title: Duodenal crypt health following exposure to Cr(VI): Micronucleus scoring, γ-H2AX immunostaining, and synchrotron X-ray fluorescence microscopy.
abstract: Lifetime exposure to high concentrations of hexavalent chromium [Cr(VI)] in drinking water results in intestinal damage and an increase in duodenal tumors in B6C3F1 mice. To assess whether these tumors could be the result of a direct mutagenic or genotoxic mode of action, we conducted a GLP-compliant 7-day drinking water study to assess crypt health along the entire length of the duodenum. Mice were exposed to water (vehicle control), 1.4, 21, or 180 ppm Cr(VI) via drinking water for 7 consecutive days. Crypt enterocytes in Swiss roll sections were scored as normal, mitotic, apoptotic, karyorrhectic, or as having micronuclei. A single oral gavage of 50mg/kg cyclophosphamide served as a positive control for micronucleus induction. Exposure to 21 and 180 ppm Cr(VI) significantly increased the number of crypt enterocytes. Micronuclei and γ-H2AX immunostaining were not elevated in the crypts of Cr(VI)-treated mice. In contrast, treatment with cyclophosphamide significantly increased numbers of crypt micronuclei and qualitatively increased γ-H2AX immunostaining. Synchrotron-based X-ray fluorescence (XRF) microscopy revealed the presence of strong Cr fluorescence in duodenal villi, but negligible Cr fluorescence in the crypt compartment. Together, these data indicate that Cr(VI) does not adversely effect the crypt compartment where intestinal stem cells reside, and provide additional evidence that the mode of action for Cr(VI)-induced intestinal cancer in B6C3F1 mice involves chronic villous wounding resulting in compensatory crypt enterocyte hyperplasia.
authors.affiliation: ToxStrategies, Inc., Katy, TX 77494, USA. Electronic address: cthompson@toxstrategies.com.
authors.forename: Chad M
authors.initials: CM
authors.lastname: Thompson
authors.affiliation: Experimental Pathology Laboratories, Sterling, VA 20166, USA. Electronic address: JWolf@epl-inc.com.
authors.forename: Jeffrey C
authors.initials: JC
authors.lastname: Wolf
authors.affiliation: BioReliance, Rockville, MD, USA. Electronic address: reem.elbekai@bioreliance.com.
authors.forename: Reem H
authors.initials: RH
authors.lastname: Elbekai
authors.affiliation: BioReliance, Rockville, MD, USA. Electronic address: madhav.paranjpe@bioreliance.com.
authors.forename: Madhav G
authors.initials: MG
authors.lastname: Paranjpe
authors.affiliation: U.S. Army Engineer Research and Development Center, Vicksburg, MS 39180, USA. Electronic address: Jennifer.M.Seiter@erdc.dren.mil.
authors.forename: Jennifer M
authors.initials: JM
authors.lastname: Seiter
authors.affiliation: U.S. Army Engineer Research and Development Center, Vicksburg, MS 39180, USA. Electronic address: Mark.A.Chappell@usace.army.mil.
authors.forename: Mark A
authors.initials: MA
authors.lastname: Chappell
authors.affiliation: Photon Sciences Department, Brookhaven National Laboratory, Upton, NY 11973, USA. Electronic address: rtappero@bnl.gov.
authors.forename: Ryan V
authors.initials: RV
authors.lastname: Tappero
authors.affiliation: ToxStrategies, Inc., Mission Viejo, CA 92692, USA. Electronic address: msuh@toxstrategies.com.
authors.forename: Mina
authors.initials: M
authors.lastname: Suh
authors.affiliation: ToxStrategies, Inc., Mission Viejo, CA 92692, USA. Electronic address: dproctor@toxstrategies.com.
authors.forename: Deborah M
authors.initials: DM
authors.lastname: Proctor
authors.affiliation: ToxStrategies, Inc., Austin, TX 78731, USA. Electronic address: abichteler@toxstrategies.com.
authors.forename: Anne
authors.initials: A
authors.lastname: Bichteler
authors.affiliation: ToxStrategies, Inc., Austin, TX 78731, USA. Electronic address: lhaws@toxstrategies.com.
authors.forename: Laurie C
authors.initials: LC
authors.lastname: Haws
authors.affiliation: ToxStrategies, Inc., Katy, TX 77494, USA. Electronic address: mharris@toxstrategies.com.
authors.forename: Mark A
authors.initials: MA
authors.lastname: Harris
mesh_terms: D000818:Animals; D002857:Chromium / Q000008:administration & dosage / Q000633:toxicity*; D004305:Dose-Response Relationship, Drug; D060766:Drinking Water; D004386:Duodenum / Q000187:drug effects* / Q000378:metabolism; D020895:Enterocytes / Q000187:drug effects / Q000378:metabolism; D005260:Female; D006657:Histones / Q000378:metabolism*; D007150:Immunohistochemistry; D008297:Male; D008822:Mice, Transgenic; D048629:Micronuclei, Chromosome-Defective / Q000139:chemically induced* / Q000706:statistics & numerical data; D008856:Microscopy, Fluorescence / Q000295:instrumentation / Q000379:methods*; D008940:Mitotic Index; D017356:Synchrotrons; D014965:X-Rays
chemical_list: D060766:Drinking Water; D006657:Histones; C475961:gamma-H2AX protein, mouse; D002857:Chromium; C074702:chromium hexavalent ion
keywords: Carcinogenesis; Cr(VI); Duodenum; Hexavalent chromium; Mode of action; Synchrotron; γ-H2AX