- Department of Neurological Surgery, Rush University Medical Center, Chicago, IL, USA
- Department of Pharmacology, Ophthalmology and Neurological Sciences, Rush University Medical Center, Chicago, IL, USA
Vincent C. Traynelis
Department of Neurological Surgery, Rush University Medical Center, Chicago, IL, USA
DOI:10.4103/2152-7806.129259Copyright: © 2014 Straus D. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
How to cite this article: Straus D, Xu S, Traynelis VC. Os odontoideum in identical twins: Comparative gene expression analysis. Surg Neurol Int 20-Mar-2014;5:37
How to cite this URL: Straus D, Xu S, Traynelis VC. Os odontoideum in identical twins: Comparative gene expression analysis. Surg Neurol Int 20-Mar-2014;5:37. Available from: http://sni.wpengine.com/surgicalint_articles/os-odontoideum-in-identical-twins-comparative-gene-expression-analysis/
Background:Os odontoideum is a well identified anomaly of the craniovertebral junction. Since its initial description, there has been a continuous debate regarding the nature of its etiology: Whether congenital or traumatic. We sought to compare the gene expression profiles in patients with congenital os odontoideum, those with traumatic os odontoideum and controls.
Methods:We have evaluated a pair of identical twins both with os odontoideum. We identified two additional patients with and four subjects without os odontoideum. We analyzed the gene expression profiles in these patients using a custom TaqMan microarray and quantitative reverse transcriptase polymerase chain reaction (qRT-PCR). The relative gene expression profiles in the two identical twins, the two nontwin patients with os odontoideum and the controls were assessed.
Results:A total of 213 genes with significantly different expression between the twin os odontoideum patients and the subjects without os odontoideum were detected. CACNG6, PHEX, CACNAD3, IL2, FAS, TUFT1, KIT, TGFBR2, and IGF2 were expressed at levels greater than 100-fold more in the twins. There were six genes with significantly different expression profiles in the twins as compared with the nontwin os odontoideum patients: CMK4, ATF1, PLCG1, TAB1, E2F3, and ATF4. There were no statistically significant differences in gene expression in the four patients with os odontoideum and the subjects without. Trends, however, were noted in MMP8, KIT, HIF1A, CREB3, PWHAZ, TGFBR1, NFKB2, FGFR1, IPO8, STAT1, COL1A1, and BMP3.
Conclusions:Os odontoideum has multiple etiologies, both traumatic and congenital and perhaps some represent a combination of the two. This work has identified a number of genes that show increased expression in a pair of twins with congenital os odontoideum and also demonstrates trends in gene expression profiles between a larger group of os odontoideum patients and non-os patients. A number of these genes are related to bone morphogenesis and maintenance.
Keywords: Anomaly, cervical spine, craniovertebral junction, identical twins, os odontoideum
Os odontoideum is a well-defined anatomic anomaly consisting of a smooth ossicle of bone separated from a shortened odontoid process.[
Patients and study design
IRB approval was obtained from Rush University Medical Center and informed consent was obtained from participants.
Two 20-year-old identical male twins were evaluated in the neurosurgery clinic at Rush University Medical Center. Neither twin had a significant history of trauma and both were active collegiate water-polo players. The clinical presentation was related to neck pain in one of the twins. Imaging revealed an os odontoideum. Flexion-extension imaging revealed an orthotopic os odontoideum with gross instability. Given this finding his brother underwent similar evaluation with comparable results. Preoperative cervical spine films are shown in Figures
Two additional, unrelated patients with an os odontoideum were subsequently treated. The first was a 49-year-old female who had a prior history of occipitocervical fusion at an outside hospital. She initially presented with hardware failure, pseudoarthrosis, and a wound infection. She did have a remote history of significant head trauma. Imaging revealed an os odontoideum, a C2-3 Klippel–Feil anomaly and a kyphotic deformity [
Four subjects with prior cervical spine radiography that clearly demonstrated normal craniovertebral junction (CVJ) anatomy—absence of os odontoideum or other congenital anomaly—were identified to serve as controls. One of these subjects was the biological mother of the identical twins with os odontoideum. This control group included a 52-year-old female, an 87-year-old female, a 49-year-old male, and a 59-year-old female.
Comparisons were utilized in a case-control model and included relative gene expression levels between the identical twins with os odontoideum and the controls without os odontoideum; between the identical twins and the unrelated patients with os odontoideum; and finally, between all patients with os odontoideum and all controls without os odontoideum.
Sample collection and RNA preparation
Blood samples were collected via venipuncture from all patients into 5 ml collection tubes containing EDTA and immediately placed on ice. Total RNA from the blood cell pellet of each sample and isolated mRNA for gene expression profiling using the miRVana RNA isolation kit (Ambion, Grand Island, NY).
mRNA qRT-PCR array assays and data analysis
A total of 1 μg of RNA of each sample was used for gene expression profiling. Gene expression profiling was performed on the HT7900 real-time PCR system (Applied Biosystems, Foster City, CA) using custom TaqMan array cards (Applied Biosystems,
To better understand the functions of the differentially expressed mRNAs, we performed a NCBI Gene search[
A comparison between the identical twins with os odontoideum (Twins) to the four control subjects (non-os) revealed 213 statistically significant differences in gene expression. The top 30 genes and a brief summary[
Comparing the Twins with the two unrelated patients with os odontoideum (non-Twin os) revealed six genes with statistically significant differences in expression. These genes and a brief summary of their functions are presented in
Comparing both the Twins and non-Twin os (Os) to the non-os group revealed no statistically significant differences in gene expression. We report the 10 genes with the greatest fold-change and an unadjusted P < 0.1 [
The identification of a pair of identical twins without a history of trauma both harboring os odontoidea of nearly identical morphology [Figures
Numerous case reports and case series attest to the rarity of this anomaly and neither the overall incidence nor the prevalence is well documented. The closest (albeit flawed) estimate is provided by Sankar et al.,[
This study identified a litany of significant differences in gene expression (213/380 genes in total) between the Twins with the non-os group. Notably, these genes have biological functionality related to bone formation and maintenance. PHEX, a gene involved in bone mineralization, was elevated 447-fold in the Twins. TUFT1, another gene related to mineralization (of enamel), was increased 109-fold. TFGBI, a gene that mediates cell–collagen interactions and is thought to be involved in endochondral bone formation, was elevated 92-fold. MMP8, a neutrophil MMP that breaks down collagen types I, II, and III, was elevated 90-fold in the Twins. RUNX2, encodes a transcription factor that induces osteoblastic differentiation and is critical in skeletal morphogenesis, was elevated 66-fold in the Twins. Though none of the expression profiles comparing the os patients to the non-os subjects held statistical significance, we found trends in genes similarly involved in bone development and maintenance. MMP8 again was elevated 95-fold; FGFR1 was elevated 25-fold; COL1A1 was elevated 15-fold and BMP3 was elevated 7-fold. Further investigation of these genes may aid in further understanding of the cellular and molecular pathogenesis of os odontoideum.
This study is compromised by the small sample size, which may, in part, be responsible for the lack of statistical significance in the trends in gene expression profile that we found between the os and non-os groups. We opted to include the gene expression profile from the Twins’ mother in order to control for potential hereditary gene expression patterns unrelated to os odontoideum. The comparison group does not contain age-matched cases, adding a potential confounding variable. There is also the limitation of data selection: There were over 200 genes with significant differences in expression profiles between the Twins and the non-os controls. The strategy employed to overcome this potential problem was to focus on those genes with the greatest magnitude of change. Though this approach allows for a concise and informative presentation of the results, it may have excluded some relevant gene profiles.
Os odontoideum has multiple etiologies, both congenital and traumatic and perhaps some cases represent a combination of the two. Further definition of each type and examination of their relative prevalence will be informative. Moreover, investigation of the relevance of this distinction as to the clinical evaluation, natural history, and treatment is appropriate. We have identified a number of genes that show increased expression in a pair of twins with congenital os odontoideum and also demonstrated trends in gene expression profiles between a larger group of os odontoideum patients and non-os patients. A number of these genes are related to bone morphogenesis and maintenance. Further investigations of the molecular biology of these genes may confer a greater understanding of this anomaly.
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