title: AHR2 morpholino knockdown reduces the toxicity of total particulate matter to zebrafish embryos.
abstract: The zebrafish embryo has been proposed as a 'bridge model' to study the effects of cigarette smoke on early development. Previous studies showed that exposure to total particulate matter (TPM) led to adverse effects in developing zebrafish, and suggested that the antioxidant and aryl hydrocarbon receptor (AHR) pathways play important roles. This study investigated the roles of these two pathways in mediating TPM toxicity. The study consisted of four experiments. In experiment I, zebrafish embryos were exposed from 6h post fertilization (hpf) until 96hpf to TPM0.5 and TPM1.0 (corresponding to 0.5 and 1.0μg/mL equi-nicotine units) in the presence or absence of an antioxidant (N-acetyl cysteine/NAC) or a pro-oxidant (buthionine sulfoximine/BSO). In experiment II, TPM exposures were performed in embryos that were microinjected with nuclear factor erythroid 2-related factor 2 (Nrf2), AHR2, cytochrome P450 1A (CYP1A), or CYP1B1 morpholinos, and deformities were assessed. In experiment III, embryos were exposed to TPM, and embryos/larvae were collected at 24, 48, 72, and 96hpf to assess several genes associated with the antioxidant and AHR pathways. Lastly, experiment IV assessed the activity and protein levels of CYP1A and CYP1B1 after exposure to TPM. We demonstrate that the incidence of TPM-induced deformities was generally not affected by NAC/BSO treatments or Nrf2 knockdown. In contrast, AHR2 knockdown reduced, while CYP1A or CYP1B1 knockdowns elevated the incidence of some deformities. Moreover, as shown by gene expression the AHR pathway, but not the antioxidant pathway, was induced in response to TPM exposure, providing further evidence for its importance in mediating TPM toxicity.
authors: Nicholas School of the Environment, Duke University, Durham, NC 27708, USA. Electronic address: andrey.massarsky@duke.edu.
authors: Andrey
authors: A
authors: Massarsky
authors: Nicholas School of the Environment, Duke University, Durham, NC 27708, USA.
authors: Audrey J
authors: AJ
authors: Bone
authors: Nicholas School of the Environment, Duke University, Durham, NC 27708, USA; School of Animal Science and Technology, Inner Mongolia Provincial Key Laboratory for Toxicants and Animal Disease, Inner Mongolia University for the Nationalities, Tongliao, Inner Mongolia 028000, China.
authors: Wu
authors: W
authors: Dong
authors: Nicholas School of the Environment, Duke University, Durham, NC 27708, USA.
authors: David E
authors: DE
authors: Hinton
authors: RAI Services Company, Winston-Salem, NC 27101, USA.
authors: G L
authors: GL
authors: Prasad
authors: Nicholas School of the Environment, Duke University, Durham, NC 27708, USA.
authors: Richard T
authors: RT
authors: Di Giulio
mesh_terms: D000818:Animals; D004625:Embryo, Nonmammalian / Q000187:drug effects*; D055785:Gene Knockdown Techniques; D060172:Morpholinos / Q000235:genetics*; D052638:Particulate Matter / Q000633:toxicity*; D018336:Receptors, Aryl Hydrocarbon / Q000235:genetics*; D015027:Zebrafish / Q000196:embryology*
chemical_list: D060172:Morpholinos; D052638:Particulate Matter; D018336:Receptors, Aryl Hydrocarbon
keywords: AHR pathway; Cigarette smoke; Oxidative stress; Total particulate matter (TPM); Zebrafish development