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Authors: C Stefan , Diana Melinte Dumitrica , Carmen Ardeleanu Central Military Emergency Hospital Carol Davila- Bucharest Victor Babes Institute, Immunohistochemistry Department

VI-th National Congress of Ophthalmology Sinaia 3-6 october 2007

Major signalling molecule:neuronsimmune systemImplications: vasodilatationcontractility neurotransmissionneurotoxicityinflammationNeurodegenerative disease: glaucomaAlzheimermultiple sclerosiscardio-cerebral-vascular diseases


L-arginine nitric oxide synthase NO (NOS)NO:half-life=a few secondssoluble in both aqueous and lipid mediareadily diffuses through the cytoplasm and membrane


VI-th National Congress of Ophthalmology Sinaia 3-6 october 2007Achike, F.I., and Kwan, C.Y. 2003. Clin Exp. Pharmacol. Physiol. 30, 605

3 NOS isoformsNOS-1 (neuronal NOS /nNOS)NOS-2 (inducible NOS /iNOS)NOS-3 (endothelial NOS /eNOS)

VI-th National Congress of Ophthalmology Sinaia 3-6 october 2007Bredt, DS, Snyder, SH (1994) Nitric oxide: a physiologic messenger molecule Annu Rev Bioche 63,175-195

VI-th National Congress of Ophthalmology Sinaia 3-6 october 2007Buerk, DG, Riva, CE, Cranstoun, SD (1996) Nitric oxide has a vasodilatory role in cat optic nerve head during flicker stimul Microvasc Re 52,13-26Neufeld, AH, Sawada, S, Becker, B. (1999) Inhibition of nitric oxide synthase-2 by aminoguanidine provides neuroprotection of retinal ganglion cells in a rat model of glaucom Proc Natl Acad Sci US 96,9944-9948Neufeld, AH (1998) New conceptual approaches for pharmacological neuroprotection in glaucomatous neuronal degeneratio J Glaucom 7,434-438

constitutiveCa dependentrole NOS1++Neurotoxicity NOS2--NeurodegenerationNOS3++NeuroprotectionVasodilatation

NOS1, NOS2: neurotoxicity, neurodegenerationNeufeld, AH, Hernandez, MR, Gonzalez, M. (1997) Nitric oxide synthase in the human glaucomatous optic nerve hea Arch Ophthalmo 115,497-503 20% of familial POAG altered NOS3 expression and NO synthesisTunny TJ, Richardson KA, Clark CV. Association study of the 5' flanking regions of endothelial-nitric oxide synthase and endothelin-1 genes in familial primary open-angle glaucoma. Clin Exp Pharmacol Physiol. 1998;25:2629

inhibition of NOS reduces choroidal blood flowSchmetterer L, Krejcy K, Kastner J, et al. The effect of systemic nitric oxide-synthase inhibition on ocular fundus pulsations in man. Exp Eye Res. 1997;64:305312


Glaucoma patients with nitrate therapy less progression of optic neuropathy and visual field lossLuksch A, Polska E, Imhof A, et al. Role of NO in choroidal blood flow regulation during isometric exercise in healthy humans. Invest Ophthalmol Vis Sci 2003;44:7349. NO: nitro vasodilatation + contractility of trabecular meshwork IOP NO: contra-apoptotic effect neuroprotection


To investigate the activity of the nitric oxide synthase (NOS) in patients with primary open angle glaucoma (POAG)


Observational, prospective, 9 months, clinical studyOne group: 9 patients with POAGInclusions criteria: no ocular or general associated pathologywithout any topical treatment in presenthomogeneous about sex and age distribution (mean age=645years, :=4:5)


Trabeculectomy (the same surgeon) NOS immunohistochymic analyse:the presencethe localisation the distribution


EnVision System (indirect polymeric immunohistochymic analyse):

10% formol fixationparaffin inclusion (58-60)4 m sectionsincubation over-night incubation with peroxidase blocking reagent (5 min)incubation with primary antibody (30 min for increased sensitivity)application of enzyme labelled polymer (incubation 5-10 min) application of the substrate chromogen (incubation 5-10 min)


brown colorationVI-th National Congress of Ophthalmology Sinaia 3-6 october 2007

VI-th National Congress of Ophthalmology Sinaia 3-6 october 2007Presence: +/ -Localisation: +/ -Distribution: +++/ ++/ +

Casepresentlocalisationlocalisationdistributiondistributionendothelial cellfibroblastendothelial cellfibroblast1+++++++2++++++3++++++4-----5+++++++6++++++7_----8++++++++9+++++++

NO - present in trabecular meshwork

NO - modulate the outflow resistance

NO future therapies


Nitric oxide donating PG F2 alpha analogues: PF-03187207 new experimental medicine for glaucoma (improved ability to lower IOP) (Nic Ox and Pfizer)


Thank you!


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