title: Ethanol exposure induces oxidative stress and impairs nitric oxide availability in the human placental villi: a possible mechanism of toxicity.
abstract: OBJECTIVE
  We undertook this investigation to explore the effects of ethanol exposure on nitric oxide synthase levels and nitric oxide release. Our hypothesis was that ethanol exposure modifies nitric oxide activity within the placenta as a result of oxidative stress.
  
  
  METHODS
  Four 10-g samples of term normal human placental villous tissue were perifused with nonrecirculating Dulbecco's modified Eagle's medium and 25-mmol/L N-[2-hydroxyethyl]piperazine-N'-[2-ethanesulfonic acid] with 0-, 50-, 100-, or 200-mmol/L ethanol. After 2 hours of exposure, tissue was removed, fixed, and frozen for analysis. Immunohistochemical analysis was performed for subtype I or neuronal nitric oxide synthase (nNOS), subtype II or inducible nitric oxide synthase (iNOS), and subtype III or endothelial nitric oxide synthase (eNOS) localization. Western blot analysis was performed for eNOS quantitation. Cyclic guanosine monophosphate and copper-zinc superoxide dismutase levels were measured by electroimmunoassay and kinetic assay, respectively. Nitric oxide release was analyzed by a Sievers nitric oxide analyzer.
  
  
  RESULTS
  Immunohistochemical examination confirmed that only eNOS was localized to the syncytiotrophoblasts. After ethanol exposure, eNOS protein expression increased 2.5- to 3.0-fold over that of the control. Tissue cyclic guanosine monophosphate content and nitric oxide release into the effluent were decreased, whereas superoxide dismutase levels were increased at higher ethanol levels (P <.05).
  
  
  CONCLUSIONS
  Ethanol exposure appears to induce oxidative stress, which may account for the decreased nitric oxide release, because nitric oxide may be shunted toward scavenging free radicals. Increased eNOS protein expression may be a response to the increased demand for nitric oxide. Decreased nitric oxide availability could adversely affect placental blood flow regulation, which could, in turn, account for the growth restriction seen in ethanol-exposed fetuses.
authors.affiliation: Department of Obstetrics and Gynecology and the Perinatal Research Laboratories, University of Wisconsin, Madison, WI 53715, USA.
authors.forename: H H
authors.initials: HH
authors.lastname: Kay
authors.forename: K M
authors.initials: KM
authors.lastname: Grindle
authors.forename: R R
authors.initials: RR
authors.lastname: Magness
mesh_terms: D015153:Blotting, Western; D002824:Chorionic Villi / Q000187:drug effects / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D000431:Ethanol / Q000009:adverse effects* / Q000633:toxicity; D005260:Female; D005317:Fetal Growth Retardation / Q000209:etiology; D006801:Humans; D007150:Immunohistochemistry; D066298:In Vitro Techniques; D009569:Nitric Oxide / Q000378:metabolism; D019001:Nitric Oxide Synthase / Q000378:metabolism; D052250:Nitric Oxide Synthase Type III; D018384:Oxidative Stress; D010920:Placenta / Q000098:blood supply / Q000187:drug effects* / Q000378:metabolism; D011247:Pregnancy
chemical_list: D009569:Nitric Oxide; D000431:Ethanol; C496314:NOS3 protein, human; D019001:Nitric Oxide Synthase; D052250:Nitric Oxide Synthase Type III