Pathomechanism of fever-induced liver failure in NBAS deficiency and treatment effect of NAC: observations in vitro and in vivo

Abstract

Pathogenic variants in neuroblastoma amplified sequence (NBAS) gene causes infantile liver failure type 2 (IFLS2; MIM 616483), characterised by recurrent episodes of liver failure triggered by febrile infections. The underlying pathophysiological mechanisms remain incompletely understood. With this work we try to shed light on the pathomechanism and propose a potential therapeutic option. METHODS: For in vitro analyses, human skin fibroblasts were obtained from three individuals with ILFS2 and one healthy control. Cells were cultivated at 37°C or 40°C. Western blots were performed to assess NBAS protein and its interaction partners. Furthermore, immunofluorescence and electron microscopy were used to examine morphological changes associated with endoplasmic reticulum (ER) stress. Apoptosis was measured using flow cytometry. The effects of N-acetylcysteine (NAC) treatment were analysed not only in cultured fibroblasts but also in vivo retrospectively in 16 affected individuals. RESULTS: We demonstrate that elevated temperature induces ER stress in fibroblasts from individuals with NBAS variants, leading to increased reactive oxygen species (ROS) production and apoptosis. Treatment with the antioxidant NAC, an established therapeutic in acetaminophen-induced liver failure, effectively mitigated oxidative stress and reduced apoptosis in vitro. In a retrospective clinical analysis, there was a trend that NAC treatment was associated with reduced severity of hepatic crises, suggesting a potential therapeutic benefit. CONCLUSIONS: Our findings link fever-induced ER stress, ROS accumulation, and apoptosis to the pathogenesis of liver failure in NBAS deficiency. NAC attenuates these cellular stress responses and may represent a promising supportive treatment option in NBAS-associated liver failure. Genetic changes to both alleles of the NBAS gene lead to, among others, recurrent episodes of fever-triggered liver failure. However, the reason why fever causes liver damage has not been understood. In this study, we examined skin cells from people with NBAS deficiency and compared them with cells from a healthy individual. When the cells were exposed to a higher temperature (similar to what happens during a fever), the NBAS-deficient cells showed signs of stress within a part of the cell called the endoplasmic reticulum (ER) and there was a higher production of reactive oxygen species (ROS). Furthermore, many of the affected cells underwent programmed cell death. We then tested the effect of an antioxidant medication (called N-acetylcysteine; NAC) used to treat certain types of liver failure. In laboratory experiments, NAC reduced the build-up of harmful ROS and protected the NBAS-deficient cells from dying. Furthermore, we reviewed clinical data from 16 people with NBAS deficiency. NAC treatment during fever-related illnesses seemed to lessen the severity of liver crises. Overall, our study suggests that fever can trigger a chain reaction in NBAS deficiency—causing cellular stress, oxidative damage and cell death—which may contribute to liver failure. NAC appears to interrupt this process and might be a useful treatment during febrile illness in people with NBAS deficiency.