完全分化的上皮的形成，并且在移植后的整个40周期间被检测到，最初为松散分散的簇，后来为明显的垂直柱。将从切除的移植物中回收的活细胞以克隆密度接种，由此形成的集落的23.3-33.3%是β-gal阳性的。此外，没有获得转基因失活的证据：从β-gal阴性的移植组织中回收的所有角质形成细胞集落也都带有lacZ转基因。这些结果表明，在体外具有长期生长特性的细胞确实在体内持续存在并形成完全分化的表皮，从而表现出干细胞的特性。
Epidermis is renewed by a population of stem cells that have been defined in vivo by slow turnover, label retention, position in the epidermis, and enrichment in beta(1) integrin, and in vitro by clonogenic growth, prolonged serial passage, and rapid adherence to extracellular matrix. The goal of this study is to determine whether clonogenic cells with long-term growth potential in vitro persist in vivo and give rise to a fully differentiated epidermis, Human keratinocytes were genetically labeled in culture by transduction with a retrovirus encoding the lacZ gene and grafted to athymic mice. Analysis of the cultures before grafting showed that 21.1-27.8% of clonogenic cells with the capacity for >30 generations were successfully transduced, In vivo, beta-galactosidase (beta-gal) positive cells participated in the formation of a fully differentiated epithelium and were detected throughout the 40-week postgraft period, initially as loosely scattered clusters and later as distinct vertical columns. Viable cells recovered from excised grafts were seeded at clonal densities and 23.3-33.3% of the colonies thus formed were beta-gal positive. In addition, no evidence of transgene inactivation mas obtained: all keratinocyte colonies recovered from grafted tissue that were beta-gal negative also tacked the lacZ transgene. These results show that cells with long-term growth properties in vitro do indeed persist in vive and form a fully differentiated epidermis, thereby exhibiting the properties of stem cells.