Date of Award
Open Access Thesis
Medical Doctor (MD)
Ahmed A. Fadiel Ph.D.
The purpose of this study was to look at the genotoxic and cytotoxic effects of benzo-α-pyrene (BαP), a chemical mutagen that is present in cigarette smoke, on the insulin-like growth factor-I (IGF-I) gene. Women who smoke during pregnancy are more likely to have a growth-restricted baby. We hypothesized that BαP exerts its effects through genotoxic and cytotoxic avenues. The cytotoxicity is manifested by chromosomal abnormalities and a decrease in the rate of cell division. The genotoxicity is manifested by changes in certain genes known to be important in mammalian fetal development such as IGF-I. IGF-I is implicated in intrauterine growth restriction (IUGR), a problem that greatly increases the risk of perinatal morbidity and mortality. To futher understand the mechanism by which BαP influences the normal growth and development of human placental cells, human placental trophoblast cells from an established immortalized cell line were utilized. Cells were cultured in appropriate media, starved (using starvation "Serum Free Medium"), and treated with two doses of BαP, 1µM (dose 1) and 5µM (dose 2). Chromosomes were prepared for cytogenetic analysis and visualized using light microscopy after Giemsa staining. Chromosomal aberrations were identified and the rate of cell division was determined through the analysis of the mitotic index for treated cells compared to a control group. To further understand the influence of BαP on the IGF-I gene expression level, RNA was extracted from control and treated cells, from which cDNA was synthesized and used for further analysis using polymerized chain reaction (PCR). The PCR results were used to better understand the genotoxicity of BαP, while chromosomal aberration analysis was used to determine the cytotoxic effects of BαP on human placental cells. Our results indicate that many chromosomal abnormalities were present in the treated groups compared to the control group. In addition, there was a significant decrease in the mitotic index of the BαP-treated cells (MI=0.3%) verses the control group (MI=0.93%), p value 0.0447. Through the PCR assay, we speculate that there is a dose-related response to BαP of the IGF-I RNA expression level, with low levels in the treated groups compared to the control group. We conclude from these results that BαP influences placental cells at both the gene and chromosome level. It also affects the cell cycle of human placental cells. It is known that smoking is deleterious for fetal development. We believe that the current study brings us closer to understanding the mechanism by which smoking can lead to fetal growth restriction.
Epperson, Brittany Albright, "The Effects of Benzo-α-Pyrene on the Insulin-like Growth Factor-I Gene" (2006). Yale Medicine Thesis Digital Library. 236.