Sagittal craniosynostosis (CS) is a pathologic condition that results in premature fusion of the sagittal suture, restricting the transverse growth of the skull leading in some cases to elevated intracranial pressure and neurodevelopmental delay. There is still much to be learned about the etiology of CS. Here, we report a case of 56-year-old male cadaver that we describe as sagittal CS with torus palatinus being an additional anomaly. The craniotomy was unsuccessful (cephalic index, CI = 56) and resulted in abnormal vertical outgrowth of the craniotomized bone strip. The histological analysis of the latter revealed atypical, noncompensatory massive bone overproduction. Exome sequencing of DNA extracted from the cadaveric tissue specimen performed on the Next Generation Sequencing (NGS) platform yielded 81 genetic variants identified as pathologic. Nine of those variants could be directly linked to CS with five of them targeting RhoA GTPase signaling, with a potential to make it sustained in nature. The latter could trigger upregulated calvarial osteogenesis leading to premature suture fusion, skull bone thickening, and craniotomized bone strip outgrowth observed in the present case.
CS is a condition that affects ~1 in 2,000–2,500 newborns and manifests itself as a premature fusion of a single or multiple cranial suture(s) leading to the deformation of a skull shape [
Therefore, the main objectives of this study were to: (i) characterize the craniofacial pathology (scaphocephaly) observed in the 56-year-old cadaver and (ii) gain insights into its genetic component by identifying the respective genetic variants through exome sequencing of DNA extracted from tissue procured from the donor’s body. A clearer understanding of the nature of the above pathology may help to better delineate the mechanism(s) responsible for its development, as a well as may improve outcomes of the specialized corrective clinical procedures.
A 56-year-old male cadaver was received through Saint Louis University (SLU) School of Medicine Gift of Body Program from an individual who had given his written informed consent. The available medical record indicated that this individual had a history of moderate mental retardation, cerebral palsy, seizure disorder, scoliosis, hydrocephalus, joint pain, mood disorder, anxiety disorder, encephalopathy and leukopenia. The cause of death was indicated as cerebral palsy. The cadaveric head was separated from the extremely contracted body and embalmed using 2 : 1 mixture of ethylene glycol and isopropyl alcohol.
The initial visual examination of the embalmed patient’s head revealed its abnormal, scaphocephalic, shape as well as a presence of bulging sagittal bone strip (Figure
(a) Physical examination of the scaphocephalic cadaver head. Superior view shows the demarcation of the displaced sagittal strip (black arrows). (b) Computed Tomography (CT) images of the cadaver head. Left: The axial view reveals a thickened skull and spaces of bone towards the posterior aspect of the skull. The long, narrow skull yielded a cranial vault index of 0.56. The brain appears to have undergone significant atrophy. Right: The coronal view shows an abnormal thinning of the skull on each side of the sagittal suture near the superior aspect of the skull. These areas likely coincide with the areas lacking bone in the axial view. (c) Increased bone thickness in the scaphocephalic skull of the individual with CS. The thickness of the frontal, parietal, occipital and temporal bones was measured in five male mesocephalic skulls (normal, dark grey) at the bony points described in [
Upon closer examination of the individual’s head it was concluded that he underwent, most likely early in infancy, a neurosurgical procedure, a sagittal strip craniotomy, with a likely effort to correct the anomalous skull shape and to reduce intracranial pressure. One of the most interesting features of the present case is an abnormal re-growth of the surgically removed bone strip and the resultant elevated vertical displacement of the skull (Figure
Examination of the exposed calvarium. (a) The exposed calvarium shows the presence of the coronal and lambdoid sutures. The vertical displacement of the sagittal strip is apparent at bregma. At the sagittal strip—parietal bone junction (dashed lines), there are areas of bridging bone and fibrous bridging tissue. (b) Internal view of the calvarium. Black arrowheads—large arachnoid granulations. (c) Torus palatinus is evident in the midline (arrow) with the epithelium reflected.
Also importantly, examination of the dural surface of the calvarium revealed several deep granular foveolae indicative of large arachnoid granulations in the sagittal strip (Figure
Physical examination of the maxillofacial features of the cadaveric head revealed a large underbite that prompted the dissection of the mandible to probe for additional abnormalities. The mandible was exposed by removing the soft tissue from the mental surface followed by bisection of the bone and tongue. This procedure revealed an exostotic hard palate (torus palatinus) and complete edentulism (Figure
Sections of bony tissue from the sagittal strip revealed areas of immature compact bone with incomplete or developing Haversian systems, whose orientation was predominately perpendicular to the section orientation (Figure
Histological analysis of the scaphocephalic calvarium. (a) The sagittal strip displays cancellous bone with variably sized osteons (white stars). Intervening medullary spaces (black star) contain typical myeloid cellular elements, but without the presence of osteoclasts. (b) The bridging bone demonstrates scattered immature Haversian systems. Areas suggestive of osteon remnants are indicated by the arrowheads. (c) An additional image through the bridging bone shows dense, confluent areas of well-formed Haversian systems characteristic of typically formed compact or cortical bone. (d) Enlarged boxed area in C shows variably sized Haversian systems of similar orientation (arrows). Portions of the image indicated by arrowheads suggest immature (woven bone) that has been replaced by newer Haversian systems resulting in the formation of compact bone.
More importantly, sections of bony tissue from surgically created margins revealed an extremely high number of osteons, with some, well-formed and others, formed incompletely (Figures
The genetic underlining of the present case was addressed by performing a genetic screen for the putative variants using NGS technology applied to DNA extracted from the respective cadaveric tissue specimen as described previously [
The sequencing of the DNA coding regions (exome) yielded 81 rare genetic variants (minor allele frequency, MAF ≤0.01) with predicted deleterious (pathological) implications (Table
Selected deleterious (pathologic) genetic variants associated with the current case of sagittal craniosynostosis.
Gene | Protein function | Variant | MAF |
---|---|---|---|
|
Rho GTPase Activating Protein 21. Functions as a GTPase-activating protein (GAP) for RHOA and CDC42. | p.Arg492Gly | 0.0021 |
|
Bone morphogenetic protein 6. Teeth development. Cartilage development. Endochondral ossification. Positive regulation of osteoblast differentiation. Positive regulation of bone mineralization. Positive regulation of chondrocyte differentiation. | p.Pro93Ser | 0.0001 |
|
Centrosomal protein of 162 kDa. Required to promote assembly of the transition zone in primary cilia. Acts by specifically recognizing and binding the axonemal microtubule. Required to mediate CEP290 association with microtubules. | p.Arg802Trp | 0.0001 |
p.Arg878Trp | |||
|
Rootletin. Major structural component of the ciliary rootlet, a cytoskeletal-like structure in ciliated cells which originates from the basal body at the proximal end of a cilium and extends proximally toward the cell nucleus (by similarity). Required for the correct positioning of the cilium basal body relative to the cell nucleus, to allow for ciliogenesis. | p.Arg637Trp | 0.0001 |
|
Dynein heavy chain 11, axonemal. Force generating protein of respiratory cilia. Produces force towards the minus ends of microtubules. Dynein has ATPase activity; the force-producing power stroke is thought to occur on release of ADP. | p.Pro2006Leu | 0.0001 |
|
GEM-interacting protein. Stimulates, in vitro and in vivo, the GTPase activity of RhoA. | p.Pro532Leu | 0.0001 |
p.Pro535Leu | |||
p.Pro561Leu | |||
|
InaD-like protein also known as PATJ. Negative regulator of Wnt signaling. Blocks DFz1 activity in the planar cell polarity pathway (PCP) in cooperation with atypical PKC. Fzd/PCP pathway represents the noncanonical Wnt signaling. | p.Glu1499Lys | 0.0099 |
|
Piezo-type mechanosensitive ion channel component 1. Pore-forming subunit of a mechanosensitive nonspecific cation channel. Plays a key role in osteogenesis. Its activation commits mesenchymal stem cells to osteogenic differentiation. | p.Pro2510Leu | 0.0042 |
|
E3 ubiquitin-protein ligase RNF213. Involved in the noncanonical Wnt signaling pathway in vascular development: acts by mediating ubiquitination and degradation of FLNA and NFATC2 downstream of RSPO3, leading to the inhibition of the noncanonical Wnt signaling pathway and promoting vessel regression. | p.Trp4677Leu | 0.01 |
The present case of craniofacial malformation could be described as a single suture sagittal CS with the additional associated anatomical pathologies being torus palatinus and complete edentulism. This conclusion was made based on the measured CI of 56 (75–90 being normal) derived from the respective CT images and the mandibulotomy results (Figures
The uniqueness and importance of this case is several-fold.
The bone regeneration represents a delicate balance between the formation of new bone and its resorption. The former process is regulated by the recruitment of osteoblasts to the site of injury and their ossification while the latter process is controlled by the osteoclasts recruitment to and their activity in the bony lesion [
It should be noted, that the genetic variants described above, although being identified as deleterious/pathologic by their stringent filtering through the three specific databases [
The current case provides a unique description of the histopathological features following craniotomy of the sagittal bone strip in CS as well as important information pointing toward a potential role of sustained RhoA signaling in the development and progression of sagittal CS.
The datasets and materials used and/or analyzed during the current study are presented in the main paper and additional files.
These data were presented in part at the Annual Experimental Biology Meeting (FASEB J. (2018), 32: Suppl. 1, Abstract 776.10).
The authors declare that they have no conflicts of interest.
This study was supported by the Center for Anatomical Science and Education, SLU School of Medicine.
We gratefully acknowledge Todd Gebke (SLU Hospital) for his expert assistance with CT imaging, Caroline Murphy and Barbara Nagel (SLU) for their skillful help with the histology slides preparation as well as Dr. Paul Cliften (GTAC, Washington University in St. Louis, St. Louis, MO, USA) for his invaluable assistance with the bioinformatics analysis. We would also like to thank Dr. MariaTeresa Tersigni-Tarrant for her assistance with the calvarial bone thickness measurements and for the contribution to the histological data analysis. We are grateful to Dr. Alexander Lin (SLU School of Medicine) for his clinical review of the case and Dr. Sidney B. Eisig (Columbia University College of Dental Medicine, New York, NY, USA) for performing the mandibulotomy.
Supplementary materials includes a description of Methods used in the study. Figure S1: craniectomy of the scaphocephalic cadaveric head. Table S1: complete list of deleterious (pathologic) genetic variants associated with the current case of sagittal CS.