Characterization of lipoproteins in human placenta and fetal circulation as well as gestational changes in lipoprotein assembly and secretion in human and mouse placentas.

In the maternal circulation, apoB-containing low-density lipoproteins (LDL) and apoA1-containing high-density lipoproteins (HDL) transport lipids. The production of lipoproteins in the placenta has been suggested, but the directionality of release has not been resolved. We compared apolipoprotein concentrations and size-exclusion chromatography elution profiles of lipoproteins in maternal/fetal circulations, and in umbilical arteries/veins; identified placental lipoprotein-producing cells; and studied temporal induction of lipoprotein-synthesizing machinery during pregnancy. We observed that maternal and fetal lipoproteins are different with respect to concentrations and elution profiles. Surprisingly, concentrations and elution profiles of lipoproteins in umbilical arteries and veins were similar indicating their homeostatic control. Human placental cultures synthesized apoB100-containing LDL-sized and apoA1-containing HDL-sized particles. Immunolocalization techniques revealed that ApoA1 was present mainly in syncytiotrophoblasts. MTP, a critical protein for lipoprotein assembly, was in these trophoblasts. ApoB was in the placental stroma indicating that trophoblasts secrete apoB-containing lipoproteins into the stroma. ApoB and MTP expressions increased in placentas from the 2nd trimester to term, whereas apoA1 expression was unchanged. Thus, our studies provide new information regarding the timing of lipoprotein gene induction during gestation, the cells involved in lipoprotein assembly and the gel filtration profiles of human placental lipoproteins. Next, we observed that mouse placenta produces MTP, apoB100, apoB48 and apoA1. The expression of genes gradually increased and peaked in late gestation. This information may be useful in identifying transcription factors regulating the induction of these genes in gestation and the importance of placental lipoprotein assembly in fetal development.

2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) enhances placental inflammation.

Preterm birth is a leading cause of perinatal morbidity and mortality that is often associated with ascending infections from the lower genital tract. Recent studies with animal models have suggested that developmental exposure to the environmental toxin 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) can increase the risk of preterm birth in the offspring. How TCDD may modify placental immunity to ascending infections is unclear. Therefore, we studied the effects of TCDD treatment on basal and Escherichia coli-stimulated cytokine production by placental explants. Cultures of second-trimester placentas were treated with up to 40 nM TCDD for 72 h and then stimulated with 10(7)CFU/ml E. coli for an additional 24h. Concentrations of cytokines and PGE2 were measured in conditioned medium by immunoassay. TCDD exposure increased mRNA levels of IL-1ß by unstimulated cultures, but no effects on protein levels of this cytokine were detected. TNF-α production was unaffected by TCDD for unstimulated cultures, but pre-treatment with 40 nM TCDD significantly increased E. coli-stimulated TNF-α production. Both basal and bacteria-stimulated PGE2 and COX-2 gene expression were enhanced by TCDD pretreatment. In contrast, production of the anti-inflammatory cytokine, IL-10, was reduced by TCDD pretreatment for both unstimulated and E. coli-stimulated cultures. No effect of TCDD on the viability of the cultures was detected. These results suggest that TCDD exposure may shift immunity to enhance a proinflammatory phenotype at the maternal-fetal interface that could increase the risk of infection-mediated preterm birth.

Can carbon monoxide prevent infection-mediated preterm birth in a mouse model?

PROBLEM: Preterm birth is frequently caused by intrauterine infection and inflammation. Recent studies have demonstrated that carbon monoxide (CO), which is produced endogenously, has potent anti-inflammatory properties. Whether or not CO can prevent infection-mediated preterm birth is unknown. METHODS: Mice were assigned to one of four groups: sham infection, sham infection + CO, infection, or infection + CO. Infections were established by intra-uterine injection of Escherichia coli on day 14 of pregnancy. Animals received daily i.p. injections of 1 mL CO-saturated lactated ringers solution (LRS) or LRS alone beginning on the morning of surgery. Gestational age at delivery and litter characteristics was noted. In second experiment, animals were sacrificed 24 hrs post-surgery and tissues were harvested for cytokine analyses. RESULTS: Escherichia coli intrauterine infection increased the number of animals delivering preterm. This effect was significantly ameliorated by CO-LRS. CO-treatment also increased litter size and weights of the surviving offspring. Cytokines in the amniotic fluid and the placenta were increased by E. coli exposure, but CO had no detectible effect on E. coli-stimulated cytokine production. No effects of CO were detected in sham-infected animals. CONCLUSION: Supplemental CO improves pregnancy outcome after intrauterine infection and may function at a point downstream of, or through pathways independent of, induction of proinflammatory cytokines.

Effect of carbon monoxide on bacteria-stimulated cytokine production by placental explants.

PROBLEM: Preterm birth is frequently caused by an inflammatory response to ascending infections of the reproductive tract. Carbon monoxide (CO) has potent anti-inflammatory properties at subtoxic concentrations. Whether or not CO can modulate inflammatory responses by placental tissues is unclear. METHODS: Placental explant cultures were incubated with heat-killed Escherichia coli or Ureaplasma parvum in the presence or absence of 250 ppm CO for 24 hr. Concentrations of cytokines relative viability of the cultures were quantified. RESULTS: Escherichia coli- and U. parvum-stimulated IL-1ß production was significantly inhibited by CO supplementation. Escherichia coli-stimulated, but not U. parvum-stimulated, IFN-γ production was inhibited by CO. While CO inhibited PGE(2) production by unstimulated cells, no effects on bacteria-stimulated prostaglandin production were detected. CO had no effect on basal or E. coli-stimulated TNF-α production but enhanced TNF-α production by cultures stimulated with U. parvum. In addition, CO tended to improve the viability of the placental cultures. CONCLUSIONS: Low concentrations of CO tended to reduce proinflammatory cytokines and to promote the production of anti-inflammatory cytokines in a pathogen-specific manner. These properties suggest that CO may be useful for promoting a pro-pregnancy cytokine milieu by placental explants and may reduce the consequences of intrauterine infections.

Can oxygen tension contribute to an abnormal placental cytokine milieu?

OBJECTIVE The aim of this study was to determine whether culturing human placental explants under different oxygen tensions will alter expression of pro- and anti-inflammatory mediators. METHODS Placental explant cultures from second-trimester, elective, terminations-of-pregnancy were incubated under 21, 5, or 1% O(2) concentrations for 24 hr in the presence or absence of IL-10. Cytokine concentrations in the conditioned medium were quantified by immunoassay. RESULTS Culture of placental explants under 21, 5, or 1% O(2) concentrations produced hyperoxic (143 ± 1.6 mmHg), normoxic (37 ± 1.6 mmHg), and hypoxic (18.2 ± 1.6 mmHg) pO(2) levels for the maternal-fetal interface in the medium. Oxygen tension had profound effects on basal placental cytokine levels as well as on IL-10-stimulated cytokine production. IL-1ß and TNF-α, but not IFN-γ production, was reduced by 21% O(2) . Moreover, 21% O(2) levels increased the anti-inflammatory cytokines IL-10 and IL-13 while 1% O(2) tended to decrease the production of these cytokines. CONCLUSIONS Five percent- O(2) incubation more accurately represents in vivo pO(2) conditions at the maternal-fetal interface. Routine culture of placental explants in room air produces a superphysiologic oxygen tension that tended to increase the production of anti-inflammatory and decrease the production of pro-inflammatory cytokines. In addition, low pO(2) may reduce responsiveness of the placenta to the anti-inflammatory actions of IL-10.