Discussion
Main Findings
Our data suggest that abnormal CNVs are associated with maternal vascular lesions in placenta of stillborn fetuses. Several pathogenic CNV deletions and duplications were identified in eight-stillborn fetuses with maternal vascular lesions, involving several genes.
Interpretation
In 88% of stillborn fetuses, a direct cause or a major contributor to death was found by histopathologic examination of the placenta, showing placental abnormalities as the most common causes of death.3, 4, 9 Specifically, maternal vascular supply abnormalities were more common in preterm stillbirth cases, while fetal vascular supply abnormalities were more common among term stillbirth cases.6, 9 Moreover, a review of 120 autopsy reports of stillborn fetuses and placentas showed maternal vascular supply abnormalities in 54 (51%), fetal vascular supply abnormalities in 28 (26%), and inflammatory lesions in 13 (12%) cases as direct or major contributors of death.4, 9 Yet, many deaths in stillbirth cases remain unexplained,26-28 and our understanding of the mechanisms of placental dysfunction and stillbirth is limited.
In our study, 22q11.21 deletion, 16p13.11 duplication, and 4q32.3q35.2 and 17p13.3 deletion CNVs were identified in stillborn fetuses with maternal vascular pathological lesions. These CNVs were previously detected among stillborn fetuses in an analysis of SCRN cases,11 but not described in the context of placental pathological lesions. The deletion of 22q11.21 is pathogenic for DiGeorge/velocardiofacial syndrome.29 Pathologic placental lesions are found in DiGeorge sequence, characterized by hypoplasia in the umbilical cord arteries and widespread calcification of microthrombi in the arteries of the second and third order villous branches.30 A case study showed a copy gain of the distal region of chromosome 4 at segment 4q32.3q35.2 in a pregnant patient that presented with fetal edema and subsequent fetal loss.31 Furthermore, the duplication of 16p13.11 is implicated in multiple congenital anomalies in pediatric patients (n=1645).32 Mutations in MYH11 (myosin heavy chain 11) gene, among several genes in the 16p13.11 region, cause thoracic aortic aneurysms and/or dissections.33, 34Lastly, the 17q12 recurrent deletion syndrome is characterized by structural or functional abnormalities of the kidney.35 HNF1B (Hepatocyte nuclear factor 1B) gene, among several genes in the 17q12 region, is implicated in renal cysts and diabetes syndrome.36
In light of prior data, our findings suggest that 22q11.21, 4q32.3q35.2, and 16p13.11 CNVs may contribute to fetal death through placental abnormalities, both directly, as well as indirectly, by contributing to cardiac abnormalities. Interestingly, maternal/fetal vascular and inflammatory placental pathological lesions were common in pregnancies complicated by congenital heart defects 37 and placental insufficiency is associated with congenital heart defects in animals and humans.38-40 Further, the placental genome plays a role in mediating fetal and maternal health.41-43 As such, 16p13.11 and 17q12 CNVs described in the context of congenital anomalies and kidney function in children and adults also suggest that genetic abnormalities in the placenta may underlie mechanisms of the developmental origins of health complications in later life.38, 42 Together, these mechanisms may provide answers to causes of death in stillbirth, a basis for estimating recurrence risk, and offer insight into the developmental origins of diseases in later life.
Strengths and Limitations
Our study has several limitations. In the present analysis, we only reported pathogenic deletions and duplications spanning several genes in stillborn fetuses with maternal vascular placental pathological lesions. While only maternal vascular placental pathological lesions were associated with CNV types in our study, the small sample size in the abnormal CNVs group may have limited power to detect associations of CNV types with other placental pathological lesions. However, to date, our study included the largest sample size of stillbirths with CNVs that had placental pathological exam completed. Another limitation is that pathologists were not blinded to stillbirth or live-birth status in the SCRN study because of the need to perform both clinical and research placental examination. Furthermore, our ability to determine pathogenicity of CNVs is limited. This will improve with increasingly larger databases of normal and abnormal phenotypes. Additionally, there are limitations to microarray-based analyses, which include inability to detect truly balanced rearrangements. Microarray-based analyses also report large chromosomal regions that span several genes, such as the ones reported in our study, making it difficult to target specific mutations in genes for clinical application. Lastly, due to lack of paternal DNA, we were not able to distinguish inherited from newly occurring CNVs in the placenta or fetus. Using higher resolution technology, such as next generation sequencing of DNA from families, future studies will be able to identify genetic mutations causing placental dysfunction and stillbirth.
One of the strengths of our study is that our cohort included a geographically, racially, and ethnically diverse study population with stillbirth. In addition, participants had a complete evaluation, including fetal postmortem examination, placental pathological analysis conducted by perinatal pathologists, and maternal-fetal testing.44 These study design features provided careful phenotyping of stillbirth included in our study and maximized the validity of the present analysis.
Conclusion
Our report provides additional support for the utility of high-density microarray analysis in the detection of CNVs associated with placental pathological lesions in stillbirth cases. Further clarification of the relationship between chromosomal aberrations and placental abnormalities will be important in order to better understand the specific mechanisms leading to placental dysfunction and stillbirth. As such, these mechanisms may provide answers to causes of death in stillbirth and the basis for estimating recurrence risk.