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Problem: Preterm birth is commonly preceded by preterm labor, a syndrome that is causally linked to both intra‐amniotic infection and intra‐amniotic inflammation. However, the stereotypical cellular immune responses in these two clinical conditions are poorly understood. Method of study: Amniotic fluid samples (n = 26) were collected from women diagnosed with preterm labor and intra‐amniotic infection (amniotic fluid IL‐6 concentrations ≥2.6 ng/mL and culturable microorganisms, n = 10) or intra‐amniotic inflammation (amniotic fluid IL‐6 concentrations ≥2.6 ng/mL without culturable microorganisms, n = 16). Flow cytometry was performed to evaluate the phenotype and number of amniotic fluid leukocytes. Amniotic fluid concentrations of classical pro‐inflammatory cytokines, type 1 and type 2 cytokines, and T‐cell chemokines were determined using immunoassays. Results: Women with spontaneous preterm labor and intra‐amniotic infection had (a) a greater number of total leukocytes, including neutrophils and monocytes/macrophages, in amniotic fluid; (b) a higher number of total T cells and CD4+ T cells, but not CD8+ T cells or B cells, in amniotic fluid; and (c) increased amniotic fluid concentrations of IL‐6, IL‐1β, and IL‐10, compared to those with intra‐amniotic inflammation. However, no differences in amniotic fluid concentrations of T‐cell cytokines and chemokines were observed between these two clinical conditions. Conclusion: The cellular immune responses observed in women with preterm labor and intra‐amniotic infection are more severe than in those with intra‐amniotic inflammation, and neutrophils, monocytes/macrophages, and CD4+ T cells are the main immune cells responding to microorganisms that invade the amniotic cavity. These findings provide insights into the intra‐amniotic immune mechanisms underlying the human syndrome of preterm labor. [ABSTRACT FROM AUTHOR]

Problem: The immune cellular composition of amniotic fluid is poorly understood. Herein, we determined: 1) the immunophenotype of amniotic fluid immune cells during the second and third trimester in the absence of intra‐amniotic infection/inflammation; 2) whether amniotic fluid T cells and ILCs display different phenotypical characteristics to that of peripheral cells; and 3) whether the amniotic fluid immune cells are altered in women with intra‐amniotic infection/inflammation. Method of Study: Amniotic fluid samples (n = 57) were collected from 15 to 40 weeks of gestation in women without intra‐amniotic infection/inflammation. Samples from women with intra‐amniotic infection/inflammation were also included (n = 9). Peripheral blood mononuclear cells from healthy adults were used as controls (n = 3). Immunophenotyping was performed using flow cytometry. Results: In the absence of intra‐amniotic infection/inflammation, the amniotic fluid contained several immune cell populations between 15 and 40 weeks. Among these immune cells: (i) T cells and ILCs were greater than B cells and natural killer (NK) cells between 15 and 30 weeks; (ii) T cells were most abundant between 15 and 30 weeks; (iii) ILCs were most abundant between 15 and 20 weeks; (iv) B cells were scarce between 15 and 20 weeks; yet, they increased and were constant after 20 weeks; (v) NK cells were greater between 15 and 30 weeks than at term; (vi) ILCs expressed high levels of RORγt, CD161, and CD103 (ie, group 3 ILCs); (vii) T cells expressed high levels of RORγt; (viii) neutrophils increased as gestation progressed; and (ix) monocytes/macrophages emerged after 20 weeks and remained constant until term. All of the amniotic fluid immune cells, except ILCs, were increased in the presence of intra‐amniotic infection/inflammation. Conclusion: The amniotic fluid harbors a diverse immune cellular composition during normal and complicated pregnancies. [ABSTRACT FROM AUTHOR]

Problem: Pyroptosis, inflammatory programmed cell death, is initiated through the inflammasome and relies on the pore‐forming actions of the effector molecule gasdermin D. Herein, we investigated whether gasdermin D is detectable in women with spontaneous preterm labor and sterile intra‐amniotic inflammation or intra‐amniotic infection. Method of study: Amniotic fluid samples (n = 124) from women with spontaneous preterm labor were subdivided into the following groups: (a) those who delivered at term (n = 32); and those who delivered preterm (b) without intra‐amniotic inflammation (n = 41), (c) with sterile intra‐amniotic inflammation (n = 32), or (d) with intra‐amniotic infection (n = 19), based on amniotic fluid IL‐6 concentrations and the microbiological status of amniotic fluid (culture and PCR/ESI‐MS). Gasdermin D concentrations were measured using an ELISA kit. Multiplex immunofluorescence staining was also performed to determine the expression of gasdermin D, caspase‐1, and interleukin‐1β in the chorioamniotic membranes. Flow cytometry was used to detect pyroptosis (active caspase‐1) in decidual cells from women with preterm labor and birth. Results: (a) Gasdermin D was detected in the amniotic fluid and chorioamniotic membranes from women who underwent spontaneous preterm labor/birth with either sterile intra‐amniotic inflammation or intra‐amniotic infection, but was rarely detected in those without intra‐amniotic inflammation. (b) Amniotic fluid concentrations of gasdermin D were higher in women with intra‐amniotic infection than in those with sterile intra‐amniotic inflammation, and its expression in the chorioamniotic membranes was associated with caspase‐1 and IL‐1β (inflammasome mediators). (c) Decidual stromal cells and leukocytes isolated from women with preterm labor and birth are capable of undergoing pyroptosis given their expression of active caspase‐1. Conclusion: Pyroptosis can occur in the context of sterile intra‐amniotic inflammation and intra‐amniotic infection in patients with spontaneous preterm labor and birth [ABSTRACT FROM AUTHOR]

The cover image is based on the Original Article The immunophenotype of decidual macrophages in acute atherosis by Navleen Gill et al., DOI 10.1111/aji.13098. [ABSTRACT FROM AUTHOR]

Problem: Acute atherosis is a uteroplacental arterial lesion that is associated with pregnancy complications such as preeclampsia and preterm birth, the latter being the leading cause of perinatal morbidity and mortality worldwide. However, the immunobiology of acute atherosis is poorly understood. Method of study: Placental basal plate samples were collected from women who delivered with (n = 11) and without (n = 31) decidua basalis lesions of acute atherosis. Multicolor flow cytometry was used to quantify M1‐ and M2‐like macrophage subsets and the expression of iNOS and IL‐12 by decidual macrophages. Multiplex fluorescence staining and phenoptics were performed to localize M1‐, MOX‐, and Mhem‐like macrophages in the decidual basalis. Results: Macrophages displayed diverse phenotypes in the decidua basalis with acute atherosis. M2‐like macrophages were the most abundant subset in the decidua; yet, this macrophage subset did not change with the presence of acute atherosis. Decidual M1‐like macrophages were increased in acute atherosis, and such macrophages displayed a pro‐inflammatory phenotype, as indicated by the expression of iNOS and IL‐12. Decidual M1‐like pro‐inflammatory macrophages were localized near both transformed and non‐transformed vessels in the decidua basalis with acute atherosis. MOX and Mhem macrophages were also identified near transformed vessels in the decidua basalis with acute atherosis. Finally, monocyte‐like cells were present on the vessel wall of non‐transformed decidual vessels, indicating a possible intravascular source for macrophages in acute atherosis. Conclusion: Decidual macrophages display different phenotypes, namely M1‐like, M2‐like, MOX, and Mhem subsets. Yet, pro‐inflammatory macrophages are enriched in the decidua basalis with acute atherosis. These findings provide a molecular foundation for future mechanistic inquiries about the role of pro‐inflammatory macrophages in the pathogenesis of acute atherosis. The immunophenotype of decidual macrophages in acute atherosis. Representative images of histological staining of decidual vessels using (A) hematoxylin and eosin or (B) oil‐red O. C, Representative image of immunohistochemical staining of CD68 to detect macrophages and foam cells. Black arrows indicate foamy macrophages, and red arrows show non‐foamy macrophages. Magnification = 200×. Scale bars = 50 μ. Flow cytometric analysis of decidual macrophage subsets was performed using t‐distributed stochastic neighbor embedding (t‐SNE) shown as (D) dot plots or (E) contour plots. Red = M1‐like macrophages (CD45+CD14+ICAM‐3−CD80+ cells), blue = M2‐like macrophages (CD45+CD14+ICAM‐3−CD209+CD163+ cells), and gray = non‐M1‐/M2‐like macrophages. [ABSTRACT FROM AUTHOR]

Problem: The inflammasome is implicated in the mechanisms that lead to spontaneous preterm labor (PTL). However, whether there is inflammasome activation in the amniotic cavity of women with PTL and intra‐amniotic infection (IAI) or sterile intra‐amniotic inflammation (SIAI) is unknown. Method of study: Amniotic fluid samples were collected from women with PTL who delivered at term (n = 31) or preterm without IAI or SIAI (n = 35), with SIAI (n = 27), or with IAI (n = 17). As a readout of inflammasome activation, extracellular ASC (apoptosis‐associated speck‐like protein containing a CARD) was measured in amniotic fluid by ELISA and the expression of ASC, caspase‐1, and interleukin (IL)‐1β was detected in the chorioamniotic membranes by multiplex immunofluorescence. Acute inflammatory responses in amniotic fluid and the placenta were also evaluated. Results: (a) Amniotic fluid concentrations of ASC and IL‐6 were higher in women with PTL and IAI or SIAI than in those who delivered preterm or at term without intra‐amniotic inflammation; (b) amniotic fluid concentrations of ASC and IL‐6 were lower in women with PTL and SIAI than in those with IAI; (c) there was a significant nonlinear correlation between ASC and IL‐6 amniotic fluid concentrations; (d) the expression of inflammasome‐related proteins (ASC, caspase‐1, and IL‐1β) in the chorioamniotic membranes was increased in women with PTL and IAI or SIAI than in those who delivered preterm or at term without intra‐amniotic inflammation; (e) inflammasome activation in the chorioamniotic membranes was weaker in women with PTL and SIAI than in those with IAI; (f) women with PTL and IAI had elevated amniotic fluid white blood cell counts compared to those without this clinical condition; and (g) severe acute placental inflammatory lesions were observed in women with PTL and IAI and in a subset of women with PTL and SIAI. Conclusion: Inflammasome activation occurs in the settings of intra‐amniotic infection and sterile intra‐amniotic inflammation during spontaneous preterm labor. [ABSTRACT FROM AUTHOR]

Problem: Human β‐defensins (HBDs) are antimicrobial peptides that participate in the soluble innate immune mechanisms against infection. Herein, we determined whether HBD‐1 was present in amniotic fluid during normal pregnancy and whether its concentrations change with intra‐amniotic inflammation and/or infection. Method of Study: Amniotic fluid was collected from 219 women in the following groups: (a) midtrimester who delivered at term (n = 35); (b) term with (n = 33) or without (n = 17) labor; (c) preterm labor with intact membranes who delivered at term (n = 29) or who delivered preterm with (n = 19) and without (n = 29) intra‐amniotic inflammation and infection or with intra‐amniotic inflammation but without infection (n = 21); and (d) preterm prelabor rupture of membranes (pPROM) with (n = 19) and without (n = 17) intra‐amniotic inflammation/infection. Amniotic fluid HBD‐1 concentrations were determined using a sensitive and specific ELISA kit. Results: (a) HBD‐1 was detectable in all amniotic fluid samples; (b) amniotic fluid concentrations of HBD‐1 were changed with gestational age (midtrimester vs term no labor), being higher in midtrimester; (c) amniotic fluid concentrations of HBD‐1 were similar between women with and without spontaneous labor at term; (d) among patients with spontaneous preterm labor, amniotic fluid concentrations of HBD‐1 in women with intra‐amniotic inflammation/infection and in those with intra‐amniotic inflammation without infection were greater than in women without intra‐amniotic inflammation or infection who delivered preterm or at term; and (e) the presence of intra‐amniotic inflammation and infection in patients with pPROM did not change amniotic fluid concentrations of HBD‐1. Conclusion: HBD‐1 is a physiological constituent of amniotic fluid that is increased in midtrimester during normal pregnancy and in the presence of culturable microorganisms in the amniotic cavity. These findings provide insight into the soluble host defense mechanisms against intra‐amniotic infection. Amniotic fluid concentrations of human beta defensin‐1 (HBD‐1) in women with spontaneous preterm labor and intact membranes. Red lines indicate medians with interquartile ranges. [ABSTRACT FROM AUTHOR]