Hepatic Neuronal Nitric Oxide Synthase Impaired Hepatic Insulin Sensitivity Through Activation of p38 MAPK

Background: nNOS contains PZD domain and binds with NOS1AP. We reported previously that NOS1AP enhanced hepatic insulin activity on its PZD binding domain suggesting an involvement of nNOS in NOS1AP effect. In present study, we aimed to explore the role of nNOS on hepatic insulin activity and the mechanism underlined. Methods: Liver specific nNOS overexpression was achieved by tail-vein-injection of adenovirus-nNOS in C57BL/6 mice fed with a high-fat diet for 8 weeks. nNOS specific inhibitor (L-NPA) was used to inhibit its activity. Findings: nNOS co-localized with NOS1AP in liver. Its phosphorylation in Ser1417 increased in the livers of ob/ob and diet-induced obese (DIO) mice and mice with liver NOS1AP deletion. It decreased in mice with NOS1AP overexpression. Overexpression of nNOS in the livers of DIO mice aggravated glucose intolerance, increased intrahepatic lipid accumulation, decreased glycogen storage, and blunted insulin-induced phosphorylation of IRβ and Akt in livers. Similarly, overexpression of nNOS increased triglyceride production, decreased glucose utilization, and insulin-induced p-IRβ, p-Akt and p-GSK3β in HepG2 cells. In contrast, inhibition of nNOS with L-NPA in ob/ob mice improved glucose tolerance and increased insulin-induced phosphorylation of IRβ and Akt in livers. Moreover, overexpression of nNOS in HepG2 cells increased p-p38 MAPK expression, and inhibition of p38 MAPK with SB203580 significantly reversed the defect in insulin signaling activation. Interpretation: Hepatic nNOS inhibited insulin signaling activity through activation of p38 MAPK in obesity, suggesting a role of nNOS in the development of hepatic insulin resistance, and a potential therapeutic target for the prevention of diabetes. Funding: This work was supported by the National Natural Science Foundation of China (81670707 to C. Wang). Funding Statement: This study was supported by the National Natural Science Foundation of China (81670707 to C. Wang). Declaration of Interests: The authors declared that they have no conflict of interest. Ethics Approval Statement: The protocol was approved by the Ethics Committee of Shanghai Jiao Tong University (2018-KY-026 (K)). Informed consent was given by all subjects and principles of the Declaration of Helsinki were followed. Procedures involving the care and use of animals were carried out in accordance with the Guidelines for the Care and Use of Laboratory Animals of Shanghai Jiao Tong University and approved by the Animal Ethics Committee of the University Affiliated Sixth People’s Hospital (DWSY2011-096/DWLL2019-0274).