Bovine Peripheral Blood Monocyte Derived Dendritic Cell Culture and Identification in Vitro

ZHAN Kang; ZHAO Qianming; SUI Yannan; FENG Feifei; ZHAN Jinshun; ZHAO Guoqi

doi:10.3969/j.issn.1006-267x.2016.07.024

Chinese Journal of Animal Nutrition >

2016 , Vol. 28 >Issue 7: 2184 - 2190

DOI: https://doi.org/10.3969/j.issn.1006-267x.2016.07.024

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Bovine Peripheral Blood Monocyte Derived Dendritic Cell Culture and Identification in Vitro

ZHAN Kang ,
ZHAO Qianming ,
SUI Yannan ,
FENG Feifei ,
ZHAN Jinshun ,
ZHAO Guoqi

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College of Animal Science and Technology, Yangzhou University, Yangzhou 225009, China

Received date: 2016-01-12

Online published: 2016-07-28

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Abstract

This study aimed to induce bovine peripheral blood monocyte derived dendritic cell by granulocyte-macrophage colony stimulating factor (GM-CSF) and interleukin-4 (IL-4) cytokines, which could lay foundation and provide cell model for dairy cow mastitis treatment using cell immunotherapy. The bovine peripheral blood monocyte was acquired by lymphocyte separation medium and seeded in 6-proe plate to culture for 2 h. Then, suspended cells containing an amount of B and T cells were discarded, and adherent cells were mostly monocyte. The cells were washed for 5 times using phosphate buffer, and 2 mL culture medium containing GM-CSF and IL-4 cytokines was added to culture for 3 d. After that, 1.4 mL culture medium was discard from medium top, and 1.8 mL culture medium containing GM-CSF and IL-4 was added to culture for another 3 d. The cell was observed under microscope every day. On days 7, the cells were harvested to detect CD11c, CD14, major histocompatibility complex Ⅱ (MHCⅡ), CD40, CD80 and CD86 molecules by flow cytometry. The result showed as follows: 1) on days 2, some of the cells started to extend spikes and accompanied by pseudopodia stretching. On days 3, the spikes and pseudopodia grew more and more. On days 4 and 5, cells with spikes and pseudopodia started aggregation and fusion. On days6, most of monocytes were derived to dendritic cells, on which plenty of spikes and pseudopodia could be clearly seen. 2) CD14, CD11c, MHCⅡ, CD80 and CD86 positively expressed cells accounts for 6.8%, 65.0%, 75.9%, 2.0% and 1.2% of induced cells by flow cytometry, respectively. In conclusion, bovine peripheral blood monocytes can be derived to dendritic cells with certain purity.

Key words： cellular immune treatment; monocyte; dendritic cell; bovine mastitis

Cite this article

ZHAN Kang , ZHAO Qianming , SUI Yannan , FENG Feifei , ZHAN Jinshun , ZHAO Guoqi . Bovine Peripheral Blood Monocyte Derived Dendritic Cell Culture and Identification in Vitro[J]. Chinese Journal of Animal Nutrition, 2016 , 28(7) : 2184 -2190 . DOI: 10.3969/j.issn.1006-267x.2016.07.024

References

[1] MELLMAN I,STEINMAN R M.Dendritic cells:specialized and regulated antigen processing machines[J].Cell,2001,106(3):255-258.

[2] WOO S R,CORRALES L,GAJEWSKI T F,et al.The STING pathway and the T cell-inflamed tumor microenvironment[J].Trends in Immunology,2015,36(4):250-256.

[3] 马燕芬,宋利文,高民,等.氧化应激对围产期奶牛乳房炎的影响及其调控机制[J].动物营养学报,2015,27(3):671-676.
[4] 李幼平.树突状细胞与移植免疫耐受研究进展[J].生物医学工程学杂志,1997,14(4):383-387.
[5] WERLING D,HOPE J C,CHAPLIN P,et al.Involvement of caveolae in the uptake of respiratory syncytial virus antigen by dendritic cells[J].Journal of Leukocyte Biology,1999,66(1):50-58.
[6] SCHREINER J,KRETSCHMER D,KLENK J,et al.Staphylococcus aureus phenol-soluble modulin peptides modulate dendritic cell functions and increase in vitro priming of regulatory T cells[J].Journal of Immunology,2013,190(7):3417-3426.

[7] GUERMONPREZ P,HELFT J,CLASER C,et al.Inflammatory Flt3L is essential to mobilize dendritic cells and for T cell responses during Plasmodium infection[J].Nature Medicine,2013,19(6):730-738.

[8] STEINMAN R M,COHN Z A.Identification of a novel cell type in peripheral lymphoid organs of mice[J].Journal of Experimental Medicine,1973,137(5):1142-1162.

[9] STEINMAN R M,HEMMI H.Dendritic cells:translating innate to adaptive immunity[J].Current Topics in Microbiology and Immunology,2006,311:17-58.
[10] LEI L Y,HOSTETTER J M.Limited phenotypic and functional maturation of bovine monocyte-derived dendritic cells following Mycobacterium avium subspecies paratuberculosis infection in vitro[J].Veterinary Immunology and Immunopathology,2007,120(3/4):177-186.
[11] CRUZ-ADALIA A,RAMIREZ-SANTIAGO G,CALABIA-LINARES L C,et al.T cells kill bacteria captured by transinfection from dendritic cells and confer protection in mice[J].Cell Host & Microbe,2014,15(5):611-622.

[12] LIU Y J.Dendritic cell subsets and lineages,and their functions in innate and adaptive immunity[J].Cell,2001,106(3):259-262.

[13] BORKOWSKI T A,LETTERIO J J,FARR A G,et al.A role for endogenous transforming growth factor β1 in Langerhans cell biology:the skin of transforming growth factor β1 null mice is devoid of epidermal Langerhans cells[J].Journal of Experimental Medicine,1996,184(6):2417-2422.

[14] GALY A,TRAVIS M,CEN D Z,et al.Human T,B,natural killer,and dendritic cells arise from a common bone marrow progenitor cell subset[J].Immunity,1995,3(4):459-473.

[15] MIYAZAWA K,ASO H,HONDA M,et al.Identification of bovine dendritic cell phenotype from bovine peripheral blood[J].Research in Veterinary Science,2006,81(1):40-45.

[16] LIPSCOMB M F,MASTEN B J.Dendritic cells:immune regulators in health and disease[J].Physiological Reviews,2002,82(1):97-130.

[17] RENJIFO X,HOWARD C,KERKHOFS P,et al.Purification and characterization of bovine dendritic cells from peripheral blood[J].Veterinary Immunology and Immunopathology,1997,60(1/2):77-88.
[18] 段杰林,杨冠,尹杰,等.肠道树突状细胞的作用机制及益生菌对其影响[J].动物营养学报,2013,25(1):26-35.

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