Recent data have shown that nucleotide-binding domain leucine-rich repeat proteins (NLRs), a class of innate immune receptors that respond to pathogen attack or cellular stress, have gained increasing attention. NLRC5 (NLR family, CARD domain containing 5) is the largest member of the NLR family, which has recently been identified as a critical regulator of immune responses. Until recently, the function of NLRC5 has been a matter of debate. In this study, we explore the role of NLRC5 in cytokine secretion and the role of the nuclear factor-κB (NF-κB) signaling pathway in tumor necrosis factor-alpha (TNF-α)-induced NLRC5 expression in LX-2 cells. We demonstrated that overexpression of NLRC5 results in an upregulation of IL-6 and IL-1β secretion. On the other hand, knockdown of NLRC5 by transfecting siRNA decreased IL-6 and IL-1β secretion in LX-2 cells. Meanwhile, the results showed that pyrrolidine dithiocarbamate (PDTC) (a specific inhibitor of the NF-κB signaling pathway) inhibited NLRC5 expression and NLRC5 silencing could increase the expression levels of p65 in cell nucleus accompanied with upregulated phosphorylation of Smad3 protein levels in response to TNF-α. These results indicated that NLRC5 plays a significant role in TNF-α-enhanced cytokine (IL-6 and IL-1β) secretion of LX-2 cells and the NF-κB/Smad3 signal pathway is involved in its induction of expression.