Infection is the only proven mechanism of disease leading to premature delivery. Microbiological and histopathological studies suggest that infection-related inflammation may account for 25% to 40% of cases of preterm deliveries.201,202,203
The evidence supporting the role of infection in the onset of labor includes the following: (1) systemic maternal infection is associated with the onset of labor and these patients have a higher frequency of PTB; (2) intrauterine infection is associated with spontaneous preterm parturition; and (3) in animal models, intrauterine injection of bacteria or bacterial products leads to preterm delivery.201,202,203
Infectious processes leading to PTB can be clinically evident (ie, pyelonephritis or chorioamnionitis); subclinical, limited to the uterus,
and present as PTL or preterm PROM, resulting from bacteria that crossed through the placenta from the maternal blood and are detected in the amniotic fluid; and/or detected only after delivery in the placenta (ie, acute histologic chorioamnionitis) (Figure 49.8
The presence of bacteria or other microorganisms in the amniotic fluid, which should be sterile,213
is associated with adverse pregnancy outcomes.164,171,172,173,174,175,176,214,215,216,217,218,219,220,221,222,223,224,225
Intrauterine infection is a frequent and important mechanism of disease leading to premature contraction, PTL, and PTB.166,201,203,226,227,228,229,230,231,232,233
Intra-amniotic infection is a chronic process,234
and identification of microorganisms during mid-trimester amniocentesis in asymptomatic patients has been associated with subsequent late miscarriage,235
and even fetal demise.171,172,173,174,175,176,237
Pathogenic microorganisms can invade the amniotic fluid through several routes. The most common is ascending infection from the vagina to the cervical canal through the membranes into the amniotic cavity and subsequently infecting the
membranes and the fetus.201,238
Other routes are maternal transmission of bacteria by hematogenous239,240,241
spread through the placenta or through the membranes, and finally iatrogenic introduction of bacteria into the amniotic cavity during amniocentesis of other medical intervention242,243
Figure 49.9 Potential route of microbial invasion of the amniotic cavity. The most common is ascending infection from the vagina to the cervical canal, through the membranes into the amniotic cavity and subsequently infecting the membranes and the fetus. Other routes are maternal transmission of bacteria by hematogenous spread through the placenta or through the membranes.
(Reprinted with permission from Goldenberg RL, Culhane JF, Iams JD, Romero R. Epidemiology and causes of preterm birth. Lancet. 2008;371(9606):75-84.)
Goncalves et al226
reported that the overall prevalence of intra-amniotic infection in patients with PTL was 12.8% and about 50% were polymicrobial. The rate of microbial invasion of amniotic cavity (MIAC) in patients with PTL and intact membrane is gestational age dependent, ranging from as high as 45% at 23 to 26 weeks to 11.5% by 31 to 34 weeks of gestation.244
Thus, the earlier the gestational age in which PTL develops, the more likely that MIAC is present.244
The diagnosis of MIAC using standardized cultivation techniques is limited by the number of bacteria detected and by the interval from sample collection to bacterial growth and identification in cultures. The introduction of 16s ribosomal DNA-polymerase chain reaction (PCR) techniques has improved the detection rate of MIAC.238,245,246,247,248,249
Indeed, in a study of amniotic fluid samples from 166 patients with PTL and intact membranes, the overall rate of positive cultures was 15.06% (25/166), of them 24% (6/25) had only positive amniotic fluid cultures, 36% (9/25) had only positive amniotic fluid 16s rDNA PCR, and in 40% (10/25) both tests were positive.246
A study by Combs et al reported that patients with PTL and intact membranes had a rate of MIAC of 10.1% (31/305), of them 65% (20/31) had both tests positive, 19% (6/31) had only 16s rDNA-PCR positive, and 16% (5/31) only cultures positive.245
These observations have clinical implications since in women presenting with PTL, a positive 16s rDNA-PCR test for microbial invasion had a positive predictive value of 100% for preterm delivery <37, <32, or <25 weeks of gestation and 68% for delivery within 1 day of amniocentesis.246
Recently, a novel molecular microbiologic technique that includes 16s PCR and electrospray ionization time-of-flight mass spectrometry (PCR/ESI-MS).250
This method can yield results within 8 hours of amniotic fluid collection and therefore in time for clinical decision. The authors reported that the introduction of this method increased the detection rate of microbial of amniotic fluid by 100% and increased the positive predictive value for spontaneous preterm delivery before 37 or 32 weeks, delivery within 7 or 2 days of amniocentesis.250
Collectively these studies suggest that there is a role for the introduction of novel molecular microbiologic techniques in the clinical workup of amniotic fluid for the detection of intra-amniotic infection in patients with PTL and intact membranes.
The most common microbial organisms isolated (either by culture or by 16s PCR) from the amniotic fluid of patients with PTL and intact membranes were genital mycoplasmas; followed by Fusobacterium
species,201 Streptococcus agalactiae, Peptostreptococcus
spp., Staphylococcus aureus, Gardnerella vaginalis, Streptococcus viridans
, and Bacteroides
Bacteria in amniotic fluid can be either planktonic (free floating) or in the form of biofilm.260,261
The latter is more challenging since bacteria use biofilms to avoid the host inflammatory response and are more difficult to isolate in standard cultivation
techniques, and such infections are often underdiagnosed. Amniotic fluid sludge, which is a particulate matter identified by ultrasound near the internal os of the uterine cervix, is in fact in part of the cases a bacterial biofilm260,261
). The identification of sludge in asymptomatic patients and in those at risk for PTB, especially when accompanied by a short sonographic cervix, is associated with an increased risk for spontaneous PTL with intact membranes, PPROM of membranes, microbial invasion of amniotic fluid, PTB, and histologic chorioamnionitis.262,263
Figure 49.10 Most frequent microorganisms identified in amniotic cavity and placenta in term and preterm deliveries. PTB, preterm birth; sp., species.
(Reprinted from Payne MS, Bayatibojakhi S. Exploring preterm birth as a polymicrobial disease: an overview of the uterine microbiome. Front Immunol. 2014;5:595.)
Inflammation has a seminal role in the process of term and preterm parturition. However, while in labor at term, inflammation is a physiologic phenomenon. In the context of PTL, intra-amniotic inflammation that initiates preterm parturition is derived from pathological processes such as intra-amniotic infection or sterile inflammation.5,202,264,265,266,267,268
Figure 49.11 Transvaginal detection of amniotic fluid sludge in a patient at 28 weeks of gestation, with bulging membranes preterm labor and chorioamnionitis.
(Reprinted with permission from Costerton JW, Hassan SS, Schaudinn C, et al. What is amniotic fluid ‘sludge’? Ultrasound Obstet Gynecol. 2007;30(5):793-798.)
Microbial invasion of amniotic fluid can be either isolated (colonization) or as microbial associated intra-amniotic inflammation. This distinction has clinical significance, in a nested case control study of asymptomatic women who underwent mid-trimester genetic amniocentesis at 16 to 18 weeks of gestation269
: (1) Women who delivered preterm had higher amniotic fluid concentrations of interleukin (IL)-6 and tumor necrosis factor (TNF) than those who delivered at term; (2) mid-trimester amniotic fluid IL-6 concentration ≥ 99.3 pg/mL had a sensitivity of 89.6%, specificity of 80.2%, and RR of 11.4 (95% CI 4.8-27.0) for preterm delivery <37 weeks of gestation; (3) mid-trimester amniotic fluid IL-6 concentration ≥ 99.3 pg/mL had a sensitivity of 91.9%, specificity of 73.8%, and RR of 15.0 (95% CI 4.8-45.5) for a positive amniotic fluid culture; (4) mid-trimester amniotic fluid TNF concentrations ≥ 6.6 pg/mL had a sensitivity of 81.3%, specificity of 79.2%, and RR of 6.2 (95% CI 3.3-11.9) for preterm delivery <37 weeks of gestation; and (5) mid-trimester amniotic fluid TNF concentrations ≥ 6.3 pg/mL had a sensitivity of 78.4%, specificity of 70.1%, and RR of 4.9 (95% CI 2.4-10.0) for positive amniotic fluid culture. Of interest, among those who delivered preterm, 47.9% had a positive mid-trimester amniotic fluid culture and elevated IL-6 and 45.8% had a positive mid-trimester amniotic fluid culture and elevated TNF, while among those who delivered preterm, the corresponding rates were 11.5% and 7.3%, respectively. Of interest, the rate of PTBs with those who had a positive amniotic fluid culture with elevated IL or TNF were 4.2% and 6.3%, respectively. Nevertheless, negative amniotic fluid cultures with elevated IL-6 or TNF were present in 41.7% and 35.4% of preterm deliveries.269
This report and others164,201,202,203,215,225,245,270,271,272,273
suggest that the inflammatory process elicited by the presence of microbacteria in the amniotic cavity is the driving force that leads to preterm parturition in cases of MIAC.
In cases of microbial associated intra-amniotic inflammation, the inflammatory reaction in the amniotic fluid is triggered by the activation of pattern recognition receptors including toll-like receptors or RAGE in response to the presence of danger-associated molecular patterns (DAMPs) or pathogen-associated molecular patterns (ie, lipopolysaccharides).274
The activation of pattern recognition receptors (PPRs) by microbes or their products illicits an inflammatory response, which in turn leads to the production of cytokines (IL-6 and TNFα) and matrix degrading enzymes (matrix metalloproteinase [MMP]-8), leading to the production of prostaglandins and activation of the common pathway of parturition6
Sterile Intra-amniotic Inflammation
Inflammatory processes in the amniotic fluid in which microorganisms cannot be detected are defined as sterile intra-amniotic inflammation. This process is activated by the presence of DAMPs in the amniotic fluid. These are endogenous intracellular molecules that are exposed during unprogrammed cellular death and can serve as a preliminary sign of tissue injury. DAMPs are also referred to as alarmins, and prominent members of this family are high-mobility group box 1 (HMGB1), uric acid, IL-1α, S100 calcium binding protein B, uric acid, and cell-free DNA.6,222,224,264,265,267,275,276,277,278,279,280
The generation of DAMPs activates the PPR that illicits an inflammatory response, which in turn leads to the production of cytokines (IL-6 and TNFα) and matrix degrading enzymes (MMP-8), leading to the production of prostaglandins and activation of the common pathway of parturition (Figure 49.12
Yoon et al281
reported that for any given gestational age, the rate of intra-amniotic inflammation is higher than that of MIAC (Figure 49.13
). The clinical characteristics of patients with PTL who had sterile intra-amniotic inflammation are similar in terms of gestational age at delivery, amniocentesis to delivery interval, and adverse neonatal outcomes, to those reported in women with PTL and microbial associated intra-amniotic inflammation.221,245,281,282
Indeed, among patients with PTL, the amniotic fluid concentration of HMGB1 proteins is increased in both patients with sterile or microbial mediated intra-amniotic inflammation.283
Moreover, the amniocentesis to delivery interval of patients with PTL and sterile amniotic fluid inflammation who had amniotic fluid HMGB1 concentration ≥8.55 pg/mL was similar to that of those with microbial associated intra-amniotic inflammation, and shorter than those with sterile intra-amniotic inflammation but HMGB1 concentration <8.55 pg/mL (P
In a mice model, intra-amniotic administration of HMGB-1 leads to PTB.283 In vitro
evidence suggests a role for HMGB1 in activation of the chorioamniotic
membranes, leading to increased production of IL-6 and IL-8, and its extranuclear fraction is detected in the uterine cervix during term and PTL, suggesting a role in cervical ripening.280
Moreover, intra-amniotic infection is associated with higher amniotic fluid concentrations of HMGB-1. Other alarmins, such as IL-1α and IL-1β, are implicated in spontaneous PTB as well.
Figure 49.12 A, Cellular stress will affect cell viability and lead to the release of damage-associated molecular patterns (DAMPs) (alarmins) by the fetal membranes and the placenta. These DAMPs will then act on the placenta, uterus, cervix, and fetal membranes, inducing inflammation. B, Inflammatory mechanism of action of alarmins at the maternal-fetal interface. Following stimulation, alarmins are released from cells at the maternal-fetal interface and act on placental cells (primarily trophoblasts) and maternal myeloid cells to induce an inflammatory response and secretion of inflammatory mediators (cytokines/chemokines), which will recruit maternal immune cells. CCL2, chemokine (C-C motif) ligand 2; cffDNA, cell free fetal DNA; HMGB-1, high mobility group box 1; IL, interleukin; MSU, monosodium urate; mtDNA, mitochondrial DNA; NALP3, NLR family pyrin domain containing 3; NFκB, nuclear factor kappa-light-chain-enhancer of activated B cells; RAGE, receptor for advanced glycation endproducts; TLR, toll-like receptor.
(A and B, Reprinted with permission from Nadeau-Vallee M, Obari D, Palacios J, et al. Sterile inflammation and pregnancy complications: a review. Reproduction. 2016;152(6):R277-R292.)
Figure 49.13 The frequency of intra-amniotic inflammation and intra-amniotic infection in patients with preterm labor according to gestational age at amniocentesis. AF, amniotic fluid.
(Reprinted with permission from Yoon BH, Romero R, Moon JB, et al. Clinical significance of intra-amniotic inflammation in patients with preterm labor and intact membranes. Am J Obstet Gynecol. 2001;185(5):1130-1136.)
In addition to IL-1 and TNFα, many other cytokines and chemokines (ie, IL-6, IL-8, IL-16, IL-18, colony-stimulating factor, macrophage migration inhibitory factor, monocyte chemotactic protein-1) are involved in the inflammatory process of microbial associated or sterile intra-amniotic inflammation.274
These observations suggest that the process of intra-amniotic inflammation involves a network of cytokines and chemokines. Indeed, Romero et al demonstrated that in patients with PTL and intact membranes, microbial associated or sterile intra-amniotic inflammation are characterized by unique network of perturbed inflammatory-related protein concentration and correlations284
Maternal Antifetal Rejection
A novel inflammatory process leading to PTB is maternal antifetal rejection, a graft-versus-host response of the fetus and the mother. This process is associated with chronic placental inflammation, characterized by villitis of unknown etiology, chronic chorioamnionitis, and chronic deciduitis and is similar to the type of graft rejection observed in patients with failed heart, lung, and kidney transplant.285
A high amniotic fluid concentration of C-X-C motif ligand (CXCL)-10 observed in patients with spontaneous preterm labor was associated with the presence of lesion consistent with chronic placental inflammation (villitis of unknown etiology, chronic chorioamnionitis, and chronic deciduitis).285
This type of placental lesion is characterized by infiltration of lymphocytes, plasma cells, and histiocytes. The maternal antifetal rejection is characterized by (1) infiltration of maternal CD8+ T cells into fetal tissues; (2) the presence of antifetal human leukocyte antigen antibodies in maternal circulation; and (3) depositions of C4d in umbilical vein and syncytiotrophoblast194
). Chronic placental inflammatory lesions are the most common placental findings in late PTB but can be observed in the mid-trimester of patients who eventually have late-preterm delivery.194
Maternal antifetal rejection is associated with fetal morbidity and mortality. Among preterm neonates born between 24 and 34 weeks of gestation, those who had periventricular leukomalacia (PVL) had a higher rate of placental chronic inflammatory lesions than those without it,286
suggesting that processes associated with maternal antifetal rejection leading to PTL and PTB may be associated with long-term sequelae in the preterm infant.
Fetal Inflammatory Response Syndrome
Fetal inflammatory response syndrome (FIRS) can be considered the fetal equivalent of systemic inflammatory response syndrome, described in adults with sepsis.287,288
FIRS is characterized by systemic activation of the fetal immune system involving all major fetal systems (ie, hematopoietic, heart, brain, lungs, kidneys, adrenals, and skin).287,288,289
Additionally, preterm neonates affected in utero by FIRS have a shorter interval from cordocentesis to delivery and a higher rate of short- and long-term complication of prematurity including RDS, neonatal sepsis, pneumonia, bronchopulmonary dysplasia (BPD), intraventricular hemorrhage (IVH), PVL, NEC, and cerebral palsy than those without this syndrome.287,290,291,292
The rate of FIRS in pregnancies complicated by preterm parturition is about 39% and increases to 49.3% in fetuses delivered within 1 week from cordocentesis.287,293
Currently, there are two types of FIRS: type I and type II. Type I is considered the highest degree of intra-amniotic infection/inflammation. It describes fetuses who mounted a systemic inflammatory immune response to microorganisms that invaded the amniotic cavity. Type I FIRS is characterized by high cord blood concentrations of proinflammatory
cytokines, especially IL-6 and TNF-α. A concentration of 11 pg/mL of IL-6 detected in fetal cord blood is the cutoff for prenatal diagnosis of FIRS type I281
as it was associated with adverse neonatal outcome and complications of prematurity. The characteristic placental lesions of FIRS type I syndrome include histologic chorioamnionitis, evidence of umbilical cord inflammation (funisitis), and chorionic vasculitis. The presence of funisitis allows a postnatal diagnosis of neonates with FIRS type I and is associated with an increased risk for the subsequent development of cerebral palsy (OR 5.5; 95% CI 1.2-24.5)287,294,295
Figure 49.14 Perturbation of the cytokine network in women with preterm labor. Each node (sphere) represents 1 of the 33 analytes; its color represents the direction of concentration change between the groups (red, increased; blue, decreased; white, no change). The link (line) between two nodes represents a significantly perturbed correlation, and its color gives the direction of correlation change (red, increased; blue, decreased). The type of line denotes the nature of the link (solid line, within module link; dashed line, cross-module link). Thick radial lines separate the modules and the set of unconnected nodes. The numbers inside/outside the dotted black circles represent the node degree/average absolute difference in correlations. A, Network of perturbed inflammatory-related protein concentration correlations between sterile intra-amniotic inflammation and no intra-amniotic inflammation. B, Network of perturbed inflammatory-related protein concentration correlations between microbial associated intra-amniotic inflammation and no intra-amniotic inflammation. C, Network of perturbed inflammatory-related protein concentration correlations between microbial associated intra-amniotic inflammation and sterile intra-amniotic inflammation. CXCL, C-X-C motif chemokine ligand; GMCSF, granulocyte macrophage-colony stimulating factor; Groa, melanoma growth-stimulatory activity; HMGB-1, high mobility group box 1; IFN, interferon; IL, interleukin; IP, interferon gamma-induced protein; MCP1, monocyte chemoattractant protein 1; MI, macrophage inflammatory proteins; RANTES, regulated on activation, normal T cell expressed and secreted; TGF, transforming growth factor; TNF, tumor necrosis factor; xMCSF, macrophage colony-stimulating factor.
(Reprinted with permission from Romero R, Grivel JC, Tarca AL, et al. Evidence of perturbations of the cytokine network in preterm labor. Am J Obstet Gynecol. 2015;213(6):836.e1-836.e18.)
Figure 49.15 Maternal antifetal rejection is characterized by (A) infiltration of maternal CD8+ T cells into fetal tissues, and (B) the presence of antifetal human leukocyte antigen (HLA) antibodies in maternal circulation that transfer to the fetus.
(Reprinted with permission from Kim CJ, Romero R, Chaemsaithong P, Kim JS. Chronic inflammation of the placenta: definition, classification, pathogenesis, and clinical significance. Am J Obstet Gynecol. 2015;213(4 suppl):S53-S69.)
Type II FIRS results from maternal antifetal rejection and manifests as a unique cord blood transcriptome in the affected fetuses. The clinical manifestations of FIRS type II await further research.296
The summary of the characteristics of the two types of FIRS is depicted in Figure 49.16
Preliminary evidence suggest an allergic-like immune response (type I hypersensitivity) can be associated with PTL and delivery.5,297
Indeed, case reports of women who had severe latex allergy298
and developed uterine contraction as well as that of women who developed PTL and acute hypersensitivity reaction after ingestion of lobster meat that resolved with treatment by steroids antihistaminic medications297
support this assumption. Type I hypersensitivity is defined as immunoglobulin (Ig)E-mediated hypersensitivity, in which the exposure of these antibodies to the allergens leads to mast cell activation, degranulation, and initiation of an inflammatory response and the section of histamine and prostaglandins and other products of mast cell degranulation can induce myometrial contractility.299,300,301
The mast cell-mediated inflammatory response is characterized by the secretion
of IL-8, IL-3, and granulocyte-macrophage colony-stimulating factor (GM-CSF), which attract and activate eosinophils that enhance the inflammatory response by secreting leukotrienes, platelet activating factors, IL-4, and IL-10 as well as activation of proteases leading to tissue damage.297
Indeed, among women who presented with PTL and had diagnostic amniocentesis without evidence of MIAC, those who had >20% eosinophils in their amniotic fluid blood cell count had a lower gestational age at delivery and a higher rate of PTB <35 and <37 weeks of gestation than those who had ≤20% eosinophils in their amniotic fluid blood cell count. This suggests that a subset of patients with PTL and a high number of eosinophils in their amniotic fluid has type I hypersensitivity-mediated PTB.
Figure 49.16 The characteristics of the fetal inflammatory response syndrome (FIRS) type I and type II. Type I results from ascending inflammation, and the feature cytokine found in fetal cord blood is IL-6. FIRS type II is maternal antifetal rejection, and the characteristic chemokine found in cord blood is CXCL-10 (C-X-C motif chemokine ligand 10). ACA, acute chorioamnionitis; CCA, chronic chorioamnionitis; CDP, chronic deciduitis; VUE, villitis of unknown etiology.
(Reprinted with permission from Kim CJ, Romero R, Chaemsaithong P, Kim JS. Chronic inflammation of the placenta: definition, classification, pathogenesis, and clinical significance. Am J Obstet Gynecol. 2015;213(4 suppl):S53-S69.)