Obesity has been reported to be associated with low grade inflammatory status. In this study, we investigated the inflammatory response as well as associated signaling molecules in immune cells from diet-induced obese mice. Four-week-old C57BL mice were fed diets containing 5% fat (control) or 20% fat and 1% cholesterol (HFD) for 24 weeks. Splenocytes (1 × 10⁷ cells) were stimulated with 10 µg/mL of lipopolysaccharide (LPS) for 6 or 24 hrs. Production of interleukin (IL)-1β, IL-6, and TNF-α as well as protein expression levels of nucleotide-binding oligomerization domain (NOD)2, signal transducer and activator of transcription (STAT)3, and pSTAT3 were determined. Mice fed HFD gained significantly more body weight compared to mice fed control diet (28.2 ± 0.6 g in HFD and 15.4 ± 0.8 g in control). After stimulation with LPS for 6 hrs, production of IL-1β was significantly higher ( P = 0.001) and production of tumor necrosis factor (TNF)-α tended to be higher ( P < 0.064) in the HFD group. After 24 hrs of LPS stimulation, splenocytes from the HFD group produced significantly higher levels of IL-6 (10.02 ± 0.66 ng/mL in HFD and 7.33 ± 0.56 ng/mL in control, P = 0.005) and IL-1β (121.34 ± 12.72 pg/mL in HFD and 49.74 ± 6.58 pg/mL in control, P < 0.001). There were no significant differences in the expression levels of STAT3 and pSTAT3 between the HFD and the control groups. However, the expression level of NOD2 protein as determined by Western blot analysis was 60% higher in the HFD group compared with the control group. NOD2 contributes to the induction of inflammation by activation of nuclear factor κB. These findings suggest that diet-induced obesity is associated with increased inflammatory response of immune cells, and higher expression of NOD2 may contribute to these changes.