Resumo

Introduction and purpose: Neurofilament proteins are found in the perikarya and especially in axons throughout the central and peripheral nervous system. The purpose of this study was to characterize the distribution of neurofilaments and their mRNA expression in human retinas by using immunohistochemistry and real time-polymerase chain reaction (PCR). Methods: Human retinas were obtained from eyes previously used for cornea donation. Some retinas were cut vertically and the sections were incubated with anti-neurofilament-68 kD (NF-L), -160 kD (NF-M) and -200 kD (NF-H) and processed with standard immunoperoxidase and immunofluorescence methods. Retinal whole-mounts were incubated with an anti-PAN antibody (recognizing all three neurofilament isoforms). The number of labeled cells and their retinal distribution in the whole-mounts were established by a systematic sampling method to map the cellular distribution along the entire retina at 2 mm intervals. Quantitative real-time PCR was used to detect NF-L, NF-M and NF-H mRNA expression in peripheral and central human retina. NF-L, NF-M and NF-H mRNA levels were measured by SyBR Green qPCR assay, using ubiquitin C (UBC) and succinate dehydrogenase complex subunit A (SDHA) as reference genes. Results: Whole-mounts labeled with anti-PAN exhibited a considerable number of fibers in the nerve fiber layer (nfl), a subset of completely stained large ganglion cells in the ganglion cell layer (gcl), and a population of presumptive amacrine cells in the inner nuclear layer (inl). Those completely stained cells in the gcl were detected in highest density values around the fovea (ca. 150 cells/mm2). In the vertical sections labeled for NF-H, we only observed immunostained somata in the gcl and fibers in the nfl. NF-L was also present in stained somata in the inl and in processes in the inner plexiform layer (ipl). No immunolabeling was found with NF-M immunohistochemistry. Transcription levels of all three neurofilaments genes were significantly higher in the central retina, and NF-L was the most expressed gene in both regions. Conclusion: RNA data indicate that all three neurofilament isoforms are expressed in the human retina, although no immunolabeling was detected for the NF-M isoform. Our results suggest that the large ganglion cells labeled by the anti-PAN antibody could represent parasol cells (also known as the M-type).