Inhibition of Nitric Oxide Production Rescues LPS-Induced Fetal Abortion in Mice I. Athanassakis,* ,1 I. Aifantis,* , † A. Ranella,* K. Giouremou,* and S. Vassiliadis* *Department of Biology, University of Crete, Heraklion, Crete, Greece; and †Ho ˆpital Necker, Unite INSERM 373, Paris, France Received February 18, 1999, and in revised form April 5, 1999 In this report, we examined the involvement of the cytokines tumor necrosis factor (TNF)-, inter- feron (IFN)-, interleukin (IL)-4, and IL-10 as well as nitric oxide (NO) in the lipopolysaccharide (LPS)- induced experimental abortion model in BALB/c mice. Although in vivo administration of LPS in pregnant mice showed a 72% decrease of serum IL- 10, no significant difference in serum TNF-, IFN-, and IL-4 levels, compared to controls, could be de- tected. At the same time, a correlation of fetal abor- tion and maternal splenomegaly with an important increase of NO synthesis in the serum was obtained. Simultaneous administration of LPS and aminogua- nidine (AG; an inhibitor to NO synthase) rescued the LPS-induced fetal abortion, reduced maternal spleen weight to physiological levels, and decreased serum NO concentration to control levels. In vitro experiments showed that LPS directly induced NO production in primary placental cells and the TPOPHO-1 trophoblast cell line by stimulating the inducible isoform of NO synthase, which ultimately could be blocked by the NO synthase inhibitors AG and L-NAME. The results indicate that LPS, despite its beneficial involvement in intracellular infec- tions, participates in inflammatory/autoimmune damage during pregnancy, leading to embryo- toxicity, which is closely linked to the NO pathway. © 1999 Academic Press Key Words: nitric oxide; LPS; fetal abortion; ami- noguanidine; L-NAME; trophoblasts; TNF-; IFN-; IL-4; IL-10. Nitric oxide (NO) 2 is synthesized by conversion of oxygen and L-arginine to NO and L-citrulline by the enzyme nitric oxide synthase (NOS), which is found in three different isoforms, each with specific chro- mosomal localization and tissue distribution (1–3). The endothelial (e-NOS) and cerebral (n-NOS) iso- forms have so far been associated with blood pres- sure regulation and neurotransmission, whereas the inducible (i-NOS) isoform is suggested to be involved in macrophage defense mechanisms against infec- tions (2, 4, 5). Thus, induction of the NO pathway by cytokines such as interferon (IFN)-, tumor necrosis factor (TNF)-, and interleukin (IL)-1 has been detected in a variety of cell types, including murine macrophages (6), endothelial cells (7), and -pancreatic cells (8). Although there is evidence suggesting that NO induction is protective in vivo against intracellular infections (leishmaniasis, liste- riosis, and blood-stage malaria; 9 –11), other reports suggest that it can mediate islet -cell autoimmune 1 To whom correspondence should be addressed at Department of Biology, University of Crete, P.O. Box 2208, 714 09 Heraklion, Crete, Greece. Fax: +30 (081) 394.379. E-mail: athan@biology. uch.gr. 2 Abbreviations used: NO, nitric oxide; NOS, nitric oxide syn- thase; e, endothelial; n, cerebral; i, inducible; IFN, interferon; TNF, tumor necrosis factor; IL, interleukin; LPS, lipopolysaccha- ride; AG, aminoguanidine; DMEM, Dulbecco’s modified essential medium; FCS, fetal calf serum; PBS, phosphate-buffered saline; FITC, fluorescein isothiocyanate; ELISA, enzyme-linked immu- nosorbent assay. NITRIC OXIDE: Biology and Chemistry Vol. 3, No. 3, pp. 216 –224 (1999) Article ID niox.1999.0224, available online at http://www.idealibrary.com on 216 1089-8603/99 $30.00 Copyright © 1999 by Academic Press All rights of reproduction in any form reserved.