Objective To investigate the effect of adipose tissue-derived mesenchymal stem cell (ASC) administered either systemically or locally in a murine model of bronchiolitis obliterans. Results When compared to controls, systemic treatment with 10(6) ASCs on D0 and a second dose on D7 significantly prevented tracheal obliteration 28 days after heterotopic tracheal transplantation (median of 94 vs. 16%; P<0.01). A single dose tended towards less stenosis than controls, but did not reach statistical significance (28 vs. 94%; P = 0.054). On the contrary, repeated local injection was incapable of preventing tracheal obliteration when compared to a single injection or controls (37 vs. 71 vs. 87%). Two intravenous doses also tended to be better than two local injections (16 vs. 37%; P = 0.058), and were better than a single local dose (16 vs. 71%; P < 0.01). Conclusion A second dose of ASC, given systemically after 7 days, reduces luminal obliteration in a heterotopic tracheal transplantation model in mice, suggesting that ASC can be used to prevent obliterative bronchiolitis after lung transplantation.