Endothelial Cell-specific Activation of NOX1 Drives Obesity and Metabolic Syndrome via Skeletal Muscle Mitochondrial Dysfunction with Novel Genetic Signatures
Presently, we investigated hypothesized roles and mechanisms of cell type-specific, selective activation of different vascular NADPH oxidase (NOX) isoforms in obesity and metabolic syndrome.  Expression of NOX1 was significantly upregulated in WT mice fed high-fat diet.  Global knockout of NOX1 (NOX1-/y), rather than of NOX2/NOX4, markedly abrogated high-fat feeding induced body weight/fat mass gain, preadipocyte differentiation, fatty liver, glucose intolerance, and insulin/leptin resistance.  Intriguingly, endothelial-specific NOX1 knockout (Cdh5cre-NOX1CKO), rather than vascular smooth muscle-specific NOX1 knockout (Myh11cre-NOX1CKO), substantially alleviated these features of obesity and metabolic syndrome.  Consistently, endothelial-specific NOX1 knockin mice (Cdh5cre-NOX1CKI) fed high-fat diet displayed exaggerated metabolic disorders.  Endothelial cell (EC)-specific knockout/knockin of NOX1 was confirmed using EC washout experiments.  Food/water intakes were not different from corresponding controls in high-fat fed NOX1-/y, Cdh5cre-NOX1CKO or Cdh5cre-NOX1CKI mice, indicating no difference in energy intake.  Instead, spontaneous activity, exercise capacity, mitochondrial oxygen consumption/ATP production, skeletal muscle mitochondrial function (ROS production and swelling activity), and mitochondrial cristae structure were all substantially improved in NOX1-/y or Cdh5cre-NOX1CKO mice, indicating augmented energy expenditure attributed to preserved skeletal muscle mitochondrial function.  Supportively, Cdh5cre-NOX1CKI mice displayed deteriorated exercise capacity and skeletal muscle mitochondrial dysfunction.  Endothelium-dependent vasorelaxation was restored in high-fat fed NOX1-/y or Cdh5cre-NOX1CKO mice, confirming improved endothelial function.  RNA-seq identified 4 genes (Cntnap4, Sgsm1, Tll2 and Syt9) and 7 genes (Odf3l2, Col9a1, Cldn23, Atp5g2, Nkx6-3, Ntsr2 and Zfp69) significantly down/upregulated in high-fat fed Cdh5cre-NOX1CKO mice, among which Cntnap4 and COL9A1 linked to muscular disorders.  Importantly, we observed marked upregulation of NOX1 in isolated coronary arteries from human obese patients.  Taken together, our data for the first time establish a novel and paradigm-shifting concept that endothelial NOX1 drives systematic metabolic phenotypes, via impairment in skeletal muscle mitochondrial dysfunction with novel genetic signatures.  Tissue-specific targeting of endothelial NOX1 and novel candidate genes may prove to be robustly effective in treating obesity and metabolic syndrome. Overall design: RNAs isolated from soleus muscles of experimental mice (3 each group from 4 experimental groups of Cdh5cre-WT mice under 8 weeks regular diet, Cdh5cre-WT mice under 8 weeks  high-fat diet, Cdh5cre-NOX1CKO mice under 8 weeks regular diet, and Cdh5cre-NOX1CKO mice under 8 weeks high-fat diet)