Espana Edgar M, Sun Mei, Birk David E
Department of Molecular Pharmacology & Physiology Morsani College of Medicine, University of South Florida, Tampa, Florida, United States 2Department of Ophthalmology, Morsani College of Medicine, University of South Florida, Tampa, Florida, United States.
Department of Molecular Pharmacology & Physiology Morsani College of Medicine, University of South Florida, Tampa, Florida, United States.
Invest Ophthalmol Vis Sci. 2015 Jun;56(6):3827-37. doi: 10.1167/iovs.14-16030.
To demonstrate the presence and location of corneal endothelial progenitor cells.
Progenitor cell markers nestin, leucine-rich repeat-containing G-protein-coupled receptor 5, Sox9, and nerve growth factor receptor p75, as well as proliferation marker Ki-67, were examined on postnatal day (P)3, P30, and P90 corneas using immunofluorescence microscopy. Mice (P3) were pulsed with 5-bromo-2'-deoxyuridine (BrdU) and chased.
Cell proliferation was observed in all layers of P3 corneas. No posterior stromal cell proliferation was noted in P30 corneas. Progenitor cell markers were expressed in the P3 cornea, but were downregulated during maturation with minimal or no expression in P90 central corneas. In contrast, cells expressing progenitor markers were located exclusively at the corneal periphery at P90. Clusters of cells reactive for progenitor markers were in the endothelial and subendothelial space in the P90 peripheral cornea. Reactivity against BrdU was localized to the central and peripheral cornea at 1 week, and to the extreme periphery 3 weeks following a BrdU pulse. Cells reactive for both BrdU and progenitor markers were localized to the peripheral endothelium. At 3 weeks, cells reactive for BrdU and the progenitor markers were localized in the peripheral endothelium. Approximately, 20% to 45% of the progenitor marker positive cells also were labeled with BrdU.
During development, the murine corneal endothelium is composed of proliferating cells expressing progenitor markers. In contrast, in the mature endothelium slow-cycling cells, cells expressing progenitor markers and a subpopulation of slow-cycling cells expressing progenitor makers are restricted to the endothelial periphery.
证明角膜内皮祖细胞的存在及其位置。
使用免疫荧光显微镜，在出生后第（P）3天、P30天和P90天的角膜上检测祖细胞标志物巢蛋白、富含亮氨酸重复序列的G蛋白偶联受体5、Sox9和神经生长因子受体p75，以及增殖标志物Ki-67。对出生3天的小鼠注射5-溴-2'-脱氧尿苷（BrdU）并进行追踪观察。
在出生3天的角膜各层均观察到细胞增殖。出生30天的角膜后基质层未观察到细胞增殖。祖细胞标志物在出生3天的角膜中表达，但在成熟过程中表达下调，在出生90天的中央角膜中表达极少或无表达。相反，在出生90天，表达祖细胞标志物的细胞仅位于角膜周边。在出生90天的周边角膜中，对祖细胞标志物呈反应性的细胞簇位于内皮和内皮下间隙。BrdU反应性在BrdU脉冲后1周定位于中央和周边角膜，3周后定位于最周边。对BrdU和祖细胞标志物均呈反应性的细胞定位于周边内皮。3周时，对BrdU和祖细胞标志物呈反应性的细胞定位于周边内皮。约20%至45%的祖细胞标志物阳性细胞也被BrdU标记。
在发育过程中，小鼠角膜内皮由表达祖细胞标志物的增殖细胞组成。相比之下，在成熟内皮中，慢循环细胞、表达祖细胞标志物的细胞以及表达祖细胞标志物的慢循环细胞亚群局限于内皮周边。
