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INTEGRATED DISINFECTION BY-PRODUCTS (DBP) MIXTURES RESEARCH: GENE EXPRESSION ALTERATIONS IN PRIMARY RAT HEPATOCYTE CULTURES EXPOSED TO DBP MIXTURES FORMED BY CHLORINATION AND OZONATION/POSTCHLORINATION
Citation:
CROSBY, L. M., W. O. WARD, T. MOORE, J. E. SIMMONS, A. B. DEANGELO, AND K. T. Morgan. INTEGRATED DISINFECTION BY-PRODUCTS (DBP) MIXTURES RESEARCH: GENE EXPRESSION ALTERATIONS IN PRIMARY RAT HEPATOCYTE CULTURES EXPOSED TO DBP MIXTURES FORMED BY CHLORINATION AND OZONATION/POSTCHLORINATION. JOURNAL OF TOXICOLOGY AND ENVIRONMENTAL HEALTH. Taylor & Francis, Inc., Philadelphia, PA, 71(17):1195-1215, (2008).
Impact/Purpose:
There exisits a limited amount of data on complex mixtures of DBPs. The primary purpose of the 'Four Lab' was directed toward filling this important data gap with coordinated chemical and toxicological evaluation of environmentally realistic concentrates of complex mixtures of DBPs. This manuscript will be part of series of papers with the aim of providing comprehensive information on all phases of DBP research using concentrates derived from “real-life” conditions.
What is the impact to the field and the Agency?
The data developed from the study provide an interesting example of the use of differential gene expression array technology for monitoring the possible health effects of finished drinking water resulting from the uses of different disinfectants. They demonstrate the utility of genomic approaches in assessment and characterization of the biological activity of environmentally realistic mixtures of DBPs. The present results might suggest the implementation of further studies with improved design criteria, provide the agency information which might prove helpful in the development of risk assessments, and have a potential to directly benefit public health.
Abstract
Large scale differential gene expression analysis was used to determine biological effects of disinfected surface waters on cultured rat hepatocytes. Source water from East Fork Lake (Harsha Lake), a reservoir on the Little Miami River in Ohio, was spiked with iodide and bromide and disinfected by chlorination or ozonation/postchlorination. The treated waters were concentrated approximately 125 fold (full strength) using reverse osmosis membrane techniques. Volatile disinfection by-products (DBPs) lost during concentration were restored to the extent possible. Primary rat hepatocytes were exposed to full-strength, 1:10 and 1:20 dilutions of the concentrates for 24 hr and assayed for cytotoxicity and gene expression alterations. The full-strength concentrates were cytotoxic: The diluted samples exhibited no cytotoxicity. Differential gene expression analysis provided evidence for the underlying causes of the severe cytotoxicity observed in rat hepatocytes treated with the full strength ozonation/postchlorination concentrate (e.g., cell cycle arrest, metabolic stasis, oxidative stress, cell death). Many gene expression responses were shared among the hepatocyte cultures treated with dilutions of the ozonation/ postchlorination and chlorination concentrates. The shift in the character of the response between the full strength concentrates and the diluted samples indicated a threshold for toxicity. A small subset of gene expression changes was identified that had been observed in the response of hepatocytes to PPARs, phthalate esters, and haloacetic acids suggesting a peroxisome proliferative response.
Description:
What is the study?
This study was designed to provide data on the in vitro toxicity of water concentrates containing complex mixtures of DBPs. Rat hepatocytes in primary culture were exposed for 24 hr to full strength, 1:10 or 1:20 dilutions of chlorination or ozonation/chlorination samples. Cytotoxicity, metabolic activity, and gene expression alterations, as measured by array technology and real-time RT-PCR, were evaluated. Results were compared to known PPAR- and DEHP- (diethylhexylphthalate) activated mRNA alterations and functional changes induced by a series of haloacetic acids
Why was it done?
There exisits a limited amount of data on complex mixtures of DBPs. The primary purpose of the 'Four Lab' was directed toward filling this important data gap with coordinated chemical and toxicological evaluation of environmentally realistic concentrates of complex mixtures of DBPs. This manuscript will be part of series of papers with the aim of providing comprehensive information on all phases of DBP research using concentrates derived from “real-life” conditions.
What is the impact to the field and the Agency?
The data developed from the study provide an interesting example of the use of differential gene expression array technology for monitoring the possible health effects of finished drinking water resulting from the uses of different disinfectants. They demonstrate the utility of genomic approaches in assessment and characterization of the biological activity of environmentally realistic mixtures of DBPs. The present results might suggest the implementation of further studies with improved design criteria, provide the agency information which might prove helpful in the development of risk assessments, and have a potential to directly benefit public health.
Abstract
Large scale differential gene expression analysis was used to determine biological effects of disinfected surface waters on cultured rat hepatocytes. Source water from East Fork Lake (Harsha Lake), a reservoir on the Little Miami River in Ohio, was spiked with iodide and bromide and disinfected by chlorination or ozonation/postchlorination. The treated waters were concentrated approximately 125 fold (full strength) using reverse osmosis membrane techniques. Volatile disinfection by-products (DBPs) lost during concentration were restored to the extent possible. Primary rat hepatocytes were exposed to full-strength, 1:10 and 1:20 dilutions of the concentrates for 24 hr and assayed for cytotoxicity and gene expression alterations. The full-strength concentrates were cytotoxic: The diluted samples exhibited no cytotoxicity. Differential gene expression analysis provided evidence for the underlying causes of the severe cytotoxicity observed in rat hepatocytes treated with the full strength ozonation/postchlorination concentrate (e.g., cell cycle arrest, metabolic stasis, oxidative stress, cell death). Many gene expression responses were shared among the hepatocyte cultures treated with dilutions of the ozonation/ postchlorination and chlorination concentrates. The shift in the character of the response between the full strength concentrates and the diluted samples indicated a threshold for toxicity. A small subset of gene expression changes was identified that had been observed in the response of hepatocytes to PPARs, phthalate esters, and haloacetic acids suggesting a peroxisome proliferative response.
