ABCG2 transporter identifies a population of clonogenic human limbal epithelial cells
ABCG2 transporter identifies a population of clonogenic human limbal epithelial cells
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DOI：
10.1634/stemcells.2004-0093
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发表时间：
2005-01-01
期刊：
影响因子：
5.2
通讯作者：
Li, DQ
中科院分区：
文献类型：
--
作者：
De Paiva, CS;Chen, Z;Li, DQ
ABCG 2是ATP结合盒（ABC）转运蛋白的成员，已被鉴定为骨髓干细胞的分子决定簇，并被提议作为干细胞的通用标记物。本研究探讨了ABCG 2的表达及其作为识别人角膜缘上皮干细胞的标志物的潜力。ABCG 2表达通过免疫荧光和免疫组织化学染色、激光扫描共聚焦显微镜、流式细胞术和半定量逆转录聚合酶链反应进行评估。通过ABCG 2单克隆抗体（mAb）或Hoechst 33342染料染色的流式细胞术从原代角膜缘上皮培养物中选择的细胞进行基因表达和集落形成效率（CFE）评价。ABCG 2蛋白主要定位于角膜缘上皮基底细胞，而不定位于角膜缘基底上细胞和角膜上皮。在原代角膜缘上皮培养物中也观察到ABCG 2染色。角膜缘上皮细胞比角膜上皮细胞表达更高水平的ABCG 2和DeltaNp 63 mRNA。用ABCG 2 mAb标记，流式细胞术检测阳性细胞数为2.5%-3.0%。ABCG 2阳性细胞在3 T3成纤维细胞饲养层上表现出比ABCG 2阴性细胞更大的CFE。通过Hoechst 33342排除试验检测侧群（SP）。SP细胞ABCG 2和DeltaNp 63 mRNA的表达较非SP细胞强，CFE也较非SP细胞高。总之，这些研究结果表明，ABCG 2转运蛋白只表达的角膜缘基底细胞和ABCG 2阳性和SP细胞具有丰富的干细胞特性，首次表明，ABCG 2可以作为一个标志物，以确定推定的角膜缘上皮干细胞。
ABCG2, a member of the ATP binding cassette (ABC) transporters, has been identified as a molecular determinant for bone marrow stem cells and proposed as a universal marker for stem cells. This study investigates ABCG2 expression and its potential as a marker that identifies human limbal epithelial stem cells. ABCG2 expression was evaluated by immunofluorescent and immunohistochemical staining, laser scanning confocal microscopy, flow cytometry, and semiquantitative reverse transcription-polymerase chain reaction. Cells selected from primary limbal epithelial cultures by flow cytometry with ABCG2 monoclonal antibody (mAb) or Hoechst 33342 dye staining were evaluated for their gene expression and colony-forming efficiency (CFE). ABCG2 protein was mainly located in the basal cells of limbal epithelia but not in the limbal suprabasal and corneal epithelia. ABCG2 staining was also observed in primary limbal epithelial cultures. Limbal epithelia express higher levels of ABCG2 and DeltaNp63 mRNAs than corneal epithelia. Labeling with ABCG2 mAb yielded 2.5%-3.0% positive cells by flow cytometry. The ABCG2-positive cells exhibited greater CFE on a 3T3 fibroblast feeder layer than ABCG2-negative cells. A side population (SP) was detected by the Hoechst 33342 exclusion assay. SP cells displayed stronger expression of ABCG2 and DeltaNp63 mRNA and greater CFE than the non-SP cells. In conclusion, these findings demonstrate that ABCG2 transporter was exclusively expressed by limbal basal cells and that the ABCG2-positive and SP cells possess enriched stem cell properties, suggesting for the first time that ABCG2 could serve as a marker to identify the putative limbal epithelial stem cells.