Open Access Thesis
Metolachlor is one of the most common pre-emergent pesticides used by Iowa farmers. It is used to control broadleaf weeds and grasses in the fields of corn and soybean. It is a frequent contaminant of groundwater and surface water due to runoff after application. It has been shown to inhibit protein and chlorophyll synthesis in plant cells, but its effect on mammalian cells has not been consistently conclusive. As metolachlor and other herbicides have been shown to be associated with harmful health effects in non-target organisms it is very important to determine a mechanism of action on non-target cells, especially human cells. Previous work from our lab studying the effects of metolachlor on human cells demonstrated that low-level metolachlor exposure can inhibit growth in several human cell types such as fibroblasts, HepG2 liver cells and THP-1 alveolar monocytes and alter the expression level of several proteins involved in cell cycle control. While we have shown that metolachlor can have an effect on cellular macromolecules involved in cell growth, is not clearly understood is if there are other cellular processes, genes and proteins that may be altered by exposure to metolachlor.
Microarray analysis was used in this study to determine if there are groups or families of genes that participate in a biological process which may be altered after herbicide exposure, and if one or several specific cellular functions are affected or altered after this exposure. Microarray analysis was performed after isolating total RNA from HepG2 cells exposed to varying levels (200 ppb, 500 ppb, and 1000 ppb) of metolachlor for varying times (24 hours, 48 hours and 72 hours) and comparing expression to that of control (0 ppb) cells for each of the time points.
To demonstrate a significant amount of differential gene expression, cells needed 1000 ppb of metolachlor at 24 hours exposure. Whereas at 48 hours of exposure, differential gene expression between control and metolachlor treated cells was observed at all three concentrations, suggesting that 48 hours of exposure may be an important and perhaps optimal time point for altered gene expression between control and metolachlor treated cells. Very little level of differential gene expression was observed at 72 hours of exposure time.
Gene Ontology (GO) analysis, using the GOrilla tool of genes that were differentially expressed between control and metolachlor treated HepG2 cells showed significant enriched clusters in the processes of DNA replication and repair, cellular response to stimuli and cell cycle regulation. Metolachlor exposure for 48 hours at 500 ppb showed eight biological processes were enriched while 1000 ppb metolachlor resulted in thirteen biological processes that were enriched.
Date of Award
Master of Science
Department of Biology
Kavita R. Dhanwada
1 PDF file (viii + 170 pages)
©2017 - Navinder Paul Kaur Brar
Brar, Navinder Paul Kaur, "Differential gene expression analysis of metolachlor treated human liver cells by microarray" (2017). Electronic Theses and Dissertations. 472.
Available for download on Friday, June 22, 2018