Progesterone promotes favorable pregnancy outcomes in recurrent spontaneous, abortion by attenuating NK Cell overactivation and upregulating the cAMP/PKA/CREB signaling axis

Objective Recurrent spontaneous abortion (RSA) is a common pregnancy complication with complex etiology, in which immune dysregulation is critical. Although progesterone (P4) is widely used clinically for RSA, its mechanism in regulating NK cell function remains unclear. Methods We analyzed decidual tissues from patients with RSA and from healthy gestational controls, and established a rat model of RSA treated with graded P4. Histopathological analysis, RT-qPCR, Western blotting, RNA sequencing (RNA-Seq), and flow cytometry were employed to evaluate cytokine profiles, the proportion of CD3 − CD161 + NK cells, candidate gene expression, and activation of the cAMP/PKA/CREB signaling pathway. Results In patients with RSA and in the RSA rat model, IFN-γ and TNF-α levels were elevated, whereas VEGF levels were decreased—accompanied by an increased proportion of CD3⁻CD161⁺ NK cells. P4 treatment significantly reduced the embryo loss rate, improved the morphological structure of the decidua tissue, and effectively inhibited the expression of inflammatory cytokines and the excessive activation of CD3⁻CD161⁺NK cells. Transcriptomic analysis identified the cAMP signaling pathway as a key dysregulated pathway; cAMP/PKA/CREB signaling was suppressed in RSA but restored by P4. Mx1 was upregulated in RSA and reversed by P4, showing negative correlations with PKA/CREB activation and positive correlations with pro-inflammatory cytokines. Conclusion P4 improves the pregnancy outcome of RSA by inhibiting the excessive activation of CD3⁻CD161⁺NK cells and activating the cAMP/PKA/CREB signaling pathway; Mx1 may play a negative regulatory role in this pathway.