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D. M. Maddox, W. L. Hicks, J. K. Naggert, P. M. Nishina; Loss of Tumor Necrosis Factors Leads to Increased Accumulation of Outer Segment Debris in Mice Homozygous for a Novel Allele of Mertk (Mertknmf12). Invest. Ophthalmol. Vis. Sci. 2008;49(13):3062. doi: https://doi.org/.
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A slowly degenerative form of peripheral retinal degeneration was identified in mice from an N-ethyl-N-nitrosourea (ENU) mutagenesis screen. Affected mice demonstrate slowly progressive thinning of the peripheral outer nuclear layer. The purpose of the study was to identify the nmf12 mutation and to characterize the progression of retinal degeneration in nmf12 mutant mice.
Homozygous nmf12 mice were mated to DBA/2J and the F1 progeny were intercrossed to produce 483 F2 mice that were phenotyped by indirect ophthalmoscopy and histology. The nmf12 locus was mapped to chromosome 2 by scanning pooled DNA samples with microsatellite markers throughout the genome. A refinement of the map position was achieved by genotyping mice recombinant between flanking markers D2Mit56 (70.5 Mb) and D2Mit423 (151.2 Mb). Progression of retinal degeneration was assessed by histological, electroretinographic (ERG) and immunohistochemical studies.
The nmf12 mutation maps to mouse chromosome 2 between markers D2Mit398 (125.5 Mb) and D2Mit224 (129.1 Mb). Mertk, a candidate gene mapping to the region was sequenced and a substitution of guanine for adenine at base pair 2147 (Ensembl) was identified. This is predicted to change histidine at amino acid 716 to argenine (H716R). Western blotting confirmed a marked decrease in MERTK protein.Histologically the nmf12 mutant mice demonstrate thinning of the peripheral retina beginning at P(45). Mice as old as 2 years of age have a preserved central retina and detectable ERG signals. Immunohistochemical studies identified an increase in Tnf in nmf12 mutant mice. To determine whether this increase in Tnf accelerated degeneration in the nmf12 mice, nmf12 mutants were crossed to triple mutant mice lacking Tnf, Lta (Tnfb) and Ltb (Tnfc). Surprisingly nmf12/nmf12; Tnfa,b,c+/- and nmf12/nmf12; Tnfa,b,c-/- mice demonstrated a more severe phenotype with marked accumulation of shed outer segment debris.
The nmf12 mutation is a novel allele of Mertk. The slow nature of retinal degeneration in these mice may allow further studies to understand the pathogenesis of Mertk mutations. Additionally, studies performed with Tnfa,b,c triple null mutant mice suggest the intriguing possibility that Tnf, as well as Lta and Ltb may be inducible factors that protect the retina from debris accumulation.
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