Background: Toll-like receptor 4 participates in the process of acute heart injury. The underlying mechanisms of its protection are multifactorial, but we hypothesized that toll-like receptor-mediated autophagy control plays a vital role. The purpose of this study was to clarify the effect of autophagy on cardiac fibrosis. Methods: Cardiac fibrosis was induced by subcutaneous isoproterenol (ISO) injection, and rapamycin was simultaneously administered orally for 14 days. Animal echocardiography was then used to evaluate the success of the cardiac fibrosis model, and the mice were killed after the echocardiography examination. Results: Toll-like receptor 4 knockout (TLR4 KO) mice had better heart function than did wild-type (WT) mice (P < .05). Rapamycin treatment reduced the left ventricular ejection fraction to 23.5% (P < .05), and the collagen volume fraction of the ISO and ISO plus rapamycin groups was 5.9% and 25.9%, respectively, in TLR4 KO mice. Compared with the WT mice, Beclin 1 and autophagy were downregulated in TLR4 KO mice (P < .05); however, the ISO plus rapamycin group had higher autophagy activity than did the ISO group in TLR4 KO mice (P < .05). Conclusions: Our results suggest that TLR4 KO-induced cardioprotection against ISO-induced cardiac fibrosis is associated with reduced autophagy induction. Cardiac fibroblast autophagy participates in its own activation. The moderate inhibition of autophagic activity may be a new strategy for treating cardiac fibrosis.