Background Prevalence of dry eye is significantly inceasing in postmenopausal women than that in men, suggesting that sex hormone plays a role in the pathogenesis of dry eye. In addition, dry eye might become worse following estrogen therapy in postmenopausal women. However, whether application of androgen can ameliorate dry eye is being concerned. Objective This study was to investigate the effect of androgen on tear film of ovariectomized female rats. Methods Forty-eight 3-month-old sexually mature female Wistar rats were randomized into the normal control group, sham group, ovariectomy (OVX) model group and testosterone-injected group. OVX models were established by bilateral ovaries enucleation in the rats of the model group and testosterone-injected group, and then androgen (3.75 mg/kg) was intramuscularly injected since 5 months after OVX at 3-day interval for 6 weeks. Only intraperitoneal fat was cut off in the sham group. In 6 weeks after injection of androgen, serum androgen concentration detected and Schirmer Ⅰtest (SⅠt), tear film break-up time (BUT) were performed. The rats were sacrificed to prepare the corneal and conjunctival samples. The expression of MUC5AC in conjunctival tissue was examined by immunofluorescence staining, and the microstructure of corneal cellular surface was observed under the scanning electron microscope before and 6 weeks after application of androgen. Animals in this study were treated in accordance with Animal Experimentation Ethic Committee Guidelines of Southern Medical University and the study protocol was approved by Ethic Committee of this University. Results The mean serum testosterone concentration was (1.83±0.12)ng/ml, and SⅠt or BUT was (3.63±0.26)mm/5 minutes or (3.73±0.38)seconds, respectively, in the OVX model group, which was significantly declined in comparison with (2.56±0.14)ng/ml, (7.47±0.66)mm/5 minutes or (9.57±0.76)seconds in the normal control group (all at P=0.000). However, the serum testosterone concentration was (3.38±0.24)ng/ml, SⅠt was (6.37±0.45)mm/5 minutes and BUT was (7.54±0.55)seconds in the testosterone-injected group, which was significantly higher than that in the OVX model group (all at P=0.000). The positive staining of MUC5AC in rat conjunctival tissue weakened in the OVX model group compared with the normal control group, and the fluorescence intensity in the testosterone-injected group was stronger than that in the OVX model group. Regularly arranged microvilli and cell metabolism hiatus on the surface of corneal cells were seen in the normal control group and the sham group; while the microvilli were shorter and irregularly arranged, and the cell metabolism hiatus were disappeared in the OVX model group. However, microvilli and cell metabolism hiatus were close to normal ones in the testosterone-injected group. Conclusions Deterioration of tear secretion and instability of tear film are are probably associated with the lower serum androgen levels in castrated female rats. Systemic supplement of androgen can promote tear secretion, improve tear film stability and alleviate ocular surface damage. Key words: Dry eye/therapy; Tears/secretion; Ovariectomy; Androgens/drug therapy; Disease models, animal; Rats, Wistar; Female