Regional variation in M??ller cell expression of stem cell-associated proteins in normal adult human retina

摘要

Purpose : M??ller cells (MC) perform multiple roles critical to retinal homeostasis. More recently, MC have been proposed as putative pluripotent stem-like cells of the neural retina. This study tested the hypothesis that there is a regional variation in MC expression of stem cell-associated proteins CD117 and CD44 in adult human eyes. Methods : Five adult human eyes (50 to 59 years, Lions NSW Eye Bank) were fixed in 10% neutral buffered formalin for at least 24 h prior to dissection. For each eye, the pupil-to-optic nerve block underwent processing for immunohistochemistry. Serial sections of each eye were immunostained with CD44 (clone DF1485, Dako) and CD117 (clone C-4, Santa Cruz) according to proprietary protocols for Ventana BenchMark and Leica Bond II automated stainers, respectively. Image J software was used to assess CD44 expression in the outer human retina (from outer nuclear layer to photoreceptor outer segments), which was expressed as a % area immunostained for CD44. CD117 expression was quantified as the % CD117+ MC cell bodies in the total inner nuclear layer. An average value for each parameter was obtained from 3 fields per eye at 40x objective, and compared between macula and peripheral retinal regions. Results : CD117 is a robust marker of human MC across all regions of the human retina and is expressed throughout the MC cytoplasm, from the inner limiting membrane and endfeet to the apices in the fibre basket. MC nuclei consistently expressed CD117. In contrast, strong CD44 expression was only seen in the MC cytoplasm in the fibre basket region. The % CD117+ MC cell bodies was higher in the peripheral retina compared to the macula (p = 0.08; Mann-Whitney test). The % area immunostained for CD44 was significantly higher in peripheral retina compared with the macula (p = 0.08; Mann-Whitney test). Conclusions : CD117, stem-cell growth factor receptor, is a robust nuclear and cytoplasmic marker of human MC, irrespective of retinal location (peripheral or central). High proportion of CD117+ MC cell bodies correlated positively with CD44 expression in MC fibre basket a feature of peripheral human retina. Macular retina is characterised by proportionally less CD117+ MC cell bodies and an absence of CD44 expression in MC, suggesting this region may be more vulnerable than peripheral retina for CD44-dependent cellular responses such as oxidative stress. This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.