Methods in isolation and characterization of bovine monocytes and macrophages

Ceciliani, Fabrizio; Morales, Glauco; Matteis, Giovanna De; Grandoni, Francesco; Ferreira, Rafaela Furioso; Roccabianca, P.; Lecchi, Cristina

doi:10.1016/j.ymeth.2020.06.017

Cited by 10 publications

(10 citation statements)

References 170 publications

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“…Heat-stressed Holstein cows showed a decreasing tendency of monocyte populations in August compared with those in May. Monocytes are myeloid cells produced by the bone marrow that play important roles in the immune response, such as to infections and injuries [59]. Contrary to the results of this study, another study reported that monocyte counts increased in summer [60].…”

Section: Discussioncontrasting

confidence: 77%

Changes in Blood Metabolites and Immune Cells in Holstein and Jersey Dairy Cows by Heat Stress

Joo

Lee

Park

et al. 2021

Animals

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Owing to increasing global temperatures, heat stress is a major problem affecting dairy cows, and abnormal metabolic responses during heat stress likely influence dairy cow immunity. However, the mechanism of this crosstalk between metabolism and immunity during heat stress remains unclear. We used two representative dairy cow breeds, Holstein and Jersey, with distinct heat-resistance characteristics. To understand metabolic and immune responses to seasonal changes, normal environmental and high-heat environmental conditions, we assessed blood metabolites and immune cell populations. In biochemistry analysis from sera, we found that variety blood metabolites were decreased in both Holstein and Jersey cows by heat stress. We assessed changes in immune cell populations in peripheral blood mononuclear cells (PBMCs) using flow cytometry. There were breed-specific differences in immune-cell population changes. Heat stress only increased the proportion of B cells (CD4–CD21+) and heat stress tended to decrease the proportion of monocytes (CD11b+CD172a+) in Holstein cows. Our findings expand the understanding of the common and specific changes in metabolism and immune response of two dairy cow breeds under heat stress conditions.

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Section: Discussioncontrasting

confidence: 77%

Changes in Blood Metabolites and Immune Cells in Holstein and Jersey Dairy Cows by Heat Stress

Joo

Lee

Park

et al. 2021

Animals

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“…In total (4.4 ± 0.83) × 10 6 BBMs were recovered from each 1-mL volume of bovine whole blood obtained from five individual cattle during approximately 2 months of culturing. Since normal bovine whole blood contains monocytes/macrophages at an average concentration of 4 × 10 5 cells/mL (Ceciliani et al, 2021), our data reveal that BBMs were efficiently proliferated using the coculture of whole blood with B46 cells. It is also likely that the endothelial features of B46 cells are effective at stimulating the proliferation of BBMs under coculturing conditions, as previously reported (Tanaka et al, 1993).…”

Section: Resultsmentioning

confidence: 76%

Isolation and propagation of bovine blood‐derived macrophages using a mixed culture with bovine endothelial B46 cells

Hiramatsu,

Ikeda,

Kawaji

et al. 2023

Cell Biology International

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Macrophages are innate immune cells with multiple functions such as phagocytosis, cytokine production, and antigen presentation. Since macrophages play critical roles in some bacterial infectious diseases in cattle, including tuberculosis, paratuberculosis, and brucellosis, the in vitro culturing of bovine macrophages is useful for evaluating host–pathogen interactions at the cellular and molecular levels. We have previously reported the establishment of two immortalized bovine liver sinusoidal cell lines, endothelial B46 cells and myofibroblast‐like A26 cells (Cell Biology International, 40, 1372−1379, 2016). In this study, we investigated the use of these cell lines as feeder cells that support the proliferation of bovine blood‐derived macrophages (BBMs). Notably, the B46 cell line efficiently acts as feeder cells for the propagation of BBMs. Compared with primary cultured vascular endothelial cells, the infinite proliferation ability of B46 cells is more beneficial for preparing confluent feeder layers. In conclusion, this study provides a simple and efficient protocol for the isolation and propagation of BBMs using a primary mixed culture of bovine whole blood with B46 feeder cells. Isolated BBMs are expected to be useful for developing in vitro models for studying the interactions between bovine pathogens and host immune cells.

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“…Whole blood from 6 clinically healthy pluriparous dairy cows was collected in sterile tubes with EDTA as an anticoagulant during routine slaughtering procedures. The peripheral blood mononuclear cells (PBMCs) were purified as previously reported from 60 mL of whole blood [ 23 ]. Briefly, the tubes were centrifuged at 1,260 g for 30 min at 18°C without brake, and the buffy coat was collected and diluted in cold PBS + 2 mM EDTA (1:2 dilution).…”

Section: Methodsmentioning

confidence: 99%

Comparative secretome analysis of Staphylococcus aureus strains with different within-herd intramammary infection prevalence

et al. 2022

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Staphylococcus aureus is a major pathogen causing intramammary infection and mastitis in dairy cows. S. aureus genotypes (GT) can differ significantly in their ability to diffuse and persist in the herd; while the association of virulence gene carriage with epidemiological behavior remains unclear, a role for secreted proteins has been postulated. We characterized the secretome of six S. aureus strains belonging to two genotypes with opposite within-herd prevalence, GTB (high) and GTS (low), corresponding to sequence types (ST) 8 and 398, by high-resolution tandem mass spectrometry and differential analysis with Proteome Discoverer. Data are available via ProteomeXchange with identifier PXD029571. Out of 720 identified proteins, 98 were unique or more abundant in GTB/ST8 and 68 in GTS/ST398. GTB/ST8 released more immunoglobulin-binding proteins, complement and antimicrobial peptide inhibitors, enterotoxins, and metabolic enzymes, while GTS/ST398 released more leukocidins, hemolysins, lipases, and peptidases. Furthermore, GTB/ST8 released the von Willebrand factor protein, staphylokinase, and clumping factor B, while GTS released the staphylococcal coagulase and clumping factor A. Hence, GTB/ST8 secretomes indicated a higher propensity for immune evasion and chronicity and GTS/ST398 secretomes for cellular damage and inflammation, consistent with their epidemiological characteristics. Accordingly, GTS/ST398 secretions were significantly more cytotoxic against bovine PBMCs in vitro . Our findings confirm the crucial role of extracellular virulence factors in S. aureus pathogenesis and highlight the need to investigate their differential release adding to gene carriage for a better understanding of the relationship of S. aureus genotypes with epidemiological behavior and, possibly, disease severity.

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Methods in isolation and characterization of bovine monocytes and macrophages

Cited by 10 publications

References 170 publications

Changes in Blood Metabolites and Immune Cells in Holstein and Jersey Dairy Cows by Heat Stress

Changes in Blood Metabolites and Immune Cells in Holstein and Jersey Dairy Cows by Heat Stress

Isolation and propagation of bovine blood‐derived macrophages using a mixed culture with bovine endothelial B46 cells

Comparative secretome analysis of Staphylococcus aureus strains with different within-herd intramammary infection prevalence

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