Lipase Expression and Activity in Saliva in a Healthy Population

Abstract

P-252 Abstract: Phthalates are widely distributed environmental contaminants found in cosmetic products, children's toys and food packaging, and are known to act as “endocrine disrupters.” Phthalates (diesters) are first metabolized to more water-soluble monoesters, the major metabolites found in human urine. Phthalates not converted to the diesters are not thought to be biologically active. Lipase enzymes hydrolyze phthalates to monoesters. Monoesters have been detected in human saliva, and saliva has been shown to be active in metabolizing diethylphthalate to monoethylphthalate. We have begun a study aimed at determining the utility of using salivary lipase activity as a biomarker for biologically relevant phthalate dose. Saliva samples were collected from 85 women with informed consent and stored at –70°C. These women were enrolled in a prospective study of the effects of prenatal exposures to pesticides and endocrine disrupting chemicals on birth outcomes and neurodevelopment. As the source of lipase enzyme in saliva is not known, we have also studied the expression of five candidate lipase genes in the major salivary glands and tongue. Real time RT-PCR was carried out using two primer sets per gene and normalization to two different housekeeping genes (ALAD and beta-actin). Expression of both gastric and pancreatic lipase mRNA was demonstrated in total mRNA from human tongue and confirmed by sequencing. Gastric and pancreatic lipases were equally expressed. Higher expression levels were detected in tongue than in two major salivary glands. Expression level in tongue for both genes was about 1% of level of ALAD and 0.05% of level of beta-actin. Lipase activity and α-amylase were measured using specific colorimetric assays. Amylase was originally chosen for normalization for salivary protein. Mean lipase and amylase activities varied significantly between individuals. Mean lipase and amylase activities were 5.45 U/L (SD ±6.96) and 47.25U/ml (SD ±38.76), respectively. Correlation between lipase and amylase was low (R2=0.208), suggesting different glands of origin. Lipase activity in saliva was low, as supported by the observed level of expression of lipase genes. High inter-individual variation in the lipase activity suggests a regulatory role of environmental factors. We are studying correlations between phthalate metabolism and lipase activity to determine its potential role as a biomarker for phthalate activation.