Objective 　To explore the expression of hypoxia-inducing factor-1α (HIF-1α) and its significance in the rats with isoproterenol-induced atrial fibrosis. Methods 　Thirty male Wistar rats were randomly divided into control group, isoproterenol [Iso, 5mg/(kg.d) for 7 days] group, and Iso + sirolimus (Rapa) group. The atrial fibrosis rat model was reproduced by subcutaneous injection of high doses of Iso in Iso group and Rapa group. Rats were treated with isoproterenol plus sirolimus in Iso-si group. All the animals were sacrificed 15 days after treatment. The contents of angiotensinⅡ(AngⅡ) in rat myocardial tissue were determined by radioimmunoassay. HE staining and Masson staining were performed for the measurement of the degree of atrial fibrosis in terms of collagen volume fraction (CVF). Immunohistochemistry and Western blotting were performed to detect the expressions of HIF-1α, TGF-β1 and MMP-9. The correlations of HIF-1α, TGF-β1 and MMP-9 to CVF and expressions of HIF-1α, TGF-β1 and MMP-9 were analyzed. Results 　The AngⅡ content was significantly higher in Iso group and Rapa group than that in control group (P<0.01). No atrial fibrosis occurred in control group (CVF was 15.482%±0.837%), and the atrial fibrosis was significantly milder in Rapa group (CVF was 16.730%±1.052%) than in Iso group (CVF was 86.704%±1.928%, P<0.01). Immunohistochemistry and Western blotting revealed that expressions of HIF-1α, TGF-β1 and MMP-9 were significantly increased in Iso group compared with that of control group (P<0.01), but the expressions were significantly decreased in Rapa group compared with that in Iso group (P<0.01). The expressions of HIF-1α, TGF-β1 and MMP-9 were positively correlated with the fibrosis degree (P<0.01); the HIF-1α content was positively correlated with the contents of TGF-β1 and MMP-9 (P<0.01); the MMP-9 content was positively correlated with the TGF-β1 content (P<0.01). Conclusion 　AngⅡ, HIF-1α, TGF-β1 and MMP-9 play an important role during the development of isoproterenol-induced atrial fibrosis in rats.