Metolachlor, a commonly used herbicide in the Midwestern USA, functions by inhibiting chlorophyll and protein synthesis in target plants. Herbicide exposure has led to detrimental effects in several organisms, affecting their growth and behavior; however, its mechanism of action in nontarget organisms is not yet clear. The EPA does not currently have enforceable regulations for maximal limits allowed in drinking water. Previous growth studies from our lab have demonstrated that increasing metolachlor concentrations and increasing time of exposure results in decreased growth of liver cells. The objective of this study was to elucidate a mechanism for this decrease of HepG2 cell growth after herbicide exposure. Results show that metolachlor at environmentally relevant levels (50–100 ppb) that previously led to decreased cell number does not lead to cell death by either necrosis or apoptosis. However, it was demonstrated that the levels of the retinoblastoma protein including two of its hyperphosphorylated forms are decreased in metolachlor exposed cells possibly leading to cell cycle arrest. The levels of another protein involved in cell cycle progression, p53, a mediator in the DNA damage response of cells, was not significantly altered except at the highest level of metolachlor (1,000 ppb) and after a 72-h exposure. These results suggest that the decrease in cell number after low-level metolachlor exposure is most likely due to an alteration in the cell cycle and not due to cell death in human liver cells.
This is a preview of subscription content, log in to check access.
Buy single article
Instant access to the full article PDF.
Price includes VAT for USA
Subscribe to journal
Immediate online access to all issues from 2019. Subscription will auto renew annually.
This is the net price. Taxes to be calculated in checkout.
Chen PL, Scully P, Chew JY, Wang JY, Lee WH. Phosphorylation of the retinoblastoma gene product is modulated during the cell cycle and cellular differentiation. Cell. 1989;58:1193–8.
Chen YW, Yang YT, Hung DZ, Su CC, Chen KL. Paraquat induces lunch alveolar epithelial cell apoptosis via Nrf-2 regulated mitochondrial dysfunction and ER stress. Arch Toxicol. 2012;86:1547–58.
Dhanwada KR, Clayon ME, Deng Y. Effects of the pesticides atrazine, metolachlor and diazinon and the binary mixtures on proliferation of human fibroblasts. Intl J Global Health. 2003;2:22–36.
Dierickx PJ. Glutathinone-dependent cytotoxicity of the chloroacetanilide herbicides alachlor, metolachlor, and propachlor in rat and human hepatoma-derived cultured cells. Cell Biol Toxicol. 1999;15:325–32.
Echeverrigaray S, Gomes LH, Tavares FCA. Isolation and characterization of metolachlor-resistant mutants of Saccharomyces cervesiae. World J Micro Biotech. 1999;15:679–81.
EPA Reregistration Eligibility Decision (RED) Facts: Metolachlor EPA-738-F-95-007. USEPA prevention, pesticides and toxic substances. 1995.
Frank R, Clegg BS, Sherman C, Chapman ND. Triazine and chloroacetamde herbicides in Sydenham river water and municipal drinking water, Dresden, Ontario, Canada, 1981–1987. Arch Envir Cont Toxicol. 1990;19:319–24.
Folch J, Yeste-Valasco M, Alvira D, de la Torre AV, Bordas M, Sureda FX, et al. Evaluation of pathways involved in pentochloropheno-induced apoptosis in rat neurons. Neurotox. 2009;30:451–8.
Greenlee AR, Ellis TM, Berg RL. Low-dose agrochemicals and lawn-care pesticides induce developmental toxicity in murine preimplantation embryos. Environ Health Perspect. 2004;112:703–9.
Grisolia CK, Ferraria I. In vitro and in vivo studies demonstrate non-mutagenicity of the herbicide metolachlor. Brazilian J Gen. 1997;20:411–4.
Hartnett S, Musah S, Dhanwada KR. Cellular effects of metolachlor exposure on human liver (HepG2) cells. Chemosphere. 2013;90:1258–66.
Kale VJ, Miranda SR, Wilbanks MS, Meyer SA. Comparative cytotoxicity of alalchlor, acetochlor, and metolachlor herbicides in isolated rat and cryopreserved human hepatocytes. J Bioch Molec Tox. 2007;22:41–9.
Kolpin DW, Thurman EM, Linhart SM. Finding minimal herbicide concentrations in ground water? Try looking for their degradates. Sci of Total Environ. 2000;248:115–22.
Mitra J, Dai CY, Somasundaram K, El-Deiry WS, Satymoorthy K, Herlyn M, et al. Induction of p21 WAF1/CIP1 and inhibition of cdk2 mediated by the tumor suppressor p16 INK4a. Molec Cell Biol. 1999;19:3916–28.
Munger R, Isacson P, Hu S, Burns T, Hanson J, Lynch CF, et al. Intrauterine growth retradation in Iowa communities with herbicides-contaminated drinking water supplies. Environ Health Perspect. 1997;105:308–14.
Murillo G, Salti GI, Kosmeder JW, Pezzuto JM, Mehta RG. Deguelin inhibits the growth of colon cancer cells through the induction of apoptosis and cell cycle arrest. Eur J Cancer. 2002;38:2446–56.
Paro R, Tiboni GM, Buccione R, Rossi G, Cellini V, Canipari R, et al. The fungicide mancozeb induces toxic effects on mammalian granulosa cells. Toxicol Appl Pharmacol. 2012;260:155–61.
Pereira SP, Fernandes MAS, Martins JD, Santos MS, Moreno AJM, Vicente JAF, et al. Toxicity assessment of the herbicide metolachlor comparative effects on bacterial and mitochondrial model systems. Toxicol In Vitro. 2009;23:1585–90.
Pothuluri JV, Evans FE, Doerge DR, Churchwell MI, Cerniglia CE. Metabolism of Metolachlor by the fungus Cunninghamella elegans. Arch Environ Contam Toxicol. 1997;32:117–25.
Powell ER, Faldladdin N, Rand AD, Pelzer D, Schrunk EM, Dhanwada KR. Atrazine exposure leads to altered growth of HepG2 cells. Toxicol In Vitro. 2011;25:644–51.
Rollof B, Belluck D, Meiser L. Cytogenic effects of cyanazine and metolachlor on human lymphocytes exposed in vitro. Mut Res Lett. 1992;281:295–8.
Rought SE, Yau PM, Chuang LF, Doi RH, Chuang RY. Effect of the chlorinated hydrocarbons heptachlor, chlordane and toxaphene on retinoblastoma tumor suppressor in human lymphocytes. Toxicol Letters. 1999;104:127–35.
Rusiecki JA, Hou L, Lee WJ, Blair A, Dosemeci M, Lubin JH, et al. Cancer incidence among herbicide applicators exposed to metolachlor in the agricultural health study. Intl J Cancer. 2006;15:3118–23.
Thorpe N, Shirmohammadi A. Herbicides and nitrates in groundwater of Maryland and childhood cancers: a geographic information systems approach. J Environ Sci Health Part C. 2005;23:261–78.
Varnagy L, Budai P, Fejes S, Susan M, Fancsi T, Keseru M, et al. Toxicity and degradation of metolachlor (Dual 960EC) in chicken embryos. Commun Agric Appl Biol Sci. 2003;68:807–11.
Weinberg RA. The retinoblastoma protein and cell cycle control. Cell. 1995;81:323–30.
Wolf MC, Moore PA. Effects of the herbicide metolachlor on the perception of chemical stimuli by Orcenectus rusticus. J North Amer Bentho Soc. 2002;21:457–67.
Zhan XM, Liu HJ, Miao YG, Liu WP. A comparative study of rac- and S-metolachlor on some activities and metabolism of silkworm, Bombyx mori L. Pest Bioch Physio. 2006;85:133–8.
Zhang HS, Gavin M, Dahiya A, Postigo AA, Ma D, Luo RX, et al. Exit from G1 and S phase of the cell cycle is regulated by repressor complexes containing HDAC-Rb-hSWI/SNF and Rb-hSWI/SNF. Cell. 2000;101:79–89.
The work performed by student authors’ satisfied requirements for undergraduate research at the University of Northern Iowa. Retinoblastoma protein ELISA analysis was performed by D. Lowry, p53 analysis was completed by D. Greiner, apoptosis analysis was performed by M. Fretheim, and cytotoxicity analysis was performed by M. Ubben. All four students were completing either their BS (DL) or BA in Biology (DG, MF, and MU). Funds were provided by Summer Undergraduate Research Program from the College of Humanities, Arts and Sciences at the University of Northern Iowa as well as a grant to K. R. Dhanwada from the Center for Health Effects of Environmental Contamination at the University of Iowa. The authors would especially like to thank Dr. David McClenahan for his assistance throughout the project and for insight during the preparation of the manuscript as well as Dr. Tilahun Abebe for editing assistance with the final manuscript.
About this article
Cite this article
Lowry, D.M., Greiner, D., Fretheim, M. et al. Mechanism of metolachlor action due to alterations in cell cycle progression. Cell Biol Toxicol 29, 283–291 (2013). https://doi.org/10.1007/s10565-013-9256-z
- Cell cycle
- Retinoblastoma protein