Characterization Of Uterine NK Cells In NOD/SCID Mice During Pregnancy

ObjectivesTo characterize uterine natural killer(uNK) cells in non-obese diabetic/severe combined immunodeficiency(NOD/SCID) mice and investigate the potential role of these cells in pregnancy tolerance.Materials and MethodsSyngeneic NOD/SCID×NOD/SCID breeding scheme was established,while BALB/c×BALB/c mating combination was used as a non-immunodeficiency control. NOD/SCID mice were characterized by functional deficits in T and B lymphocytes in addition to impaired NK cell function.The two types of syngeneic pregnancy mice were groped respectively.Some pregnant females were left untreated while the others were administrated intraperitoneally with polyinosinic polycytidylic acid[poly(I:C)] or lipopolysaccharide(LPS) respectively at gestational days 8.5 and 9.5,with the phosphate buffer solution(PBS)-administrated mice as the control group.The mice were killed at day 13.5 of gestation and the resorption rate of embryos was calculated.Mononuclear cells were isolated from peripheral blood(PB),placenta and decidua, DX5⁺ cells were separated using microbead-conjugated anti-mouse DX5(CD49b) and magnetic affinity cell sorting separation columns.Then detected the phenotype and the expression of cytokines by flow cytometry.Peripheral blood mononuclear cells were harvested from non-pregnant or pregnant BALB/c and NOD/SCID mice,and DX5⁺ cells were purified by magnetic affinity cell sorting.Aliquots of cells were cultured in vitro,with the stimulation of poly(I:C),LPS, the combination of poly(I:C) and LPS,or left untreated.These cells were cultured in the presence or absence of inhibitor,detected TLR-agonists on IFN-γproduction in DX5⁺ cells and TLR3.TLR4.TLR8 expression. Detected NK cell cytotoxicity by Standard ⁵¹Cr release assay and analyzed the expression of iNOS in placental NK cells by flow cytometer.ResultsThe dominant subset of uNK cells in NOD/SCID mice were DX5(CD49b)⁺,asialo ganglio-N-tetraosylceramide(ASGM1)⁺,CD25⁺,CD122⁺,Thy-1(CD90)^hi,c-kit (CD117)^hi and interleukin(IL)-10⁺.In addition,the percentage of interferon(IFN)-γ⁺ subset was slightly increased in response to selected TLR-agonists in the NOD/SCID, whereas the corresponding percentage in BALB/c could be increased dramatically. Such an effect could be abrogated by inhibitors including LY294002,SP600125 and PD98059.The significant increase of IFN-γ⁺ NK cell percentage in BALB/c was concomitant with the increase of embryo resorption rate.In contrast,the resorption rate in NOD/SCID mice was not significantly increased upon the induction of polyinosinic polycytidylic acid[poly(I:C)]or lipopolysaccharide(LPS).As expected,the NK cells from NOD/SCID mice displayed a detectable but lower cytotoxicity than BALB/c,as determined by standard ⁵¹Cr release assay.In addition,the uNK cells from NOD/SCID mice also displayed a hyposensitivity to LPS-induced production of inducible nitric oxide synthase(iNOS).ConclusionsA considerable percentage of immature NK cells were detected at the feto-maternal interface in NOD/SCID mice.These cells were hyposensitive to the stimulation of selected TLR-agonists.Such a status appeared to be beneficial for the maintenance of pregnancy.