Identification Of A Mouse Model Of TTR Gene Gly83Arg Mutation In A Family With Hereditary Vitreous Amyloidosis

Objective: To identify whether the mouse model with Gly83 Arg using gene targeting technique is a stable model for studying the Gly83 Arg family hereditary vitreous amyloidosis and to confirm whether the mutant point is the genetic character of the disease.Methods: The SPF third-generation mice(NEO-,FLP-,TTR+/-)including 6 males and 9females were housed.10 C57BL/6 mice were in the control group.Intraperitoneal injection of 2.5% chloral hydrate(0.1ml/25g)once a week.Compound tropicamide eye drops were used for mydriasis.Mice were sacrificed immediately after vitreous opacity,then liver tissues were for Gene sequencing.The mice were randomly selected in the experimental group(Gly83Arg,n=4)and control group(C57BL/6,n=4).The samples were taken from hearts,brains,livers,kidneys and eyeballs for paraffin sections,then some of them were detected by Congo red staining and polarized light,and the others for immunohistochemistry.The expression of TTR gene mRNA was detected by fluorescence quantitative PCR.The expression of TTR protein was detected by Western Blot.Results: Missense mutation in the codon encoding glycine 83 were confirmed in the model mice,which mutated from GGC to CGC.The vitreous bodies of TTR mutant mice were positive for Congo red staining and polarized light detection,otherwise,livers,kidneys,hearts and brains were negative.Immunohistochemical results showed that the livers of TTR control group were positive and the other tissues were negative.The vitreous bodies of model mice were positive and the other tissues were negative.The results of fluorescence quantitative PCR showed that the expression of TTR mRNA in the controls were higher than in the model mice,and this difference was statistically significant(t=3.030,P=0.023).The results of Western Blot showed that the expression of TTR protein in the controls were higher than in the model mice,and this difference was statistically significant(t=3.224,P=0.018).Conclusion: 1.Gly83 Arg of TTR is a molecular character of familial hereditary vitreous amyloidosis and is manifested only in the eye.2.Mouse(C57BL/6)model of TTR Gly83 Arg was established successfully,and the strain of mice can be used for the study of family hereditary vitreous amyloidosis.