Fig. 5: 6ʹ-SLN ligands on the endometrial cells engaged Siglec-9 on Met-5A cells

a To confirm binding between 6ʹ-SLN and Siglec-9, cell lysates from Met-5A cells were subjected to a pull-down assay using biotin-labeled 6ʹ-SLN and incubated with avidin beads. Proteins binding to the avidin beads were resolved and subjected to western blot analysis. b Met-5A cells were incubated with siRNA targeting Siglec-9 for 24 h. Expression of Siglec-9 in Met-5A cells was assessed by RT-PCR and western blot analysis. c siSiglec-9-transfected Met-5A cells were cultured to a confluent monolayer. Tracker™ Green CMFDA-labeled Ishikawa cells were added to the monolayer and incubated for 20 min. Adhered Ishikawa cells were manually counted, and the results are expressed as the fold-difference relative to the control (mean ± SD). d Ishikawa cells were treated with TGF-β1 (10 ng/mL) for 48 h. Met-5A cells were incubated with neutralizing anti-Siglec-9 antibody (2 μg/mL) 24 h before the adhesion assay. Fluorescence-labeled Ishikawa cells were added onto Met-5A cells. After 20 min of incubation and then washing, attached cells were manually counted, and the results are expressed as the fold-difference relative to the control. ** p < 0.01 and *** p < 0.001 compared with the control group. ^## p < 0.01 and ^### p < 0.001 compared with the TGF-β1-treated group