mTOR complexes promote peripheral nerve damage and Schwann cell injury in diabetes

Abstract

Background: Diabetic Neuropathy (DN) is one of the microvascular complications associated with diabetes. The clinical manifestations of DN include sensorimotor loss, paresthesia and altered pain and thermal perception. Hyperglycemia has been reported to be one of the triggers that induce peripheral nerve injury, characterized by myelin abnormalities, axonal atrophy and blunted regenerative potential. Nevertheless, the pathways that lead to the pathophysiology of diabetes-induced peripheral nerve injury has yet to be elucidated. Our group has identified reactive oxygen species (ROS) as a final key mediator in a number of diabetic complications, but their role in the etiology of DN has not been previously described. One of the major sources of ROS is the NADPH Oxidases (NOX) family, which we and others have shown to be involved in the pathophysiology of diabetic complications. More importantly, the crosstalk of NADPH oxidases with downstream or upstream signaling molecules is at the heart of the deleterious effect seen in diabetic complications. Aim: The following study aims to investigate the role of NOX1 enzyme as sources of oxidative stress in myelinating Schwann cells (SCs) and sciatic nerves in the diabetic milieu. Furthermore, the crosstalk of the NADPH oxidases with the mTOR signaling pathway and the role of this signaling axe in SCs and sciatic nerve injury will be identified. Methods: Mouse Schwann Cell (MSC80) cell line are used for in vitro studies and the C57-BL6, NOD-FVB, and Swiss Webster mice are used for the in vivo studies. Western blot and PCR analyses are used to assess expression of myelin proteins, NOXs, mTOR complexes associated proteins in addition to their signaling effectors. In vitro silencing of NOX and mTOR is also performed by transfection to further elucidate the role of the NOX-mTOR axe in diabetic peripheral injury. NADPH oxidase activity assay is used to assess NADPH-induced intracellular ROS production alongside DHE staining, and HPLC in SCs and sciatic nerves. Cellular apoptoti