Establishment,identification and application of induction culture method of mono-nuclear macrophages from cow bone marrow
10.16303/j.cnki.1005-4545.2024.08.13
- VernacularTitle:奶牛骨髓源单核巨噬细胞诱导培养方法建立鉴定及应用
- Author:
Yu WANG
1
,
2
;
Xiaolin YANG
;
Lili GUO
;
Pengfei GONG
;
Jingze WU
;
Wei MAO
;
Shuangyi ZHANG
;
Bo LIU
;
Jinshan CAO
Author Information
1. 内蒙古农业大学兽医学院,内蒙古呼和浩特 010018
2. 农业农村部动物疾病临床诊疗技术重点实验室,内蒙古呼和浩特 010018
- Keywords:
cow;
bone marrow derived macrophages;
isolation culture identification;
PGD2
- From:
Chinese Journal of Veterinary Science
2024;44(8):1674-1681
- CountryChina
- Language:Chinese
-
Abstract:
In order to establish the isolation,culture and identification method of cow bone marrow-derived macrophages,three different media(RPMI-1640,DMEM,DMEM/F12)were added with 20％fetal bovine serum(FBS),2.4％chlorine-streptomycin,1.2％glutamine(Gln),and M-CSF(20 ng/mL),respectively,to induce the monocytes extracted from the bone marrow of dairy cows to become macrophages.The induced M0 macrophages were polarized into M1-type macrophages by adding lipopolysaccharide(LPS).The morphology of macrophages was observed by optical mi-croscope at day 1,4 and 7,and the differences of differentiated macrophages between the three media were compared.The effects of prostaglandin D2(PGD2)-DP2 receptor pathway on the secre-tion of cytokines(IL-6,TNF-α)induced by Escherichia coli and phagocytosis of macrophages were also investigated.The results showed that the morphological changes of cells cultured in the medium of RPMI-1640 were the most obvious and the number was large.A large number of char-acteristic markers of mononuclear macrophages were detected(M0 markers:CD1 1b,CD14;M1 markers:CD11b,CD80)expression,M0 and M1 macrophage purity were 79.9％and 93.5％,re-spectively.COX-2 and H-PGDS gene expressions were significantly increased in E.coli group com-pared with the blank control group.The secretion of PGD2also increased significantly(P＜0.000 1).DP2 receptor inhibitors(CAY10471,CAY10595)could significantly inhibit the secretion of E.coli in-duced pro-inflammatory cytokines(IL-6,TNF-α)and significantly enhance the killing effect of macrophages on E.coli.The above results showed that the induced cells had the characteristic mor-phology and immunophenotype of macrophages.E.coli can induce the production of PGD2 in mac-rophages,and the PGD2-DP2 pathway regulates the secretion of cytokines in E.coli infected macro-phages.
