Gestational Stage—Specific Regulation of NK and T cell Compositions by Trafficking and Proliferation 2254964

Abstract Introduction Decidual natural killer (NK) and T cells undergo compositional changes during pregnancy, yet the mechanisms regulating their origin and maintenance remain unclear.Using T-bet fate mapping mice(B6.RosaRFP;Tbx21-Cre) with serial intravascular labeling (SIVL) and BrdU incorporation, we tracked how trafficking and local proliferation coordinate lymphocyte homeostasis across gestation Methods Non-pregnant and pregnant mice at GD7-8 and GD12-14, received BrdU intraperitoneally and sequential αCD45 tail-vein injections (AF488 at 24 hours and AF647 at 5 minutes) before sacrifice. Decidual Leukocytes were isolated, stained with αCD45 (APC/Fire810) and lineage markers for B, T, NK, and classified as continuously circulating (CC), recent emigrant (RE), recent immigrant (RI) or unlabeled cells as tissue localized (TL) Results SIVL showed that at GD7-8, TL fraction of CD4+, CD8+ and NKs declined (P = 0.001, P = 0.0007, P = 0.001) with increased CC CD4+andCD8+ T cells (P = 0.01,0.02), and RI NKs (P = 0.0001), versus virgin. TL reduction was smaller for NK than CD4+ and CD8+ T cells (P = 0.008, 0.001). TL NKs showed elevated proliferation(P = 0.004, vs virgin) limited to decidua and absent in splenic NKs or decidual T cells, suggesting local expansion may sustain early TL NK pool. By GD12-14, NK proliferation sharply decreased, coinciding with further reduction of TL NK fraction (P < 0.0001,P=.004 vs GD7-8), while CD4+ and CD8+ T cells began proliferating(P = 0.006, 0.01,vs virgin), with stronger CD4+ expansion (P = 0.03) and a higher CD4+/CD8+ ratio(P < 0.0001,vs virgin). Interestingly, proliferation at this stage involved T-bet independent NK and CD8+ cells, suggesting a shift to less cytotoxic profile. However, CC, RI, TL and RE fractions within CD4+andCD8+T cells remain comparable. Conclusion Our findings reveal a gestational stage-specific coordination of trafficking and proliferation that maintains decidual immune homeostasis, marked by early NK proliferation and mid-gestation T cell expansion reflecting on CD4+/CD8+ ratio. Funding Source ECEB and R21AI175753 Topic Categories Mucosal and Regional Immunology (MUC)