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Rouzbeh Amini, Neda Rashidi, Syril Dorairaj, Anup Pant, Mickey Shah, Ge Zhang, Vineet Thomas; Iris Stromal Cell Nuclear Aspect Ratio Alters After Pharmacologically-induced Miosis and Mydriasis. Invest. Ophthalmol. Vis. Sci. 2018;59(9):2016. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
The irides in primary angle-closure glaucoma (PACG) patients express significantly higher level of COL1A1 gene in comparison to those in open angle glaucoma patients (Clin Experiment Ophthalmol, 44:684–92). In addition, COL1A1 gene expression is directly related to the extracellular matrix stiffness and cell nuclei deformation (Science 341:1240104). The purpose of this study was to examine how much the nuclei of normal irides deform when pupil diameter changes.
Porcine eyes were obtained from a local slaughterhouse. Irides were dissected and isolated in three groups: (i) miosis was induced using 1% pilocarpine, (ii) mydriasis was induced using 2.5% phenylephrine and 1% tropicamide, and (iii) flaccid irides were used as control. No experiment was conducted later than 4 hours post mortem. After necessary steps for fixing using 4% paraformaldehyde, paraffin embedding, and sectioning (7 µm thickness), samples were stained with DAPI. Fluorescent images were taken using a confocal microscope (Olympus FV1000). ImageJ (NIH, Bethesda, MD) was used to quantify the the nuclear aspect ratio, a metric for nuclear deformation, defined as the ratio of the semi-major axis to the semi-minor axis of an ellipse fitted to each cell nucleus. In all three groups, nuclear aspect ratios were calculated (n = 300 (100 cells per 3 irides) for each group).
The stromal cell nuclei deformed to more elongated configurations after drug stimulation as compared to control flaccid irides (typical examples in Fig. 1a–1c). In particular, nuclear aspect ratios were 1.75 ± 0.53 (mean ± standard deviation), 2.19 ± 0.79, and 2.18 ± 0.78 for the flaccid, miosis, and mydriasis groups, respectively. Both in the miosis group and in the mydriasis group, the nuclear aspect ratios were significantly higher than that of the control group (p<0.01, t-test, Fig.1d).
Our study showed that iris stromal cell nuclei deformed significantly when pupil diameter changed due to miosis or mydriasis. Recent studies have shown that irides are stiffer in PACG patients (IOVS, 2015 May: 6139). Iris stiffness is expected to influence stromal cell nuclei deformation and potentially alter the mechanical regulation of their genome. The outcomes of the current experiments combined with multi-scale computational models can be used to predict cellular deformation following extracellular matrix stiffening in the irides of PACG patients.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.
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