PURPOSE: Excess of iron (Fe) in lung allografts is involved in the pathogenesis of the bronchiolitis obliterans syndrome (BOS). Fe is deposited inside lysosomes of lung macrophages (LMs). When challenged by oxidants, such Fe triggers lysosomal damage and cell death and promotes inflammation. BOS is prevented by azithromycin (AZM), which accumulates inside lysosomes and interferes with the lysosomal pool of Fe. Therefore, we assessed lysosomal leakage in LMs harvested from lung transplant patients and healthy subjects and tested the ability of AZM to protect lysosomes and macrophages against oxidative stress.
METHODS: LMs were harvested from 5 lung transplant patients without AZM-treatment, 1 lung transplant patient with on-going AZM-treatment and 6 healthy subjects without medications. Cultures of LMs and murine J774 macrophages (the latter pre-treated with AZM or not) were oxidatively stressed by a stable concentration of H2O2 (120 mikroM). Lysosomal leakage, assessed as the increase of cytosolic green acridine orange (AO) fluorescence (AO is released from leaking lysosomes into the cytosol during oxidant challenge), was evaluated at the end of the 1 h oxidant challenge. Cell death was evaluated at 10 h after ended oxidative stress.
RESULTS: LMs from lung transplant patients not treated with AZM exhibited significantly higher AO-values before and after oxidative stress than LMs from healthy subjects. In contrast, the AO-value of LMs from the AZM-treated lung transplant patient was strikingly lower before and after oxidant challenge. The frequency of dead cells was proportional to the degree of lysosomal leakage. AZM efficiently prevented lysosomal leakage in cultures of oxidatively stressed J774 macrophages.
CONCLUSIONS: This study presents evidence that lysosomes of LMs from lung transplants are much more sensitive to oxidant challenge than those from healthy controls.
CLINICAL IMPLICATIONS: This study provides a rationale for the use of AZM to prevent BOS in lung transplants; it is possible that the protective effect of AZM on the small airways is mediated by a protection of lysosomes and cells against Fe-catalyzed oxidative damage.
DISCLOSURE: The following authors have nothing to disclose: Hans Persson, Linda Vainikka
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