Modeling Phenotypic Heterogeneity of Glycogen Storage Disease Type 1a Liver Disease in Mice by Somatic CRISPR/CRISPR-associated protein 9-Mediated Gene Editing.

BACKGROUND AND AIMS: Patients with glycogen storage disease type 1a (GSD-1a) primarily present with life-threatening hypoglycemia and display severe liver disease characterized by hepatomegaly. Despite strict dietary management, long-term complications still occur, such as liver tumor development. Variations in residual glucose-6-phosphatase (G6PC1) activity likely contribute to phenotypic heterogeneity in biochemical symptoms and complications between patients. However, lack of insight into the relationship between G6PC1 activity and symptoms/complications and poor understanding of the underlying disease mechanisms pose major challenges to provide optimal health care and quality of life for GSD-1a patients. Currently available GSD-1a animal models are not suitable to systematically investigate the relationship between hepatic G6PC activity and phenotypic heterogeneity or the contribution of gene-gene interactions (GGIs) in the liver. APPROACH AND RESULTS: To meet these needs, we generated and characterized a hepatocyte-specific GSD-1a mouse model using somatic CRISPR/CRISPR-associated protein 9 (Cas9)-mediated gene editing. Hepatic G6pc editing reduced hepatic G6PC activity up to 98% and resulted in failure to thrive, fasting hypoglycemia, hypertriglyceridemia, hepatomegaly, hepatic steatosis (HS), and increased liver tumor incidence. This approach was furthermore successful in simultaneously modulating hepatic G6PC and carbohydrate response element-binding protein, a transcription factor that is activated in GSD-1a and protects against HS under these conditions. Importantly, it also allowed for the modeling of a spectrum of GSD-1a phenotypes in terms of hepatic G6PC activity, fasting hypoglycemia, hypertriglyceridemia, hepatomegaly and HS. CONCLUSIONS: In conclusion, we show that somatic CRISPR/Cas9-mediated gene editing allows for the modeling of a spectrum of hepatocyte-borne GSD-1a disease symptoms in mice and to efficiently study GGIs in the liver. This approach opens perspectives for translational research and will likely contribute to personalized treatments for GSD-1a and other genetic liver diseases.

Quantification of Biventricular Function and Morphology by Cardiac Magnetic Resonance Imaging in Mice with Pulmonary Artery Banding.

Right ventricular (RV) function and failure are major determinants of outcome in acquired and congenital heart diseases, including pulmonary hypertension. Assessment of RV function and morphology is complex, partly due to the complex shape of the RV. Currently, cardiac magnetic resonance (CMR) imaging is the golden standard for noninvasive assessment of RV function and morphology. The current protocol describes CMR imaging in a mouse model of RV pressure load induced by pulmonary artery banding (PAB). PAB is performed by placing a 6-0 suture around the pulmonary artery over a 23 G needle. The PAB gradient is determined using echocardiography at 2 and 6 weeks. At 6 weeks, the right and left ventricular morphology and function is assessed by measuring both end-systolic and end-diastolic volumes and mass by ten to eleven cine slices 1 mm thick using a 9.4 T magnetic resonance imaging scanner equipped with a 1,500 mT/m gradient. Representative results show that PAB induces a significant increase in RV pressure load, with significant effects on biventricular morphology and RV function. It is also shown that at 6 weeks of RV pressure load, cardiac output is maintained. Presented here is a reproducible protocol for the quantification of biventricular morphology and function in a mouse model of RV pressure load and may serve as a method for experiments exploring determinants of RV remodeling and dysfunction.