In this study, we have found that CAM, chronic villitis, immature placental villi, SGA, and maternal overweight/obesity are independently associated with risk for stillbirth at term.
We found CAM in 65.1% of stillborn cases and 40.7% in liveborn controls. Other studies have shown prevalence of CAM in stillbirths ranging between 57% to 96% [14
] and in term liveborns from 5% to 68% [13
]. The reason for these discrepancies between different studies is not entirely clear. The fact that the majority of studies claiming high proportions of CAM were from developing countries may suggest some kind of underlying geographical/environmental factor. Other explanations may include differences in the definition of stillbirth and histological CAM, interobserver variability between reporting pathologists, small sample sizes, or selection of material. The high frequency of CAM in our control group might be additionally related to the selection criterion being liveborn, and not uncomplicated (“healthy”) pregnancy, in contrast to several other reports. Our study is one of the largest studies in the literature and provides a comprehensive view of the stillbirth population in the major Stockholm area.
A high frequency of CAM, vasculitis and/or funisitis could be due to the duration of established contractions or the time interval between broken membranes and delivery. This information was not available for all in the study group. A subanalysis of half of the vaginally delivered cases and controls did not show any difference concerning time between rupture of membranes and delivery but there was a significant difference regarding time from established contraction to delivery, with longer labour in the control group (P < 0.05, data not presented).
Although CAM is generally believed to represent an ascending microbial infection [14
], our results of blood culture from the heart of the neonate failed to show a bacterial etiology of this placental inflammation. One reason for this might be that blood samples were only cultured for aerobic and anaerobic bacteria. The presence of other microorganisms, such as ureaplasma or mycoplasma, which have been related to stillbirth in other studies [1
], may have eluded detection. Several studies have questioned the efficacy of culture alone for detecting bacteria [21
] suggesting the need for supplemental molecular techniques (i.e., PCR).
The high risk of stillbirth correlated to severe CAM point and to the importance of clarifying the background of CAM since it is obviously a threat to the fetus in term pregnancy.
Vasculitis and funisitis are histological markers of the fetal inflammatory response syndrome and are risk markers for neurological impairment [4
] and decreased psychomotor development [6
]. However, the relation between fetal inflammatory response syndrome and stillbirth is less well understood. In a recent work, Gordon et al. [25
] found that absence of a fetal inflammatory response was strongly associated with unexplained antepartum death. Our results are consistent with these data. Although vasculitis and/or funisitis were significantly more common in the placentas from stillborn than from liveborn neonates in our study, the risk of stillbirth was not elevated in the multivariate logistic regression analysis. Further studies are needed to elucidate whether fetal inflammatory response, exemplified by the presence of inflammation in fetal vessels and umbilical cord in the placenta, as well as cytokine release, is unrelated to the mechanisms of stillbirth or even confers some kind of protective action.
Chronic villitis has received less attention than CAM, especially in stillbirth. Villitis affects 5–15% of all third trimester placentas [7
] and has been associated with adverse neurologic outcomes, IUGR, stillbirth [4
], premature delivery, and high maternal BMI [7
Syridou et al. recently showed [5
] that the presence of chronic villitis in stillbirth placentas was associated with increased detection of viral DNA, especially in advanced gestational ages, implicating a link between viral infection and the pathogenesis of stillbirth. However, in the majority of cases, the etiology of chronic villitis remains unknown, often referred to as villitis of unknown etiology (VUE, [7
]). Myerson et al. [26
] demonstrated that the majority of lymphocytes in VUE are maternal T cells, and the process is associated to significant destruction of syncytiotrophoblast; this may contribute to the breakdown of the local placental barrier and the graft-versus-host-like invasion of maternal cells into the villi. In our study, villitis, independent of the extent, was clearly overrepresented in stillbirth placentas (18.3% compared to 5.1% in controls) and villitis (<1% or ≥1%) had approximately a four-fold risk for stillbirth in term pregnancies. Our data do not provide any clues regarding the underlying mechanism, although it appears to not be a direct effect, that is, not related to extent of reduction of placental parenchyma. Rather, it might involve some subtle immunologic imbalance with possible secretion of regulatory cytokines or other factors in parallel with fetal inflammatory response.
Villous immaturity, otherwise referred to as villous maturation defect, retarded villous maturation or terminal villous insufficiency, mainly involving a defect in villous vascularisation, is an often underrecognized entity [27
]. The histopathological diagnosis is generally considered difficult, and a high rate of interobserver variability has been documented [28
]. Nevertheless, Stallmach et al. clearly showed that this type of placental defect can be a cause of fetal hypoxia and is linked to highly increased risk for stillbirth, especially during the end of pregnancy [27
]. Our results confirm these observations, showing a more than six-fold risk of stillbirth. Villous immaturity has been associated to maternal diabetes [27
] and trisomy 21 [29
]. Among our stillbirth placentas showing villous immaturity, two (11%) were connected to diabetes, a figure similar to the findings of Stallmach et al. [27
], and additional two had trisomy 21; in the remaining cases, the defect must be regarded as “idiopathic.”
SGA showed a sevenfold risk of stillbirth. SGA is a known risk factor for stillbirth [30
] and can include infants with intrauterine growth retardation and be related to maternal smoking and mothers with illness as for example preeclampsia [31
]. In this study, we did not find an elevated risk for stillbirth among smoking mothers or mother with severe illness or pregnancy complication. The lack of difference might be due to the small amount of smokers and mothers with severe illness or pregnancy complications that might lead to a type 2 error related to inadequate power in this study. Another explanation can be that fetuses with these risks die in earlier gestation.
Earlier studies have reported that high BMI is a risk factor for stillbirth [32
]. In this study, we found a two-fold risk for stillbirth if BMI >24.9. High BMI is an increasing problem globally and is associated with several diseases as preeclampsia and diabetes [33
]. Even in healthy overweight and obese women, the risk for term stillbirth is increased [32
]. The reason for that is unclear and needs to be further investigated.
There is always a risk for bias in case-control studies, for example, recall or observer bias. Since the cases in this study were included after the stillbirth was identified and most of the information was collected from the maternity medical records, we do not think that recall bias is a big problem in this study, even though the risk cannot be ignored. The pathologist was not blinded for the groups, implying a risk for observer bias. To decrease that risk, a structured protocol for placental examination was used. By using a graded scale for CAM the risk for inter- and intra-observer variability can be further reduced [10
]. We could exclude the risk for interobserver variability since all placental examinations were performed by only one pathologist. A disadvantage of this approach is that the reproducibility of placental findings is not validated, but we considered that outside the scope of the present study.