Immunomodulatory Effects of Bacterial Toll-like Receptor Ligands on the Phenotype and Function of Milk Immune Cells in Dromedary Camel

Abstract

(1) Toll-like receptors (TLR) are a family of pattern recognition receptors that sense distinct molecular patterns of microbial origin. Although the immune cell composition of camel milk has been recently described, host–pathogen interaction studies in the camel mammary gland are still scarce. The present study aimed to use a whole milk stimulation assay for investigating the modulatory effect of selected Toll-like receptor (TLR) ligands on the phenotype and function of milk immune cells. (2) Methods—camel milk samples (n = 7) were stimulated in vitro with the TLR4 ligand LPS or the TLR2/1 ligand Pam3CSK4, and separated milk cells were evaluated for stimulation-induced shape change, the expression of cell surface markers, phagocytosis, apoptosis, ROS production, and NETosis. Stimulation with PMA was used as a control stimulation. (3) Results—all stimulants induced shape change in milk cells, change in the expression of several cell markers, and increased cell apoptosis and NETosis. In addition, stimulation with Pam3CSK4 and PMA was associated with enhanced ROS production, while only PMA stimulation resulted in enhanced bacterial phagocytosis by milk immune cells. (4) Conclusions—our data indicates selective modulating effects of the TLR ligands LPS and Pam3CSK4 on camel milk phagocytes. These results may have implications for the use of synthetic TLR agonists as immunomodulatory adjuvants of the immune response to intra-mammary vaccines against mastitis pathogens.

NETosis and Calcium influx in Dromedary Camel Neutrophils after in vitro Toll-like Receptor Stimulation

“…Such studies are still lacking for camel neutrophils. In a recent study, activation of TLR-4, TLR-2/1, and TLR-7/8, but not of TLR-3, resulted in the activation of camel neutrophils with stimulation-induced shape change and modulation of activation markers expression (Hussen et al, 2023b).…”

“…TLRs are membrane and intracellular pattern recognition receptors interacting with PAMPs (Newton and Dixit, 2012;Zindel and Kubes, 2020). Although few recent studies analyzed the impact of some TLR-agonists on some functions of neutrophils in the dromedary camel (Hussen et al, 2023a;Hussen et al, 2023b), many questions still exist regarding the modulatory effect of TLR stimulation on neutrophils phenotype and function. Especially the potential of bacterial and viral TLR-agonists to induce NETformation or Ca 2+ influx in camel neutrophils has not been investigated so far.…”

“…The TLR stimulation was performed as previously described (Hussen et al, 2023b). The TLR ligands lipopolysaccharide (LPS), Pam3CSK4, R848, and Poly IC were purchased from Invivogen (San Diego, USA).…”

Interaction of Specific Monoclonal Antibodies with Leukocyte Antigens in Camels

“…Monoclonal antibodies (mAbs) play a crucial role in the process of identifying immune cells and monitoring their distribution and mobilization in distinct tissues such as the lungs, the ovary, and the lymph node (Gunnes et al, 2003;Maecker et al, 2012;Rivers et al, 2022). In order to avoid the laborious procedure of generating species-specific mAbs, researchers frequently employ cross-reactive mAbs (Farady et al, 2009;Irani et al, 2016;Grandoni et al, 2020;Grandoni et al, 2023;Hussen et al, 2023a;Hussen et al, 2023b). Over the past few years, several studies have been performed to test the cross-reactivity of mAbs against leukocyte antigens sourced from humans or other veterinary animals with their corresponding camel antigens (Hussen et al, 2017;Hussen and Schuberth, 2020;.…”

“…The list of mAbs and the target cluster of differentiation antigen molecules have been recently reviewed (Hussen and Schuberth, 2020). The identification of such mAbs has been employed in several important studies on camel immune cells (Hussen et al, 2020a;Hussen et al, 2020b;Hussen et al, 2021;Hussen et al, 2023a;Hussen et al, 2023b).…”