Anti-IFNA1 Monoclonal antibody, PE/Cy7®

Infection of ruminants and humans with Fasciola gigantica is attracting increasing attention due to its economic impact and public health significance. However, little is known of innate immune responses during F. gigantica infection. Here, we investigated the expression profiles of genes involved in Toll-like receptors (TLRs) and NOD like receptors (NLRs) signaling pathways in buffaloes infected with 500 F. gigantica metacercariae. Serum, liver and peripheral blood mononuclear cell (PBMC) samples were collected from infected and control buffaloes at 3, 10, 28, and 70 days post infection (dpi). Then, the levels of 12 cytokines in serum samples were evaluated by ELISA. Also, the levels of expression of 42 genes, related to TLRs and NLRs signaling, in liver and PBMCs were determined using custom RT2 Profiler PCR Arrays. At 3 dpi, modest activation of TLR4 and TLR8 and the adaptor protein (TICAM1) was detected. At 10 dpi, NF-kappa B1 and Interferon Regulatory Factor signaling pathways were upregulated along with activation of TLR1, TLR2, TLR6, TLR10, TRAF6, IRF3, TBK1, CASP1, CD80, and IFNA1 in the liver, and inflammatory response with activated TLR4, TLR9, TICAM1, NLRP3, CD86, IL-1B, IL-6, and IL-8 in PBMCs. At 28 dpi, there was increase in the levels of cytokines along with induction of NLRP1 and NLRP3 inflammasomes-dependent immune responses in the liver and PBMCs. At 70 dpi, F. gigantica activated TLRs and NLRs, and their downstream interacting molecules. The activation of TLR7/9 signaling (perhaps due to increased B-cell maturation and activation) and upregulation of NLRP3 gene were also detected. These findings indicate that F. gigantica alters the expression of TLR5 and NLRs genes to evade host immune defenses. Elucidation of the roles of the downstream effectors interacting with these genes may aid in the development of new interventions to control disease caused by F. gigantica infection.

Introduction: IFNA1 (interferon alpha) is a key cytokine regulating the activity of numerous immune cells. Plasmacytoid dendritic cells (pDCs) as natural interferon-producing cells play critical roles as sensors of pathogens and link innate to adaptive immunity. CpG motifs within DNA sequences activating toll-like receptor 9 (TLR9) are the main stimuli eliciting IFNA1 secretion from pDCs. Adrenergic substances are capable of differentially modulating the response from various immune cells. Hence, the aim of this study was to examine how adrenoceptor stimulation influences TLR9-induced IFNA1 secretion from human pDCs. Methods: PBMCs generated from human whole blood and pDCs enriched from buffy coats were stimulated with LPS and CpG-ODN 2336 in the presence or absence of epinephrine and different adrenoceptor antagonists. Secretion of TNF and IFNA1 was measured by ELISA. Flow cytometry was used to determine efficacy of pDC enrichment and adrenoceptor expression of PBMC subsets. The influence of modified IFNA1 secretion on NK cell activity was evaluated using a colorimetric tumor cell lysis assay. Results: TLR9-induced IFNA1 secretion as well as TLR4-induced TNF secretion from PBMCs was dose-dependently attenuated by coincubation with epinephrine. Combination with different specific adrenoceptor antagonists revealed that this effect was mediated by the adrenoceptor b2 (ADRB2). Since flow cytometric analysis could exclude the presence of ADRB2 on pDCs, highly enriched pDCs lacked any visible impact of adrenoceptor stimulation on TLR9-induced IFNA1 release. Combination of pDCs with PBMCs restored the effect, even when they were separated by a permeable membrane. Suppression of TLR9-mediated IFNA1 secretion from PBMCs by adrenoceptor stimulation reduced the lytic activity of NK cells on K562 tumor cells. Conclusion: We provide insights into the underlying mechanisms of the interrelation between immune responses and pharmacological agents widely used in clinical practice. Our results have implications for the future treatment of human patients, in which the endogenous immune response plays a pivotal role, such as during viral infections, inflammatory diseases and cancers.