ABSTRACT
The corneal epithelium is renowned for high regenerative potential, which is dependent on the coordinated function of its diverse progenitor subpopulations. However, the molecular pathways governing corneal epithelial progenitor differentiation are incompletely understood. Here, we identify a highly proliferative limbal epithelial progenitor subpopulation characterized by expression of basal cell adhesion molecule (BCAM) that is capable of holocone formation and corneal epithelial sheet generation. BCAM-positive cells can be found among ABCB5-positive limbal stem cells (LSCs) as well as among ABCB5-negative limbal epithelial cell populations. Mechanistically, we show that BCAM is functionally required for cellular migration and differentiation and that its expression is regulated by the transcription factor p63. In aggregate, our study identifies limbal BCAM expression as a marker of highly proliferative corneal epithelial progenitor cells and defines the role of BCAM as a critical molecular mediator of corneal epithelial differentiation.
Subject(s)
Epithelium, Corneal , Limbus Corneae , Cell Differentiation , Cells, Cultured , Cornea , Epithelial Cells/metabolism , Limbus Corneae/metabolism , Stem Cells/metabolismABSTRACT
Glioblastoma multiforme (GBM) is a malignant brain tumor with a poor prognosis resulting from tumor resistance to anticancer therapy and a high recurrence rate. Compelling evidence suggests that this is driven by subpopulations of cancer stem cells (CSCs) with tumor-initiating potential. ABC subfamily B member 5 (ABCB5) has been identified as a molecular marker for distinct subsets of chemoresistant tumor-initiating cell populations in diverse human malignancies. In the current study, we examined the potential role of ABCB5 in growth and chemoresistance of GBM. We found that ABCB5 is expressed in primary GBM tumors, in which its expression was significantly correlated with the CSC marker protein CD133 and with overall poor survival. Moreover, ABCB5 was also expressed by CD133-positive CSCs in the established human U-87 MG, LN-18, and LN-229 GBM cell lines. Antibody- or shRNA-mediated functional ABCB5 blockade inhibited proliferation and survival of GBM cells and sensitized them to temozolomide (TMZ)-induced apoptosis in vitro Likewise, in in vivo human GBM xenograft experiments with immunodeficient mice, mAb treatment inhibited growth of mutant TP53, WT PTEN LN-229 tumors, and sensitized LN-229 tumors to TMZ therapy. Mechanistically, we demonstrate that ABCB5 blockade inhibits TMZ-induced G2/M arrest and augments TMZ-mediated cell death. Our results identify ABCB5 as a GBM chemoresistance marker and point to the potential utility of targeting ABCB5 to improve current GBM therapies.
Subject(s)
ATP Binding Cassette Transporter, Subfamily B , Antibodies, Neoplasm/pharmacology , Apoptosis/drug effects , Brain Neoplasms , Drug Resistance, Neoplasm/drug effects , G2 Phase Cell Cycle Checkpoints/drug effects , Glioblastoma , M Phase Cell Cycle Checkpoints/drug effects , Neoplasm Proteins , RNA, Small Interfering , Temozolomide/pharmacology , ATP Binding Cassette Transporter, Subfamily B/antagonists & inhibitors , ATP Binding Cassette Transporter, Subfamily B/genetics , ATP Binding Cassette Transporter, Subfamily B/metabolism , Animals , Brain Neoplasms/drug therapy , Brain Neoplasms/genetics , Brain Neoplasms/metabolism , Brain Neoplasms/pathology , Cell Line, Tumor , Drug Resistance, Neoplasm/genetics , Female , Glioblastoma/drug therapy , Glioblastoma/genetics , Glioblastoma/metabolism , Glioblastoma/pathology , Humans , Mice , Neoplasm Proteins/antagonists & inhibitors , Neoplasm Proteins/genetics , Neoplasm Proteins/metabolism , RNA, Small Interfering/genetics , RNA, Small Interfering/pharmacology , Xenograft Model Antitumor AssaysABSTRACT
PURPOSE: To identify factors associated with isolation yields of ATP-binding cassette (ABC) superfamily member B5 (ABCB5)-positive limbal stem cells (LSCs) from human cadaveric donor eyes. METHODS: Whole eye globes were obtained from the Saving Sight eye bank, Kansas City, MO and the CorneaGen eye bank, Seattle, WA. ABCB5-positive LSCs were sorted by flow cytometry upon anti-ABCB5 monoclonal antibody staining within one week after donor death. The yields of live limbal epithelial cells in their entirety and of isolated pure ABCB5-positive LSC subsets were correlated with variables contained in the eye donors' medical information. RESULTS: The mean isolation yield of live limbal epithelial cells and ABCB5-positive LSCs per donor eye was (340,000⯱â¯160,000 and 2,608⯱â¯1,842 respectively, mean⯱â¯SD). Stepwise regression analysis showed that cardiac disease-related death was the strongest negative predictor of the ABCB5-positive LSC isolation yield (pâ¯=â¯0.01). While we observed a trend for an age-related decline in the yield of ABCB5-positive LSCs, a statistically significant association could not be established (2% decrease/year, pâ¯=â¯0.11). Additionally, despite a trend for decreased isolation yields of total live limbal epithelial cells isolated from single donors with a longer time between death and tissue processing (pâ¯=â¯0.04), this did not affect the yields of purified ABCB5-positive LSC, which was independent of increasing time between death and tissue processing (pâ¯=â¯0.50). CONCLUSIONS: Our study identifies cardiac disease-related death as a donor variable significantly associated with lower ABCB5-positive LSC isolation yields.
