Peripheral Nerve Protein Expression and Carbonyl Content in N,N-Diethlydithiocarbamate Myelinopathy

Abstract

Human exposure to dithiocarbamates results from their uses as pesticides, in manufacturing and as pharmaceutical agents. Neurotoxicity is an established hazard of dithiocarbamate exposure and has been observed in both humans and experimental animals. Previous studies have shown that the neurotoxicity of certain dithiocarbamates, including N,N-diethyldithiocarbamate (DEDC), disulfiram, and pyrrolidine dithiocarbamate can manifest as a primary myelinopathy of peripheral nerve. Because increased levels of copper in peripheral nerve and elevated levels of lipid peroxidation products accompany DEDC induced lesions it has been suggested that disruption of copper homeostasis and increased oxidative stress may contribute to myelin injury. To further assess the biological impact of DEDC-mediated lipid peroxidation in nerve the changes in protein expression levels resulting from DEDC exposure were determined. In addition, protein carbonyl content in peripheral nerve was also determined as an initial assessment of protein oxidative damage in DEDC neuropathy. Rats were exposed to DEDC by intraabdominal osmotic pumps for 8 weeks and proteins extracted from sciatic nerves of DEDC exposed animals and from non-exposed controls. The comparison of protein expression levels using two dimensional difference gel electrophoresis demonstrated significant changes in 56 spots of which 46 were identified by MALDI-TOF/MS. Among the proteins showing increased expression were three isoforms of glutathione transferase, important for detoxification of reactive αβ,-unsaturated aldehydes generated from lipid peroxidation. The increased expression of one isoform, glutathione transferase pi, was localized to the cytoplasm of Schwann cells using immunohistochemistry. Immunoassay for nerve protein carbonyls demonstrated a significant increase of approximately 2 fold for the proteins isolated from the DEDC exposed rats. These data support the ability of DEDC to promote protein oxidative damage in peripheral nerve and to produce sufficient lipid peroxidation in either myelin or another component of the Schwann cell to elicit a protective cellular response to oxidative stress.

Copper accumulation and lipid oxidation precede inflammation and myelin lesions in N,N-diethyldithiocarbamate peripheral myelinopathy

“…Previous proteomic investigations of peripheral nerve have shown the greatest increases in protein expression to be associated with members of the IgG class (Viquez et al, 2007). In particular IgG2a demonstrated increases greater than six fold.…”

“…Previous studies have reported increased levels of copper and lipid oxidation to be associated with dithiocarbamate induced neurotoxicity (Calviello, 2005;Delmaestro, 1995;Tonkin, 2004) and correlative data have shown a relationship between copper levels in peripheral nerve and the severity of myelin injury produced by pyrrolidine dithiocarbamate and N,Ndiethyldithiocarbamate (DEDC) in rodent models (Valentine et al, 2006;Valentine et al, 2007). Additionally, proteomic analysis of sciatic nerve proteins using this animal model has demonstrated significant elevations in expression levels of glutathione transferase isozymes within Schwann cells consistent with activation of the antioxidant response element pathways subsequent to DEDC exposure (Viquez et al, 2007). These observations coupled with the affinity of dithiocarbamates for copper have led to the hypothesis that dithiocarbamates bind copper and generate a lipophilic complex that accumulates and promotes lipid oxidation within myelin membranes leading to a subsequent myelinopathy.…”

“…Up regulation of the antioxidant response element pathway may have contributed to the subsequent decrease in these parameters. Evidence for activation of the antioxidant response element by DEDC has been provided by the observed elevations in expression levels of several isoforms of glutathione transferase subsequent to DEDC exposure (Viquez et al, 2007). Additionally, if the observed lipid oxidation is dependent upon redox active copper species, increased expression of copper chaperones or metallothioneins may have also diminished the level of redox active copper within nerve subsequent to the 2-week time point.…”

Nitrogen Substituent Polarity Influences Dithiocarbamate-Mediated Lipid Oxidation, Nerve Copper Accumulation, and Myelin Injury

“…A previous study also found a trend toward elevation of zinc in the liver of DEDC treated rats (12), however, no treatment related lesions in the liver or elevations in aspartate amino transferase were found in any of the animals examined. Previous studies have demonstrated that elevations in lipid peroxidation and protein expression levels of antioxidant enzymes in peripheral nerve accompanied DEDC administration (14, 32). Increased expression of glutathione transferase isoforms has been detected in nerve by proteomic analyses; and the observation that elevations of peripheral nerve MDA and F 2 -isoprostanes preceded the onset of lesions suggested that lipid peroxidation could contribute to the development of myelin lesions.…”

Linking protein oxidation to environmental pollutants: Redox proteomic approaches

“…Higher protein carbonyls levels were found, for instance, in corkwing wrasse (Symphodus melops) captured near a disused copper mine where water and sediments are contaminated with heavy metals [44]. Also, copper accumulation and oxidative damage have been linked to toxicity of dithiocarbamate fungicides in rats [45,46].…”