Intratracheal recombinant surfactant protein d prevents endotoxin shock in the newborn preterm lamb

Abstract

Rationale: The susceptibility of neonates to pulmonary and systemic infection has been associated with the immaturity of both lung structure and the immune system. Surfactant protein (SP) D is a member of the collectin family of innate immune molecules that plays an important role in innate host defense of the lung.

Objectives: We tested whether treatment with recombinant human SP-D influenced the response of the lung and systemic circulation to intratracheally administered Escherichia coli lipopolysaccharides.

Methods: After intratracheal lipopolysaccharide instillation, preterm newborn lambs were treated with surfactant and ventilated for 5 h.

Measurement: Survival rate, physiologic lung function, lung and systemic inflammation, and endotoxin level in plasma were evaluated.

Main results: In control lambs, intratracheal lipopolysaccharides caused septic shock and death associated with increased endotoxin in plasma. In contrast, all lambs treated with recombinant human SP-D were physiologically stable and survived. Leakage of lipopolysaccharides from the lungs to the systemic circulation was prevented by intratracheal recombinant human SP-D. Recombinant human SP-D prevented systemic inflammation and decreased the expression of IL-1beta, IL-8, and IL-6 in the spleen and liver. Likewise, recombinant human SP-D decreased IL-1beta and IL-6 in the lung and IL-8 in the plasma. Recombinant human SP-D did not alter pulmonary mechanics following endotoxin exposure. Recombinant human SP-D was readily detected in the lung 5 h after intratracheal instillation.

Conclusions: Intratracheal recombinant human SP-D prevented shock caused by endotoxin released from the lung during ventilation in the premature newborn.

Figures

Figure 1.

Kaplan-Meier plot of recombinant human surfactant protein D (rhSP-D) treated group and control group. In the control group, only 20% of the lambs survived until the end of the 5-h study period. In contrast, all lambs treated with rhSP-D survived. p

Figure 2.

(A) rhSP-D prevents systemic spread of lipopolysaccharides (LPS) from the lung. Intratracheal endotoxin was detected in circulation and was increased over time in the control group. rhSP-D decreased plasma endotoxin concentration during the 5-h study. (B) Treatment with rhSP-D prevented the endotoxin shock. Systolic blood pressure was maintained at normal level of premature newborn in rhSP-D–treated groups. In contrast, blood pressure gradually decreased in the control group after 3 h of age. *p < 0.05 versus control.

Figure 3.

(A) Blood pH was maintained with rhSP-D treatment. Although LPS treatment associated with decreased blood pH, treatment with rhSP-D maintained pH and prevented prenatal endotoxin-induced shock. (B) Blood base excess (BE) was altered by intratracheal LPS. Intratracheal LPS induced metabolic acidosis and rhSP-D treatment prevented the low BE and endotoxin shock.

Figure 4.

Sequential measurement of Pco2 and ventilatory pressure. (A) Endotracheal LPS caused an increase in Pco2 after 3 h of age. Pco2 was maintained in group treated with rhSP-D. (B) Ventilatory pressure (PIP-PEEP) used to maintain target tidal volume was similar for both groups. *p < 0.05 versus control.

Figure 5.

Proinflammatory cytokine expression. (A, B) IL-1β, IL-6, and IL-8 mRNAs in spleen and liver were increased in control lambs after intratracheal LPS instillation. Proinflammatory cytokine mRNAs in spleen and liver were decreased by rhSP-D. (C) Endotracheal LPS increased IL-1β, Il-6, and IL-8 mRNAs in the lung. Expression of IL-1β was decreased in the group treated with rhSP-D. (D) IL-8 concentrations in plasma were increased in the control group. Plasma IL-8 concentrations were kept low by rhSP-D treatment. *p < 0.05 versus control.

Figure 6.

Lung morphology with hematoxylin and eosin staining (A, B) and immunohistochemistry of IL-8 (C, D) and IL-1β (E, F). In both control and rhSP-D groups there are increased granulocyte and positively stained inflammatory cells for IL-8 and IL-1β. The inflammatory cells immunostained for IL-8 and IL-1β were decreased by intratracheal rhSP-D treatment.

Figure 7.

Lung function was not affected by rhSP-D treatment. (A) Dynamic lung compliance, calculated from Vt, PIP-PEEP, and body weight during ventilation, and (B) deflation limb of static lung pressure–volume curve were similar.

Figure 8.

High levels of rhSP-D were detected in bronchoalveolar lavage fluid (BALF) by Western blot 5 h after endotracheal rhSP-D instillation (animals #6 and 7). BALF from control lambs (animals #1 and 2) did not show any rhSP-D.