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BAC-TRAP technology in neurotoxicology: assessing the astrocyte response to MPTP-induced damage in the ALDH1L1 BAC-TRAP mouse.

Miller-DB; Kelly-KA; O'Callaghan-JP
Toxicologist 2014 Mar; 138(1):95
A central problem in neurotoxicology is detecting the selective and unpredictable damage to specific cells produced by toxic agents and mixtures. Evaluating astrogliosis overcomes this problem because reactive astrocytes show the location of toxicant-induced damage occurring anywhere in the CNS. Enhanced expression of GFAP is a hallmark of reactive astrocytes; however, few other astrogliosis biomarkers are known. Heintz & Greengard (2008) introduced BAC-TRAP (translating ribosome affinity purification) technology that allows the characterization of the actively translating transcriptome of a particular cell type. For example, ALDH1L1 is an astrocyte specific enzyme thought to be a housekeeping gene. Thus, ALDH1L1 BAC-TRAP mice can be used to characterize the transcriptome of astrocytes under various conditions. To begin to characterize additional biomarkers of astrogliosis occurring in response to neurotoxic damage ALDH1L1 BAC-TRAP mice were given a single 12.5 mg/kg s.c. dose of MPTP, a well characterized dopaminergic neurotoxicant that induces significant astrogliosis, Striatal tissue was obtained at 12, 24, and 48 hrs following a single s.c. dosage of saline or 12.5 mg/kg MPTP. Striatal tissue was subjected to TRAP utilizing an eGFP antibody that only binds to actively translating RNA in astrocytes. Changes in the actively translating RNA induced by MPTP damage were determined by microarray (Illumina MouseWG-8 v2 Expression BeadChip) and the dataset interrogated using Ingenuity Pathway Analysis (IPA). MPTP induced robust transcriptome changes in genes previously identified as astrocyte specific (e.g., 403, 399, 804 fold increases in TIMP1 at 12, 24, 48 hrs, respectively) as well as others not previously considered astrocyte-specific (e.g., 219, 203, 3.69 fold increases in PHOX2A at 12, 24, 48 hrs). Our data indicate the BAC-TRAP technology can be used to identify additional biomarkers of astrogliosis and will aid in characterizing various astrocyte phenotypes.
Toxicology; Neurotoxicity; Models; Cell-function; Cellular-function; Toxic-effects; Biomarkers; In-vivo-study; Neurotoxins; Enzymes; Laboratory-animals; Animals; Genes; Neurological-system; Neurological-reactions; Cell-damage
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The Toxicologist. Society of Toxicology 53rd Annual Meeting and ToxExpo, March 23-27, 2014, Phonex, Arizona