Amelioration of Diabetic Neuropathy by TAT-Mediated Enhanced Delivery of Metallothionein and SOD

Abstract

Because diabetic neuropathy (DN) appears to result from oxidative stress in neuronal tissues, antioxidant treatment should counteract the condition. Metallothionein (MT) and superoxide dismutase (SOD) are free-radical scavengers, but their ability to cross biological membranes is limited. Applying cell penetrating peptide technologies, we made Tat-MT and Tat-SOD constructs and tested their ability to protect PC12 cells, as surrogates of peripheral nerve cells, from various forms of oxidative damage. Tat-MT and Tat-SOD were successfully delivered to PC12 cells, and the intracellular activities of MT and SOD increased in line with the amount of protein delivered. These agents inhibited cellular damage and apoptotic signaling caused by three different types of injuries (high glucose, hypoxia, and advanced glycation end product injury). We also examined transduction of Tat-MT and Tat-SOD into Otsuka Long-Evans Tokushima fatty rats. A single ip injection of Tat-MT and Tat-SOD resulted in increased radical scavenging activity and decreased apoptosis, by inhibiting nuclear factor κB and MAPK signaling. Continuous treatment resulted in improved myelination of sciatic nerves and delayed the clinical development of DN. We conclude that effective delivery of a combination antioxidant treatment may facilitate the repair of damage in patients with DN.

Huntington’s Disease-Induced Cardiac Disorders Affect Multiple Cellular Pathways

“…Furthermore, manipulation of SOD was associated with levels of oxidative stress in several neurodegenerative diseases [76, 77]. SOD overexpression has also been shown to reduce diabetic cardiomyopathy and neurodegeneration by ROS [78]. …”

Reappraisal of metallothionein: Clinical implications for patients with diabetes mellitus

“…Metallothionein is able to scavenge a wide range of ROS, including O 2 − and H 2 O 2 , at higher efficiencies than other antioxidants. The recent development of protein transduction technology has made it possible to deliver MT and SOD into several cloned cells (H9C2, INS‐1, H4IIE and PC12 cells) and various primary cultured cells (rat and mouse cardiomyocytes, rat and mouse islets of Langerhans, and rat renal mesangial cells) in vitro . Normally, the transduction of MTs into different cells is inefficient.…”

“…A single i.p. injection of Tat–MT resulted in the delivery of these antioxidants to the nerves, pancreas, liver, heart, and brain, with effects persisting for as long as 4 days after the transduction . Increased radical scavenging activity and decreased apoptosis via the mitochondrial and ER pathways were found in the various tissues after continuous, prolonged (16 weeks), and periodic (at every 3 days) treatment with Tat‐MT, which delayed development of diabetes and its complications clinically without any visible side effects …”

“…However, after supplying exogenous SOD or its mimics to animals or humans, this compensatory induction of catalase may not occur quickly enough to remove the secondary generation of H 2 O 2 . As an alternative, we chose to deliver MT and SOD at the same time . The protective effects of combined antioxidant treatment against glucolipotoxicity and other ROS‐ and RNS‐induced injuries were superior to either treatment alone, as well as other exogenous antioxidant treatments .…”

“…It has been reported that increased OS, resulting from various mechanisms associated with ROS‐ and RNS‐induced injury, is accompanied by activation of MAPK, a decrease in AMPK expression, and possibly by activation of Wnt signaling pathways (Fig. ) . In addition to the classical involvement of MAPK, various nutrient‐sensing pathways are important in the development of diabetic complications including diabetic neuropathy and nephropathy .…”