title: Measurement of In Vivo Three-Dimensional Corneal Cell Density and Size Using Two-Photon Imaging in C57BL/6 Mice.
abstract: To measure the cell size and cell density in five layers of the central cornea in the widely used inbred C57BL/6 mouse strain using in vivo three-dimensional (3D) two-photon (2PH) imaging.
  
  
  
  Corneas were scanned using a 2PH laser scanning fluorescence microscope after staining with plasma membrane stain and Hoechst 33342. Good quality 3D images were selected for the cell density and cell size analysis. Cell density was determined by counting the cell nuclei in a predefined cube of 3D images. Cell size measurements, including cell surface area, cell volume, nuclear surface area and nuclear volume, were automatically quantified using the Imaris software. The cell and nuclear surface-area-to-volume ratio (S:V ratio) and the cell nuclear-cytoplasmic ratio (N:C ratio) were calculated.
  
  
  
  The highest cell density was observed in the basal epithelium and the lowest in the posterior stroma. The highest cell surface area was found in the anterior stroma, and the highest cell volume was observed in the superficial epithelium. The lowest cell surface area and cell volume were both found in the basal epithelium. The highest S:V ratio was observed in the basal epithelium and the lowest in the superficial epithelium. The highest cell nuclear surface area and volume were both observed in the superficial epithelium and the lowest in the basal epithelium. The highest cell nuclear S:V ratio was observed in the basal epithelium and the lowest in the superficial epithelium. The highest N:C ratio was found in the basal epithelial cells and the lowest in the posterior keratocytes.
  
  
  
  We are the first to quantify the cell density and size parameters, including cell surface area and volume, cell nuclear surface area and volume, and the S:V ratio, in the five layers of the central cornea. These data provide important cell morphology features for the study of corneal physiology, pathology and disease in mice, particularly in C57BL/6 mice.
authors.affiliation: a Henan Eye Institute, Department of Ophthalmology, Henan Eye Hospital, Henan Provincial People's Hospital and People's Hospital of Zhengzhou University , Zhengzhou , People's Republic of China and.
authors.forename: Hongmin
authors.initials: H
authors.lastname: Zhang
authors.affiliation: a Henan Eye Institute, Department of Ophthalmology, Henan Eye Hospital, Henan Provincial People's Hospital and People's Hospital of Zhengzhou University , Zhengzhou , People's Republic of China and.
authors.forename: Siyu
authors.initials: S
authors.lastname: He
authors.affiliation: a Henan Eye Institute, Department of Ophthalmology, Henan Eye Hospital, Henan Provincial People's Hospital and People's Hospital of Zhengzhou University , Zhengzhou , People's Republic of China and.
authors.forename: Susu
authors.initials: S
authors.lastname: Liu
authors.affiliation: a Henan Eye Institute, Department of Ophthalmology, Henan Eye Hospital, Henan Provincial People's Hospital and People's Hospital of Zhengzhou University , Zhengzhou , People's Republic of China and.
authors.forename: Yanting
authors.initials: Y
authors.lastname: Xie
authors.affiliation: a Henan Eye Institute, Department of Ophthalmology, Henan Eye Hospital, Henan Provincial People's Hospital and People's Hospital of Zhengzhou University , Zhengzhou , People's Republic of China and.
authors.forename: Guoming
authors.initials: G
authors.lastname: Chen
authors.affiliation: a Henan Eye Institute, Department of Ophthalmology, Henan Eye Hospital, Henan Provincial People's Hospital and People's Hospital of Zhengzhou University , Zhengzhou , People's Republic of China and.
authors.forename: Junjie
authors.initials: J
authors.lastname: Zhang
authors.affiliation: a Henan Eye Institute, Department of Ophthalmology, Henan Eye Hospital, Henan Provincial People's Hospital and People's Hospital of Zhengzhou University , Zhengzhou , People's Republic of China and.
authors.forename: Shengtao
authors.initials: S
authors.lastname: Sun
authors.affiliation: b Penn State Hershey Eye Center, Penn State Milton S. Hershey Medical Center, Penn State College of Medicine , Hershey , PA , USA.
authors.forename: David
authors.initials: D
authors.lastname: Liang
authors.affiliation: a Henan Eye Institute, Department of Ophthalmology, Henan Eye Hospital, Henan Provincial People's Hospital and People's Hospital of Zhengzhou University , Zhengzhou , People's Republic of China and.
authors.forename: Liya
authors.initials: L
authors.lastname: Wang
mesh_terms: D000818:Animals; D050541:Bowman Membrane / Q000166:cytology; D002452:Cell Count; D048429:Cell Size; D003315:Cornea / Q000166:cytology*; D003319:Corneal Stroma / Q000166:cytology; D004728:Endothelium, Corneal / Q000166:cytology; D019573:Epithelium, Corneal / Q000166:cytology; D021621:Imaging, Three-Dimensional; D051379:Mice; D008810:Mice, Inbred C57BL; D018613:Microscopy, Confocal / Q000379:methods*; D023421:Models, Animal
keywords: C57BL/6 mice; Cell density; cell size; cornea; in vivo three-dimensional