Proceedings of The Physiological Society

Physiology 2016 (Dublin, Ireland) (2016) Proc Physiol Soc 37, PCB046

Poster Communications

Alveolar macrophages express the bone morphogenetic protein antagonist gremlin-1

L. Mthunzi1, M. Kubica1, U. Knaus1, P. McLoughlin1

1. School of Medicine, University College Dublin, Dublin, Ireland.

Background Pulmonary hypoxia secondary to lung disease results in inflammation and the subsequent development of hypoxia-induced pulmonary hypertension (HPH). We have previously shown that the bone morphogenetic protein antagonist, gremlin-1, is lung-selectively upregulated at levels of hypoxia equivalent to those found in hypoxic lung diseases1. Additionally, we have shown that gremlin-1 plays a key role in the development of HPH by increasing pulmonary vascular resistance2. Macrophages play a critical role in the development of HPH although their exact mechanism of action remains to be elucidated3. Preliminary data suggests that the macrophage may be an important source of gremlin-1 during the development of HPH. We hypothesize that macrophages are sources of gremlin-1 in the hypoxic lung and that macrophage-derived gremlin-1 plays a role in the development of HPH. In this study we sought to: confirm gremlin-1 upregulation in the hypoxic lung, investigate whether gremlin-1 is produced by macrophages and assess whether hypoxia regulates expression of gremlin-1 by macrophages. Methods Adult male C57Bl6/J mice were used for all experiments. All experiments were carried out under licence from the animal research ethics committee of University College Dublin. Mice were maintained in normoxia (21% O2) or hypoxia (10% O2) for indicated time points then sedated by inhalation of isofluorane and subsequently killed by intraperitoneal injection of sodium pentobarbitone (200 mg/kg). Bronchoalveolar lavage was performed on mice to isolate alveolar macrophages. Lungs from mice were removed and rapidly flash frozen in liquid nitrogen. Bone marrow-derived macrophages (BMDMs) were generated using standard protocols with L-929 cell conditioned medium. Results Expression of gremlin-1 mRNA was significantly increased in lungs isolated from hypoxic mice (n=6) compared to normoxic controls (n=6). Immunostaining demonstrated that alveolar macrophages isolated by bronchoalveolar lavage are sources of gremlin-1. In addition, alveolar macrophages isolated from hypoxic lungs (n=6) display increased gremlin-1 immunostaining compared to normoxic controls (n=6). Furthermore, gremlin-1 mRNA expression is increased in alveolar macrophages isolated from hypoxic lungs (n=6) compared to normoxic controls (n=6). BMDMs display gremlin-1 immunostaining and mRNA expression but unlike alveolar macrophages, BMDMs do not increase gremlin-1 levels in response to hypoxia. Conclusions Our findings establish alveolar macrophages as sources of gremlin-1 in the lung. Furthermore, alveolar macrophages increase gremlin-1 expression in the lung in response to hypoxia. Absence of a hypoxia-induced increase in gremlin-1 levels in BMDMs suggests that this response may be tissue specific. Future work will investigate the role of macrophage-derived gremlin-1 in the development of HPH.

Where applicable, experiments conform with Society ethical requirements