This study sought to investigate whether Ganoderma lucidum polysaccharide (GLP) has a protective effect on lipopolysaccharide (LPS)-induced inflammatory injury to mammary epithelial HC-11 cells and to characterize the mechanism involved. Cell viability was assessed using the cell counting kit 8 (CCK-8) method, tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and IL-1β levels were measured by enzyme linked immunosorbent assay (ELISA), and IκBa, p65 NF-κB and STAT3 mRNA were determined using quantitative reverse transcription PCR (qRT-PCR), p65 and STAT3 protein expression were determined using Western blotting, respectively. GLP was shown to inhibit LPS-induced TNF-α, IL-6, and IL-1β production (P < 0.01 or P < 0.05), GLP was also shown to increase IκBα mRNA expression (P < 0.01), decrease p65 and STAT3 mRNA expression (P < 0.01 or P < 0.05), and decrease p-p65, p65, p-STAT3, and STAT3 protein expression in breast epithelial cells (P < 0.01 or P < 0.05). The findings suggest that GLP inhibits nuclear factor kappa-B (NF-κB) and signal transducers and activators of transcription (STAT) signaling by preventing IκBα degradation and p65 and STAT3 phosphorylation. This results in lower LPS-induced TNF-α, IL-6, and IL-1β production and prevents inflammatory cell injury.