A G1528C Hadha knock-in mouse model recapitulates aspects of human clinical phenotypes for long-chain 3-hydroxyacyl-CoA dehydrogenase deficiency.

Long chain 3-hydroxyacyl-CoA dehydrogenase deficiency (LCHADD) is a fatty acid oxidation disorder (FAOD) caused by a pathogenic variant, c.1528 G > C, in HADHA encoding the alpha subunit of trifunctional protein (TFPα). Individuals with LCHADD develop chorioretinopathy and peripheral neuropathy not observed in other FAODs in addition to the more ubiquitous symptoms of hypoketotic hypoglycemia, rhabdomyolysis and cardiomyopathy. We report a CRISPR/Cas9 generated knock-in murine model of G1528C in Hadha that recapitulates aspects of the human LCHADD phenotype. Homozygous pups are less numerous than expected from Mendelian probability, but survivors exhibit similar viability with wildtype (WT) littermates. Tissues of LCHADD homozygotes express TFPα protein, but LCHADD mice oxidize less fat and accumulate plasma 3-hydroxyacylcarnitines compared to WT mice. LCHADD mice exhibit lower ketones with fasting, exhaust earlier during treadmill exercise and develop a dilated cardiomyopathy compared to WT mice. In addition, LCHADD mice exhibit decreased visual performance, decreased cone function, and disruption of retinal pigment epithelium. Neurological function is affected, with impaired motor function during wire hang test and reduced open field activity. The G1528C knock-in mouse exhibits a phenotype similar to that observed in human patients; this model will be useful to explore pathophysiology and treatments for LCHADD in the future.

A Ketogenic & Low-Protein Diet Slows Retinal Degeneration in rd10 Mice.

Purpose: Treatments that delay retinal cell death regardless of genetic causation are needed for inherited retinal degeneration (IRD) patients. The ketogenic diet is a high-fat, low-carbohydrate diet, used to treat epilepsy, and has beneficial effects for neurodegenerative diseases. This study aimed to determine whether the ketogenic diet could slow retinal degeneration. Methods: Early weaned, rd10 and wild-type (WT) mice were placed on either standard chow, a ketogenic diet, or a ketogenic & low-protein diet. From postnatal day (PD) 23 to PD50, weight and blood ß-hydroxybutyrate levels were recorded. Retinal thickness, retinal function, and visual performance were measured via optical coherence tomography, electroretinography (ERG), and optokinetic tracking (OKT). At PD40, serum albumin, rhodopsin protein, and phototransduction gene expression were measured. Results: Both ketogenic diets elicited a systemic induction of ketosis. However, rd10 mice on the ketogenic & low-protein diet had significant increases in photoreceptor thickness, ERG amplitudes, and OKT thresholds, whereas rd10 mice on the ketogenic diet showed no photoreceptor preservation. In both rd10 and WT mice, the ketogenic & low-protein diet was associated with abnormal weight gain and decreases in serum albumin levels, 27% and 56%, respectively. In WT mice, the ketogenic & low-protein diet was also associated with an ∼20% to 30% reduction in ERG amplitudes. Conclusions: The ketogenic & low-protein diet slowed retinal degeneration in a clinically relevant IRD model. In WT mice, the ketogenic & low-protein diet was associated with a decrease in phototransduction and serum albumin, which could serve as a protective mechanism in the rd10 model. Although ketosis was associated with protection, its role remains unclear. Translational Relevance: Neuroprotective mechanisms associated with the ketogenic & low-protein diet have potential to slow retinal degeneration.

Suppression of cGMP-Dependent Photoreceptor Cytotoxicity With Mycophenolate Is Neuroprotective in Murine Models of Retinitis Pigmentosa.

Purpose: To determine the effect of mycophenolate mofetil (MMF) on retinal degeneration on two mouse models of retinitis pigmentosa. Methods: Intraperitoneal injections of MMF were administered daily in rd10 and c57 mice starting at postoperative day 12 (P12) and rd1 mice starting at P8. The effect of MMF was assessed with optical coherence tomography, immunohistochemistry, electroretinography, and OptoMotry. Whole retinal cyclic guanosine monophosphate (cGMP) and mycophenolic acid levels were quantified with mass spectrometry. Photoreceptor cGMP cytotoxicity was evaluated with cell counts of cGMP immunostaining. Results: MMF treatment significantly delays the onset of retinal degeneration and cGMP-dependent photoreceptor cytotoxicity in rd10 and rd1 mice, albeit a more modest effect in the latter. In rd10 mice, treatment with MMF showed robust preservation of the photoreceptors up to P22 with associated suppression of cGMP immunostaining and microglial activation; The neuroprotective effect diminished after P22, but outer retinal thickness was still significantly thicker by P35 and OptoMotry response was significantly better up to P60. Whereas cGMP immunostaining of the photoreceptors were present in rd10 and rd1 mice, hyperphysiological whole retinal cGMP levels were observed only in rd1 mice. Conclusions: Early treatment with MMF confers potent neuroprotection in two animal models of RP by suppressing the cGMP-dependent common pathway for photoreceptor cell death. The neuroprotective effect of MMF on cGMP-dependent cytotoxicity occurs independently of the presence of hyperphysiological whole retinal cGMP levels. Thus our data suggest that MMF may be an important new class of neuroprotective agent that could be useful in the treatment of patients with RP.