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Objective: Pyroptosis is an inflammatory form of programmed cell death that is mediated by the activation of the inflammasome and depends on the pore-forming function of gasdermin D. Therefore, the detection of gasdermin D represents in vivo evidence of pyroptosis. We recently showed that there is intra-amniotic inflammasome activation in spontaneous labor at term; however, evidence of pyroptosis is lacking. The objectives of this study were to investigate (1) whether gasdermin D is detectable in the amniotic fluid of women who delivered at term; (2) whether amniotic fluid gasdermin D concentrations are associated with the process of spontaneous labor at term; and (3) whether gasdermin D is expressed in the chorioamniotic membranes from these patients.Methods: This retrospective cross-sectional study included amniotic fluid samples from 41 women who underwent spontaneous labor at term (n = 17) or delivered at term without labor (n = 24). As a readout of pyroptosis, gasdermin D was determined in amniotic fluid samples using a specific and sensitive ELISA kit. The 90th percentile of amniotic fluid gasdermin D concentrations was calculated among women without spontaneous labor at term (reference group). The association between high amniotic fluid gasdermin D concentrations (≥90th percentile in the reference group) and spontaneous labor at term was tested using the Fisher's exact test. A p value <.05 was considered significant. Multiplex immunofluorescence staining and phenoptics (multispectral imaging) were performed to determine gasdermin D expression in the chorioamniotic membranes and to colocalize this protein with the inflammasome-related molecules caspase-1 and interleukin-1β.Results: (1) Gasdermin D is present in the amniotic fluid of women who delivered at term; (2) the 90th percentile of amniotic fluid gasdermin D concentrations in women who delivered at term without spontaneous labor was 3.4 ng/mL; (3) the proportion of women with amniotic fluid gasdermin D concentrations above the threshold was higher in those who underwent term labor than in those who delivered at term without labor; (4) amniotic fluid concentrations of gasdermin D > 3.4 ng/mL were significantly associated with the presence of spontaneous labor in women who delivered at term (odds ratio 6.0, p-value .048); and (5) the protein expression of gasdermin D is increased in the chorioamniotic membranes of women who underwent spontaneous labor at term and is colocalized with caspase-1 and IL-1β.Conclusions: Gasdermin D is increased in the amniotic fluid and chorioamniotic membranes of women who underwent spontaneous labor at term compared to those without labor. These data provide evidence implicating pyroptosis in the mechanisms that lead to the sterile inflammatory process of term parturition. [ABSTRACT FROM AUTHOR]

Objective: Human β-defensin-3 (HBD-3) has a broad spectrum of antimicrobial activity, and activity and, therefore, plays a central role in host defense mechanisms against infection. Herein, we determined whether HBD-3 was a physiological constituent of amniotic fluid during midtrimester and at term and whether the concentration of this defensin was increased in amniotic fluid of women with spontaneous preterm labor and intact membranes and those with preterm prelabor rupture of membranes (pPROM) with intra-amniotic inflammation or intra-amniotic infection.Methods: Amniotic fluid was collected from 219 women in the following groups: (1) midtrimester who delivered at term (n = 35); (2) with or without spontaneous labor at term (n = 50); (3) spontaneous preterm labor with intact membranes who delivered at term (n = 29); (4) spontaneous preterm labor with intact membranes who delivered preterm with or without intra-amniotic inflammation or intra-amniotic infection (n = 69); and (5) pPROM with or without intra-amniotic infection (n = 36). Amniotic fluid HBD-3 concentrations were determined using a sensitive and specific ELISA kit.Results: (1) HBD-3 is a physiological constituent of amniotic fluid; (2) the amniotic fluid concentration of HBD-3 did not change with gestational age (midtrimester versus term not in labor); (3) amniotic fluid concentrations of HBD-3 were higher in women with spontaneous labor at term than in those without labor; (4) in the absence of intra-amniotic inflammation, amniotic fluid concentrations of HBD-3 were similar between women with spontaneous preterm labor who delivered preterm and those who delivered at term; (5) among patients with spontaneous preterm labor who delivered preterm, amniotic fluid concentrations of HBD-3 were greater in women with intra-amniotic infection than in those without this clinical condition; (6) among patients with spontaneous preterm labor, amniotic fluid concentrations of HBD-3 were higher in women with intra-amniotic inflammation or intra-amniotic infection who delivered preterm than in those without these clinical conditions who delivered at term; and (7) women with pPROM and intra-amniotic infection had higher median amniotic fluid concentrations of HBD-3 than those without this clinical condition.Conclusion: Human β-defensin-3 is a physiological constituent of amniotic fluid and increases during the process of labor at term. Amniotic fluid concentrations of HBD-3 were increased in women with spontaneous preterm labor with intact membranes or pPROM with intra-amniotic inflammation or intra-amniotic infection, indicating that this defensin participates in the host defense mechanisms in the amniotic cavity against microorganisms or danger signals. These findings provide insight into the soluble host defense mechanisms against intra-amniotic inflammation and intra-amniotic infection. [ABSTRACT FROM AUTHOR]

Objective: Upon inflammasome activation, the adaptor protein of the inflammasome ASC (apoptosis-associated speck-like protein containing a CARD) forms intracellular specks, which can be released into the extracellular space. The objectives of this study were to investigate whether (1) extracellular ASC is present in the amniotic fluid of women who delivered at term; (2) amniotic fluid ASC concentrations are greater in women who underwent spontaneous labor at term than in those who delivered at term in the absence of labor; and (3) amniotic epithelial and mesenchymal cells can form intracellular ASC specks in vitro. Methods: This retrospective cross-sectional study included amniotic fluid samples from 41 women who delivered at term in the absence of labor (n = 24) or underwent spontaneous labor at term (n = 17). Amniotic epithelial and mesenchymal cells were also isolated from the chorioamniotic membranes obtained from a separate group of women who delivered at term (n = 3), in which ASC speck formation was assessed by confocal microscopy. Monocytes from healthy individuals were used as positive controls for ASC speck formation (n = 3). Results: (1) The adaptor protein of the inflammasome ASC is detectable in the amniotic fluid of women who delivered at term; (2) amniotic fluid ASC concentration was higher in women who underwent spontaneous labor at term than in those who delivered at term without labor; and (3) amniotic epithelial and mesenchymal cells are capable of forming ASC specks and/or filaments in vitro. Conclusion: Amniotic fluid ASC concentrations are increased in women who undergo spontaneous labor at term. Amniotic epithelial and mesenchymal cells are capable of forming ASC specks, suggesting that these cells are a source of extracellular ASC in the amniotic fluid. These findings provide in vivo evidence that there is inflammasome activation in the amniotic cavity during the physiological process of labor at term. [ABSTRACT FROM AUTHOR]