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Loss of Ikbkap Causes Slow, Progressive Retinal Degeneration in a Mouse Model of Familial Dysautonomia

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ABSTRACT

Familial dysautonomia (FD) is an autosomal recessive congenital neuropathy that is caused by a mutation in the gene for inhibitor of kappa B kinase complex-associated protein (IKBKAP). Although FD patients suffer from multiple neuropathies, a major debilitation that affects their quality of life is progressive blindness. To determine the requirement for Ikbkap in the developing and adult retina, we generated Ikbkap conditional knockout (CKO) mice using a TUBA1a promoter-Cre (Tα1-Cre). In the retina, Tα1-Cre expression is detected predominantly in retinal ganglion cells (RGCs). At 6 months, significant loss of RGCs had occurred in the CKO retinas, with the greatest loss in the temporal retina, which is the same spatial phenotype observed in FD, Leber hereditary optic neuropathy, and dominant optic atrophy. Interestingly, the melanopsin-positive RGCs were resistant to degeneration. By 9 months, signs of photoreceptor degeneration were observed, which later progressed to panretinal degeneration, including RGC and photoreceptor loss, optic nerve thinning, Müller glial activation, and disruption of layers. Taking these results together, we conclude that although Ikbkap is not required for normal development of RGCs, its loss causes a slow, progressive RGC degeneration most severely in the temporal retina, which is later followed by indirect photoreceptor loss and complete retinal disorganization. This mouse model of FD is not only useful for identifying the mechanisms mediating retinal degeneration, but also provides a model system in which to attempt to test therapeutics that may mitigate the loss of vision in FD patients.

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Activation of microglia was observed in the Ikbkap CKO optic nerves. Maximum-intensity z-stack projections of longitudinal optic nerve sections were stained with microglial/macrophage marker Iba1 in 9-month control (A) and mutant (B) optic nerves. Representative images are shown. Microglia in control optic nerves had thin ramified branches (arrowheads in Aʹ inset); microglia in mutant optic nerves had ameboid morphology (arrows in Bʹ) indicative of inflammatory response. White boxes in A and B indicate areas imaged at higher magnification in Aʹ and Bʹ, respectively. (C) The number of Iba1-positive pixels that met the criteria was counted and divided by the total number of pixels in the volume of the area, as described in Materials and Methods. The data show that mutant microglia occupied increased areas in the optic nerve, suggesting the presence of inflammatory response. *p = 0.05 with a two-tailed t-test (n = 6 for control and n = 5 for mutant). Scale bars, 100 μm (A and B) and 50 μm (Aʹ and Bʹ).
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Figure 6: Activation of microglia was observed in the Ikbkap CKO optic nerves. Maximum-intensity z-stack projections of longitudinal optic nerve sections were stained with microglial/macrophage marker Iba1 in 9-month control (A) and mutant (B) optic nerves. Representative images are shown. Microglia in control optic nerves had thin ramified branches (arrowheads in Aʹ inset); microglia in mutant optic nerves had ameboid morphology (arrows in Bʹ) indicative of inflammatory response. White boxes in A and B indicate areas imaged at higher magnification in Aʹ and Bʹ, respectively. (C) The number of Iba1-positive pixels that met the criteria was counted and divided by the total number of pixels in the volume of the area, as described in Materials and Methods. The data show that mutant microglia occupied increased areas in the optic nerve, suggesting the presence of inflammatory response. *p = 0.05 with a two-tailed t-test (n = 6 for control and n = 5 for mutant). Scale bars, 100 μm (A and B) and 50 μm (Aʹ and Bʹ).

Mentions: We also analyzed the optic nerves (RGC axon bundle) of 6- and 9-month mutant and control mice. Although the circumference of the optic nerves was slightly smaller in both 6- and 9-month mutant optic nerves compared with controls, the difference was not significant (data not shown). A microglial/macrophage marker, Iba1 (Ito et al., 2001), showed no difference in microglial number and morphology in the 6-month mutant and control optic nerves (data not shown). However, by 9 months, the optic nerves of mutant animals demonstrated evidence of inflammation (Fig. 6; Ito et al., 2001). Whereas control optic nerves showed microglia with thin ramified branches, mutant optic nerves had microglia with ameboid morphology (Fig. 6A, B). Our analysis shows increased infiltration of activated microglia in the optic nerves of mutant mice (Fig. 6C).


Loss of Ikbkap Causes Slow, Progressive Retinal Degeneration in a Mouse Model of Familial Dysautonomia
Activation of microglia was observed in the Ikbkap CKO optic nerves. Maximum-intensity z-stack projections of longitudinal optic nerve sections were stained with microglial/macrophage marker Iba1 in 9-month control (A) and mutant (B) optic nerves. Representative images are shown. Microglia in control optic nerves had thin ramified branches (arrowheads in Aʹ inset); microglia in mutant optic nerves had ameboid morphology (arrows in Bʹ) indicative of inflammatory response. White boxes in A and B indicate areas imaged at higher magnification in Aʹ and Bʹ, respectively. (C) The number of Iba1-positive pixels that met the criteria was counted and divided by the total number of pixels in the volume of the area, as described in Materials and Methods. The data show that mutant microglia occupied increased areas in the optic nerve, suggesting the presence of inflammatory response. *p = 0.05 with a two-tailed t-test (n = 6 for control and n = 5 for mutant). Scale bars, 100 μm (A and B) and 50 μm (Aʹ and Bʹ).
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Figure 6: Activation of microglia was observed in the Ikbkap CKO optic nerves. Maximum-intensity z-stack projections of longitudinal optic nerve sections were stained with microglial/macrophage marker Iba1 in 9-month control (A) and mutant (B) optic nerves. Representative images are shown. Microglia in control optic nerves had thin ramified branches (arrowheads in Aʹ inset); microglia in mutant optic nerves had ameboid morphology (arrows in Bʹ) indicative of inflammatory response. White boxes in A and B indicate areas imaged at higher magnification in Aʹ and Bʹ, respectively. (C) The number of Iba1-positive pixels that met the criteria was counted and divided by the total number of pixels in the volume of the area, as described in Materials and Methods. The data show that mutant microglia occupied increased areas in the optic nerve, suggesting the presence of inflammatory response. *p = 0.05 with a two-tailed t-test (n = 6 for control and n = 5 for mutant). Scale bars, 100 μm (A and B) and 50 μm (Aʹ and Bʹ).
Mentions: We also analyzed the optic nerves (RGC axon bundle) of 6- and 9-month mutant and control mice. Although the circumference of the optic nerves was slightly smaller in both 6- and 9-month mutant optic nerves compared with controls, the difference was not significant (data not shown). A microglial/macrophage marker, Iba1 (Ito et al., 2001), showed no difference in microglial number and morphology in the 6-month mutant and control optic nerves (data not shown). However, by 9 months, the optic nerves of mutant animals demonstrated evidence of inflammation (Fig. 6; Ito et al., 2001). Whereas control optic nerves showed microglia with thin ramified branches, mutant optic nerves had microglia with ameboid morphology (Fig. 6A, B). Our analysis shows increased infiltration of activated microglia in the optic nerves of mutant mice (Fig. 6C).

View Article: PubMed Central - HTML - PubMed

ABSTRACT

Familial dysautonomia (FD) is an autosomal recessive congenital neuropathy that is caused by a mutation in the gene for inhibitor of kappa B kinase complex-associated protein (IKBKAP). Although FD patients suffer from multiple neuropathies, a major debilitation that affects their quality of life is progressive blindness. To determine the requirement for Ikbkap in the developing and adult retina, we generated Ikbkap conditional knockout (CKO) mice using a TUBA1a promoter-Cre (Tα1-Cre). In the retina, Tα1-Cre expression is detected predominantly in retinal ganglion cells (RGCs). At 6 months, significant loss of RGCs had occurred in the CKO retinas, with the greatest loss in the temporal retina, which is the same spatial phenotype observed in FD, Leber hereditary optic neuropathy, and dominant optic atrophy. Interestingly, the melanopsin-positive RGCs were resistant to degeneration. By 9 months, signs of photoreceptor degeneration were observed, which later progressed to panretinal degeneration, including RGC and photoreceptor loss, optic nerve thinning, Müller glial activation, and disruption of layers. Taking these results together, we conclude that although Ikbkap is not required for normal development of RGCs, its loss causes a slow, progressive RGC degeneration most severely in the temporal retina, which is later followed by indirect photoreceptor loss and complete retinal disorganization. This mouse model of FD is not only useful for identifying the mechanisms mediating retinal degeneration, but also provides a model system in which to attempt to test therapeutics that may mitigate the loss of vision in FD patients.

No MeSH data available.


Related in: MedlinePlus