The carotid axis revisited

The aim was to determine the variations in the level of origin of carotid bifurcation and diameters of the common, internal, and external carotid arteries which is clinically important for several interventional procedures. Therefore, 165 human embalmed corpses were dissected. The data collected were analyzed using the Chi square-test and the Pearson correlation test. The results of previous studies have been reviewed. In relation to the level of the carotid bifurcation, taking as a reference point the hyoid bone, the values ranged from 4 cm below the hyoid body to 2.5 cm above the body of the hyoid, being the average height—0.33 cm, with a standard deviation of 1.19 cm. The right carotid bifurcation was established at a higher level (x = − 0.19 cm.) than the left one (x = − 0.48 cm.) (p = 0.046). On the contrary, no significant gender differences could be observed. The arterial calibres of the common and internal carotid arteries were higher in male than female. In the internal carotid artery (X = 0.76 cm.), the left was greater than the right (X = 0.72 cm.) (P = 0.047). However, no differences in the distribution of the calibre of the external carotid artery were found neither by side nor gender. Variations in the level of bifurcation and calibres of carotid arteries are relevant for interventional radiology procedures and head and neck surgeries. Knowledge of these anatomical references might help clinicians in the interpretation of the carotid system.


Material and methods
A total sample of 165 embalmed body donors to science belonged to the Department of Anatomy, University of Cambridge, UK, were dissected. The individuals had given their written informed consent prior to death for their use for scientific and educational purposes and donated their bodies. According to National Law, scientific institutions (in general Institutes, Departments or Divisions of Medical Universities) are entitled to receive the body after death mainly by means of a specific legacy, which is a special form of last will and testament. No bequests were accepted without the donor having registered their legacy and been given appropriate information upon which to make a decision based upon written informed consent (policy of ethics); therefore, an ethics committee approval was waived 26 . These body-donors have been previously dissected by preclinical students and completed by the authors. The gender distribution was 74 male and 91 female body-donors, with an age range of 60 to 103 years. Clinical histories were available, in no case containing any reference to vascular surgical interventions.
Based upon the completeness of the neck structures after dissection, 4 different samples of the available bodydonors were taken into account in this study following these parameters: (1) the calibre of the common carotid artery 103 cases (47 male and 55 female); (2) the calibre of the internal carotid artery 139 cases (63 male and 77 female); (3) the calibre of the external carotid artery 206 cases (95 male and 111 female) and (4) the height of carotid bifurcation in 141 cases (61 male and 80 female).
Distances and external diameters were measured with callipers. These measurements were verified by a second investigator. Statistical comparisons were made using the chi-square test, with a value of p < 0.05 taken as statistically significant and the Pearson correlation test. Previous published results were carefully reviewed and compared with this sample.
Ethical approval. All procedures performed in studies involving human participants were in accordance with the ethical standards of the institutional and/or national research committee and with the 1964 Helsinki declaration and its later amendments or comparable ethical standards.
Informed consent. Informed consent was obtained from all individual participants included in the study.
According to Spanish and Austrian National Law, scientific institutions (in general Institutes, Departments or Divisions of Medical Universities) are entitled to receive the body after death mainly by means of a specific legacy, which is a special form of last will and testament. No bequests are accepted without the donor having registered their legacy and been given appropriate information upon which to make a decision based upon written informed consent (policy of ethics); therefore, an ethics committee approval is not necessary.

Results
In order to improve the clarity, the results obtained have been exposed following the order proposed in the "Material and Methods" section.
1) In relation to the calibre of the common carotid artery, it has been studied in a sample made up of 103 cases (47 in male and 56 in female). The values obtained for the calibre of the common carotid artery ranged from 0.7 to 1.3 cm. The average diameter obtained was 0.97 cm with a standard deviation of 0.14 cm (Fig. 1). When analyzing the diameter of the common carotid artery depending on the side, no statistically significant differences were observed (p > 0.05). On the contrary, when carrying out the analysis considering the gender variable, these differences have been observed with a significance of p = 0.001. The average arterial calibre of the common carotid artery was higher in male (X = 1.01 cm.) than in female (X = 0.93 cm.). 2) The calibre of the internal carotid artery variable has been studied in a sample made up of 139 cases, 63 in male and 76 in female. The values found for the internal carotid artery ranged from 0.4 cm to 1.8 cm. The average diameter obtained was 0.74 cm. with a standard deviation of 0.16 cm (Fig. 2). When analyzing the diameter of the internal carotid artery according to the side, the diameter of the left internal carotid artery (X = 0.76 cm) was greater than the diameter of the right internal carotid artery (X = 0.72 cm) being statistically significant (p = 0.047). In the case of gender, differences have been observed with a marked tendency towards statistical significance (p = 0.54). The arterial calibre of the internal carotid artery was greater in male (x = 0.77 cm) than in female (x = 0.71 cm).
3) The calibre of the external carotid artery has been studied in a sample consisting of 206 cases, 95 in male and 111 in female. The values found for the external carotid artery ranged from 0.3 to 0.9 cm. The average diameter obtained was 0.49 cm. with a standard deviation of 0.08 cm (Fig. 3.). In the distribution of this arterial calibre by side and gender, statistically significant differences have not been observed (p > 0.05). Both the internal and external carotid arteries originate from the common carotid artery, specifically from the carotid bifurcation; therefore, a correlation study has been performed between the diameters of the common carotid artery and the internal carotid artery and between the diameters of the common carotid artery and the external carotid artery.
The application of the Pearson correlation test has shown that there was no clear correlation between the diameters of the common and internal carotid artery (Fig. 4), but a tendency towards correlation between the diameters of the common carotid artery and the external carotid artery was verified (p = 0.113) (Fig. 5).
Statistically significant association has been obtained in the correlation between the diameters of the internal and external carotid arteries (p = 0.003) (Fig. 6).  www.nature.com/scientificreports/ 4) In relation to the level of the carotid bifurcation, this variable has been studied in a sample consisting of 141 cases, 61 in male and 80 in female. The level, which has been taken as a reference point, was the body of the hyoid bone. The values obtained for the level of the carotid bifurcation ranged from 4 cm. below the hyoid body and 2.5 cm. above the body of the hyoids. The average height was-0.33 cm., with a standard deviation of 1.19 cm. (Figs. 7, 8A,B). The Klippel-Feil anomaly, typically associated with a low carotid bifurcation, was not found in our sample size.
In the comparative study between sides, a statistical significance of p = 0.046 has been verified, establishing the right carotid bifurcation at a higher point (x = − 0.19 cm.) than the left carotid bifurcation (x = − 0.48 cm.). On the contrary, no significant differences have been observed in relation to the distribution by gender (p > 0.05).

Discussion
Most of the references consulted in this work regarding these arteries refer mainly to anatomical variations in relation to their origin, course, or disposition.
Regarding the calibre of these arteries, the results of the studies, as a whole, presented a great variability since they depended on the technique with which it has been studied (Conventional Angiography, Computerized      24 , was carried out from a morphometric analysis with a digital caliper of the luminal diameter of 86 carotid systems that were corroborated with the results of an ECO Doppler study. In our series, these measurements were performed extraluminally with a caliper in a variable sample depending on the artery studied. Comparing both series, a certain similarity has been observed in the diameters of the internal and external carotid arteries, taking into account that the differences obtained could be in relation to the different measurement performed -digital vs. manual-and with the thickness of the arterial wall. However, regarding the difference observed at the level of the common carotid artery-32 mm-these reasons did not justify this huge difference, nor could any reference in relation to the thickness explain it. In addition, and although it is another artery, the external carotid, in comparison with a radiological study, such as that of Czerwinski et al. 14 did not obtain such irregular results, since the average diameter obtained by this author for this artery was 0.57 cm (n = 240). Delving into our results with regard to the common carotid artery, this study verified that, with a sample of 103 cases, the values obtained ranged from 0.7 cm and 1.3 cm,   www.nature.com/scientificreports/ and the average diameter was 0.97 cm with a standard deviation of 0.14 cm. Therefore, the obtained values were grouped with little dispersion around the mean value in a sample whose size was greater than 100 cases. Based on the fact that the external and internal carotid arteries originated from the common carotid artery, a correlation study has been performed between these arteries. No statistically significant correlations between the common and internal carotid arteries and the common and external carotid arteries (p > 0.05) have been observed. Curiously, when studying the correlation between the internal and external carotid arteries, this was established in a statistically significant way (p = 0.003), which together with the results of the previous comparison with another author, could lead us to think about the possibility of a measurement error of the common carotid artery that should be confirmed with a larger sample size in future studies.
The diameter of the carotid arteries has been analyzed in relation to the side and gender with the following conclusions:  www.nature.com/scientificreports/ • The average diameter of the common carotid artery was higher in male (X = 1.01 cm.) than in female (X = 0.93 cm.) (P = 0.001). We did not observe significant differences regarding the side (p > 0.05). • The arterial calibre of the internal carotid artery was greater in male (x = 0.77 cm.) than in female (x = 0.71 cm.) With a p very close to statistical significance (p = 0.05). Also, the diameter of the left internal carotid artery (X = 0.76 cm.) was greater than the diameter of the right internal carotid artery (X = 0.72 cm.) (P = 0.047). • On the contrary, in the distribution of the diameter of the external carotid artery by side and gender, no statistically significant differences have been found (p > 0.05).
In relation to the anatomical level at which the carotid bifurcation occurs, two reference points are classically taken. An anterior one that corresponds to the upper edge of the thyroid cartilage 1,7,12 and a posterior one, corresponding to the 4th cervical vertebra 3,14 . The upper edge of the thyroid cartilage would be equivalent to the upper edge of the greater horns of the thyroid cartilage 4 .
In most cases the choice of one or the other is in relation to the method chosen when making the measurements. Using a surgical approach, the most common option is the anterior anatomical margin given its greater accessibility. However, when measurements are made using radiological techniques, the most feasible option is the anatomical relief of the vertebral column and bone framework, being, on the other hand, more stable regarding the age variable 3 .
In our case with a study based on body-donor dissection, and an entire sample with an age greater than 65 years, being homogeneous at the age variable, the reference level was the hyoid bone body.
The comparison of our results with those of the consulted bibliography which used a body-donor sample is exposed in Table 2.
In terms of embryogenesis, the first two aortic arches are commonly lost. However, it could happen that the persistent of the dorsal parts of the second arches becomes the root portion of the stapedial artery on each side or the persistent of connection of the dorsal and ventral parts of the aortic arches give rise to the hypoglossal artery 27,28 . The third and left fourth arches are retained, becoming the root portion of the internal carotid artery and the arcus aortic respectively. On the other hand, both the ventral and dorsal aortae beyond the portion of the third arch are preserved, the former gave rise to the stem of the external carotid directed cranially; some cases of external carotid artery agenesia have been reported 29 . The latter gave rise to the second part of the internal carotid artery, whereas the ventral aorta between the third and fourth arches becomes the stem of the common carotid artery 27 . The movement of the heart downward towards the thorax dragging the carotid arteries could be more or less extensive justifying the different level of division 30 . The asymmetry, right-left, of the blood flow justify differences of caliber observed among the carotid arteries. The collateral and terminal branches of the external carotid artery appear in human embryos of 14 mm to 17 mm by sprouting of the main external carotid trunk 27 . However, there have been reported cases where the collateral arteries arise from the internal or common carotid arteries 10,31,32 .
In our study, the percentage of cases in which the carotid bifurcation occurs at a higher level than the reference level taken previously has been clearly higher (20.5% vs. 3.5% for Quain et al. 7 , 3.3% for Al-Rafiah et al. 8 , and 0% for Lucev et al. 12 , Lo et al. 9 , and Mompeó et al. 18 ). Our results have been more similar to those of McAfee et al. 4 , who also observed the possibility of high bifurcations more frequently in a series of 140 hemi sides, since in up to 82% of cases the bifurcation was established in a 2.5 cm area inferior to the branch of the jaw.
In the distribution of the height of the carotid bifurcation according to the side and gender, unlike Smith and Larsen et al. 19 , in our series this height has been established at a higher level on the right side than on the left (p = 0.046) and, unlike Adachi 2 , no differences regarding gender have been observed (p > 0.05).
Therefore, a precise knowledge of possible variation of the carotid axis morphology could help clinicians planning neck or facial surgery (laryngectomy, thyroidectomy), thrombo-endarterectomy, tumor angiogenesis, severe stenosis, aneurysms etc. [33][34][35][36][37][38][39][40] . The surgeon should bear in mind, just before starting any process, the www.nature.com/scientificreports/ normal, variations and possible anomalies of the carotid axis. These variations and anomalies are present in approximately 1.2% of cases. The normal anatomy, variations or anomalies could be detected easily by imaging techniques as preoperative conventional CT angiography, 3D CT angiography magnetic resonance angiography, digital subtraction angiography, doppler ultrasonography, etc. [33][34][35][36][37][38][39][40] . The correlation between imaging diagnosis techniques and anatomical studies based on cadaveric samples might be important for accuracy in planning (mini-)invasive carotid techniques. It allows a precise identification of variations or anomalous of the carotid arteries, thereby reducing the possible risk of intraoperative surgery 34,35,40 .

Conclusions
Variations in the calibres of the carotid arteries (for the common carotid artery 0.97 cm in average, for the internal 0.74 cm in average, and 0.49 cm for the external, respectively) and the carotid bifurcation level (reference point the hyoid bone with an average height of-0.33 cm, right carotid bifurcation at a higher level (x = − 0.19 cm.) than the left one (x = − 0.48 cm.)) are relevant for interventional radiology procedures and head and neck surgeries. A precise knowledge of the anatomical variations could help clinicians in the interpretation of the carotid system allowing them different possibilities in terms of diagnosis and treatment.
Therefore, we should take into account the anatomical variations in the appearance of the carotid system in radio diagnostic images and during surgical approaches, since this variability in the location or size of these structures could have negative consequences for our future patients.

Data availability
The datasets generated during and/or analysed during the current study are available from the corresponding author on reasonable request.