Citrin/Mitochondrial Glycerol-3-phosphate Dehydrogenase Double Knock-out Mice Recapitulate Features of Human Citrin Deficiency

Abstract

Citrin is the liver-type mitochondrial aspartate-glutamate carrier that participates in urea, protein, and nucleotide biosynthetic pathways by supplying aspartate from mitochondria to the cytosol.Citrin also plays a role in transporting cytosolic NADH reducing equivalents into mitochondria as a component of the malate-aspartate shuttle. In humans, loss-of-function mutations in the SLC25A13 gene encoding citrin cause both adult-onset type II citrullinemia and neonatal intrahepatic cholestasis, collectively referred to as human citrin deficiency. Citrin knock-out mice fail to display features of human citrin deficiency. Based on the hypothesis that an enhanced glycerol phosphate shuttle activity may be compensating for the loss of citrin function in the mouse, we have generated mice with a combined disruption of the genes for citrin and mitochondrial glycerol 3-phosphate dehydrogenase. The resulting double knock-out mice demonstrated citrullinemia, hyperammonemia that was further elevated by oral sucrose administration, hypoglycemia, and a fatty liver, all features of human citrin deficiency. An increased hepatic lactate/pyruvate ratio in the double knock-out mice compared with controls was also further elevated by the oral sucrose administration, suggesting that an altered cytosolic NADH/NAD ؉ ratio is closely associated with the hyperammonemia observed. Microarray analyses identified over 100 genes that were differentially expressed in the double knock-out mice compared with wild-type controls, revealing genes potentially involved in compensatory or downstream effects of the combined mutations. Together, our data indicate that the more severe phenotype present in the citrin/mitochondrial glycerol-3-phosphate dehydrogenase double knock-out mice represents a more accurate model of human citrin deficiency than citrin knock-out mice.

Citrin deficiency as a cause of chronic liver disorder mimicking non-alcoholic fatty liver disease

“…A large amount of citrate is converted to acetyl-CoA by ATP-citrate lyase, further leading to overproduction of fatty acids in hepatocytes ( Figure 3C). In fact, elevated mRNA levels of ATP-citrate lyase have been confirmed in livers of citrin/mitochondrial glycerol-3-phosphate dehydrogenase double knockout mouse, a suitable model of human citrin deficiency [12]. The mRNA levels of CD36 (fatty acid translocase) were also elevated in these mouse livers [12], suggesting increased fatty acid uptake capacity into hepatocytes.…”

“…In fact, elevated mRNA levels of ATP-citrate lyase have been confirmed in livers of citrin/mitochondrial glycerol-3-phosphate dehydrogenase double knockout mouse, a suitable model of human citrin deficiency [12]. The mRNA levels of CD36 (fatty acid translocase) were also elevated in these mouse livers [12], suggesting increased fatty acid uptake capacity into hepatocytes. Based on these findings, enhancement of de novo lipogenesis and fatty acid uptake into hepatocytes are considered to be key events in the development of hepatic steatosis in patients with citrin deficiency.…”

“…Generally speaking, the prognosis of patients with CTLN2 is not favorable after the onset of encephalopathy, and liver transplantation is required for severe CTLN2 which is refractory to various ammonia-lowering and neuroprotective therapies. Furthermore, certain types of conventional treatments for brain edema, such as intravenous administration of hyperosmotic and high sugar solutions, have been reported to aggravate encephalopathy [2,10,11], and the mechanism of toxicity of carbohydrate overload for citrin deficiency has been recently clarified using mouse models [12]. Thus, early diagnosis of citrin deficiency and CTLN2 and appropriate treatment might improve prognosis.…”

Novel two-step derivation method for the synchronous analysis of inherited metabolic disorders using urine

“…Citrin has several functions, which include participation in the biosynthesis of urea, protein and nucleic acids (35), and the provision of adequate NADH for mitochondrial membranes (36). Therefore, citrin dysfunction affects synthetic or catabolic pathways, leading to a series of metabolic dysfunctions and varying degrees of clinical manifestations (37).…”

Blood glucose and insulin and correlation of SLC25A13 mutations with biochemical changes in NICCD patients

“…5,10,19 Retrospective analysis of biochemical indices in 75 NICCD cases carried out by Ohura et al 20 in Japan had found 18 cases (24%) with hypoglycemia, and concluded that hypoglycemia is common among neonatalonset NICCD. By using gene knockout technology, Saheki et al 21 who established mice with SLC25A13 deficiency had shown hypoglycemia after fasting, in comparison to wild type mice. In mainland China, Zhang et al 22 also discussed this issue, in her study of 20 NICCD cases, hypoglycemia was found in NICCD patients compared to a control group (P < 0.05) with the mean value of (2.78 AE 0.63) mmol/L.…”