Elsevier

Free Radical Biology and Medicine

Volume 49, Issue 6, 15 September 2010, Pages 1036-1045
Free Radical Biology and Medicine

Original Contribution
Role of 12/15-lipoxygenase in nitrosative stress and peripheral prediabetic and diabetic neuropathies

https://doi.org/10.1016/j.freeradbiomed.2010.06.016Get rights and content

Abstract

This study evaluated the role of 12/15-lipoxygenase, which converts arachidonic acid to 12(S)- and 15(S)-hydroxyeicosatetraenoic acids, in nitrosative stress in the peripheral nervous system and peripheral prediabetic and diabetic neuropathies. The experiments were performed in C57BL6/J mice made diabetic with streptozotocin or fed a high-fat diet and in human Schwann cells cultured in 5.5 or 30 mM glucose. 12/15-Lipoxygenase overexpression and activation were present in sciatic nerve and spinal cord of diabetic and high-fat diet-fed mice, as well as in human Schwann cells cultured in high concentrations of D-, but not L-glucose. 12/15-Lipoxygenase inhibition by cinnamyl-3,4-dihydroxy-α-cyanocinnamate (8 mg kg 1 day 1 sc, for 4 weeks after 12 weeks without treatment) alleviated the accumulation of nitrated proteins in the sciatic nerve and spinal cord, and large and small nerve fiber dysfunction, but not intraepidermal nerve fiber loss. 12/15-Lipoxygenase gene deficiency alleviated nitrosative stress and nerve conduction deficit, but not small sensory fiber neuropathy, in high-fat diet-fed mice. In conclusion, 12/15-lipoxygenase is implicated in nitrosative stress and peripheral neuropathy in mouse models of type 1 and early type 2 diabetes. Its presence in human Schwann cells and upregulation by high glucose suggest a potential involvement in human disease.

Section snippets

Reagents

Unless otherwise stated, all chemicals were of reagent-grade quality and were purchased from Sigma Chemical (St. Louis, MO, USA). Cinnamyl-3,4-dihydroxy-α-cyanocinnamate (CDC) was obtained from Enzo Life Sciences International (Plymouth Meeting, PA, USA). Mouse monoclonal (clone 1A6) anti-nitrotyrosine (NT) antibody for Western blot analysis of nitrated proteins was purchased from Millipore (Billerica, MA, USA). Rabbit polyclonal (clone H-100) anti-LO antibody for Western blot analysis was

Results

In experiment 1, LO fluorescence was clearly identifiable in endothelial (Fig. 1a) and Schwann (Fig. 1b) cells of mouse sciatic nerve.

In experiment 2, the initial (before STZ administration) body weights were similar in all experimental groups (Table 1). Weight gain during the 16-week study was lower in both untreated and CDC-treated diabetic mice than in the nondiabetic control group. CDC treatment did not affect weight gain in either control or diabetic mice. Initial (after STZ

Discussion

It has been known for at least a decade that diabetes is associated with disturbances in arachidonic acid metabolism in peripheral nerve [35]. Arachidonic acid is metabolized by cyclooxygenases (COX)-1 and -2, 5- and 12/15-LOs, and cytochrome P450 epoxygenase. Diabetes does not affect rat sciatic nerve COX-1 expression, and the findings with the nonselective COX inhibitor flurbiprofen suggest that COX-1 is essentially required for normal peripheral nerve function [36]. In contrast, sciatic

Acknowledgments

The study was supported by National Institutes of Health Grants DK074517 and DK077141 and American Diabetes Association Research Grant 7-08-RA-102 (both to I.G.O.). The Cell Biology and Bioimaging Core utilized in this work is supported in part by COBRE (NIH P20 RR021945) and CNRU (NIH 1P30-DK072476) center grants from the National Institutes of Health. The authors thank Drs. Douglas E. Wright from the University of Kansas Medical Center (Kansas City, KS, USA) and Gary L. Pittenger from Eastern

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