Nicotinamide Nucleotide Transhydrogenase (NNT) is essential for adrenal steroidogenesis: clinical and in vitro lessons


 
 
 Nicotinamide nucleotide transhydrogenase (NNT) acts as an antioxidant defense mechanism. NNT mutations cause familial glucocorticoid deficiency (FGD). How impaired oxidative stress disrupts adrenal steroidogenesis remains poorly understood.
 
 
 
 To ascertain the role played by NNT in adrenal steroidogenesis.
 
 
 
 The genotype-phenotype association of a novel pathogenic NNT variant was evaluated in a boy with FGD. Under basal and oxidative stress (OS) induced conditions, transient cell cultures of the patient’s and controls wild type (WT) mononuclear blood cells were used to evaluate antioxidant mechanisms and mitochondrial parameters [reactive oxygen species (ROS) production, reduced glutathione (GSH), and mitochondrial mass]. Using CRISPR/Cas9, a stable NNT gene knockdown model was built in H295R adrenocortical carcinoma cells to determine the role played by NNT in mitochondrial parameters and steroidogenesis. NNT immunohistochemistry was assessed in fetal and post-natal human adrenals.
 
 
 
 The homozygous NNT p.G866D variant segregated with the FGD phenotype. Under basal and OS conditions, p.G866D homozygous mononuclear blood cells exhibited increased ROS production, and decreased GSH levels and mitochondrial mass when compared to WT NNT cells. In line, H295R NNT knocked-down cells presented impaired NNT protein expression, increased ROS production, decreased the mitochondrial mass, as well as the size and the density of cholesterol lipid droplets. NNT knockdown affected steroidogenic enzyme expression, impairing cortisol and aldosterone secretion. In human adrenals, NNT is abundantly expressed in the transition fetal zone and in zona fasciculata.
 
 
 
 Together, these studies demonstrate the essential role of NNT in adrenal redox homeostasis and steroidogenesis.
