Department of Pediatrics, University Medical Center Groningen, University of Groningen, Groningen, The Netherlands.
Section of Metabolic Diseases, Beatrix Children's Hospital, University Medical Center Groningen, Groningen, The Netherlands.
Hepatology. 2021 Nov;74(5):2491-2507. doi: 10.1002/hep.32022. Epub 2021 Aug 15.
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.
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.
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.
糖原贮积病 1a 型（GSD-1a）患者主要表现为危及生命的低血糖，并伴有肝肿大为特征的严重肝脏疾病。尽管进行了严格的饮食管理，但仍会出现长期并发症，如肝肿瘤的发展。葡萄糖-6-磷酸酶（G6PC1）活性的差异可能导致患者生化症状和并发症的表型异质性。然而，缺乏对 G6PC1 活性与症状/并发症之间关系的深入了解，以及对潜在疾病机制的理解不足，给 GSD-1a 患者提供最佳的医疗保健和生活质量带来了重大挑战。目前可用的 GSD-1a 动物模型并不适合系统地研究肝 G6PC 活性与表型异质性之间的关系，也不适合研究基因-基因相互作用（GGI）在肝脏中的作用。
为了满足这些需求，我们使用体细胞核移植 CRISPR/Cas9 介导的基因编辑技术创建并表征了一种肝细胞特异性 GSD-1a 小鼠模型。肝 G6pc 的编辑使肝 G6PC 活性降低了 98%，导致生长不良、空腹低血糖、高三酰甘油血症、肝肿大、肝脂肪变性（HS）和肝脏肿瘤发生率增加。这种方法还成功地同时调节肝 G6PC 和碳水化合物反应元件结合蛋白，后者是一种在 GSD-1a 中被激活并在这些条件下防止 HS 的转录因子。重要的是，它还允许在肝 G6PC 活性、空腹低血糖、高三酰甘油血症、肝肿大和 HS 方面对 GSD-1a 表型进行建模。
总之，我们证明了体细胞核移植 CRISPR/Cas9 介导的基因编辑允许在小鼠中对一系列肝细胞携带的 GSD-1a 疾病症状进行建模，并有效地研究肝脏中的 GGI。这种方法为转化研究开辟了前景，并可能有助于 GSD-1a 和其他遗传性肝脏疾病的个体化治疗。
