HLA-G Does Not Mediate the Natural Killer Cell Suppression Associated with Endometriosis

Abstract

Objectives: The immunological mechanisms that support persistence and proliferation of ectopic endometrial implants within the peritoneal cavity of women with endometriosis are unknown. However, evidence of inhibition in natural killer (NK) cell function has been described in several reports of this syndrome. In the current study we tested the hypothesis that a nonclassical major histocompatibility antigen, HLA-G, was responsible for NK cell suppression.Design: Nested case-control study of women with and without laparoscopic evidence of endometriosis.Materials and Methods: Peritoneal fluid specimens from 5 women with revised AFS stage I-IV endometriosis and 5 age-matched normal controls without laparoscopic evidence of endometriosis were tested for the presence of HLA-G protein. Endometriosis and normal endometrial biopsies (2 each) were directly evaluated for HLA-G or used to prepare stromal cell cultures for this purpose. The expression of HLA-G in peritoneal fluid, tissue and isolated cell culture lysates and supernatants was determined by immunoblotting with a specific monoclonal antibody.Results: Using the Western blotting assay, we were unable to detect HLA-G in specimens of peritoneal fluid from normal or endometriosis subjects. By contrast, JEG-3 cell lysates demonstrated the expected 37 kD immunopositive band. Neither normal endometrium nor endometriosis-derived biopsies contained detectable HLA-G protein. Under basal culture conditions neither normal endometrial nor endometriosis stromal cells expressed HLA-G protein. Even following incubation with TNF-α and IFN-γ the cells failed to express detectable HLA-G protein.Conclusions: HLA-G protein was not detectable in peritoneal fluid specimens of endometriosis patients or controls. Moreover, ectopic and eutopic endometrial tissues and stromal cells do not express HLA-G. NK cell inhibition in endometriosis must be mediated by factors other than HLA-G.These studies were supported in part by a grant from the Deutsche Forschungsgemeinschaft (DH) and the NIH/NICHD, through cooperative agreement U54-HD37321, as part of the Specialized Cooperative Centers Program in Reproduction Research (RNT). Objectives: The immunological mechanisms that support persistence and proliferation of ectopic endometrial implants within the peritoneal cavity of women with endometriosis are unknown. However, evidence of inhibition in natural killer (NK) cell function has been described in several reports of this syndrome. In the current study we tested the hypothesis that a nonclassical major histocompatibility antigen, HLA-G, was responsible for NK cell suppression. Design: Nested case-control study of women with and without laparoscopic evidence of endometriosis. Materials and Methods: Peritoneal fluid specimens from 5 women with revised AFS stage I-IV endometriosis and 5 age-matched normal controls without laparoscopic evidence of endometriosis were tested for the presence of HLA-G protein. Endometriosis and normal endometrial biopsies (2 each) were directly evaluated for HLA-G or used to prepare stromal cell cultures for this purpose. The expression of HLA-G in peritoneal fluid, tissue and isolated cell culture lysates and supernatants was determined by immunoblotting with a specific monoclonal antibody. Results: Using the Western blotting assay, we were unable to detect HLA-G in specimens of peritoneal fluid from normal or endometriosis subjects. By contrast, JEG-3 cell lysates demonstrated the expected 37 kD immunopositive band. Neither normal endometrium nor endometriosis-derived biopsies contained detectable HLA-G protein. Under basal culture conditions neither normal endometrial nor endometriosis stromal cells expressed HLA-G protein. Even following incubation with TNF-α and IFN-γ the cells failed to express detectable HLA-G protein. Conclusions: HLA-G protein was not detectable in peritoneal fluid specimens of endometriosis patients or controls. Moreover, ectopic and eutopic endometrial tissues and stromal cells do not express HLA-G. NK cell inhibition in endometriosis must be mediated by factors other than HLA-G. These studies were supported in part by a grant from the Deutsche Forschungsgemeinschaft (DH) and the NIH/NICHD, through cooperative agreement U54-HD37321, as part of the Specialized Cooperative Centers Program in Reproduction Research (RNT).