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Matthew Halton Smith, LaKisha Buie, Terete Borras; Generation of a Viable Matrix GLA Knock-In-Cre (MGP.KI.Cre) Mouse Reveals MGP Eye Expression Uniquely Targeted to the Trabecular Meshwork and Sclera Region Supporting the Optic Nerve Head. Invest. Ophthalmol. Vis. Sci. 2014;55(13):5665.
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© ARVO (1962-2015); The Authors (2016-present)
MGP is the tenth most abundant gene in the human trabecular meshwork (TM) tissue. MGP is a well-established inhibitor of vascular calcification. Knockout mice die at 6 wks due to massive arterial calcification making evaluation of the eye difficult. Our goal is to understand the role of MGP in the eye in vivo. We seek to determine localization of MGP expression by the generation of a viable MGP.KI.Cre mouse and crosses with ROSA26loxPSTOP.reporter mice.
The MGP WT gene of a C57BL/6 mouse was replaced with an engineered MGP by homologous recombination in C57BL/6N ES cells. The target vector was constructed by retrieving MGP DNA arms, 5.1 kb (5’,-1,972/+3,162) and 3.2 kb (3’,+3,043 post-polyA) from C57 BAC RP23-108P5. Sequences for an IRES, Cre-recombinase and neomycin gene (FTR1 flanked) were inserted between the MGP DNA termination codon and polyA. Chimeras were crossed with B6 albino mice for germ line transmission selection and with B6.ROSA26.FLP1 to remove the neomycin gene. Two MGPKI.cre/wt founders were intercrossed to generate homozygous MGPKI.cre/KI.cre. Mice from these progenies were crossed with B6.ROSA26.loxPSTOP.LacZ and B6.ROSA26.loxPSTOP.tomato. Eyes were enucleated and processed for X-gal staining and fluorescence histological evaluation.
The MGP.KI.Cre allele produces active MGP WT protein, making mice viable. Histological analyses of 4 wks old eyes from MGP.KI.Cre.LacZ litters show intense and very specific staining of the TM and posterior subchoroidal sclera, especially on the sclera region supporting the ONH. Cornea, iris, lens, ciliary body and retina were not stained. Outside the eye, deep staining was observed in the aorta and cartilage, as expected. Mice from the MGP.KI.Cre.tomato litters had their hands, feet and snout visible red.
Because TM outflow facility and ONH biomechanical scleral stress are key parameters in glaucoma, the simultaneous targeted expression of MGP to both regions suggests the high relevance of this gene in the development of the disease. Inhibition of calcification, and thus of hardening of the tissue, could represent an important common contributor to their normal physiology. This mouse offers a unique opportunity to study factors affecting together the TM and ONH, and to potentially search for a shared treatment of glaucoma.
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