Phenotypic and functional analysis of bovine peripheral blood dendritic cells before parturition by a novel purification method

Abstract

Dendritic cells (DCs) are specialized antigen presenting cells specializing in antigen uptake and processing, and play an important role in the innate and adaptive immune response. A subset of bovine peripheral blood DCs was identified as CD172a+/CD11c+/MHC (major histocompatibility complex) class II + cells. Although DCs are identified at 0.1%–0.7% of peripheral blood mononuclear cells (PBMC), the phenotype and function of DCs remain poorly understood with regard to maintaining tolerance during the pregnancy. All cattle used in this study were 1 month before parturition. We have established a novel method for the purification of DCs from PBMC using magnetic‐activated cell sorting, and purified the CD172a+/CD11c+ DCs, with high expression of MHC class II and CD40, at 84.8% purity. There were individual differences in the expressions of CD205 and co‐stimulatory molecules CD80 and CD86 on DCs. There were positive correlations between expression of cytokine and co‐stimulatory molecules in DCs, and the DCs maintained their immune tolerance, evidenced by their low expressions of the co‐stimulatory molecules and cytokine production. These results suggest that before parturition a half of DCs may be immature and tend to maintain tolerance based on the low cytokine production, and the other DCs with high co‐stimulatory molecules may already have the ability of modulating the T‐cell linage.

Precise Delineation and Transcriptional Characterization of Bovine Blood Dendritic-Cell and Monocyte Subsets

“…Bovine DC subsets have been addressed in numerous studies, looking at blood ( 30 , 32 – 37 , 39 ), afferent lymph ( 25 – 31 ), lymph nodes ( 30 ), spleen ( 59 ), skin ( 29 ) and intestine ( 60 ). Sei et al ( 35 ) described three DC subsets in blood of cattle after depletion for CD3, IgM, CD14, and CD11b, defining pDC as CD4 + MHC-II − , and two subsets of cDC as being CD4 − MHC-II + and either positive or negative for CD11c.…”

“…While monocyte research in cattle is relatively young, some efforts have been made in the last two decades to functionally characterize bovine DC subsets, mainly in afferent lymph ( 25 – 31 ), but also in blood ( 30 , 32 – 37 ). High expression of CD205 has been described to allow identification of DC in bovine afferent lymph ( 38 ).…”

Prevention of mastitis in multiparous dairy cows with a previous history of mastitis by oral feeding with probiotic Bacillus subtilis

“…To detect dendritic cells (DCs), a negative selection was first performed to remove T cells, B cells, and monocytes from the PBMC according to methods noted in previous studies (Miyazawa et al, 2006; Zhuang et al, 2018). After blocking with control goat and rat IgG, PBMC were stained with a mixture of anti‐CD3 (MM1A), anti‐B cells (BAQ44A) and anti‐CD14 (CAM36A) antibodies for 30 min at 4°C, followed by an incubation with microbeads‐conjugated secondary antibodies.…”

“…A subset of the bovine peripheral blood DCs was identified as CD172a + /CD11c + /MHC class II + cells in the CD3 À /B-B2 À /CD14 À population (Miyazawa et al, 2006;Zhuang et al, 2018). However, it is well-known that CD11c is highly expressed on monocytes, macrophages (Mø) and natural killer (NK) cells, and that CD172a + / CD11c + cells possibly include a subset of T cells, B cells, NK cells and monocytes/Mø.…”