Brumley, Sarah G.1,3, Wadkins, David2,3; Kuehn, Markus H.2,3
1University of Iowa, Biology Department; 2University of Iowa Department of Ophthalmology and Visual Sciences; 3VA Center for the Prevention and Treatment of Visual Loss, Iowa City, IA
Purpose: The trabecular meshwork (TM) of the eye is critical in maintaining aqueous humor outflow and intraocular pressure. The cellular density in the TM decreases with age and is particularly low in eyes with glaucoma. TM cells are thought to be derived from a population of stem cells, referred to as TM stem cells (TMSC), found in the ‘insert zone’ that is located anterior to the TM. We hypothesized that reduced cellularity of TM cells is due to loss of TMSC. In order to test this hypothesis, we determined the number of TMSC and TM cells in the eyes of young eye donors, as well as in glaucoma eyes and age-matched controls.
Methods: Human eyes were obtained from young donors (n=3), healthy older donors (n=6), and those with glaucoma (n=7) and fixed in paraformaldehyde and embedded in paraffin. From each eye 5 sagittal section were obtained and incubated with antibodies directed against Nestin and ABCG2. The number of double positive cells in the insert zone and the trabecular meshwork was determined using fluorescence microscopy by an investigator blinded to the status of the sample. The cellularity of the TM was determined on the same sections by counting DAPI positive nuclei.
Results: As expected, eyes of younger donors contained the largest number of TM cells (69.3 ±5.8 per section) while those of older, but healthy, donors contained more than glaucomatous eyes (59.8±6.5 vs 50.4±2.6; p=0.019). Likewise, we detected the largest number of TMSC in young eyes (5.84±1.58), significantly higher than in both healthy older eyes (2.79± 0.98, p=0.0012), or those with glaucoma (1.03± 0.16, p=0.0001 when compared to young eyes and p=0.012 when compared to age matched controls). Overall, we found a modest correlation between the number of TM cells and that of TMSC (R2=0.53).
Conclusions: Our data indicate a clear decline in the number of TMSC with age and a further reduction in eyes with glaucoma. This finding is consistent with the notion that the degeneration or loss of stemness of TMSC is the cause of reduced TM cellularity which, in turn, is associated with the development of elevated intraocular pressure.
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