We studied whether the therapeutic efficacy of the antifibrotic agent cis-4-hydroxy-L-proline (cHyp) in preventing bleomycin-induced pulmonary fibrosis in rats is enhanced by intratracheal delivery in liposomes. Dual-radiolabeled liposomes were used to study the distribution and stability of liposomes after intratracheal instillation. Lung retention was > 20% 1 wk after intratracheal instillation of 9 mumol phospholipid, and liposomes were intact as indicated by the ratio of the lipid and aqueous-phase markers remaining unchanged. For the fibrosis study, groups of rats were instilled with 1.2 U bleomycin (Bleo) and treated 1 and 2 wk later by single intratracheal instillation of test compounds. The control group received 0.3 ml saline (Bleo/sal). The treated groups received 9 mumol phospholipid in 0.3 ml of the following liposome preparations: empty liposomes (Bleo/lip), liposomes and 100 mg/kg of free unencapsulated cHyp (Bleo/lip/cHyp), and 100 mg/kg of liposome-encapsulated cHyp (Bleo/lip-cHyp). At 3 wk, fibrosis (mg hydroxyproline/g weight lung) by groups was as follows: control, 2.6 +/- 0.1 (SEM); Bleo/sal, 3.2 +/- 0.1, Bleo/lip, 3.2 +/- 0.1, and Bleo/lip/cHyp, 3.1 +/- 0.1, p < 0.05 compared with control; Bleo/lip-cHyp, 2.6 +/- 0.1, p < 0.05 compared with Bleo/sal, n = 3 to 6. Histologic grading of fibrosis did not show decreased fibrosis in the Bleo/lip-cHyp group, probably because of the focal nature of the fibrotic lesions. We conclude that cHyp encapsulated in liposomes prevents bleomycin-induced fibrosis by biochemical measurements. Delivery of antifibrotic agents to the lung in carrier vehicles promotes retention and may enhance their efficacy in treating bleomycin-induced pulmonary fibrosis.