Effect of porcine respiratory coronavirus infection on lipopolysaccharide recognition proteins and haptoglobin levels in the lungs

Abstract

Porcine respiratory coronavirus (PRCV) potentiates respiratory disease and proinflammatory cytokine production in the lungs upon intratracheal inoculation with lipopolysaccharide (LPS) at 1 day of infection. This study aimed to quantify LPS-binding protein (LBP), CD14 and haptoglobin in the lungs throughout a PRCV infection. LBP and CD14 recognize LPS and enhance its endotoxic activity, whereas haptoglobin dampens it. Gnotobiotic pigs were inoculated intratracheally with PRCV (n = 34) or saline (n = 5) and euthanized 1-15days post inoculation (DPI). Virus was detected in the lungs from 1 to 9DPI. Cell-associated CD14 in lung tissue increased up to 15 times throughout the infection, due to an increase in highly CD14+ monocyte-macrophages from 1 to 12DPI and CD14+ type 2 pneumocytes from 7 to 9DPI. LBP and soluble CD14 levels in bronchoalveolar lavage fluids were elevated from 1-12DPI, with up to 35- and 4-fold increases, respectively. Haptoglobin levels increased significantly (x4.5) at 7DPI. In addition, we found that PRCV could sensitize the lungs to LPS throughout the infection, but the response to LPS appeared less enhanced at the end of infection (7DPI). The marked increases in LBP, CD14 and haptoglobin were not correlated with the extent of the LPS response.

Fig. 1

Kinetic profiles of virus titers (A), cell-associated CD14 in lung tissue (B), soluble CD14 in BAL fluids (C), LBP in BAL fluids (D) and haptoglobin in BAL fluids (E) throughout a PRCV infection. The left part of the graph shows data of PBS control pigs euthanized from 1 to 15 DPI. Each dot corresponds to one pig and the solid line represents the mean. The dotted line represents the detection limit. Means marked with an asterisk differ significantly (P < 0.05) from those of the PBS control pigs.

Fig. 2

Double immunofluorescence staining for CD14 and cytokeratin 18 in the lung tissue of PBS- and PRCV-inoculated pigs. Panel A (bar = 50 μm) demonstrates that PBS-inoculated pigs contain few highly CD14-positive cells in the lung tissue. At 1 DPI, there is a massive accumulation of highly CD14-positive monocyte-like cells near the bronchioli. At 7 DPI, the alveolar tissue contains numerous clusters of highly CD14-positive monocyte-macrophages. At this time point of infection, part of the type 2 pneumocytes also stain positive for CD14, as indicated by the yellow color in the merge picture. Panel B (bar = 10 μm) illustrates in more detail the CD14 signal on a cytokeratin 18-positive pneumocyte.

Fig. 3

Kinetic profiles of soluble CD14 (A), LBP (B) and haptoglobin (C) in sera throughout a PRCV infection.