Recurrent spontaneous abortion (RSA) brings tremendous difficulties to clinical treatment and prognosis. Pre-B-cell leukemia homeobox 1 (PBX1), as a functional transcription factor, involves in the regulation of cell apoptosis and proliferation. However, the underlying mechanism of PBX1 in RSA treatment has not been explored. We established a lipopolysaccharide (LPS)-induced abortion model and detected PBX1 expression with real-time PCR, western blot and immunohistochemistry. The PBX1-overexpressed adenovirus (AV-oePBX1) was infected into trophoblast cells treated with LPS to define the function of PBX1 on cell apoptosis, inflammation and NF-κB pathway. A luciferase reporter assay was conducted to validate the transcription regulation of PBX1 on transmembrane and ubiquitin like domain containing 1 (TMUB1). Compared to women with normal abortion, PBX1 was downregulated in the placental villous tissues of RSA patients. The placental tissues of LPS-treated mice also manifested notably reduction of PBX1 at mRNA and protein levels. PBX1 overexpression alleviated inflammation and apoptosis of trophoblast cells. Substantially, PBX1 was negatively correlated with TMUB1, which was highly expressed in RSA patients and LPS-treated mice. Moreover, PBX1 bound to TMUB1 promoter and inhibited its transcription. Interestingly, exogenous TMUB1 abolished the effects of PBX1 on apoptosis, inflammation, and NF-κB signal pathway. In total, PBX1 attenuated cell apoptosis and inflammation, and suppressed NF-κB signal pathway induced by LPS through downregulating TMUB1 transcription. Therefore, PBX1 may be developed as a novel target for clinical treatment of RSA.