Lipopolysaccharide-induced premature cervical ripening and inflammation in pregnant mice can be alleviated by nicotine

Abstract

Background Clinical treatment of preterm birth (PTB) is usually dependent on inhibiting uterine contractions. Such therapy is only effective in the short-term and cannot fundamentally low the incidence of PTB. Premature cervical maturation is another important cause of PTB; it is associated with invasion of macrophages, which produce proinflammatory cytokines. We previously found that activation of the alpha-7 nicotinic acetylcholine receptor (α7nAChR) by nicotine alleviates the systemic inflammatory response in murine models of lipopolysaccharide (LPS)-induced PTB, but the underlying mechanisms remain unclear. Methods Cervical tissues were collected from women in preterm and term labor. An animal model of PTB was established by administering 25 µg/kg lipopolysaccharide (LPS) to normal pregnant C57BL/6 mice on gestational day 16 (GD16). PTB animals received 1 mg/kg of α7nAChR agonist nicotine or nicotine combined with 1 µg/kg α7nAChR antagonist α-bungarotoxin (α-BGT) from GD14 to GD17. The gestational age, the rates of preterm birth and the survival rate of pups were recorded. Immunofluorescence staining were utilized for the quantification of α7nAChR expression on cervical macrophages and changes in macrophage polarization, RT-PCR was used to determine the mRNA levels of M1 and M2 macrophage biomarkers. Elisa assay was used to detect changes in fetal inflammation. Immunofluorescence staining and western blotting were used to investigate the signaling pathways underlying nicotine’s promoting LPS-induced conversion of M1 macrophage to M2 in the cervix of PTB animals. Results α7nAChR expression on cervical macrophages and cervical collagen content was decreased in PTB patients and murine models of LPS-induced PTB. The number of M1 cervical macrophages, other pro-inflammatory mediators in the cervix and fetal inflammation was unregulated in pregnant mice following LPS administration. Nicotine treatment reduced the rate of PTB, decreased fetal inflammation, improved maternal and neonatal outcome; nicotine also increased polarization of cervical macrophages toward the anti-inflammatory M2 phenotype possibly by suppressing activation of the MAP kinases JNK and ERK1/2 and nuclear translocation of NF-κB and rescuing the inhibited JAK2/STAT3 and PI3K/AKT pathways. The effects of nicotine were reversed by the selective α7nAChR antagonist α-bungarotoxin (α-BGT). Conclusions Our findings indicate that nicotine prevents LPS-induced cervical ripening and inflammatory response by inducing M2 macrophage polarization, and nicotine may serve as potential anti-premature cervical maturation agents for treatment of PTB.