Abstract
Purpose: :
To analyze expression of the lymphatic endothelial cell (LEC) marker podoplanin in the mur-ine, bovine and human retina.
Methods: :
Human retinas were obtained from donor eyes rejected from cornea transplantation. Bovine eyes were obtained from a local abattoir, murine eyes were freshly enucleated from C57/Bl6 mice priorly sacrificed by sub-lethal narcotization and cervical dislocation. Retina samples were prepared as described elsewhere and fixed in PFA for immunhistofluorescence or Ito`s fixative for immunoelectron microscopy. Sections of cryo- or paraffin-embedded retinas were analyzed for expression of Podoplanin, CRALBP, and PKCalpha alone or in combination. For 3D modeling z-stacks were produced by confocal microscopy using a Zeiss LSM5 Pascal, and image analysis and editing were done with an evaluation version of the Imaris 7.1.1 software.
Results: :
Podoplanin labeling was detected on the surface of cellular processes spanning from the inner limiting membrane throughout the outer nuclear layer (ONL) in all analyzed species. Nuclei of these immunopositive cells were located in the inner nuclear layer (INL) as demonstrated by DAPI labeling. An antibody against the Müller cell marker CRALBP labeled the same cell structures as Podoplanin, whereas PKCalpha - a marker for bipolar and amacrine cells - did not colocalize with Podoplanin or CRALBP. Confocal microscopy and subsequent 3D modeling suggested that the CRALBP signal was located within the cytoplasm of the Müller cells whereas the Podoplanin label was restricted to their cell membrane.
Conclusions: :
Podoplanin might be a novel marker for adult retinal Müller cells. Its distinct functional role, however, is to be elucidated in subsequent studies.
Keywords: retina • Muller cells • microscopy: light/fluorescence/immunohistochemistry