Background: Pregnancy is characterized by an inflammatory-like process and this may be exacerbated in preeclampsia. The heme oxygenase (HO) enzymes generate carbon monoxide (CO) that induces blood vessel relaxation and biliverdin that acts as an endogenous antioxidant. Materials and Methods:We examined the expression and localization of HO-1 and HO-2 in normal and preeclamptic placenta using reverse transcription polymerase chain reaction (RT-PCR), RNase protection assay, immunoblotting and immunohistochemistry. In addition, the effect of HO activation on tumor necrosis factor-alpha (TNF␣) induced placental damage and on feto-placental circulation was studied. Results: We provide the first evidence for the role of HO as an endogenous placental factor involved with cytoprotection and placental blood vessel relaxation. HO-1 was significantly higher at term, compared with first trimester placentae indicating its role in placental vascular development and regulation. HO-1 predominantly localized in the extravascular connective tissue that forms the perivascular contractile sheath around the developing blood vessels. HO-2 was localized in the capillaries, as well as the villous stroma, with weak staining of trophoblast. Induction of HO-1 caused a significant attenuation of TNF␣mediated cellular damage in placental villous explants, as assessed by lactate dehydrogenase leakage (p Ͻ 0.01). HO-1 protein was significantly reduced in placentae from pregnancies complicated with preeclampsia, compared with gestationally matched normal pregnancies. This suggests that the impairment of HO-1 activation may compromise the compensatory mechanism and predispose the placenta to cellular injury and subsequent maternal endothelial cell activation. Isometric contractility studies showed that hemin reduced vascular tension by 61% in U46619-preconstricted placental arteries. Hemininduced vessel relaxation and CO production was inhibited by HO inhibitor, tin protoporphyrin IX. Conclusions: Our findings establish HO-1 as an endogenous system that offers protection against cytotoxic damage in the placenta, identifies the HO-CO pathway to regulate feto-placental circulation and provides a new approach to study the disease of preeclampsia.

Human placental development involves coordinated angiogenesis and trophoblast outgrowth that are compromised in intrauterine growth restriction (IUGR). As Tie-2(؊/؊) mice exhibit growth retardation and vascular network malformation, the expression of Tie-2 and its ligands, angiopoietin-1 (Ang-1) and angiopoietin-2 (Ang-2), were investigated in human placenta from normal pregnancies and those complicated by severe IUGR. Ribonucleotide protection assays showed no significant change in the expression of Ang-2 mRNA between gestationally matched normal and IUGR placentas; however, immunoblots revealed that Ang-2 protein was significantly decreased in IUGR, suggesting that this may contribute to the abnormal development of the villous vasculature. In situ hybridization studies showed that Ang-1 and Tie-2 were detected in the cyto/syncytiotrophoblast bilayer in first-trimester placenta, whereas Ang-2 mRNA was restricted to the cytotrophoblast, suggesting their role in trophoblast function. At term, Ang-1 mRNA and immunoreactive protein were restricted to the paravascular tissues of the primary stem villi, supporting its role in vessel maturation. In contrast, Ang-2 was expressed throughout the term villous core, perhaps to permit the developing placental vascular network to remain in a state of fluidity. As these studies also revealed that trophoblast, in addition to endothelial cells, expressed Tie-2 receptors, we investigated the potential role of Ang-1/Ang-2 on trophoblast proliferation, migration, and the release of NO. Successful placentation requires the development of a low-impedance uteroplacental circulation after transformation of the maternal intramyometrial portion of the spiral arterioles by trophoblast invasion.1 Hemochorial placentation is also dependent on the establishment and maintenance of a competent fetoplacental vascular network formed by the processes of vasculogenesis and branching (first and second trimesters) and nonbranching (third trimester) angiogenesis.

Notch signaling is an evolutionarily conserved mechanism, used to regulate cell fate decisions. Four Notch receptors have been identified in man (Notch‐1 to ‐4). In this study, semiquantitative reverse transcription polymerase chain reaction (RT‐PCR) and immunohistochemistry were used to examine the expression pattern of Notch receptor genes in whole adult human liver and isolated liver cell preparations. All 4 receptors were expressed in the adult liver, with no significant differences in the levels of Notch‐1, ‐2, and ‐4 messenger RNA (mRNA) between normal and diseased liver. However, Notch‐3 expression appeared to be increased in diseased tissue. The distribution of Notch‐1 and ‐4 in normal tissue was similar, with Notch‐1 also detectable at low levels in the sinusoidal endothelium. Notch‐2 expression was more widely distributed, and detectable in hepatocytes, medium‐sized bile ducts, and the sinusoidal endothelium. Notch‐3 expression was seen on hepatocytes, with weaker expression detectable in portal veins, hepatic arteries, and the sinusoids. In normal liver tissue Notch‐1, ‐2, and ‐3 were found to be coexpressed on bile duct epithelium; however, with the exception of Notch‐3 in primary sclerosing cholangitis (PSC) livers, expression was absent on proliferating ductules in all disease states examined. Interestingly, the expression of Notch‐2 and ‐3 was associated with numerous small vessels within the portal tract septa of diseased tissue. The absence of Notch receptor expression on proliferating bile ductules and its presence on neovessels suggests that Notch signaling may be important for normal bile duct formation and the aberrant neovascularization seen in diseased liver tissue.