Abstract 1733 Neonatal Pulmonology II: Oxidant and Inflammatory Lung Injury Poster Symposium, Tuesday, 5/4

Background: Proinflammatory cytokines such as IL-8 have been implicated in the pathogenesis of bronchopulmonary dysplasia. In a rat model of neonatal chronic lung injury induced by hyperoxia, neutrophil chemokines CINC-1 and MIP-2 (rat IL-8 analogs) showed increased expression with age and hyperoxia. The receptor for these chemokines, CXCR2, is homologous to the predominant human and rabbit IL-8 receptor, IL-8rb, also known as CXCR2. Acute inflammation induced by LPS has been shown to downregulate IL-8rb. In other studies, IL-8rb is transcriptionally regulated. Objective: We sought to test whether CXCR2 mRNA expression in rat lung was inhibited in this model of neonatal hyperoxia induced lung injury, and whether anti-cytokine antibody treatment altered CXCR2 mRNA abundance, indicating effects on transcriptional regulation. Method: Newborn rats were exposed to air or 95%O2 for up to 8 days (d). Some oxygen exposed pups were treated on d 3 and 4 with 1(O26+1), or 5 (O26+ 5)µg/kg of anti-CINC-1 antibody or diluent control to reduce inflammation, and killed at 6 d. After exposure, animals were killed and lungs were lavaged and perfused with buffered saline. RNA was extracted and CXCR2 mRNA was measured by semiquantitative RT-PCR at 25 cycles, using 32P-dCTP incorporation. cDNA transcripts were electrophoresed, and dried gels were detected using a phosphorimager. Images were analyzed with ImageQuant v 1.2. Myeloperoxidase was measured in lung homogenates spectrophotometrically to assess the amount of accumulated neutrophils in lavaged, perfused lung. CXCR2 mRNA pixel values were normalized to 6 d air control. Myeloperoxidase values are expressed as mean ± S.E.M, and were normalized to 6 day air control. 3 or 4 animals were analyzed for each condition. (Table) Results: Although CXCR2 mRNA accumulation increased with hyperoxia at 6d and 8d, neutrophil accumulation was greater, therefore CXC/MPO (mRNA/neutrophil) was only 0.3 and 0.5 × control at 6d and 8d 95%O2. Anti-CINC-1reduced CXCR2 mRNA and MPO in parallel, as reflected in the CXC/MPO ratios of 0.7 and 0.8. Conclusion: CXCR2 mRNA declines in neutrophils which accumulate in hyperoxia treated newborn lung, possibly due to hyperoxia effects on transcription or message stability. CXCR2 mRNA declines in lung from anti-CINC-1 treated hyperoxia exposed rat pups, in parallel with the decline in neutrophils as measured by myeloperoxidase. Downregulation of the CXCR2 receptor may serve to modulate inflammation as it progresses.

Table 1 No caption available
Full size table