Lactobacillus plantarum is a well-documented probiotic that has been used in clinical trials for the regulation of the immune system and treatment of gastrointestinal diseases. In this study, we evaluated the effects of L. plantarum cell lysates on the immune regulation through the in vitro and in vivo studies. L. plantarum lysates were prepared by sonication method, and we observed that the repetition of disruption step increased indicator components within the bacterial lysates. Indicator components might affect TNF-α production. L. plantarum lysates did not induce TNF-α production, while LPS-induced TNF-α production was dramatically inhibited in a sonication-dependent manner in THP-1 cells. Oral administration of L. plantarum lysates effectively attenuated the horny layer formation and decreased epidermal thickening in NC/Nga mice skin. The damage to barrier function after the 8 weeks oral administration was reduced by L. plantarum lysates as compared to that in the atopic dermatitis (AD) mice. Further study revealed that L. plantarum lysates polarized Th1 response via induction of IL-12 and IFN-γ production and inhibition of IL-4 and IgE production in NC/Nga mice. Together, our results suggest that L. plantarum lysates are remarkable material for host homeostasis and it could be used for the treatment of inflammatory diseases.