Background
A complex community of microorganisms in the gastrointestinal (GI) tract, known as the gut microbiota, exerts major effects on gene expression and cytokine profile. Extracellular vesicles (EVs) which are produced by bacteria could be sensed by Toll like receptors (TLRs). The interaction between gut microbiota and TLRs affects homeostasis and immune responses. In this study, we evaluated TLR9 gene expression and cytokines level in Caco-2 cell line treated with Lactobacillus casei as one of the gut microbiota and its EVs.
Conclusions
L. casei and its EVs have pivotal role in the cross talk between gut microbiota and the host especially in the modulation of the immune system. This study shows for the first time the increasing level of anti-inflammatory cytokines by EVs released by L. casei. Based on the last studies on immunomodulatory effect of EVs on immune cells and our results in cell line level, we postulate that L. casei derived EVs could be possible candidates for the reduction of immune responses.
Methods
In the present study, L. casei derived EVs was extracted via ultracentrifugation. The quality control assessment included the evaluation of physicochemical characteristics of EVs. For the treatment of Caco-2 cell line, L. casei and its EVs (100 and 150 μg/mL) were used. In addition, qRT-PCR assay was carried out to evaluate the mRNA expression of TLR9 gene. ELISA assay was also performed to determine the levels of IFNγ, TNF-α, GM-CSF, IL-1α, IL-1β, IL-2, IL-4, IL-6, IL-8, IL-12, IL-17A, and IL-10 cytokines.
Results
The results showed that L. casei slightly increased TLR9 gene expression in the Caco-2 cell line. It was also found that EVs at concentrations of 100 and 150 μg/mL could significantly decrease TLR9 gene expression. Furthermore, L. casei significantly increased IL-10 and IFNγ levels. Based on the findings, the level of IL-17A, as a proinflammatory cytokine, decreased by L. casei. Both concentrations of EVs decreased the level of IFNγ, while increasing the concentrations of IL-4 and IL-10. EVs from L. casei could modulate immune responses in the Caco-2 cell line. Both EVs and L. casei activated the expression and secretion of several cytokines. Conclusions: L. casei and its EVs have pivotal role in the cross talk between gut microbiota and the host especially in the modulation of the immune system. This study shows for the first time the increasing level of anti-inflammatory cytokines by EVs released by L. casei. Based on the last studies on immunomodulatory effect of EVs on immune cells and our results in cell line level, we postulate that L. casei derived EVs could be possible candidates for the reduction of immune responses.