Immunochemical Detection of Hepatic Cocaine-Protein Adducts in Mice
详细信息 查看全文
- 作者：Florence M. Ndikum-Moffor ; John W. Munson ; Nagraj K. Bokinkere ; Jennifer L. Brown ; Nigel Richards ; and Stephen M. Roberts
- 刊名：Chemical Research in Toxicology
- 出版年：1998
- 出版时间：March 1998
- 年：1998
- 卷：11
- 期：3
- 页码：185 - 192
- 全文大小：170K
- 年卷期：v.11,no.3(March 1998)
- ISSN：1520-5010
文摘
Cocaine is capable of producing hepatic necrosis inlaboratory animals and humans. Studiesin mice indicate that N-oxidative metabolism of cocaine is required forhepatotoxicity and havesuggested that toxicity may result from the adduction of proteins bycocaine-reactivemetabolites. To aid in identifying protein targets forcocaine-reactive metabolites, an antibodywas raised in rabbits immunized with cocaine linked via the tropanenitrogen to a carrierprotein (bovine serum albumin). Hepatic proteins fromcocaine-treated mice (ICR males, 50mg of cocaine/kg of body weight, ip) and saline-treated controls wereprepared from wholeliver homogenate or following subcellular fractionation, and Westernblot analyses of hepaticproteins using this antibody were conducted following one- andtwo-dimensional SDS-PAGE.Analysis of liver homogenate from cocaine-treated mice revealedmajor protein targets withapproximate molecular masses of 20 kDa (pI = 6.0), 44 kDa(two proteins with pI's of 5.0 and7.0), 52-54 kDa (pI = 4.5), and 64 kDa (pI= 5.5). These specific protein targets were shownto be localized in the mitochondria and microsomes. Several minorbands of immunoreactivitywere also seen in mice treated with cocaine, but not in saline-treatedcontrols. Pretreatmentof mice with the P450 inhibitor SKF 525A diminished or eliminated theformation of thesecocaine-protein adducts. Liver sections from cocaine-treatedmice immunostained using theantibody indicated the presence of cocaine-adducted proteins in thecentrilobular and midzonalregions of the lobule, corresponding to areas of hepatocyte swellingand necrosis. This studyindicates that reactive metabolites from cocaine bind to discreteproteins in specific regions ofthe liver, consistent with a role for protein adduction in cocainehepatotoxicity.