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neonatal mice. In the current study, the team took an in vitro approach: they isolated cerebral cortex cells from rat fetuses, cultured the cells, and exposed them to various concentrations of PBDE-99. “If you expose in vivo, you will probably have indirect effects via different kinds of endocrine pathways [and so on], which will probably affect the end phenotype,” explains Alm. “So
Flame retardants are added to consumer products such as carpets, upholstery, and computers to slow the rate of ignition and the spread of flames. That sounds like a good idea, but scientists have discovered that many of these chemicals may be neurotoxicants that affect motor and cognitive functions. To investigate the effects of one such flame retardant, polybrominated diphenyl ether-99 (PBDE-99), on brain cells, Henrik Alm and colleagues at Uppsala University (Sweden), Stockholm University, and the University of Southern Denmark conducted an in vitro proteomics study. They report the results in JPR (DOI 10.1021/pr900723c). Surprisingly, they found that high and low doses of the chemical sometimes have dramatically different effects on the proteome. One problem with PBDEs is Varied responses. Brain cells treated with various that they “do not remain bound concentrations of PBDE-99 displayed strikingly different protein profiles. to a product but slowly leak into our air, dust, and water and eventually enter our food and here, we wanted to take all of that bodies,” says Mohamed Abou-Elwafa away and just look at direct effects on Abdallah of Birmingham University the neurons.” (U.K.). Of particular concern are the Cortical cells were treated with 3, 10, high levels found in infants and or 30 µM PBDE-99 for 24 h. Proteins young children. were extracted and analyzed with 2D PBDEs are common environmental difference gel electrophoresis (known contaminants in the U.S. and Canada, as DIGE). In a finding consistent with where strict regulations promoting their the team’s previous studies, several use were enacted years ago. Now that proteins were differentially regulated in the chemicals have been implicated as PBDE-treated cells compared with conpotential developmental neurotoxicants, trols. Most of these proteins play a role however, most forms have been banned in forming the cytoskeleton; cytoskelin a few U.S. states and in the EU. In etal proteins are necessary for the deaddition, the only U.S. manufacturer of velopment of various neuronal strucPBDEs stopped producing them in 2004. tures. “If you disrupt this molecular Despite these steps, the chemicals are interplay, it is likely to have major efstill a problem because they persist in fects on brain function and brain arthe environment and accumulate in the chitecture,” explains Alm. food chain. When the researchers took a closer In their previous work, Alm and collook at the data, they realized that only leagues conducted in vivo proteomic 9 out of 292 identified proteins were studies of the effects of PBDE-99 in
10.1021/pr901169z
© 2010 American Chemical Society
common among all of the PBDE treatments. Only one protein was common between the 3 and 10 µM groups, whereas the 3 and 10 µM groups had 28 and 26 proteins in common with the 30 µM group, respectively. Alm notes that this result shows that PBDE-99 has different effects at low and high concentrations. Lucio Costa of the University of Washington says that this is a key observation with possible implications for risk assessment. “The results of this paper point to the fact that one should use caution when extrapolating from higher concentrations to lower, environmentally relevant exposure levels,” he explains. Abdallah states that another important piece of information derived from the study is that it “sheds light on the neurotoxic effects of PBDEs at low, sub-cytotoxic levels, which are comparable to the actual levels reported in mammals under normal exposure conditions.” The researchers also treated a set of cells with 30 µM PBDE-99 for 1 h and compared these data to those obtained at 24 h, as well as a 1 h control. “We reasoned that the effects you can see after one hour are probably due to posttranslational modifications rather than transcripts,” says Alm. In statistical analyses, the proteins in the 30 µM/1 h sample already were differentially regulated compared with proteins in the other groups. Staining three of the 2D gels with a phosphoprotein stain revealed that several proteins were phosphorylated in that sample. As expected, this study is leading Alm and colleagues toward additional experiments. They plan to follow up on some of the proteins found in the study with MALDI imaging in vivo, and they will also investigate epigenetic modifications that result from PBDE exposure. —Katie Cottingham HENRIK ALM
Flame retardants have different effects at high and low doses
Journal of Proteome Research • Vol. 9, No. 3, 2010 1193
