Generation of dendritic cells from human monocytes to study immune response against Burkholria pseudomallei

Abstract

Burkholderia pseudomallei

(B. pseudomallei) is a major causative agent of melioidosis in southeast Asia and

northern Australia. Clinical manifestations range from asymptomatic, sub-acute and chronic infections to acute

sepsis. However, the mechanism of immune response to

B. pseudomallei is known very little. This study aimed to

generate monocyte derived dendritic cells (MoDcs) from human peripheral blood samples as a model to study

immune response to

B. pseudomallei. By using immune-magnetic bead sorting, monocytes (CD14+ cells) were

isolated from pheripheral blood and the purity of obtained monocytes was approximately 95%. Moreover, the

monocytes differentiated into ็immature้ DC by GM-CSF and IL-4 activation as evidenced by their surface

phenotypes assayed by flow cytometry eg. HLA-DR and CD11c upregulation whilst CD14 downregulation. When

MoDCs were collected and stimulated with heat killed-

B. pseudomallei or lipopolysaccharide from E. coli for

24 or 48 hours, the results indicated the highest levels of IL-6 induced by

B. pseudomallei and it was comparable

between 24 and 48 hours. In conclusion, we demonstrated the generation of MoDCs which could be used for further

study of immune responses to

B. pseudomallei and other pathogens.