Figure 4
Figure 4. MHC class I Abs cause ALI in a 2-event in vivo model. (A) Rats were injected with either NS or LPS intraperitoneally 2 hours prior to transfusion and then infused with mouse IgG (0.3 mg/kg, control) or MHC class I Abs (0.075-4.5 mg/kg OX18 or OX27), followed by Evans blue dye (EBD), which binds to albumin. Rats were humanely killed 6 hours after transfusion, a BAL was performed, and ALI was measured as the percentage of EBD leak into the BAL from the plasma (% BAL EBD/% plasma EBD, y-axis), which is depicted as a function of the treatment group (x-axis). Rats that were treated with NS (■) did not evidence significant EBD leak for any treatment group including rats transfused with 4.5 mg/kg OX27. In addition, rats injected with LPS () and infused with mouse IgG did not demonstrate EBD leak, compared with the LPS/NS controls, and neither did LPS-treated rats infused with 0.15 mg/kg OX18 or 0.075 mg/kg OX27. However, rats injected with LPS then infused with 0.30 to 0.45 mg/kg OX18 and 0.15 to 0.30 mg/kg OX27 had a significant amount of lung injury compared with LPS/NS and LPS/mouse IgG (*P < .05 from LPS/NS; †P < .05 from LPS/mouse IgG, n = 4). (B) Rats were injected with saline (NS) or LPS 2 hours prior to the infusion of 0.15 mg/kg OX27. After completion of the experimental protocol, the rats were humanely killed, and the lungs were snap-frozen and fixed, embedded in paraffin, and sectioned. All images were shot at 40×, and the sections were stained with H&E (i,iii) or immunohistochemistry was performed (ii,iv) to localize rat PMNs (red) and to demonstrate the pulmonary parenchyma by staining the membranes green (WGA linked to Cy-3) and the nuclei blue (bis-benzamide). Rat lungs from animal injected with NS and then infused with OX27 demonstrated normal pulmonary histology without evidence of ALI (i) with a few PMNs visible in the lung (arrow, ii). In contrast, in rats injected with LPS and then infused with OX27 there are obvious signs of ALI with pulmonary edema, hyaline membrane formation, and arcuate inflammation with PMNs (iii). The immunohistochemistry demonstrated widespread PMN infiltration throughout the lung parenchyma as shown by the red color further demarcated by the white arrows (iv; n = 2).

MHC class I Abs cause ALI in a 2-event in vivo model. (A) Rats were injected with either NS or LPS intraperitoneally 2 hours prior to transfusion and then infused with mouse IgG (0.3 mg/kg, control) or MHC class I Abs (0.075-4.5 mg/kg OX18 or OX27), followed by Evans blue dye (EBD), which binds to albumin. Rats were humanely killed 6 hours after transfusion, a BAL was performed, and ALI was measured as the percentage of EBD leak into the BAL from the plasma (% BAL EBD/% plasma EBD, y-axis), which is depicted as a function of the treatment group (x-axis). Rats that were treated with NS (■) did not evidence significant EBD leak for any treatment group including rats transfused with 4.5 mg/kg OX27. In addition, rats injected with LPS () and infused with mouse IgG did not demonstrate EBD leak, compared with the LPS/NS controls, and neither did LPS-treated rats infused with 0.15 mg/kg OX18 or 0.075 mg/kg OX27. However, rats injected with LPS then infused with 0.30 to 0.45 mg/kg OX18 and 0.15 to 0.30 mg/kg OX27 had a significant amount of lung injury compared with LPS/NS and LPS/mouse IgG (*P < .05 from LPS/NS; †P < .05 from LPS/mouse IgG, n = 4). (B) Rats were injected with saline (NS) or LPS 2 hours prior to the infusion of 0.15 mg/kg OX27. After completion of the experimental protocol, the rats were humanely killed, and the lungs were snap-frozen and fixed, embedded in paraffin, and sectioned. All images were shot at 40×, and the sections were stained with H&E (i,iii) or immunohistochemistry was performed (ii,iv) to localize rat PMNs (red) and to demonstrate the pulmonary parenchyma by staining the membranes green (WGA linked to Cy-3) and the nuclei blue (bis-benzamide). Rat lungs from animal injected with NS and then infused with OX27 demonstrated normal pulmonary histology without evidence of ALI (i) with a few PMNs visible in the lung (arrow, ii). In contrast, in rats injected with LPS and then infused with OX27 there are obvious signs of ALI with pulmonary edema, hyaline membrane formation, and arcuate inflammation with PMNs (iii). The immunohistochemistry demonstrated widespread PMN infiltration throughout the lung parenchyma as shown by the red color further demarcated by the white arrows (iv; n = 2).

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