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Inducible nitric oxide synthase, Nos2, does not mediate optic neuropathy and retinopathy in the DBA/2J glaucoma model.

Libby RT, Howell GR, Pang IH, Savinova OV, Mehalow AK, Barter JW, Smith RS, Clark AF, John SW - BMC Neurosci (2007)

Bottom Line: The expression of Nos2 in the optic nerve head was analyzed at both the RNA and protein levels at different stages of disease pathogenesis.Optic nerve head Nos2 RNA levels varied and increased during moderate but decreased at early and severe stages of disease.Additionally, neither Nos2 deficiency nor aminoguanidine had any detectable affect on the glaucomatous optic nerve damage.

View Article: PubMed Central - HTML - PubMed

Affiliation: The Jackson Laboratory, Bar Harbor, ME 04609, USA. Richard_Libby@URMC.rochester.edu

ABSTRACT

Background: Nitric oxide synthase 2 (NOS2) contributes to neural death in some settings, but its role in glaucoma remains controversial. NOS2 is implicated in retinal ganglion cell degeneration in a rat glaucoma model in which intraocular pressure (IOP) is experimentally elevated by blood vessel cauterization, but not in a rat glaucoma model where IOP was elevated by injection of hypertonic saline. To test the importance of NOS2 for an inherited glaucoma, in this study we both genetically and pharmacologically decreased NOS2 activity in the DBA/2J mouse glaucoma model.

Methods: The expression of Nos2 in the optic nerve head was analyzed at both the RNA and protein levels at different stages of disease pathogenesis. To test the involvement of Nos2 in glaucomatous neurodegeneration, a allele of Nos2 was backcrossed into DBA/2J mice and the incidence and severity of glaucoma was assessed in mice of each Nos2 genotype. Additionally, DBA/2J mice were treated with the NOS2 inhibitor aminoguanidine and the disease compared to untreated mice.

Results: Optic nerve head Nos2 RNA levels varied and increased during moderate but decreased at early and severe stages of disease. Despite the presence of a few NOS2 positive cells in the optic nerve head, NOS2 protein was not substantially increased during the glaucoma. Genetic deficiency of Nos2 or aminoguanidine treatment did not alter the IOP profile of DBA/2J mice. Additionally, neither Nos2 deficiency nor aminoguanidine had any detectable affect on the glaucomatous optic nerve damage.

Conclusion: Glaucomatous neurodegeneration in DBA/2J mice does not require NOS2 activity. Further experiments involving various models are needed to assess the general importance of Nos2 in glaucoma.

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NOS2 deficiency does not alter glaucomatous neurodegeneration. (A, B) There were no gross anatomical differences between young Nos2+/+ (A) and Nos2-/- (B) nerves. (C) Eventually 80% of the optic nerves of DBA/2J mice develop severe optic nerve degeneration (Nos2+/+). (D) The complete absence of NOS2 did not prevent glaucomatous neurodegeneration as many Nos2-/- nerves had severe axonal damage. (E) Nos2 genotype had no effect on the distribution of nerve damage. Numbers of nerves by age, genotype and disease state were: (given as total number analyzed, genotype, number of that genotype with mild, moderate, severe glaucoma) 3.5 months 6 Nos2+/+ 6,0,0; 7 Nos2+/- 7,0,0;11 Nos2-/- 11,0,0; 10–12 months 53 Nos2+/+ 23,3,27; 53 Nos2+/- 21,9,23; 56 Nos2-/- 17,3,36 (P value for all comparisons > 0.3). F) Average counts of RGC layer neurons from Nos2 mice of each genotype with either severe glaucomatous damage or no detectable glaucomatous damage (total number of cells of the 8 fields counted per retina; see methods). Nos2 genotype conferred no differential protective effect to RGCs in eyes with different degrees of axon loss. RGCs comprise 45–50% of cells in the RGC layer [71] and so almost all RGCs were lost in mice with severe axon loss independent of Nos2 genotype. Percentage of surviving cells; Nos2+/+, 54.4% ± 1.5; Nos2+/-, 54.6% ± 1.5; Nos2-/-, 58.0% ± 2.5; P values for all comparisons > 0.28. Number of retinal flat mounts counted, (given as genotype, mild, severe) Nos2+/+ 9, 7; Nos2+/- 12, 8; Nos2-/- 8, 13. Bar = 100 μm.
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Figure 3: NOS2 deficiency does not alter glaucomatous neurodegeneration. (A, B) There were no gross anatomical differences between young Nos2+/+ (A) and Nos2-/- (B) nerves. (C) Eventually 80% of the optic nerves of DBA/2J mice develop severe optic nerve degeneration (Nos2+/+). (D) The complete absence of NOS2 did not prevent glaucomatous neurodegeneration as many Nos2-/- nerves had severe axonal damage. (E) Nos2 genotype had no effect on the distribution of nerve damage. Numbers of nerves by age, genotype and disease state were: (given as total number analyzed, genotype, number of that genotype with mild, moderate, severe glaucoma) 3.5 months 6 Nos2+/+ 6,0,0; 7 Nos2+/- 7,0,0;11 Nos2-/- 11,0,0; 10–12 months 53 Nos2+/+ 23,3,27; 53 Nos2+/- 21,9,23; 56 Nos2-/- 17,3,36 (P value for all comparisons > 0.3). F) Average counts of RGC layer neurons from Nos2 mice of each genotype with either severe glaucomatous damage or no detectable glaucomatous damage (total number of cells of the 8 fields counted per retina; see methods). Nos2 genotype conferred no differential protective effect to RGCs in eyes with different degrees of axon loss. RGCs comprise 45–50% of cells in the RGC layer [71] and so almost all RGCs were lost in mice with severe axon loss independent of Nos2 genotype. Percentage of surviving cells; Nos2+/+, 54.4% ± 1.5; Nos2+/-, 54.6% ± 1.5; Nos2-/-, 58.0% ± 2.5; P values for all comparisons > 0.28. Number of retinal flat mounts counted, (given as genotype, mild, severe) Nos2+/+ 9, 7; Nos2+/- 12, 8; Nos2-/- 8, 13. Bar = 100 μm.

Mentions: Some optic nerves from DBA/2J mice begin to show signs of glaucomatous neurodegeneration at 9 months of age [40]. The incidence and severity of glaucoma then increases rapidly over the next months, but not all mice develop glaucomatous nerve damage. Complete deficiency of NOS2 did not prevent homozygous mutant mice from developing severe glaucoma (Fig. 3C, D, E), showing that NOS2 is not required for glaucomatous axon degeneration in the optic nerve. Additionally, NOS2 deficiency did not lessen the amount of glaucomatous optic nerve degeneration in mice that were 10–12 months of age (Fig. 3E).


Inducible nitric oxide synthase, Nos2, does not mediate optic neuropathy and retinopathy in the DBA/2J glaucoma model.

Libby RT, Howell GR, Pang IH, Savinova OV, Mehalow AK, Barter JW, Smith RS, Clark AF, John SW - BMC Neurosci (2007)

NOS2 deficiency does not alter glaucomatous neurodegeneration. (A, B) There were no gross anatomical differences between young Nos2+/+ (A) and Nos2-/- (B) nerves. (C) Eventually 80% of the optic nerves of DBA/2J mice develop severe optic nerve degeneration (Nos2+/+). (D) The complete absence of NOS2 did not prevent glaucomatous neurodegeneration as many Nos2-/- nerves had severe axonal damage. (E) Nos2 genotype had no effect on the distribution of nerve damage. Numbers of nerves by age, genotype and disease state were: (given as total number analyzed, genotype, number of that genotype with mild, moderate, severe glaucoma) 3.5 months 6 Nos2+/+ 6,0,0; 7 Nos2+/- 7,0,0;11 Nos2-/- 11,0,0; 10–12 months 53 Nos2+/+ 23,3,27; 53 Nos2+/- 21,9,23; 56 Nos2-/- 17,3,36 (P value for all comparisons > 0.3). F) Average counts of RGC layer neurons from Nos2 mice of each genotype with either severe glaucomatous damage or no detectable glaucomatous damage (total number of cells of the 8 fields counted per retina; see methods). Nos2 genotype conferred no differential protective effect to RGCs in eyes with different degrees of axon loss. RGCs comprise 45–50% of cells in the RGC layer [71] and so almost all RGCs were lost in mice with severe axon loss independent of Nos2 genotype. Percentage of surviving cells; Nos2+/+, 54.4% ± 1.5; Nos2+/-, 54.6% ± 1.5; Nos2-/-, 58.0% ± 2.5; P values for all comparisons > 0.28. Number of retinal flat mounts counted, (given as genotype, mild, severe) Nos2+/+ 9, 7; Nos2+/- 12, 8; Nos2-/- 8, 13. Bar = 100 μm.
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Figure 3: NOS2 deficiency does not alter glaucomatous neurodegeneration. (A, B) There were no gross anatomical differences between young Nos2+/+ (A) and Nos2-/- (B) nerves. (C) Eventually 80% of the optic nerves of DBA/2J mice develop severe optic nerve degeneration (Nos2+/+). (D) The complete absence of NOS2 did not prevent glaucomatous neurodegeneration as many Nos2-/- nerves had severe axonal damage. (E) Nos2 genotype had no effect on the distribution of nerve damage. Numbers of nerves by age, genotype and disease state were: (given as total number analyzed, genotype, number of that genotype with mild, moderate, severe glaucoma) 3.5 months 6 Nos2+/+ 6,0,0; 7 Nos2+/- 7,0,0;11 Nos2-/- 11,0,0; 10–12 months 53 Nos2+/+ 23,3,27; 53 Nos2+/- 21,9,23; 56 Nos2-/- 17,3,36 (P value for all comparisons > 0.3). F) Average counts of RGC layer neurons from Nos2 mice of each genotype with either severe glaucomatous damage or no detectable glaucomatous damage (total number of cells of the 8 fields counted per retina; see methods). Nos2 genotype conferred no differential protective effect to RGCs in eyes with different degrees of axon loss. RGCs comprise 45–50% of cells in the RGC layer [71] and so almost all RGCs were lost in mice with severe axon loss independent of Nos2 genotype. Percentage of surviving cells; Nos2+/+, 54.4% ± 1.5; Nos2+/-, 54.6% ± 1.5; Nos2-/-, 58.0% ± 2.5; P values for all comparisons > 0.28. Number of retinal flat mounts counted, (given as genotype, mild, severe) Nos2+/+ 9, 7; Nos2+/- 12, 8; Nos2-/- 8, 13. Bar = 100 μm.
Mentions: Some optic nerves from DBA/2J mice begin to show signs of glaucomatous neurodegeneration at 9 months of age [40]. The incidence and severity of glaucoma then increases rapidly over the next months, but not all mice develop glaucomatous nerve damage. Complete deficiency of NOS2 did not prevent homozygous mutant mice from developing severe glaucoma (Fig. 3C, D, E), showing that NOS2 is not required for glaucomatous axon degeneration in the optic nerve. Additionally, NOS2 deficiency did not lessen the amount of glaucomatous optic nerve degeneration in mice that were 10–12 months of age (Fig. 3E).

Bottom Line: The expression of Nos2 in the optic nerve head was analyzed at both the RNA and protein levels at different stages of disease pathogenesis.Optic nerve head Nos2 RNA levels varied and increased during moderate but decreased at early and severe stages of disease.Additionally, neither Nos2 deficiency nor aminoguanidine had any detectable affect on the glaucomatous optic nerve damage.

View Article: PubMed Central - HTML - PubMed

Affiliation: The Jackson Laboratory, Bar Harbor, ME 04609, USA. Richard_Libby@URMC.rochester.edu

ABSTRACT

Background: Nitric oxide synthase 2 (NOS2) contributes to neural death in some settings, but its role in glaucoma remains controversial. NOS2 is implicated in retinal ganglion cell degeneration in a rat glaucoma model in which intraocular pressure (IOP) is experimentally elevated by blood vessel cauterization, but not in a rat glaucoma model where IOP was elevated by injection of hypertonic saline. To test the importance of NOS2 for an inherited glaucoma, in this study we both genetically and pharmacologically decreased NOS2 activity in the DBA/2J mouse glaucoma model.

Methods: The expression of Nos2 in the optic nerve head was analyzed at both the RNA and protein levels at different stages of disease pathogenesis. To test the involvement of Nos2 in glaucomatous neurodegeneration, a allele of Nos2 was backcrossed into DBA/2J mice and the incidence and severity of glaucoma was assessed in mice of each Nos2 genotype. Additionally, DBA/2J mice were treated with the NOS2 inhibitor aminoguanidine and the disease compared to untreated mice.

Results: Optic nerve head Nos2 RNA levels varied and increased during moderate but decreased at early and severe stages of disease. Despite the presence of a few NOS2 positive cells in the optic nerve head, NOS2 protein was not substantially increased during the glaucoma. Genetic deficiency of Nos2 or aminoguanidine treatment did not alter the IOP profile of DBA/2J mice. Additionally, neither Nos2 deficiency nor aminoguanidine had any detectable affect on the glaucomatous optic nerve damage.

Conclusion: Glaucomatous neurodegeneration in DBA/2J mice does not require NOS2 activity. Further experiments involving various models are needed to assess the general importance of Nos2 in glaucoma.

Show MeSH
Related in: MedlinePlus