Malondialdehyde-protein adducts in the spleens of aniline-treated rats: Immunochemical detection and localization

M Khan, X. Wu, Ghulam Ansari, Paul J. Boor

Research output: Contribution to journalArticle

28 Citations (Scopus)

Abstract

Previously it was reported that aniline exposure in rats induces increased lipid peroxidation and formation of malondialdehyde (MDA)-protein adducts in the spleen. In order to further elucidate the role of MDA-protein adducts in the splenic toxicity of aniline, studies were conducted to detect and localize these adducts in the spleen. Rabbit polyclonal antisera to MDA-keyhole limpet hemocyanin were employed for immunohistochemical localization and Western blot analyses of MDA-protein adducts in the spleens of aniline-treated (65 mg/kg/d aniline in the drinking water for 30 d) and control rats. For immunohistochemical localization of MDA-protein adducts in the spleen, a new approach using alkaline phosphatase-fast red (red color) to demonstrate bound primary antibodies was adopted, providing a sharper and increased contrast compared to horseradish peroxidase-diaminobenzidine (brown color) methodology. This new approach allowed us to differentiate the changes in aniline-treated spleens, which had extensive brownish deposits of iron proteins. Spleens from aniline-treated rats showed intense staining for these adducts in the red pulp areas (where iron was also localized), especially within the sinusoidal macrophages. Spleens from control rats showed only mild staining for adducts and only traces of iron. Western blot analyses of splenic microsomal proteins from aniline-treated and control rats showed the presence of 13 different MDA-modified proteins. However, 26-, 32-, and 44-kD proteins were more prominent in the aniline-treated rats. The colocalization of MDA-protein adducts with iron in the red pulp of the spleen suggests that iron-catalyzed lipid peroxidation leading to formation of MDA-protein adducts could be a potential mechanism for splenic toxicity of aniline.

Original languageEnglish (US)
Pages (from-to)93-102
Number of pages10
JournalJournal of Toxicology and Environmental Health - Part A
Volume66
Issue number1
DOIs
StatePublished - Jan 10 2003

Fingerprint

Aniline
Malondialdehyde
Rats
Spleen
Proteins
protein
Rat control
Iron
iron
Lipids
Lipid Peroxidation
Pulp
Toxicity
Color
Western Blotting
lipid
aniline
detection
Staining and Labeling
toxicity

ASJC Scopus subject areas

  • Environmental Science(all)
  • Environmental Chemistry
  • Public Health, Environmental and Occupational Health
  • Pollution
  • Toxicology
  • Health, Toxicology and Mutagenesis

Cite this

@article{fe45fe4cc8844a9c8e6ec15006fdae6c,
title = "Malondialdehyde-protein adducts in the spleens of aniline-treated rats: Immunochemical detection and localization",
abstract = "Previously it was reported that aniline exposure in rats induces increased lipid peroxidation and formation of malondialdehyde (MDA)-protein adducts in the spleen. In order to further elucidate the role of MDA-protein adducts in the splenic toxicity of aniline, studies were conducted to detect and localize these adducts in the spleen. Rabbit polyclonal antisera to MDA-keyhole limpet hemocyanin were employed for immunohistochemical localization and Western blot analyses of MDA-protein adducts in the spleens of aniline-treated (65 mg/kg/d aniline in the drinking water for 30 d) and control rats. For immunohistochemical localization of MDA-protein adducts in the spleen, a new approach using alkaline phosphatase-fast red (red color) to demonstrate bound primary antibodies was adopted, providing a sharper and increased contrast compared to horseradish peroxidase-diaminobenzidine (brown color) methodology. This new approach allowed us to differentiate the changes in aniline-treated spleens, which had extensive brownish deposits of iron proteins. Spleens from aniline-treated rats showed intense staining for these adducts in the red pulp areas (where iron was also localized), especially within the sinusoidal macrophages. Spleens from control rats showed only mild staining for adducts and only traces of iron. Western blot analyses of splenic microsomal proteins from aniline-treated and control rats showed the presence of 13 different MDA-modified proteins. However, 26-, 32-, and 44-kD proteins were more prominent in the aniline-treated rats. The colocalization of MDA-protein adducts with iron in the red pulp of the spleen suggests that iron-catalyzed lipid peroxidation leading to formation of MDA-protein adducts could be a potential mechanism for splenic toxicity of aniline.",
author = "M Khan and X. Wu and Ghulam Ansari and Boor, {Paul J.}",
year = "2003",
month = "1",
day = "10",
doi = "10.1080/15287390306464",
language = "English (US)",
volume = "66",
pages = "93--102",
journal = "Journal of Toxicology and Environmental Health - Part A: Current Issues",
issn = "1528-7394",
publisher = "Taylor and Francis Ltd.",
number = "1",

}

TY - JOUR

T1 - Malondialdehyde-protein adducts in the spleens of aniline-treated rats

T2 - Immunochemical detection and localization

AU - Khan, M

AU - Wu, X.

AU - Ansari, Ghulam

AU - Boor, Paul J.

PY - 2003/1/10

Y1 - 2003/1/10

N2 - Previously it was reported that aniline exposure in rats induces increased lipid peroxidation and formation of malondialdehyde (MDA)-protein adducts in the spleen. In order to further elucidate the role of MDA-protein adducts in the splenic toxicity of aniline, studies were conducted to detect and localize these adducts in the spleen. Rabbit polyclonal antisera to MDA-keyhole limpet hemocyanin were employed for immunohistochemical localization and Western blot analyses of MDA-protein adducts in the spleens of aniline-treated (65 mg/kg/d aniline in the drinking water for 30 d) and control rats. For immunohistochemical localization of MDA-protein adducts in the spleen, a new approach using alkaline phosphatase-fast red (red color) to demonstrate bound primary antibodies was adopted, providing a sharper and increased contrast compared to horseradish peroxidase-diaminobenzidine (brown color) methodology. This new approach allowed us to differentiate the changes in aniline-treated spleens, which had extensive brownish deposits of iron proteins. Spleens from aniline-treated rats showed intense staining for these adducts in the red pulp areas (where iron was also localized), especially within the sinusoidal macrophages. Spleens from control rats showed only mild staining for adducts and only traces of iron. Western blot analyses of splenic microsomal proteins from aniline-treated and control rats showed the presence of 13 different MDA-modified proteins. However, 26-, 32-, and 44-kD proteins were more prominent in the aniline-treated rats. The colocalization of MDA-protein adducts with iron in the red pulp of the spleen suggests that iron-catalyzed lipid peroxidation leading to formation of MDA-protein adducts could be a potential mechanism for splenic toxicity of aniline.

AB - Previously it was reported that aniline exposure in rats induces increased lipid peroxidation and formation of malondialdehyde (MDA)-protein adducts in the spleen. In order to further elucidate the role of MDA-protein adducts in the splenic toxicity of aniline, studies were conducted to detect and localize these adducts in the spleen. Rabbit polyclonal antisera to MDA-keyhole limpet hemocyanin were employed for immunohistochemical localization and Western blot analyses of MDA-protein adducts in the spleens of aniline-treated (65 mg/kg/d aniline in the drinking water for 30 d) and control rats. For immunohistochemical localization of MDA-protein adducts in the spleen, a new approach using alkaline phosphatase-fast red (red color) to demonstrate bound primary antibodies was adopted, providing a sharper and increased contrast compared to horseradish peroxidase-diaminobenzidine (brown color) methodology. This new approach allowed us to differentiate the changes in aniline-treated spleens, which had extensive brownish deposits of iron proteins. Spleens from aniline-treated rats showed intense staining for these adducts in the red pulp areas (where iron was also localized), especially within the sinusoidal macrophages. Spleens from control rats showed only mild staining for adducts and only traces of iron. Western blot analyses of splenic microsomal proteins from aniline-treated and control rats showed the presence of 13 different MDA-modified proteins. However, 26-, 32-, and 44-kD proteins were more prominent in the aniline-treated rats. The colocalization of MDA-protein adducts with iron in the red pulp of the spleen suggests that iron-catalyzed lipid peroxidation leading to formation of MDA-protein adducts could be a potential mechanism for splenic toxicity of aniline.

UR - http://www.scopus.com/inward/record.url?scp=0037427872&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0037427872&partnerID=8YFLogxK

U2 - 10.1080/15287390306464

DO - 10.1080/15287390306464

M3 - Article

C2 - 12587293

AN - SCOPUS:0037427872

VL - 66

SP - 93

EP - 102

JO - Journal of Toxicology and Environmental Health - Part A: Current Issues

JF - Journal of Toxicology and Environmental Health - Part A: Current Issues

SN - 1528-7394

IS - 1

ER -