Study Of Gene Expression Patterns In Rats’ Offsprings Induced By Pregnant Benzo[a]Pyrene Exposure

BackgroundBenzo[a]pyrene(BaP) is one of the most representative chemicals of polycyclic aromatic hydrocarbon (PAHs) and is the main pollutant of the production and living environment. BaP mainly comes from tobacco smoke, cooking oil, pyrolysis and incomplete combustion of coal, oil, wood and other organic matters.BaP enters the body through the air, food, drinking water and other ways. Now, the carcinogenicity, immune toxicity, neurotoxicity of BaP has been widely reported. Although the effect of BaP on birth outcome has been epidemiological confirmed, but the specific biological effect and mechanism need further clear. In recent years, gene chips were used in the carcinogenicity mechanism research of BaP. Some synergistic genes, such as oncogenes, tumor suppressor genes, cell cycle related genes, DNA synthesis genes, and DNA repair genes, were found. In this study, the gene chips were used to investigate the gene expression patterns of offspring’ liver cells induced by prenatal BaP exposure, and further investigate the possible mechanisms of adverse effect of BaP on the growth and development the offspring.Objectives1. To investigate the effects of BaP exposure during pregnancy on the growth development of infants2. To study the gene expression patterns in rats’ livers of infant rats after BaP exposure during pregnancy and explore the important gene and signaling pathways in the toxic mechanism of BaP3. To study the expression levels of apoptosis related proteins Bcl-xl and Bax in offspring liver after BaP exposure in pregnant ratsMethods1. Pregnant SD rats were randomly divided into four groups:vehicle control (corn oil) and BaP treatment groups (0.75mg/kg,1.5mg/kg and3mg/kg)(n=8). BaP solutions were given by gastric infusion from the third day to the17th day of pregnancy.2. The pregnant rats’weights in different days (0,4th,7th, and19th day) were measured. After delivery the body length, tail length and weight of offspring were observed.3. The level of BaP in blood of the pregnant rats and the offsprings were measured by HPLC.4. After delivery the offspring’s liver were taken to detect the gene expression by RatRef-12chip. Different expression variations were identified. The expression pattern of the genes relate to the dose of prenatal BaP exposure were analyzed. All the differently expressed genes involved in the signaling pathway were analyzed. The core genes and key signal pathways of the differently expressed gene network were also analyzed.5. Realtime PCR method was used to detect the expression levels of core genes to verify the results of gene chips.6. Western blot method was used to detect the Bcl-xl and Bax protein levels in the liver of offsprings.Results1. With the increased level of BaP during pregnancy, the growth indicators of the newborn pups such as body weight, length, and tail length have decreased, but there was no significant difference between the groups (P>0.05).2. In the medium-dose group, the level of BaP in pregnant rats and the offsprings were respectively. In high-dose group, the level of BaP in pregnant rats and the offsprings were respectively. In low-dose group, no BaP was detected both in pregnant rats and the offsprings.3. After prenatal BaP exposure1,232genes with different expression variations were identified. To further narrow down the target genes which harbor great significance among the1,232genes and relate to the dose of prenatal BaP exposure, we chose to use the26model profiles to summarize the expression pattern of the genes. Among the26patterns,6expression patterns of genes were identified with significant P-value (P<0.05). All the differently expressed genes involved in the signaling pathway mainly focus on:growth and development, toxicant metabolism and inflammation. The data from gene network analysis demonstrate that four genes, such as Rela, Cyp2c13, Mapk8and Plcg1are in the core of the network.4. Realtime PCR results showed that the expression level od Cyp2cl3gene were up-regulated, and the expression of Gstol、Rela、Bcl21、Plcg1and Mapk8genes were down-regulated. The results were in accordance with the results of gene chips.5. Western blot results showed that compared with the control group the Bcl-xl protein expression of offspring liver significantly decreased in the middle and high dose group, while Bax protein expression were significantly increased, the differences were statistically significant (P<0.05or P<0.01). The ratio of Bcl-xl/Bax also decreased with the increase of BaP exposure.Conclusion1. Gene expression patterns of offspring’liver cells were influenced by prenatal BaP exposure. Some key genes and signal pathways were also found. The study provides an important clue for the toxicity and mechanisms of the prenatal BaP exposure on the growth and development of offsprings.2. With the increase of BaP exposure in pregnancy, the ratio of apoptosis related protein Bcl-xl/Bax in newborn liver decreased, suggesting that apoptosis may be one of the mechanisms in the BaP toxic effects to offsprings.