Pulmonary fibrosis is a major cause of death in patients with acute respiratory distress syndrome (ARDS). Our previous study revealed that lipopolysaccharide (LPS) challenge could lead to mouse lung fibroblast proliferation. Additionally, inhibition of autophagy in lung fibroblasts was also reported to be crucial during the process of pulmonary fibrosis. However, the correlation between proliferation and inhibition of autophagy of lung fibroblasts and the underlying mechanism remain unknown. In this study, we report that autophagy was inhibited in mouse lung fibroblasts after LPS challenge, and was accompanied by activation of the PI3K-Akt-mTOR signaling pathway. Treating mouse lung fibroblasts with LPS resulted in mTOR and Akt phosphorylation, p62 up-regulation, and significant down-regulation of autophagosomes, which could be reversed by PI3K-Akt inhibitors (Ly294002) or mTOR inhibitors (rapamycin, RAPA). Furthermore, either LPS or hydroxychloroquine (HCQ), an autophagy inhibitor, could promote mouse lung fibroblast proliferation, which could be reversed by RAPA application. The present research therefore reveals that LPS promotes lung fibroblast proliferation through autophagy inhibition via activation of the PI3K-Akt-mTOR pathway.