Retinal deficiency of MMACHC does not lead to an ocular phenotype in mice

Abstract

Purpose : Methylmalonic aciduria with homocystinuria (cobalamin deficiency cblC type) is a severe but rare disease, caused by mutations in the MMACHC gene. cblC deficiency is multi-symptomatic and often includes ophthalmic manifestations such as macular degeneration, optic nerve pallor and vascular changes that may progress to complete blindness. Cobalamin supplementation can improve some symptoms, but the ocular phenotype remains progressive, suggesting that it might result from a local dysfunction of MMACHC in the retina. Here, we tested the potential ocular consequences of a pan-retinal Mmachc knockout in mice.

Methods : Expression of human MMACHC was tested in the central and peripheral retina and retinal pigment epithelium (RPE) of donor eyes and of mice by real-time PCR. A retina-specific knockout of Mmachc (retina^ΔMmachc) was generated using Pax6-Cre mice to delete E3 and E4 of Mmachc in a novel Mmachc^flox/flox mouse line. Knockout of Mmachc was confirmed on the genomic level by PCR and on the RNA level by in situ hybridization and real-time PCR. Retinal function was tested by electroretinography, Tissue integrity was investigated by light microscopy and immunofluorescence of tissue sections as well as by fundus imaging, optical coherence tomography and angiography. Metabolites were measured by LC-MS/MS and reversed-phase HPLC.

Results : MMACHC was stably expressed in human retina and RPE up to 91 years of age and in mice from day 1 to 365. In retina^ΔMmachc mice, expression of Mmachc was absent in the peripheral retina but was not affected in the central retina or the RPE. The overall reduction of Mmachc expression in the retina of retina^ΔMmachc mice was 76±4%. Retinas of knockout mice showed an elevation of cblC-related metabolites including methylmalonic acid (5.6-fold) and homocysteine (1.6-fold) that are known to accumulate also in the serum of patients. However, retinal morphology and function in knockouts was normal up to 1 year of age.

Conclusions : Our data indicate that Mmachc deficiency in the neuronal retina does not lead to ocular impairments in mice. It may be possible, that other supporting cells that do not carry the knockout allele, such as endothelial cells or the RPE, may help to nourish the cells of the retina to compensate them for their lack of Mmachc.

This is a 2021 ARVO Annual Meeting abstract.