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 andnorthern Australia. Clinical manifestations range from asymptomatic, sub-acute and chronic infections to acutesepsis. However, the mechanism of immune response to B. pseudomallei is known very little. This study aimed togenerate monocyte derived dendritic cells (MoDcs) from human peripheral blood samples as a model to studyimmune response to B. pseudomallei. By using immune-magnetic bead sorting, monocytes (CD14+ cells) wereisolated from pheripheral blood and the purity of obtained monocytes was approximately 95%. Moreover, themonocytes differentiated into ็immature้ DC by GM-CSF and IL-4 activation as evidenced by their surfacephenotypes assayed by flow cytometry eg. HLA-DR and CD11c upregulation whilst CD14 downregulation. WhenMoDCs were collected and stimulated with heat killed- B. pseudomallei or lipopolysaccharide from E. coli for24 or 48 hours, the results indicated the highest levels of IL-6 induced by B. pseudomallei and it was comparablebetween 24 and 48 hours. In conclusion, we demonstrated the generation of MoDCs which could be used for furtherstudy of immune responses to B. pseudomallei and other pathogens.