Intrauterine fractures are a rare finding in routine prenatal imaging studies. This condition can be secondary to maternal trauma, genetic disorders of the skeleton, as well as other predisposing maternal metabolic and vascular disorders [1
]. Genetic disorders that have previously been reported to cause intrauterine fractures include osteogenesis imperfecta (OI), osteopetrosis, hypophosphatasia and Ehlers–Danlos syndrome (EDS) type VII with a genetic mutation of type I collagen [1
]. Other acquired factors that may increase the risk of intrauterine skeletal fragility include vascular compromise of the fetal skeleton and maternal metabolic abnormalities [1
OI is the most common hereditary bone fragility disorder associated with abnormalities in type I collagen causing a variety of clinical manifestations [2
]. At least 90% of OI patients have autosomal dominant mutations in COL1A1
genes, while the minority of the patients had other mutations that affect structural integrity of the skeleton [2
]. Interestingly, it has been shown in some populations that a significant number of patients with clinical features of OI tested negative for known causative genetic mutations for OI [3
]. These observations suggest that when an infant presents with a history of a fracture or fractures with a negative genetic test for OI, there are likely causes besides nonaccidental trauma, including other causative genetic disorders of the structural components of the skeleton, resulting in bone fragility.
Like OI, Ehlers–Danlos Syndrome (EDS) is a genetic disorder of the collagen–elastin matrix. However, unlike OI, most forms of EDS, including EDS, hypermobility type (hEDS), are not associated with mutations of either COL1A1
genes, and the causative genetic mutation of hEDS is still unknown [5
]. Many of the physical manifestations of OI are very similar to those observed in patients with EDS, including capillary fragility, joint hypermobility and bone fragility in infants and adults [5
]. Other clinical manifestations of hEDS include chronic pain, mast cell hypersensitivity, gastroparesis, chronic fatigue, dysautonomia, and anxiety among other associated symptoms. The management of hEDS includes treatment of acute manifestations such as joint dislocation, attenuation of chronic symptoms, and prevention of acute and chronic complications [6
We report a male infant who had multiple fractures in utero consistent with OI features. Genetic testing for OI was negative. The patient’s mother had been previously diagnosed with hEDS. We identified a homozygous initiator codon loss-of-function mutation in the CCDC134 gene along with other possible predisposing genetic variants, including the homozygous variants of the genes CCDC134, COL15A1 and ZPFM, and the heterozygous variants of the genes MYH3, BCHE, AUTS2 and ZFPM1. We discussed how the combination of these variants may cause the complex phenotype of OI and EDS features.
The patient, at 32w1d of gestation, was found to have intrauterine growth retardation (IUGR), a decreased thoracic size, short limbs and multiple fractures by high-resolution ultrasonography (Figure 1
, Figure 2
and Figure 3
). The ultrasound performed by a licensed technician and interpreted by a board-certified radiologist showed a placenta posterior that was normal in appearance. A subsequent fetal MRI revealed a deformity of the thoracic cage and micromelia with suspected bilateral fractures of the proximal femurs and the right upper extremities. These findings suggested a variant of OI or a syndrome resembling OI.
The male infant was born at 40w1d gestational age via C-section due to prolonged labor with no significant complications at Boston Medical Center. Placental examination was unremarkable. The patient’s birth weight of 3030 g was at the 19th percentile and his length was 45 cm (below the 3rd percentile). The occipitofrontal head circumference was 36 cm (77th percentile). After birth, the radiologic findings of fractures were consistent with the ultrasonography during pregnancy. These included multiple rib fractures and bilateral fractures of the proximal humeral and femoral diaphysis (Figure 4
, Figure 5
and Figure 6
). He was then transferred to Boston Children’s Hospital for bisphosphonate therapy, further management, and genetic testing. A subsequent genetic test for OI was negative for pathogenic variants.
This is the first case report of an infant with multiple intrauterine fractures of long bones and anterior and posterior ribs consistent with a phenotype of OI, who tested negative for genetic mutations of type I collagen (i.e., COL1A1 and COL1A2 genes). This was confirmed by our whole-genome sequencing evaluation. Our clinical evaluation of the mother was consistent with her diagnosis of hEDS. Although it can be difficult to determine if an infant has EDS hypermobility type since there is no genetic test for this condition, the infant had many physical characteristics and medical history conditions associated with EDS hypermobility type. These included blue sclerae, mast cell hypersensitivity, gastroparesis symptoms, excessive joint hypermobility, and greater skin translucency and elasticity than would be expected for a six-month-old child. Whole-genome sequencing of the infant and his parents’ DNA did not reveal any pathologic mutations known to cause OI or classical EDS. However, whole-genome sequencing of the infant revealed potentially pathogenic variants associated with osteogenesis and bone development, including the homozygous variants of the genes CCDC134, COL15A1 and ZFPM1, and the heterozygous variants of the genes MYH3, BCHE and AUTS2. Based on these findings, it can be concluded that the multiple intrauterine fractures in this infant may be caused by the combination of these genetic variants. Of note, although there is no history of consanguinity in the family, this possibility cannot be excluded based on the observed identical rare variants.
The underlying pathophysiology of the multiple intrauterine fractures observed in this infant is thought to be primarily mediated by the dysregulated ERK-MAPK pathway in the osteoprogenitors, which has been shown to be essential for skeletal development and homeostasis [9
]. The CCDC134
gene encodes the coiled-coil domain containing 134 (CDCC134) secretory protein that inhibits the intracellular ERK-MAPK pathway by inhibiting transcriptional activity of ELK1 and phosphorylation of ERK and JNK/SAPK [10
]. Dubail et al. [3
] reported that the homozygous loss-of-function mutation at the initiator codon in the CDCC134
gene caused bone fragility in three patients who presented with clinical features of OI which did not respond to bisphosphonate therapy. Subsequent functional studies confirmed that this loss-of-function genetic mutation of CDCC134
leads to the absence of the CDCC134 protein, which induced the phosphorylation of ERK and inhibited the expression of OPN
(osteopontin) and COL1A1
, thereby leading to reduced mineralization in the osteoblasts of the patients [3
]. It should be noted that one of the patients in the Dubail paper was found to have fractures at birth. It is unclear if these were new fractures from the birth process or healing fractures, in which case the phenotype would be similar to our patient, who had as many as 23 in utero/fetal fractures.
While the mutation of CDCC134
is likely the most important causative factor for in utero skeletal fragility in our patient, it should be noted that this genetic mutation has been previously reported to have highly variable phenotypic expressivity, including clinical features of bone fragility/OI in childhood and adulthood [3
]. It is therefore probable that other genetic variants identified in our patient may have contributed to the development of a more severe form of skeletal fragility that led to intrauterine fractures. These include the homozygous variants of COL15A1
and the heterozygous variants of MYH3, BCHE
. Although the exact mechanisms by which the genetic variations in these genes affect early-life skeletal development are still unclear, there is evidence that these genes are involved in the osteogenic process and maintenance of the healthy mineralized skeleton.
encodes the alpha chain of type XV collagen that is recently known as novel bone extracellular matrix protein [11
]. This protein plays an essential role in the early stage of the osteogenic process and has been implicated in bone mineralization by influencing the deposition of minerals into the matrix [11
encodes the zinc finger protein, FOG family member 1. This protein interacts with the transcription factor GATA2 in the osteogenic lineage and was shown to be essential for trabecularization and the mechanical strength of the bone [12
encodes the protein myosin heavy chain 3, a major contractile protein in the skeletal muscle [13
]. Genetic mutations in this gene are associated with congenital arthrogryposis syndromes and spondylocarpotarsal synostosis syndrome, a rare group of skeletal dysplasias [14
]. The relationship between MYH3
and osteogenesis is thought to be related to its function in regulating transforming growth factor-β activity in the sclerotome [15
encodes the enzyme butyrylcholinesterase which degrades acetylcholine in addition to acetylcholinesterase. This enzyme is expressed by the osteoblast-like cells and is involved in regulating the number of osteoclasts and bone microarchitecture [16
]. Finally, AUTS2
encodes the autism susceptibility candidate 2 protein that is involved in neural migration and neurogenesis. It has been reported that mutations in this gene are associated with neurological and skeletal abnormalities [17
], suggesting the possible link of this gene to skeletal development.
It is of particular interest that the mother was clinically diagnosed with hEDS and the infant therefore had a 50% chance of acquiring it from her. He demonstrated physical findings consistent with hEDS, including joint hypermobility, intense blue sclera, and increased translucency and elasticity of the skin. It is still to be determined whether the infant will continue to demonstrate signs and symptoms of hEDS later in his life, as clinical evaluation of joint hypermobility, physical exam findings and medical history in infants are questioned as to their reliability [18
EDS and OI are connective tissue disorders involving the collagen–elastin matrix that have overlapping clinical features, including bone fragility [19
]. Among the 13 subtypes of EDS with different phenotypes, the most common subtype is hEDS, which is associated with joint hypermobility, increased elasticity of the skin, and fragility of the capillaries and skeleton [18
]. While there are genetic tests for some subtypes of EDS, no genetic test has been developed for diagnosing hEDS [2
]. EDS has, however, been shown to be associated with an increased risk of fractures in adults, especially of the vertebrae, independent of bone mineral density [20
]. In addition, there has been a report of 67 infants with fragility fractures and a concurrent family history of hEDS [21
]. Since genetic mutations directly responsible for skeletal fragility in EDS have yet to be identified, the association of EDS with fragility fractures in infants needs more investigation [22
]. At the same time, it is recognized that EDS and OI present with similar clinical features, including joint hypermobility and vascular fragility [2
]. This concept of overlapping clinical features of EDS and OI is supported by the case reports of the coexistence of OI and EDS and is strengthened by our report [21
Based on this notion, there may be common genetic mutations that explain bone fragility and joint hypermobility in EDS and possibly OI. Our case report suggests that the genetic mutation in CCDC134 is one of the possibilities. Although this hypothesis is opposed by the fact that the father had no joint hypermobility, it is not uncommon for a genetic disease to have variable clinical manifestations. Further studies are required to investigate whether the heterozygous mutation of CCDC134 is associated with joint hypermobility.
This case report has major implications for the approach to the diagnosis of nonaccidental trauma (child abuse) in children with multiple fractures. The findings of multiple fractures with various stages of healing with a negative genetic test for OI, Menkes disease and glutaric acidemia type 1 have been accepted by some geneticists and child abuse experts to be sufficient to support a diagnosis of nonaccidental trauma [27
]. The findings in this case imply that the genetic variants involved in skeletal development and fragility are not limited to the current panel of genetic tests and thus raise a question on the validity of the current recommendations. This is consistent with previous reports showing that a large number of patients with clinical features of OI tested negative for known causative genetic mutations for OI [3
]. Our report documents impressive in utero fractures of both arms and both legs with follow-up X-rays after birth documenting these fractures as well as anterior and posterior rib fractures in various stages of healing. If this mother had brought in her son for medical care later in his infancy without prior diagnosis of in utero fractures, these X-ray findings would almost certainly have resulted in the diagnosis of nonaccidental trauma since the infant tested negative for OI and other metabolic causes for infantile skeletal fragility, including Vitamin D deficiency. This would likely have resulted in the removal of the infant (and any siblings) from the home and accusations of felony child abuse against the parents.
This scenario was documented to have occurred in three nonaccidental trauma index cases where at least one parent was documented to have EDS and the infant had medical and physical evidence for the same bone fragility disorder [21
]. Similar to our infant who had multiple fractures identified in utero and in infancy and who was vitamin D sufficient, a 2-month-old infant presented with 10 fractures, including a femoral fracture and anterior rib fractures, which were diagnosed as caused by nonaccidental trauma due to the fact that the infant tested negative for OI and the fractures were in various stages of healing. The infant’s mother had EDS and the infant manifested clinical signs for the same genetic disorder. No other metabolic abnormalities were observed in the infant and the infant was found to be vitamin D sufficient. The parents were charged with felony child abuse and the infant removed from their care. After further consideration, the child was returned to the parents [21
These cases raise serious concern about the present criteria that are used for the diagnosis of nonaccidental trauma in infants who present with multiple fractures. It should be acknowledged that the current diagnostic panel of bone fragility disorder represents only “known” genetic variants, not all possible genetic variants. This notion is supported by the observation by Dubail et al. [3
] that 25 of the 350 patients with clinical features of bone fragility consistent with OI remained without molecular diagnosis as they tested negative for OI and related bone fragility disorders. One should consider adding CDCC134 to the genetic panel for bone fragility disorders. This also supports the urgent need for further investigations to identify additional causative genetic variants for skeletal fragility, including yet to be identified genes associated with a well-recognized bone fragility disorder associated with a genetic defect of the collagen–elastin matrix: EDS.
There are certain limitations of this case report that should be acknowledged. First, a case report generally provides a relatively low level of evidence compared with other study designs. Further studies with a more robust study design are warranted to confirm our observation. Second, we were unable to acquire bone tissue and measure the expression of the CCDC134
gene in our reported case. However, our results revealing the identical genetic mutation of CDCC134
in our infant with intrauterine fractures has verified the functional study by Dubail et al. [3
] that a dysregulated ERK-MAPK pathway due to CDCC134 deficiency is involved in the pathogenesis of skeletal fragility in these patients. Finally, it should be noted that the CCDC134
gene is also important in regulating collagen synthesis and promoting proliferation and activation of cytotoxic T lymphocytes [28
]. We did not observe any other extra-skeletal abnormality in this infant and the information on T cell function is unavailable. A long-term follow-up of immune function is therefore warranted.