The importance of innate immunity lies not only in directly confronting pathogenic and non-pathogenic insults but also in instructing the development of an efficient adaptive immune response. The Nlrp3 inflammasome provides a platform for the activation of caspase-1 with the subsequent processing and secretion of IL-1 family members. Given the importance of IL-1 in a variety of inflammatory diseases, understanding the role of Nlrp3 inflammasome in the initiation of innate and adaptive immune responses cannot be overstated. This review examines recent advances in inflammasome biology with an emphasis on its roles in sterile inflammation and triggering of adaptive immune responses.

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BackgroundMacrophages play essential roles in both innate and adaptive immune responses. Bacteria require endotoxin, a complex lipopolysaccharide, for outer membrane permeability and the host interprets endotoxin as a signal to initiate an innate immune response. The focus of this study is kinetic and global transcriptional analysis of the chicken macrophage response to in vitro stimulation with endotoxin from Salmonella typhimurium-798.ResultsThe 38535-probeset Affymetrix GeneChip Chicken Genome array was used to profile transcriptional response to endotoxin 1, 2, 4, and 8 hours post stimulation (hps). Using a maximum FDR (False Discovery Rate) of 0.05 to declare genes as differentially expressed (DE), we found 13, 33, 1761 and 61 DE genes between endotoxin-stimulated versus non-stimulated cells at 1, 2, 4 and 8 hps, respectively. QPCR demonstrated that endotoxin exposure significantly affected the mRNA expression of IL1B, IL6, IL8, and TLR15, but not IL10 and IFNG in HD 11 cells. Ingenuity Pathway Analysis showed that 10% of the total DE genes were involved in inflammatory response. Three, 9.7, 96.8, and 11.8% of the total DE inflammatory response genes were significantly differentially expressed with endotoxin stimulation at 1, 2, 4 and 8 hps, respectively. The NFKBIA, IL1B, IL8 and CCL4 genes were consistently induced at all times after endotoxin treatment. NLRC5 (CARD domain containing, NOD-like receptor family, RCJMB04_18i2), an intracellular receptor, was induced in HD11 cells treated with endotoxin.ConclusionsAs above using an in vitro model of chicken response to endotoxin, our data revealed the kinetics of gene networks involved in host response to endotoxin and extend the known complexity of networks in chicken immune response to Gram-negative bacteria such as Salmonella. The induction of NFKBIA, IL1B, IL8, CCL4 genes is a consistent signature of host response to endotoxin over time. We make the first report of induction of a NOD-like receptor family member in response to Salmonella endotoxin in chicken macrophages.

BackgroundNLRC5 is a member of the CARD domain containing, nucleotide-binding oligomerization (NOD)-like receptor (NLR) family, which recognizes pathogen-associated molecular patterns (PAMPs) and initiates an innate immune response leading to inflammation and/or cell death. However, the specific role of NLRC5 as a modulator of the inflammatory immune response remains controversial. It has been reported to be a mediator of type I IFNs, NF-kB, and MHC class I gene. But no study on NLRC5 function has been reported to date in chickens. In the current study, we investigated the role of NLRC5 in the regulation of IFNA, IFNB, IL-6, and MHC class I in the chicken HD11 macrophage cell line, by using RNAi technology. HD11 cells were transfected with one of five siRNAs (s1, s2, s3, negative-siRNA, or a mixture of s1, s2, s3-siRNAs). After 24 hours, cells were exposed to LPS or poly (I:C) or a vehicle control. Gene expression of NLRC5, IFNA, IFNB, IL-6, and MHC class I at 2, 4, 6, and 8 hours post stimulation (hps) was quantified by qPCR.ResultsThe expression of NLRC5, IFNA, IFNB, and IL-6 genes in negative irrelevant transfection controls was up-regulated at 2 hps after LPS treatment compared to the vehicle controls. S3-siRNA effectively knocked down NLRC5 expression at 4 hps, and the expression of IFNA and IFNB (but not IL-6 and MHC class I) was also down-regulated at 4 hps in s3-siRNA transfected cells, compared to negative irrelevant transfection controls. Stimulation by LPS appeared to relatively restore the decrease in NLRC5, IFNA, and IFNB expression, but the difference is not significant.ConclusionsFunctional characterization of chicken NLRC5 in an in vitro system demonstrated its importance in regulating intracellular molecules involved in inflammatory response. The knockdown of NLRC5 expression negatively mediates gene expression of IFNA and IFNB in the chicken HD11 cell line; therefore, NLRC5 likely has a role in positive regulation of IFNA and IFNB expression. No direct relationship was found between NLRC5 knockdown and IL-6 and MHC class I expression. Future studies will further clarify the roles of NLRC5 and other NLRs in infectious diseases of chickens and may increase the efficacy of antiviral vaccine design.

Vitamin E modulates the immune response, in part by reducing inflammation. The bacterial component lipopolysaccharide (LPS) can induce an inflammatory response in chickens. The objective of this study was to evaluate immunomodulatory effects of dietary type and level of vitamin E on response of broilers to LPS. One-day-old broiler males (n=96) were placed in a vitamin E-type (synthetic, natural) × vitamin E level (22, 220 IU/kg)×LPS (LPS, saline) block design. At 22 d, LPS (or saline) was injected subcutaneously. Spleens were harvested for RNA isolation at 3 and 24 h postinjection. Relative levels of RNA expression were measured for the immune-related genes: avian β defensin 10 (AvBD10), interleukin 6 (IL6), interferon-γ (IFN-γ), inducible nitric oxide synthase (iNOS), interleukin 10 and transforming growth factor- β1 (TGF-β1). Avian β defensin 10 and iNOS are innate antimicrobial proteins. Interleukin 6 and IFN-γ are pro-inflammatory cytokines, whereas interleukin 10 and transforming growth factor-β1 are anti-inflammatory cytokines. There were significantly higher splenic levels of IL6, IFN-γ, iNOS, and IL10 RNA expression at 3 h postinjection in chickens receiving LPS than in chickens 24 h post-LPS injection or saline-injected birds at either time. These data suggest that LPS induced an immune response that was regulated by both pro- and anti-inflammatory cytokines. Birds fed natural-type (versus synthetic) vitamin E had a significantly lower LPS-induced inflammatory response, as indicated by lower IL6 RNA expression levels, suggesting a protective effect from natural-type vitamin E when a chicken encounters a bacterial component.