Abstract
Brucella abortus is a Gram-negative, facultative intracellular bacterium that causes brucellosis, a worldwide zoonotic disease leading to undulant fever in humans and abortion in cattle. The immune response against this bacterium relies on the recognition of microbial pathogen-associated molecular patterns, such as lipoproteins, lipopolysaccharides, and DNA; however, the immunostimulatory potential of B. abortus RNA remains to be elucidated. Here, we show that dendritic cells (DCs) produce significant amounts of IL-12, IL-6, and IP-10/CXCL10, when stimulated with purified B. abortus RNA. IL-12 secretion by DCs stimulated with RNA depends on TLR7 while IL-6 depends on TLR7 and partially on TLR3. Further, only TLR7 plays a role in IL-12 production induced by B. abortus infection. Moreover, cytokine production in DCs infected with B. abortus or stimulated with bacterial RNA was reduced upon pretreatment with MAPK/NF-κB inhibitors. By confocal microscopy, we demonstrated that TLR7 is colocalized with B. abortus in LAMP-1+ Brucella-containing vacuoles. Additionally, type I IFN expression and IP-10/CXCL10 secretion in DCs stimulated with bacterial RNA were dependent on TLR3 and TLR7. Our results suggest that TLR3 and TLR7 are not required to control Brucella infection in vivo, but they play an important role on sensing B. abortus RNA in vitro.
Highlights
The innate immunity is important to the initial recognition of pathogens, the development of a neutralizing response, and mobilization of the adaptive immunity [1]
To determine whether TLR3 and TLR7 are required to induce proinflammatory cytokine production in dendritic cells (DCs) stimulated with B. abortus, cells from C57BL/6, TLR3 KO, TLR7 KO, and MyD88 KO were cultured with B. abortus (MOI 100:1) and after 24 h the supernatants were tested for production of IL-12, IL-6, and TNF-α by ELISA
DCs from C57BL/6, TLR3 KO, TLR7 KO, and MyD88 KO were stimulated with purified B. abortus RNA and after 24 h the supernatants were tested for production of IL-12, IL-6, and TNF-α
Summary
The innate immunity is important to the initial recognition of pathogens, the development of a neutralizing response, and mobilization of the adaptive immunity [1]. It was reported that bacteria from group B Streptococcus (GBS) potently induced type I IFN in conventional DCs (cDCs) by a mechanism that required TLR7, MyD88, and the transcription factor IRF-1 These molecules colocalized with bacterial products in degradative vacuoles bearing lysosomal markers, linking lysosomal recognition of bacterial RNA with a robust IFN response [11]. Deshmukh et al reported that the recognition of ssRNA from GBS and other Gram-positive bacteria by macrophages and monocytes depends on the adaptors MyD88 and UNC93B, but not TLRs [14] It was demonstrated by Gratz et al that in cDCs and macrophages stimulated with Streptococcus pyogenes RNA, type I IFN was induced in the absence of TLR3, TLR7, and TLR9 [15]
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