Di(2-ethylhexyl) phthalate Is a Highly Potent Agonist for the Human Constitutive Androstane Receptor Splice Variant CAR2

The human constitutive androstane receptor (CAR, CAR1) regulates the expression of genes involved in xenobiotic metabolism in the liver. The CAR gene uses multiple alternative splicing events during pre-mRNA processing, thereby enhancing the CAR transcriptome. Previous reports have identified two prominent human CAR variants, CAR2 and CAR3, that possess four- and five-amino acid insertions in their ligand binding domains, respectively. Unlike the constitutively active reference form of the receptor, we now demonstrate that CAR2 is a ligand-activated receptor and comprises approximately 30% of the reference transcript level in human liver tissues in human hepatocytes. Furthermore, we identify the common plasticizer, di(2-ethylhexyl) phthalate (DEHP), as a highly potent and uniquely selective agonist of CAR2. Results from reporter transactivation and mammalian two-hybrid assays reveal that DEHP activates CAR2 at low nanomolar concentrations, results further supported by analysis of CAR target gene expression in primary human hepatocytes. In addition, comparative genomic analyses show that the typical mouse, rat, and marmoset models of DEHP toxicity cannot accurately profile potential human toxicity because of these species' inability to generate a CAR2-like transcript. The discovery that CAR2 is an ultimate human DEHP receptor identifies a novel pathway modulating human DEHP toxicity with potential clinical implications for a subset of patients undergoing critical care medical interventions

Title Page Di(2-ethylhexyl) phthalate is a highly potent agonist for the human constitutive androstane receptor splice variant, CAR2 MOL 53702 2 Running Title Page Running Title: DEHP is a highly potent agonist for the human CAR splice variant, CAR2

Abbreviations: CAR, constitutive androstane receptor; DEHP, di(2-ethylhexyl) phthalate; PXR, pregnane X receptor MOL 53702 3 Abstract The human constitutive androstane receptor (CAR, CAR1) regulates the expression of genes involved in xenobiotic metabolism in the liver. The CAR gene utilizes multiple alternative splicing events during pre-mRNA processing, thereby enhancing the CAR transcriptome. Previous reports have identified two prominent human CAR variants, CAR2 and CAR3, possessing 4-and 5-amino acid insertions in their ligand binding domains, respectively. Unlike the constitutively active reference form of the receptor, we now demonstrate that CAR2 is a ligand activated receptor and comprises approximately 30% of the reference transcript level in human liver tissues in human hepatocytes. Further, we identify the common plasticizer, di(2-ethylhexyl) phthalate (DEHP), as a highly potent and uniquely selective agonist of CAR2. Results from reporter transactivation and mammalian two-hybrid assays reveal that DEHP activates CAR2 at low nanomolar concentrations, results further supported by analysis of CAR target gene expression in primary human hepatocytes. In addition, comparative genomic analysis show that the typical mouse, rat and marmoset models of DEHP toxicity can not accurately profile potential human toxicity due to these species inability to generate a CAR2-like transcript. The discovery that CAR2 is an ultimate human DEHP receptor identifies a novel pathway modulating human DEHP toxicity with potential clinical implications for a subset of patients undergoing critical care medical interventions. MOL 53702

Feasibility of Low Radiation Dose Retrospectively-Gated Cardiac CT for Functional Analysis in Adult Congenital Heart Disease.

AbstractBackgroundThe use of cardiac computed tomography (CT) in the evaluation of adult congenital heart disease patients is limited due to concerns of high radiation doses. The purpose of this study was to prospectively assess whether low radiation dose cardiac CT is feasible to evaluate ventricular systolic function in adults with congenital heart disease.MethodsThe study group included 30 consecutive patients with significant congenital heart disease who underwent a total of 35 ECG-gated cardiac CT scans utilizing a 320-detector row CT scanner. Each study included a non-contrast scan and subsequent contrast-enhanced retrospectively-gated acquisition. Effective radiation dose was estimated by multiplying the dose length product by a k-factor of 0.014mSv/mGycm.ResultsThe mean age of the patients was 34.4±8.9years, 60% were men, and mean body mass index was 24.2±4.3kg/m2. A majority of patients (n=28, 93.3%) had contraindications to cardiac MRI. A tube potential of 80kV was used in 27 (77.1%) of the contrast-enhanced scans. The mean signal-to-noise and contrast-to-noise ratios were 11.5±3.9 and 10.3±3.7, respectively. The median radiation dose for non-contrast and contrast-enhanced images were 0.1mSv (0.07–0.2mSv) and 0.94mSv (0.5–2.1mSv), respectively. All 35 CT scans were successfully analyzed for ventricular systolic function.ConclusionsA low radiation contrast-enhanced, retrospectively-gated cardiac CT with a median radiation dose of less than 1mSv was successful in evaluating ventricular systolic function in 30 consecutive adult congenital heart disease patients who underwent a total of 35 scans