Human GLUD1 (Glutamate dehydrogenase 1, mitochondrial) ELISA Kit

Liver fibrosis is the excessive accumulation of extracellular matrix proteins, which is mainly caused by accumulation of activated hepatic stellate cells (HSCs). The mechanisms of activation and proliferation of HSCs, two key events after liver damage, have been studied for many years. Here we report a novel pathway to control HSCs by regulating glutamine metabolism. We demonstrated that the proliferation of HSCs is critically dependent on glutamine that is used to generate alpha-ketoglutarate (alpha-KG) and non-essential amino acid (NEAA). In addition, both culture- and in vivo-activated HSCs have increased glutamine utilization and increased expression of genes related to glutamine metabolism, including GLS (glutaminase), aspartate transaminase (GOT1) and glutamate dehydrogenase (GLUD1). Inhibition of these enzymes, as well as glutamine depletion, had a significant inhibitory effect on HSCs activation. In addition to providing energy expenditure, conversion of glutamine to proline is enhanced. The pool of free proline may also be increased via downregulation of PDX expression. Hedgehog signaling plays an important role in the regulation of glutamine metabolism, as well as TGF-beta 1, c-Myc, and Ras signalings, via transcriptional upregulation and repression of key metabolic enzymes in this pathway. Finally, changes in glutamine metabolism were also found in mouse liver tissue following CCI4-induced acute injury. Conclusion: Glutamine metabolism plays an important role in regulating the proliferation and activation of HSCs. Strategies that are targeted at glutamine metabolism may represent a novel therapeutic approach to the treatment of liver fibrosis.

Activating mutations in the GLUD1 gene, which encodes glutamate dehydrogenase (GDH), result in the hyperinsulinism-hyperammonemia syndrome. GDH is an allosterically regulated enzyme responsible for amino acid-mediated insulin secretion via the oxidative deamination of glutamate to 2-oxoglutarate, leading to ATP production and insulin release. This study characterizes a novel combination of mutations in GLUD1 found in a neonate who presented on the first day of life with severe hypoglycemia, hyperammonemia, and seizures. Mutation analysis revealed a novel frameshift mutation (c.37delC) inherited from the asymptomatic mother that results in a truncated protein and a de novo activating mutation (p.S445L) close to the GTP binding site that has previously been reported. GTP inhibition of GDH enzyme activity in 293T cells expressing the p.S445L or wild-type GDH showed that the half-maximal inhibitory concentration (IC50) for GTP was approximately 800 times higher for p.S445L compared to wild type. GTP inhibition of GDH activity in lymphoblasts from the patient, from a heterozygote for the p.S445L mutation, and in wild-type lymphoblasts showed that the IC50 for GTP of the patient was approximately 200 times that of wild type and 7 times that of heterozygote. However, while the patient had a loss of GTP inhibition of GDH that was more severe than that of heterozygotes, the patient's clinical phenotype is similar to typical heterozygous mutations of GDH. This is the first time we have observed a functionally homozygous activating mutation of GDH in a human.