Boris W. Kramer

Pulmonary and systemic endotoxin tolerance in preterm fetal sheep exposed to chorioamnionitis.

In a model of human chorioamnionitis, fetal sheep exposed to a single injection, but not repeated injections, of intra-amniotic endotoxin develop lung injury responses. We hypothesized that repeated exposure to intra-amniotic endotoxin induces endotoxin tolerance. Fetal sheep were given intra-amniotic injections of saline (control) or Escherichia coli LPS O55:B5 (10 mg) either 2 days (2-day group, single exposure), 7 days (7-day group, single exposure), or 2 plus 7 days (2- and 7-day repeat exposure) before preterm delivery at 124 days gestation (term = 150 days). Endotoxin responses were assessed in vivo in the lung and liver, and in vitro in monocytes from the blood and the lung. Compared with the single 2-day LPS exposure group, the (2 plus 7 days) repeat LPS-exposed lambs had: 1) decreased lung neutrophil and monocyte inducible NO synthase (NOSII) expression, and 2) decreased lung cytokine and liver serum amyloid A3 mRNA expression. In the lung, serum amyloid A3 mRNA expression decreased in the airway epithelial cells but not in the lung inflammatory cells. Unlike the single 7-day LPS exposure group, peripheral blood and lung monocytes from the repeat-LPS group did not increase IL-6 secretion or hydrogen peroxide production in response to in vitro LPS. Compared with controls, TLR4 expression did not change but IL-1R-associated kinase M expression increased in the monocytes from repeat LPS-exposed lambs. These results are consistent with the novel finding of endotoxin tolerance in preterm fetal lungs exposed to intra-amniotic LPS. The findings have implications for preterm infants exposed to chorioamnionitis for both responses to lung injury and postnatal nosocomial infections.

Thymic changes after chorioamnionitis induced by intraamniotic lipopolysaccharide in fetal sheep

OBJECTIVE Regulatory T lymphocytes mediate homeostasis of the immune system and differentiate under the control of the transcription factor FoxP3 in the fetal thymus. We asked whether fetal inflammation caused by chorioamnionitis would modulate thymus development. STUDY DESIGN Fetal sheep were exposed to an intraamniotic injection of 10 mg lipopolysaccharide at 5 hours, 1 day, 2 days, or 5 days before delivery at 123 gestation days. Cord blood lymphocytes, plasma cortisol, and thymus weight were measured. Glucocorticoid receptor-, activated caspase-3-, Ki-67-, proliferating cell nuclear antigen-, nuclear factor-kappaB-, and FoxP3-positive cells were immunohistochemically evaluated in thymus. RESULTS Intraamniotic lipopolysaccharide exposure decreased the number of circulating lymphocytes by 40% after 1 day. Thymus-to-body weight ratios were reduced in all lipopolysaccharide groups by a maximum of 40% at 5 days. Lipopolysaccharide exposure modestly increased plasma cortisol concentration, increased nuclear factor-kappaB immunostaining in fetal thymus and reduced the number of FoxP3-positive cells by 40% at 1 day. CONCLUSION Intraamniotic exposure to lipopolysaccharide induced thymic changes and influenced thymic FoxP3 expression.

Intravenous lipopolysaccharide-induced pulmonary maturation and structural changes in fetal sheep

BACKGROUND Antenatal pulmonary inflammation is associated with reduced risk for respiratory distress syndrome but with an increased risk for bronchopulmonary dysplasia (BPD) with impaired alveogenesis. OBJECTIVE We hypothesized that fetal systemic inflammation induced by intravenous (IV) lipopolysaccharide (LPS) would affect lung development in utero. STUDY DESIGN Twenty-one fetal sheep were instrumented (107 days gestational age). Control fetuses received saline (N = 12) and 9 in the study group received 100 ng of LPS IV 3 days after surgery. Animals were assessed for lung maturation and structure after 3 (N = 5) and 7 (N = 4) days. RESULTS Interleukin-6 concentration increased in the bronchoalveolar lavage more than 40-fold 3 days after LPS IV. Processing of pro-surfactant protein (SP)-B to mature SP-B and increased SP-B concentrations were shown 7 days after LPS IV. Deposition of elastin fibers at sites of septation was disturbed within 3 days after LPS IV. CONCLUSION Lung maturation and disturbed lung structure occurred after short-term exposure to fetal inflammation and suggests new targeted therapies for BPD.

All-Trans Retinoic Acid and Intra-Amniotic Endotoxin-Mediated Effects on Fetal Sheep Lung

All‐trans retinoic acid (RA) is a potent modulator of lung development. Chorioamnionitis, which is frequently associated with preterm birth, causes fetal lung inflammation and improves lung function but also results in alveolar simplification and microvascular injury. Endotoxin‐mediated chorioamnionitis reduces RA concentration in the fetal lung to 16% of control values. We hypothesized that administration of RA to the fetus before induction of chorioamnionitis would preserve septation of the distal airspaces. Time‐mated ewes with singletons were assigned to receive a fetal intramuscular treatment with 20,000 IU of RA in olive oil (or olive oil only) 3 hr prior to intra‐amniotic injection of endotoxin (20 mg, E. coli 055:B5) or saline, at 124‐day gestational age and 7 days after the fetal treatment. The right cranial lung lobe was processed for morphometric analysis. RA treatment did not affect chorioamnionitis‐induced fetal and systemic inflammation or interleukin‐8 concentrations in lung tissue. RA administration alone did not alter lung structure. Relative to control lungs (5 ± 3 mL/kg), lung volume increased similarly with endotoxin (22 ± 4 mL/kg) or RA plus endotoxin (20 ± 3 mL/kg; P < 0.05). Alveolar wall thickness was 4.2 ± 0.3 μm after endotoxin‐induced chorioamnionitis, 6.0 ± 0.4 μm in controls (P < 0.05 versus endotoxin) and 5.5 ± 0.2 μm after RA and endotoxin (P < 0.05 versus control, n.s. versus endotoxin). The ratio of airspace versus tissue was 4.6 ± 0.3 in endotoxin‐induced chorioamnionitis, 2.1 ± 0.3 in controls and 4.1 ± 0.5 after RA and endotoxin. We conclude that fetal treatment with RA did not prevent inflammation‐induced alveolar simplification. Anat Rec, 2008.

Lung hypoplasia in a patient with del(2)(q33-q35) demonstrated by chromosome microdissection.

We report on a 17-month-old girl with multiple malformations, including lung hypoplasia, multiple ventricular septal defects, craniofacial anomalies, and malrotation of the intestine. Moreover, the patient showed Robin sequence, developmental delay, as well as pre- and postnatal growth retardation. Postnatal cytogenetic analysis revealed an interstitial deletion on the long arm of chromosome 2. Microdissection and reverse chromosome painting of the aberrant chromosome 2 as well as FISH with a panel of chromosome 2q band-specific YACs mapped the deletion to 2q33-q35. Lung hypoplasia has not been described so far in patients with del(2)(q33-q35). A review of previously reported patients showed variable phenotypes apparently due to different deleted chromosomal segments.