Technical Abstract: Chronic inflammation is associated with obesity and insulin resistance. However, the underlying mechanisms are not fully understood. Pattern recognition receptors Toll-like receptors and Nucleotide-oligomerization domain containing proteins play critical roles in innate immune response. Here we report that activation of nucleotide binding oligomerization domain containing protein 1 (NOD1) in adipocytes impairs insulin signaling and glucose uptake. The mRNA level of NOD1 is markedly increased in differentiated murine 3T3-L1 adipocytes and human primary adipocyte culture upon adipocyte conversion. Stimulation of NOD1 with a synthetic ligand Tri-DAP induces proinflammatory chemokine MCP-1, RANTES and cytokine TNF-a and MIP-2 (human IL-8 homolog) and IL-6 mRNA expression in 3T3-L1 adipocytes in a time and dose dependent manner. Similar proinflammatory profiles are observed in human primary adipocyte culture stimulated with Tri-DAP. Moreover, NOD1 activation suppresses insulin signaling as revealed by attenuated phosphorylation of Akt and downstream target GSK3a/3ß and subsequent insulin-induced glucose uptake in 3T3-L1 adipocytes. Together, our results suggest that NOD1 may play an important role in adipose inflammation and insulin resistance.