title: Estrogen blocks 3-nitropropionic acid-induced Ca2+i increase and cell damage in cultured rat cerebral endothelial cells.
abstract: Systemic administration of 3-nitropropionic acid (3-NPA, a mycotoxin) induces brain damage accompanied by disturbance in the blood-brain barrier (BBB). Since the endothelial cells are important components of the BBB and the first target of a systemic intoxication, in the present study, the effect of 3-NPA on primary cultured rat brain endothelial cells (rBECs) was examined by studying intracellular Ca(2+) ([Ca(2+)](i)) response using imaging techniques with fura-2. rBECs were prepared using a method of Kis et al. [Eur. J. Pharmacol. 368 (1999) 35-42] and Szabo et al. [Neurobiology 5 (1997) 1-16]. Almost all cells were immunoreactive to antibody against the factor VIII-related antigen (von-Willebrand factor). They showed a typical dose-dependent increase of [Ca(2+)](i) in response to ATP or bradykinin. Low concentrations of 3-NPA (1.7 mM, 3.4 mM) caused no changes, and a medium concentration (6.8 mM) increased the [Ca(2+)](i) gradually and progressively, and the increase was reversed incompletely back to the resting level after washing. A high concentration (13.6 mM) increased the [Ca(2+)](i) irreversibly. These elevations of [Ca(2+)](i) were absent in a Ca(2+)-free medium. In endothelial cells treated with 17beta-estradiol (above 10(-5) M) or with a selective estrogen receptor modulator, tamoxifen (5 x 10(-7) M), no elevation of [Ca(2+)](i) was observed with 3-NPA treatment. The response to ATP was impaired after application of 3-NPA, but it was preserved by cotreatment with 17beta-estradiol or tamoxifen. An estrogen receptor antagonist ICI 182,780 inhibited these effects by 17beta-estradiol or tamoxifen. Lysosomal neutral red uptake and TUNEL experiments revealed the necrotic but not apoptotic cell death at least in this acute stage. Data indicate that a medium to high concentration of 3-NPA induces damage on rBECs as revealed by an accumulation of [Ca(2+)](i), but the damage was protected by cotreatment with 17beta-estradiol or tamoxifen, suggesting that estrogen may be protective for the brain vascular damage via estrogen receptor.
authors.affiliation: Department of Nephrourology, Nagoya City University, Graduate School of Medical Sciences Kawasumi 1, Mizuhoku, Mizuhocho, Nagoya 467-8601, Japan. mipo@med.nagoya-cu.ac.jp
authors.forename: Mihoko
authors.initials: M
authors.lastname: Mogami
authors.forename: Hideki
authors.initials: H
authors.lastname: Hida
authors.forename: Yutaro
authors.initials: Y
authors.lastname: Hayashi
authors.forename: Kenjiro
authors.initials: K
authors.lastname: Kohri
authors.forename: Yuji
authors.initials: Y
authors.lastname: Kodama
authors.forename: Cha
authors.initials: C
authors.lastname: Gyun Jung
authors.forename: Hitoo
authors.initials: H
authors.lastname: Nishino
mesh_terms: D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D000959:Antihypertensive Agents / Q000008:administration & dosage / Q000494:pharmacology*; D001920:Bradykinin / Q000494:pharmacology; D001921:Brain / Q000187:drug effects / Q000378:metabolism*; D002118:Calcium / Q000032:analysis / Q000378:metabolism*; D020013:Calcium Signaling / Q000187:drug effects; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D004730:Endothelium, Vascular / Q000737:chemistry / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D004958:Estradiol / Q000031:analogs & derivatives* / Q000494:pharmacology; D004965:Estrogen Antagonists / Q000494:pharmacology; D000077267:Fulvestrant; D007150:Immunohistochemistry; D020287:In Situ Nick-End Labeling; D009336:Necrosis; D009574:Nitro Compounds; D011422:Propionates / Q000008:administration & dosage / Q000494:pharmacology*; D051381:Rats; D017208:Rats, Wistar; D013629:Tamoxifen / Q000494:pharmacology; D014841:von Willebrand Factor / Q000378:metabolism
chemical_list: D000959:Antihypertensive Agents; D004965:Estrogen Antagonists; D009574:Nitro Compounds; D011422:Propionates; D014841:von Willebrand Factor; D013629:Tamoxifen; D000077267:Fulvestrant; D004958:Estradiol; D000255:Adenosine Triphosphate; C015392:3-nitropropionic acid; D001920:Bradykinin; D002118:Calcium