Objective To observe splenic T cell apoptosis and XIAP-associated factor 1 (XAF1), FAS, and tumor necrosis factor (TNF)-α protein expression levels in rats with acute lung injury (ALI), and to determine their roles in the protective effect of Yifei Jianpi formula. Methods Sixty male specific pathogen-free SD rats were divided randomly into blank, model, positive control, and high-, medium-, and low-dose Yifei Jianpi formula groups. Rats in the positive control group were given 0.5 g/kg dexamethasone by gavage, and rats in the high-, medium-, and low-dose Yifei Jianpi formula groups were given 12, 6, and 3 g/kg Yifei Jianpi formula by gavage, respectively. Rats in the model and blank groups were given equal amounts of saline by gavage. All medications were administered once a day for 14 days. Lung function testing was carried out in all rats. We observed the imaging characteristics of the lungs and changes in the organ index and lung tissue wet/dry weight (W/D) in each group, and detected the pathological changes in lung tissues by hematoxylin-eosin staining. Splenic T-cell subpopulations (CD4⁺/CD8⁺) and apoptosis of splenic T-cells were detected by flow cytometry and XAF1, FAS, and TNF-α protein expression levels in the spleen were detected by Western Blot. Results Rats in the model group showed reduced lung function, decreased spleen and thymus organ indexes, and significantly higher W/D of lung tissue (P<0.01). In addition, they had inflammatory exudation and alveolar rupture in the lung tissue, accompanied by thickening of the lung texture and large areas of ground-glass shadows, with a significant decrease in T-cell subsets (CD4⁺/CD8⁺) and significant increases in XAF1, FAS, and TNF-α proteins, and in the rate of T-cell apoptosis (P<0.01). Yifei Jianpi formula significantly reduced the W/D spleen of rat lung tissues, significantly increased the thymus organ index (P<0.05, P<0.01) and the T-cell subpopulation (CD4⁺/CD8⁺), and significantly decreased the protein expression levels of XAF1, FAS, and TNF-α, and the T-cell apoptosis rate (P<0.05,P<0.01). Conclusions ALI induced up-regulation of XAF1, FAS, and TNF-α protein expression and T-cell apoptosis in the spleen of rats, and Yifei Jianpi formula may protect against ALI by down-regulating these factors.