This study investigated the effects of hypoxic and hyperosmotic stresses on the matrix metalloproteinases 2 and 9 (MMP-2, MMP-9) and their tissue inhibitors 2 and 1 (TIMP-2, TIMP-1) respectively in cultured lung fibroblasts from broiler chickens. Isolated and sub-cultured primary lung fibroblasts from 1-day-old broilers were exposed to hypoxia and hyperosmosis. The presence of the enzymes and the effects of hypoxia and hyperosmosis on the enzymes in cultured lung fibroblasts, and in the culture media, were evaluated with the streptavidin-biotin-peroxidase method and western blotting using commercially available primary monoclonal antibodies. Short (6 hours) and long term (12 hours) hypoxia, hypoxia + hyperosmosis (6 hours) and hyperosmosis (6 hours) triggered the release of both MMP-2 and TIMP-1 in experimental groups when compared with their controls. Neither MMP-9 nor TIMP-2 antibodies stained the fibroblast cells kept in normoxic, hypoxic or hyperosmotic conditions.