Citation:Li M,Huang JF,Li Y,Shen J,Yang LJ,Chen Q,Zhang QP,Yi XN.Dynamic changes of inducible nitric oxide synthase expression in rat’s retina and its role on blood-retinal barrier injury after acute high intraocular pressure.Int J Ophthalmol 2022;15(7):1053-1061,doi:10.18240/ijo.2022.07.03
Dynamic changes of inducible nitric oxide synthase expression in rat’s retina and its role on blood-retinal barrier injury after acute high intraocular pressure
Received:June 09, 2021  Revised:February 21, 2022
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DOI:10.18240/ijo.2022.07.03
Key Words:inducible nitric oxide synthase  acute high intraocular pressure  blood-retinal barrier  ZO-1
Fund Project:Supported by the National Natural Science Foundation Project (No.81660217); National College Students Innovation and Entrepreneurship Training Program Project (No.201911810004).
                       
AuthorInstitution
Min Li College of Basic Medicine and Life Sciences, Hainan Medical University, Key Laboratory of Tropical Brain Research and Translation of Hainan Province, Haikou , Hainan Province, China; Department of Human Anatomy and Neurobiology, School of Basic Medical Sciences, Central South University, Changsha , Hunan Province, China
Ju-Fang Huang Department of Human Anatomy and Neurobiology, School of Basic Medical Sciences, Central South University, Changsha , Hunan Province, China
Yi Li Grade 18 Clinical Bachelor of Clinical College of Hainan Medical University, Haikou , Hainan Province, China
Jing Shen Department of Ophthalmology, the First Affiliated Hospital of Hainan Medical University, Haikou , Hainan Province, China
Lu-Jia Yang Grade 18 Clinical Bachelor of Clinical College of Hainan Medical University, Haikou , Hainan Province, China
Qian Chen Grade 18 Clinical Bachelor of Clinical College of Hainan Medical University, Haikou , Hainan Province, China
Quan-Peng Zhang College of Basic Medicine and Life Sciences, Hainan Medical University, Key Laboratory of Tropical Brain Research and Translation of Hainan Province, Haikou , Hainan Province, China
Xi-Nan Yi College of Basic Medicine and Life Sciences, Hainan Medical University, Key Laboratory of Tropical Brain Research and Translation of Hainan Province, Haikou , Hainan Province, China
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Abstract:
      AIM: To clarify the role of inducible nitric oxide synthase (iNOS) in blood-retinal barrier (BRB) injury after acute high intraocular pressure (IOP) in rats.

    METHODS: Forty-two Sprague-Dawley (SD) rats were randomized into 7 groups [control (Cont), 3, 6, 12, 24, 48, and 72h, n=6]. Except Cont group, other groups’ retina tissue was obtained at corresponding time points after a model of acute high IOP have been established in rats. The expression of iNOS and tight junction protein zonula occludens (ZO)-1 was detected by Western blotting. Evans blue (EB; 3% ) was injected into the great saphenous vein to detect the leakage of EB by spectrophotometer. Nine rats were divided into Cont, 6h, 12h groups, the expression of iNOS was localized by immunofluorescence. In order to verify the role of iNOS in the damage to BRB, thirty-six rats were randomly divided into 4 groups [Cont, Cont+inhibitor (Inh), 6h and 6h+Inh, n=9]. After treatment with the iNOS-specific inhibitor 1400W, the expression of iNOS and ZO-1 and the leakage of BRB were detected again.

    RESULTS: The immunofluorescence results showed that the expression of iNOS was observed in the Cont group and 6h group, but not in the 12h group. iNOS was mainly expressed in the retinal nerve fiber layer, ganglion cell layer and inner nuclear layer and that it did not colocalize with the retinal ganglion cell marker NeuN but was co-expressed with the vascular endothelial cell marker CD31. Western blotting showed that in the early period (3h, 6h) after acute high IOP, the expression of iNOS was upregulated, then the down-regulation of iNOS were tested in the follow-up timing spots. ZO-1 expression showed a continuous down-regulation after 6h. The quantitative results for EB showed that the amount of EB leakage began to increase at 3h after acute high IOP. At 6h, the leakage of EB was lower, but at 12h, the leakage of EB was highest, after which it gradually recovered but remained higher than that in the Cont group. The expression of iNOS was down-regulated after 1400W treatment. ZO-1 expression was not significantly changed in the Cont+Inh group and the 6h group, and significantly down-regulated in the 6h+Inh group, and the leakage of EB was significantly increased after 1400W treatment.

    CONCLUSION: These results suggest that the upregulation of iNOS expression in the early stage after acute high IOP may have a protective effect on BRB injury.

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