IQGAP2 expression In mi-124 treated macrophages exp2

Peripheral blood was collected from 20 healthy piglets via jugular vein and mixed with an anticoagulant. The blood sample was diluted in a ratio of 1:1 with sterile normal saline. The peripheral blood mononuclear cells were isolated using Percoll. The cells were transferred to a 6-well cell culture plate. After adhered to well surface and washed, the cells were stimulated with GM-CSF (20 ng/mL) and IL-4 (10 ng/mL) for six days to obtain fully differentiated macrophages. Afterwards, the macrophages were transfected with IQGAP2 siRNA, miR-124, and antisense miR-124. At 0, 6, and 12 h, the flow cytometry assay was used to detect the number of IQGAP2-positive cells. The differentiated porcine macrophages in each group were infected with Salmonella in logarithmic growth stage by adding 5 mL of Salmonella solution (106 cfu). Cells and bacteria were incubated at 37 °C, 5% CO2 for 2 h, rinsed with phosphate-buffered saline (PBS) three times, and treated with 100 ?g/mL gentamicin for 2 h. Subsequently, the cells were collected after continued culture in antibiotic-free medium. The cells infected with Salmonella at 0, 6, and 12 h post infection were collected. The expression of IQGAP2 and the number of IQGAP2-positive cells among the Salmonella-infected cells was determined by flow cytometry.