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. 2016 Apr 4;2016:8468549. doi: 10.1155/2016/8468549

Figure 4.

Figure 4

HLA-G expression mediates the HLSC activity on NKs. (a) Representative (n = 4) cytofluorimetric analysis of HLA-G expression on HLSC membrane and after intracytoplasmic staining, using the specific MEMG/9 antibody (native form for human HLA-G1). (b) Western Blot analyses confirmed the presence of HLA-G in HLSCs at different passages of culture (p4, p6, and p8). (c) Soluble HLA-G is present in HLSC supernatants, as indicated by Western Blot after immune-precipitation with specific antibody. HLA-G protein expression was detected through mAb 4H84, resulting in specific band corresponding to the expected molecular weight of 39 kDa. (d) Representative (n = 4) cytofluorimetric analysis of CD107a expression on NKs in degranulation assay, after 4 days of incubation with HLSCs in the presence or the absence of specific blocking antibody against HLA-G (87-G). HLSCs, via the secretion of HLA-G reduced NK degranulation, as a specific neutralizing anti-HLA-G antibody restored degranulation activity to levels similar to those following incubation of NKs with K562 cells alone (CD56+CD107+: 87 ± 5% in NK CTRL, 5.5 ± 4% in NKs cocultured with HLSCs, and 90 ± 5.3% in NKs cocultured with HLSCs in the presence of blocking antibody).