Architecture of head and neck soft tissues and associated entheses: An exploration of sexual dimorphism in, and population differences between, New Zealand and Thai individuals

Abstract

Understanding the musculoskeletal anatomy of soft tissues of the head and neck is important for surgical applications, biomechanical modelling and management of injuries, such as whiplash. Additionally, analysing sex and population differences in cervical anatomy can inform how biological sex and population variation may impact these anatomical applications. Although some muscles of the head and neck are well‐studied, there is limited architectural information that also analyses sex and population variation, for many small cervical soft tissues (muscles and ligaments) and associated entheses (soft tissue attachment sites). Therefore, the aim of this study was to present architectural data (e.g., proximal and distal attachment sites, muscle physiological cross‐sectional area, ligament mass, enthesis area) and analyse sex and population differences in soft tissues and entheses associated with sexually dimorphic landmarks on the cranium (nuchal crest and mastoid process) and clavicle (rhomboid fossa). Through the dissection and three‐dimensional analysis of 20 donated cadavers from New Zealand (five males, five females; mean age 83 ± 8 years; range 67–93 years) and Thailand (five males, five females; 69 ± 13 years; range 44–87 years), the following soft tissues and their associated entheses were analysed: upper trapezius, semispinalis capitis and the nuchal ligament (nuchal crest); sternocleidomastoid, splenius capitis and longissimus capitis (mastoid process); the clavicular head of pectoralis major, subclavius, sternohyoid and the costoclavicular (rhomboid) ligament (rhomboid fossa). Findings indicate that although muscle, ligament and enthesis sizes were generally similar to previously published data, muscle size was smaller for six of the eight muscles in this study, with only the upper trapezius and subclavius demonstrating similar values to previous studies. Proximal and distal attachment sites were largely consistent with the current research. However, some individuals (six of 20) had proximal upper trapezius attachments on the cranium, with most attaching solely to the nuchal ligament, contrasting with existing literature, which often describes attachment to the occipital bone. With respect to sexual dimorphism, the Thai sample exhibited more sex differences in muscle size than the New Zealand sample, but for enthesis size (area), both samples had the same amount of statistically significant sex differences (5 of 10). Additionally, some significant population differences were found when comparing muscle and enthesis size data between the New Zealand and Thai samples. Despite these findings, no sex or population differences were found for ligament size (mass) in either group. This paper presents new architectural data for several understudied areas of the head and neck, as well as providing analyses on sex and population differences, two areas that have limited representation in anatomy.

This paper presents architectural data and analyses sex and population differences in muscles, ligaments and entheses related to sexually dimorphic areas of the cranium and clavicle in New Zealand and Thai cadavers. The Thai sample was more sexually dimorphic in muscle size than the New Zealand sample, although both had similar sex differences for enthesis size. This study explores understudied areas of the head and neck and can be useful in applications such as surgery and injury management.

1. INTRODUCTION

Anatomical dissection research of the head and neck musculature often focuses on large, superficial structures such as the upper trapezius (Bayoglu et al., 2017; Borst et al., 2011; Johnson et al., 1994; Kamibayashi & Richmond, 1998; Van Ee et al., 2000) and sternocleidomastoid (Bayoglu et al., 2017; Borst et al., 2011; Kamibayashi & Richmond, 1998; Kennedy et al., 2017; Van Ee et al., 2000) with deep muscles, such as subclavius, receiving little attention (Bayoglu et al., 2017). The greater number of dissection studies on upper trapezius and sternocleidomastoid may be due to their superficial location, size or possibly their role in neck pain and dysfunction, such as whiplash associated disorder or concussion (Alsalaheen et al., 2019; Elliott et al., 2014; Reddy et al., 2021; Uthaikhup et al., 2017). However, a detailed understanding of muscle architecture (fascicle length, physiological cross‐sectional area [PCSA], pennation angle, attachment sites/entheses) (Lieber & Fridén, 2001; Vasavada et al., 2011) of deeper, smaller muscles like subclavius or longissimus capitis is also essential from a functional perspective (Keidan et al., 2021; Vasavada et al., 2011), as well as for informing surgical applications (Clark et al., 2019; Kadri & Al‐Mefty, 2007; Lee et al., 2014) and biomechanical modelling (Kamibayashi & Richmond, 1998; Östh et al., 2017; Van Ee et al., 2000; Zheng et al., 2013). Additionally, enthesis data can improve understanding of how soft tissues interact with the skeleton in archaeological contexts, such as studying entheseal changes on the dry bone to understand past populations (Churchill & Morris, 1998; Gresky et al., 2016; Hawkey & Merbs, 1995; Henderson et al., 2017; Lieverse et al., 2009; Molnar, 2006; Niinimäki, 2011; Noldner & Edgar, 2013; Villotte et al., 2016; Villotte & Knusel, 2013; Weiss et al., 2012; Wilczak, 1998; Wilczak et al., 2017). Furthermore, within the cervical region, ligament morphology (e.g., size, entheses) is understudied, with limited data on the nuchal ligament, for example (Humphreys et al., 2003).

Anatomical parameters can be influenced by factors such as sex, population and age (Alsalaheen et al., 2019; Frontera et al., 2000; Kim et al., 2021). Sexual dimorphism (Williams & Carroll, 2009) is evident in some cervical muscles, with males generally exhibiting larger muscle sizes than females (Alsalaheen et al., 2019; Keidan et al., 2021; Kennedy et al., 2017; Mayoux‐Benhamou et al., 1995; Rankin et al., 2005; Valera‐Calero et al., 2020). However, sex differences are typically not explored in anatomical studies and further knowledge of this may contribute to a better understanding of the variation between sexes in the risk and management of neck injuries (Alsalaheen et al., 2019), identifying the role that sex differences may play in clinical and functional applications (Keidan et al., 2021), and recognising variation between male and female sexes, not gender (Christensen et al., 2014, 2019; Hollimon, 2017; Konigsberg & Hens, 1998), although it is important to acknowledge that biological sex is not a simple dichotomy (Garofalo & Garvin, 2020).

Understanding population differences within anatomical data is also important to ensure that published information is applicable to diverse populations around the world. Interestingly, population variation is largely understudied in anatomy (Kim et al., 2021), despite its strong representation in the anthropological literature (Bidmos & Asala, 2004; de Paiva & Segre, 2003; Franklin et al., 2006; Orish et al., 2014; Suazo et al., 2009; Tallman, 2019; Walker, 2008). Anatomical studies on a range of populations are common in some areas, such as studies of anatomical variation in the sternocleidomastoid (Anıl et al., 2017; Cherian & Nayak, 2008; de Amorim et al., 2010; Dupont et al., 2018; Fazliogullari et al., 2010; Ferreira‐Arquez, 2018; Heo et al., 2020; Kim et al., 2015; Natsis et al., 2009; Oh et al., 2019; Raikos et al., 2012; Saha et al., 2014; Sirasanagandla et al., 2012). However, larger studies often do not address the role population differences may play in anatomical variation and many do not specify the population(s) represented in their studies (Johnson et al., 1994; Kamibayashi & Richmond, 1998; Van Ee et al., 2000). Although some dissection studies provide architectural data on muscles of the head and neck (Bayoglu et al., 2017; Borst et al., 2011; Kamibayashi & Richmond, 1998; Van Ee et al., 2000), it is clear that further research is needed on a broader range of cervical muscles, ligaments and associated entheses that also consider sex and population variation.

This research is part of a larger interdisciplinary project examining anthropological and anatomical relationships of three sexually dimorphic skeletal landmarks used for sex estimation in anthropology: the nuchal crest and mastoid process of the cranium, and the rhomboid fossa of the clavicle. Although these landmarks are relevant in anthropological sex estimation—where robust traits are associated with males and gracile features with females (Buikstra & Ubelaker, 1994; Rogers et al., 2000; Walker, 2008)—the associated soft tissues of the head and neck are not well understood as they relate to these skeletal traits. Therefore, these structures were chosen not only to test these relationships but also because they represent different muscles and ligaments that move and/or stabilise the head and neck (Houseman et al., 2000; Sinnatamby & Last, 2011; Standring, 2021), which may contribute to overall skeletal robusticity. Although interdisciplinary research across anatomy and anthropology is important (Mello‐Gentil & Souza‐Mello, 2021; Rissech, 2021), no existing research has assessed sexual dimorphism and population variation of the soft tissues and entheses of the head and neck. Therefore, this paper aims to present architectural data and analyse sex and population differences in cervical soft tissues (muscles and ligaments) and entheses related to these cranial and clavicular landmarks.

2. METHODS

Architectural data were collected from the dissection and three‐dimensional (3D) scanning of New Zealand and Thai cadaveric samples (by JSD, PhD student with 4 years of experience in dissection). For the cranial landmarks, three soft tissues associated with the nuchal crest (upper trapezius, semispinalis capitis, nuchal ligament) and mastoid process (sternocleidomastoid, splenius capitis, longissimus capitis) were included. For the rhomboid fossa, the clavicular head of the pectoralis major, subclavius, sternohyoid and the costoclavicular (rhomboid) ligament, which attaches directly to the rhomboid fossa (Tubbs et al., 2009) were examined.

2.1. Samples

Twenty recently deceased individuals bequeathed to the Department of Anatomy, University of Otago, New Zealand (10) and Khon Kaen University, Thailand (10) were used for this study. The New Zealand embalmed cadaver sample included five females and five males (mean age 83 ± 8 years, range 67–93 years), and the Thai fresh cadaver sample also had five females and five males (mean age 69 ± 13 years, range 44–87 years). Information about population affinity, beyond generalised geographic regions, was not available; however, generally the New Zealand sample represented people of European descent and the Thai individuals were of Asian descent. Ethical approval was obtained from the University of Otago Ethics Committee (reference: H18/113) and Khon Kaen University (reference: HE621296).

Individuals from the New Zealand sample were embalmed with a phenoxyethanol‐based solution (Crosado et al., 2020). Thai individuals were unembalmed and kept in a freezer between dissection sessions to slow decomposition, given 30 min to reach room temperature prior to dissection, and were fully dissected within 3 days. Although there may be limitations to comparing data from cadaveric samples with different preservation processes, previous studies suggest the differences in tissue size may be minimal (Crosado et al., 2020; Cutts, 1988) and the importance of exploring population variation in this study outweighed these limitations.

Exclusion criteria encompassed factors that prevented data collection such as previous surgical intervention, pathological conditions, antemortem trauma and postmortem damage. Based on this, there were two exclusions: (1) the right clavicular head of pectoralis major from a New Zealand male, which exhibited extensive postmortem damage and (2) the right subclavius enthesis from a Thai male, which was unable to be accurately identified due to a complete, but fully remodelled, fracture of the mid‐clavicle.

2.2. Dissection

Architectural data collected for muscles included attachment sites, size (fascicle length, volume, PCSA), and fascicle pennation angle (Lieber & Fridén, 2001; Vasavada et al., 2011). Data for ligaments included length, width (proximal and distal, where applicable) and mass (Gu et al., 2020; Tubbs et al., 2009). Additionally, enthesis surface area was measured digitally using 3D scans. As the skeletal landmarks were used as the main points of reference, attachments associated with the landmarks are referred to as proximal and attachments elsewhere are labelled distal. Prior to the start of data collection, dissection of the left deltoid muscle from a New Zealand male was completed to establish the protocol for this study.

Eight muscles and two ligaments (one bilateral and one unilateral) were dissected on both right and left sides progressing from superficial (upper trapezius, sternocleidomastoid and pectoralis major) to deep (splenius capitis, longissimus capitis, semispinalis capitis and the nuchal ligament). Following their dissection, the head and clavicles were removed to dissect the sternohyoid, subclavius and costoclavicular ligament.

Each muscle was dissected by fascicle, defined as a bundle of muscle fibres that attach at a common tendinous location (Lieber & Fridén, 2001). After removal, fascicles were trimmed of tendinous material and excess fascia, and length and mass were recorded. To collect muscle size data, PCSA was calculated using the formula PCSA = fascicle volume/length (Johnson et al., 1994; Kennedy et al., 2017; Langenderfer et al., 2004; Vasavada et al., 2011; Verstappen, 2015). Mass was converted to volume, per the muscle density constant of mass/1.0576 g/cm² described in Klein Breteler et al. (1999). The PCSA values of each fascicle were totalled to give the overall PCSA of each muscle, and total muscle volume was calculated by summing the converted volume of each fascicle.

The pennation angle of individual fascicles was measured by placing a transparent plastic protractor over the muscle, to determine the angle at which each fascicle lay in relation to the axis of force for the muscle (Vasavada et al., 2011), which often followed the midline of the muscle. This was done prior to fascicle removal, and the axis of force was estimated based on the attachment sites of each muscle (Kamibayashi & Richmond, 1998). Pennation angle can be used to calculate PCSA, but its inclusion accounts for possible loss of force due to the angle of a muscle (Vasavada et al., 2011). In this study, muscle size is of greater interest than force and, therefore, pennation angle was not included in PCSA calculations, in line with previous dissection studies (Johnson et al., 1994, Kennedy et al., 2017, Langenderfer et al., 2004, Vasavada et al., 2011, Verstappen, 2015).

Ligaments were removed in their entirety, and their length, width, and mass were measured. In the case of the nuchal ligament, which does not exhibit a uniform rectangular shape, both proximal and distal widths were measured.

2.3. Enthesis area measurements

During removal of each muscle or ligament, the attachment sites were outlined with a grease pencil to facilitate 3D scanning and digital enthesis area measurements. All entheses associated with the skeletal landmarks (landmark entheses) were outlined (Figure 1), and some distal attachment sites (distal entheses) were also outlined: the clavicular attachment of the upper trapezius, the clavicular and manubrial attachments of the sternocleidomastoid (Figure 1), the clavicular head of pectoralis major on the humerus, and the sternohyoid on the manubrium, if present. The remaining distal entheses (semispinalis capitis, the nuchal ligament, splenius capitis, longissimus capitis, subclavius and the costoclavicular ligament) were not included, due to small attachment areas such as the spinous or transverse processes.

FIGURE 1.

Example from a New Zealand male with (a) outlined landmark entheses associated with the mastoid process (lateral view of right mastoid process) and (b) outlined distal entheses associated with sternocleidomastoid (anterior view of medial clavicles and manubrium). A, anterior; I, inferior; L, left; P, posterior; R, right; S, superior.

The equipment for the 3D scans included an iPad Pro and a Structure Sensor 3D Scanner (Occipital Inc; Model SA17), which was used in conjunction with the application Scanner—Structure SDK. Once scans of the relevant entheses had been obtained, they were uploaded to Blender (Version 2.80, Blender Foundation, Netherlands) and the area of each enthesis outline was digitally measured. This provided a surface area of each outlined enthesis in a 3D space to account for the various morphological features of the underlying bone. This method was validated to ensure the 3D scans were to scale and that Blender provided accurate area measurements.

The intra‐observer reliability of the enthesis area measurements taken in Blender was assessed using repeat measurements from six individuals (three Thai—two males and one female—and three New Zealand—two females and one male). The second set of measurements was taken 6 months following initial data collection and were repeated by digitally remeasuring the area of all outlined entheses from the original 3D scans, with the investigator (JSD) blinded to previous digital outlines and measurements.

2.4. Statistical analyses

Means and standard deviations were calculated in Microsoft Excel for architectural data collected from muscles, ligaments and entheses. The data for muscle PCSA, ligament mass and enthesis area were then normalised for height using the formula of normalised value = original data point/height ² (Jaric, 2002). This provided analysis of the data with the size variable removed, which can offer a useful perspective when analysing information across sexes and populations. Both standard and normalised data are presented; however, as size differences are a main factor of sexual dimorphism (Cabo et al., 2012; Kurki, 2011; Spradley & Stull, 2018), analyses using standard data are emphasised. Data were then transferred to SPSS (IBM Corp, Version 27) with non‐parametric Mann–Whitney U tests (Altman et al., 1983) used to analyse differences between muscle, ligament or enthesis size, and sex and population. The level of statistical significance was set at p < 0.05.

The intra‐observer reliability of enthesis area measurements taken in Blender was assessed by intraclass correlation coefficients (ICCs). Excellent reliability was indicated by ICC values over 0.9, good reliability ranged from 0.75 to 0.9, moderate from 0.5 to 0.75, and poor reliability was less than 0.5 (Koo & Li, 2016).

3. RESULTS

This study identified the architectural morphology and sex and population differences in New Zealand and Thai cadaveric samples for muscles (Table 1), ligaments (Table 2) and entheses (Tables 3 and 4) associated with the nuchal crest, mastoid process and rhomboid fossa. Architectural data included proximal and distal attachment sites, size and any observed anatomical variation. Preliminary testing found no statistically significant differences between sides and, therefore, data from the right and left sides were combined for analyses of sex and population differences.

TABLE 1.

Muscle architectural data and sex and population differences for New Zealand and Thai individuals.

Muscle	Sample	Sex	N	Architecture				Sex differences		Population differences
				Fascicle length (cm)	Muscle volume (cm3)	Muscle PCSA (cm2)	Pennation angle (°)	p	p Normalised	p	p Normalised
Upper trapezius	New Zealand	M	10	11.7 (2.1)	22.7 (4.2)	2.1 (0.6)	0–50	0.912	0.393	0.231	0.049
		F	10	10.7 (0.7)	22.8 (11.4)	2.2 (1.0)	0–50
	Thailand	M	10	13.9 (1.1)	43.5 (13.0)	3.2 (1.1)	0–40	0.004	0.011
		F	10	11.8 (1.2)	22.1 (9.0)	1.9 (0.7)	0–50
Semispinalis capitis	New Zealand	M	10	12.8 (2.0)	24.6 (5.9)	2.0 (0.5)	0–30	0.190	0.529	0.989	0.277
		F	10	11.5 (1.7)	20.6 (6.6)	1.9 (0.6)	0–40
	Thailand	M	10	14.6 (0.8)	39.6 (9.9)	2.8 (0.8)	0–25	<0.001	<0.001
		F	10	12.8 (1.1)	18.1 (2.1)	1.4 (0.2)	0–20
Sternocleidomastoid	New Zealand	M	10	14.2 (1.4)	17.6 (2.8)	1.2 (0.1)	0–20	0.684	0.063	0.904	0.114
		F	10	12.5 (0.7)	15.6 (4.8)	1.3 (0.5)	0–20
	Thailand	M	10	15.7 (1.0)	33.0 (8.3)	2.1 (0.6)	0–25	<0.001	<0.001
		F	10	14.2 (1.2)	13.3 (2.6)	0.9 (0.1)	0–20
Splenius capitis	New Zealand	M	10	10.4 (1.1)	12.5 (2.6)	1.2 (0.2)	0–20	0.739	0.853	0.529	0.086
		F	10	10.0 (0.9)	11.3 (4.9)	1.1 (0.4)	0–10
	Thailand	M	10	13.1 (0.7)	23.4 (6.8)	1.8 (0.5)	0	<0.001	<0.001
		F	10	12.4 (0.8)	10.9 (2.5)	0.9 (0.2)	0
Longissimus capitis	New Zealand	M	10	9.8 (3.3)	3.3 (2.1)	0.3 (0.1)	0–10	0.190	1.000	0.142	0.008
		F	10	10.4 (1.5)	2.7 (1.0)	0.3 (0.1)	0
	Thailand	M	10	14.0 (2.5)	5.6 (1.8)	0.4 (0.1)	0	0.063	0.436
		F	10	11.8 (3.1)	3.5 (0.9)	0.3 (0.1)	0
Pectoralis major (clavicular head)	New Zealand	M	9	14.5 (1.7)	26.1 (12.0)	1.8 (0.8)	0–20	0.004	0.065	0.901	0.569
		F	10	14.2 (0.9)	15.4 (6.8)	1.1 (0.4)	0–10
	Thailand	M	10	16.0 (1.8)	29.2 (6.3)	1.9 (0.4)	0–10	<0.001	<0.001
		F	10	14.7 (1.6)	13.9 (4.5)	0.9 (0.2)	0
Sternohyoid	New Zealand	M	10	10.7 (0.7)	3.5 (0.7)	0.3 (0.1)	0	0.190	0.739	0.165	0.698
		F	10	10.2 (1.1)	2.9 (0.9)	0.3 (0.1)	0
	Thailand	M	10	13.8 (1.2)	4.6 (1.3)	0.3 (0.1)	0	0.001	0.009
		F	10	12.3 (1.2)	2.4 (0.6)	0.2 (0.04)	0
Subclavius	New Zealand	M	10	7.5 (1.7)	3.2 (0.8)	0.4 (0.1)	0	0.105	0.796	0.383	1.000
		F	10	7.4 (1.1)	2.6 (1.3)	0.3 (0.1)	0
	Thailand	M	10	7.3 (2.0)	3.5 (2.1)	0.5 (0.2)	0	0.007	0.029
		F	10	7.0 (1.1)	1.9 (0.8)	0.3 (0.1)	0

Note: Bold values = statistical significance (p < 0.05). Mean values and (SD) presented for length, volume and PCSA; range provided for fascicle pennation angle; data from right and left sides combined. Results from both standard and height‐normalised data are shown.

Abbreviations: F, female; M, male; N, number of muscles; PCSA, physiological cross‐sectional area.

TABLE 2.

Ligament architectural data and sex and population differences for New Zealand and Thai individuals.

Ligament	Sample	Sex	N	Architecture				Sex differences		Population differences
				Length (cm)	Width (cm)		Mass (g)
					Proximal	Distal		p	p Normalised	p	p Normalised
Nuchal ligament	New Zealand	M	5	13.3 (1.4)	3.8 (1.3)	1.0 (0.4)	6.3 (1.6)	0.841	0.310	0.393	0.165
		F	5	12.1 (1.9)	2.8 (0.9)	0.7 (0.3)	7.2 (3.1)
	Thailand	M	5	14.0 (1.5)	3.0 (1.2)	1.3 (0.3)	8.9 (3.0)	0.222	0.690
		F	5	12.3 (1.7)	2.9 (0.7)	1.0 (0.6)	6.9 (2.3)
Costoclavicular ligament	New Zealand	M	10	3.8 (1.0)	1.8 (0.6)		2.5 (1.3)	0.280	0.631	0.242	0.383
		F	10	3.1 (0.4)	1.5 (0.4)		1.8 (1.0)
	Thailand	M	10	3.0 (0.7)	1.6 (0.4)		1.7 (0.8)	0.529	0.912
		F	10	2.8 (0.6)	1.5 (0.3)		1.5 (0.8)

Note: Bold values = statistical significance (p < 0.05). Mean values and SD presented for standard data. Data from the right and left sides combined for the costoclavicular ligament. Results from both standard and height‐normalised data are shown.

Abbreviations: F, female; M, male; N, number of ligaments.

TABLE 3.

Enthesis architectural data and sex and population differences for landmark entheses in New Zealand and Thai individuals.

Enthesis	Sample	Sex	N	Architecture	Sex differences		Population differences
				Enthesis area (cm2)	p	p Normalised	p	p Normalised
Upper trapezius Cranial enthesis	New Zealand	M	1	0.5	0.500	0.500	0.762	0.61
		F	3	2.7 (1.7)
	Thailand	M	6	2.6 (1.0)	—	—
		F	0	—
Semispinalis capitis Cranial enthesis	New Zealand	M	10	8.8 (1.4)	0.089	0.529	0.414	0.231
		F	10	7.9 (0.8)
	Thailand	M	10	8.4 (1.4)	0.218	0.631
		F	10	7.7 (1.1)
Nuchal ligament Cranial enthesis	New Zealand	M	5	3.2 (0.6)	0.841	0.421	0.796	0.631
		F	5	3.2 (1.4)
	Thailand	M	5	2.8 (0.8)	0.421	0.095
		F	5	3.3 (0.9)
Sternocleidomastoid Cranial enthesis	New Zealand	M	10	12.8 (2.7)	0.009	0.123	0.174	0.529
		F	10	9.0 (2.2)
	Thailand	M	10	10.3 (2.9)	0.280	0.796
		F	10	9.0 (2.6)
Splenius capitis Cranial enthesis	New Zealand	M	10	4.9 (1.2)	0.052	0.579	0.327	0.758
		F	10	3.8 (1.3)
	Thailand	M	10	3.9 (0.7)	0.579	0.684
		F	10	4.0 (1.5)
Longissimus capitis Cranial enthesis	New Zealand	M	10	1.8 (0.6)	0.015	0.052	0.052	0.192
		F	10	1.3 (0.3)
	Thailand	M	10	1.4 (0.3)	0.029	0.075
		F	10	1.0 (0.3)
Pectoralis major (clavicular head) Clavicle enthesis	New Zealand	M	9	11.3 (7.1)	0.006	0.017	0.792	0.351
		F	10	5.4 (0.8)
	Thailand	M	10	7.7 (1.3)	0.005	0.143
		F	10	5.8 (1.2)
Sternohyoid Clavicle enthesis	New Zealand	M	9	4.6 (2.1)	0.604	0.780	0.012	0.042
		F	10	4.0 (1.2)
	Thailand	M	10	3.2 (1.0)	0.237	0.573
		F	8	2.6 (0.9)
Subclavius Clavicle enthesis	New Zealand	M	10	7.9 (2.6)	0.190	0.393	0.011	0.026
		F	10	5.6 (2.1)
	Thailand	M	9	6.0 (2.0)	0.028	0.053
		F	10	4.0 (1.3)
Costoclavicular ligament Clavicle enthesis	New Zealand	M	10	4.1 (1.9)	0.035	0.075	0.028	0.046
		F	10	2.6 (0.6)
	Thailand	M	10	2.8 (0.7)	0.004	0.023
		F	10	1.8 (0.5)

Note: Bold values = statistical significance (p < 0.05). Mean values and SD presented for standard data. Data from the right and left sides combined, where applicable. Results from both standard and height‐normalised data are shown.

Abbreviations: F, female; M, male; N, number of entheses.

TABLE 4.

Enthesis architectural data and sex and population differences for distal entheses in New Zealand and Thai individuals.

Enthesis	Sample	Sex	N	Architecture	Sex differences		Population differences
				Enthesis area (cm2)	p	p Normalised	p	p Normalised
Upper trapezius Clavicle enthesis	New Zealand	M	10	10.5 (3.1)	0.280	0.579	0.678	0.383
		F	10	9.5 (1.6)
	Thailand	M	10	10.6 (1.5)	0.063	0.190
		F	10	8.8 (2.0)
Sternocleidomastoid Clavicle enthesis	New Zealand	M	10	3.4 (1.3)	0.529	0.481	0.211	0.046
		F	10	3.1 (1.0)
	Thailand	M	10	5.2 (1.4)	0.002	0.003
		F	10	2.8 (1.3)
Sternocleidomastoid Manubrium enthesis	New Zealand	M	10	2.3 (0.8)	0.052	0.353	0.758	0.127
		F	10	1.6 (0.5)
	Thailand	M	10	2.0 (0.6)	0.481	0.853
		F	10	1.9 (0.5)
Pectoralis major (clavicular head) Humerus enthesis	New Zealand	M	9	6.4 (2.0)	<0.001	0.113	0.175	0.813
		F	10	4.0 (0.4)
	Thailand	M	10	4.7 (1.0)	0.315	0.912
		F	10	3.9 (1.2)
Sternohyoid Manubrium enthesis	New Zealand	M	4	3.8 (3.0)	1.000	1.000	0.159	0.14
		F	6	2.5 (0.8)
	Thailand	M	8	4.4 (2.0)	0.368	0.368
		F	4	3.4 (1.4)

Note: Bold values = statistical significance (p < 0.05). Mean values and SD are presented for standard data. Data from the right and left sides combined. Results from both standard and height‐normalised data are shown.

Abbreviations: F, female; M, male; N, number of entheses.

3.1. Muscles

3.1.1. Upper trapezius

The proximal attachments of the upper trapezius were predominantly non‐skeletal, and, in many instances, the muscle attached to the nuchal ligament without extending to the occiput (Figure 2). Of the 20 individuals, only six had attachments on the cranium, which ranged from small areas near the external occipital protuberance to broader areas along the occipital bone, situated at or above the superior nuchal line. Proximal attachments at the nuchal ligament ranged from the level of C1 to C7, with one individual (New Zealand female) exhibiting a bony attachment at the C7 and T1 spinous processes.

FIGURE 2.

Posterior view of the neck and head of a New Zealand female exhibiting cranial attachment of the upper trapezius (a, b) and a second New Zealand female demonstrating a lack of cranial attachment (c, d). (a, c) The trapezius prior to removal, and (b, d) the cranium following removal of the muscle, with cranial attachments outlined in black, if present (b). I, inferior; L, left; R, right; S, superior.

The distal border of the upper trapezius was defined by its clavicular attachment, most commonly at the superior and posterior aspects of the lateral clavicle. This attachment extended as far as the middle clavicle (one New Zealand female) or the muscle attached at only the posterolateral clavicle (one New Zealand male). Additionally, two small fascicles branched from the upper trapezius muscle and ran along the posterior clavicle, connecting medially on the bone (one New Zealand male) (Figure 3).

FIGURE 3.

Anterior view of the right clavicle from a New Zealand male exhibiting anatomical variation of the right upper trapezius. White arrows indicate fascicles branching from the distal attachment and extending medially along the posterior clavicular. I, Inferior; L, lateral; M, medial; S, superior.

Muscle PCSA of the upper trapezius ranged from 1.9 cm² for Thai females to 3.2 cm², for Thai males, with New Zealand male and female values in between, and nearly the same (2.1 and 2.2 cm², respectively). Pennation angles were similar across all groups, ranging from 0–50° with the exception of Thai males, with a range of 0–40°.

3.1.2. Semispinalis capitis

Proximally, semispinalis capitis consistently attached inferior to the nuchal crest and often directly at the superior nuchal line, extending from the external occipital protuberance to the lateral area of the occipital bone. In some individuals, this muscle is also attached to the nuchal ligament (one New Zealand male) and the surrounding deep fascia (New Zealand female, Thai female, Thai male).

Distally, semispinalis capitis attached to transverse processes, from C2 to T7. In seven individuals (New Zealand male and female, three Thai males, two Thai females), some fascicles were also attached to the spinous processes, and one New Zealand male exhibited a unique attachment to the nuchal ligament, at the level of C4.

The mean PCSA of semispinalis capitis ranged from 1.4 cm² (Thai females) to 2.8 cm² (Thai males), with New Zealand males and females in the mid‐range. Pennation angle had the largest range for New Zealand females (0–40°) and smallest for Thai females (0–20°).

3.1.3. Sternocleidomastoid

The sternocleidomastoid proximal attachment was relatively consistent across individuals, covering the mastoid process (anterior aspect, superior to inferior) and extending posteriorly onto the occiput at the superior nuchal line. In one Thai female, a portion of the muscle attached to the deep fascia at the occipital bone on both sides.

Distally, the sternocleidomastoid attached on both the superior medial clavicle and the superior aspect of the anterior manubrium; however, there were two instances (one New Zealand male, one Thai female) where the clavicular attachments also extended to the posterior aspect of the bone.

Mean PCSA ranged from 0.9 cm² (Thai females) to 2.1 cm² (Thai males). Pennation angle was the same for all groups at 0–20°, except for Thai males at 0–25°.

3.1.4. Splenius capitis

For all individuals, the splenius capitis proximal attachment was directly inferior to the sternocleidomastoid and extended the length of the mastoid process (posterior superior aspect to anterior inferior tip) onto the occiput at, or inferior to, the superior nuchal line.

The distal attachment for most individuals was along the nuchal ligament (between C2 and C7) and at the spinous processes (C7 to T3). However, five individuals (New Zealand: two males, one female; Thai: one male, one female) had no bony attachments, and two (New Zealand female, Thai male) had vertebral attachments on only one side.

Thai females had the smallest mean PCSA (0.9 cm²) and Thai males had the largest (1.8 cm²), with New Zealand males and females in the middle range. In 17 individuals, splenius capitis was not pennated, but pennation angle ranged between 0 and 20° for one New Zealand male, and 0–10° for two New Zealand females.

3.1.5. Longissimus capitis

Proximally, longissimus capitis consistently attached to the mastoid process (posterior and inferior aspects), inferior to splenius capitis. Distally, this muscle attached to cervical and thoracic transverse processes (C2 through T7), although variation was evident. Nine individuals (five New Zealand and four Thai) had attachments at only the cervical vertebrae (C3 to C7) and one (Thai female) at only the thoracic vertebrae (T1 and T3–T7).

The mean muscle PCSA of longissimus capitis was mostly consistent across sex and population (0.3 cm²), with the exception of Thai males (0.4 cm²). This muscle was only pennated in one New Zealand male (range 0–10°, left side).

3.1.6. Pectoralis major (clavicular head)

The clavicular head of pectoralis major was defined by its proximal attachment at the clavicle, generally on the anterior medial aspect. However, in four New Zealand individuals, the muscle extended further laterally or along the superior or inferior aspects of the clavicle. The distal attachment was consistent for all individuals at the anterior proximal shaft of the humerus, along the crest of the lesser tubercle (the medial lip of the bicipital groove) (White et al., 2011), just above the midline.

The mean PCSA for the clavicular head of the pectoralis major was higher in Thai males (1.9 cm²) compared with females (0.9 cm²), with values for the New Zealand individuals in the middle. Pennation angle ranged between 0 and 20°, with New Zealand males exhibiting the largest range (0–20°) and Thai females none (0°).

3.1.7. Sternohyoid

Sternohyoid attached proximally to the anterior portion of the hyoid body in all individuals. Distally, this muscle generally attached at the clavicle and manubrium, but nine individuals across all groups exhibited attachments at only the clavicle. Additionally, in one New Zealand male, the sternohyoid solely attached to the manubrium on the right side and the clavicle and manubrium on the left. The sternohyoid had consistency in size across individuals, with a mean PCSA of 0.3 cm² for all groups, except the Thai females (0.2 cm²), and no pennation.

3.1.8. Subclavius

Proximally, the subclavius is generally attached along the length of the inferior middle clavicle. However, additional scapular attachments were noted (Figure 4), where five individuals (one New Zealand male, two Thai males and two Thai females) had unilateral or bilateral attachments at the coracoid process. The distal attachment was consistently at the superior aspect of the first rib. Thai males had the highest mean PCSA (0.5 cm²), with females from both samples exhibiting the lowest values (0.3 cm²). This muscle was not pennated.

FIGURE 4.

Posterior view of the right neck and shoulder region from a Thai female showing the anatomical variation of the right subclavius with an additional attachment to the scapula (white arrow). I, inferior; L, left; R, right; S, superior

3.1.9. Sex and population differences

Statistically significant sex differences were evident for nearly all muscles, but mostly in the Thai sample (Table 1). Except longissimus capitis, the mean PCSA of all muscles was significantly larger in Thai males compared with Thai females, for both standard and normalised data. In the New Zealand sample, the clavicular head of pectoralis major (standard data) was the only muscle that was significantly larger in males compared with females (p = 0.004).

There were no statistically significant differences between populations for standard PCSA data (Table 1). When normalised for height, the upper trapezius and longissimus capitis exhibited population differences, with mean PCSA significantly larger in the Thai sample.

3.2. Ligaments

3.2.1. Nuchal ligament

The nuchal ligament was consistently attached superiorly at the external occipital protuberance, extending along the occipital bone to the foramen magnum. Additional attachments were also noted at the posterior arch of C1 (eight individuals across both samples) and the C2 spinous processes (three Thai males and two Thai females).

Inferiorly, the most consistent attachment was the C7 spinous process (16 of 20 individuals); however, one of these individuals (Thai male) had an additional attachment at both C6 and C7. Additionally, three males (two New Zealand and one Thai) had inferior attachments at only C6 and one Thai female at only C5. The mean mass of the nuchal ligament ranged from 6.3 g for New Zealand males to 8.9 g for Thai males, while females from both samples represented the mid‐range.

3.2.2. Costoclavicular ligament

Individuals across all groups had the same proximal attachment of the costoclavicular ligament (inferior medial clavicle). The ligament attached distally at the superior manubrium, although in one Thai female, this attachment encompassed both the manubrium and the first rib. Mean ligament mass ranged between 1.5 and 2.5 g, with New Zealand individuals exhibiting the highest values and Thai individuals the lowest.

3.2.3. Sex and population differences

There were no significant differences across sex or population for ligament mass for either the nuchal or costoclavicular ligaments (Table 2).

3.3. Entheses

3.3.1. Landmark entheses

For area measurements of landmark entheses on the cranium (Table 3), little data were available for the upper trapezius, due to limited skeletal attachments (10 total), although, for most of the observed attachments, mean enthesis area measurements were similar at 2.6 cm² (Thai males) and 2.7 cm² (New Zealand females). For semispinalis capitis, females exhibited the smallest landmark entheses and males the largest, with mean areas ranging from 7.7 cm² (Thai females) to 8.8 cm² (New Zealand males). The nuchal ligament landmark enthesis had similar area measurements across all groups (3.2 to 3.3 cm²) with the exception of Thai males (2.8 cm²). New Zealand males had a larger sternocleidomastoid landmark enthesis (12.8 cm²) than the others (Thai males 10.3 cm², both female groups 9.0 cm²). Additionally, New Zealand males had the largest areas for both splenius capitis (4.9 cm²) and longissimus capitis (1.8 cm²) landmark entheses, and the female groups had the smallest, although splenius capitis was slightly larger in Thai females (4.0 cm²; New Zealand 3.8 cm²), and longissimus capitis was larger in New Zealand females (1.3 cm²; Thai 1.0 cm²).

Area measurements for clavicular landmark entheses were largest for the New Zealand male group, for most entheses (Table 3). For the clavicular head of the pectoralis major, areas ranged from 5.4 cm² (New Zealand females) to 11.3 cm² (New Zealand males), with the Thai group representing the middle values. Furthermore, the entheseal area for sternohyoid was smallest in the Thai and largest in the New Zealand samples (range of 2.6 to 4.6 cm²). For subclavius and the costoclavicular ligament entheses, Thai females had the smallest areas (subclavius 4.0 cm²; costoclavicular ligament, 1.8 cm²) and New Zealand males the largest (subclavius 7.9 cm²; costoclavicular ligament 4.1 cm²).

3.3.2. Distal entheses

For distal entheses (Table 4), the area of the upper trapezius distal enthesis on the clavicle ranged from 8.8 cm² (females, both samples) to 10.6 cm² (males, both samples). The distal attachments of sternocleidomastoid were larger in males than females (New Zealand and Thai) for both the clavicle (range 2.8–5.2 cm²) and manubrium (1.6–2.3 cm²) entheses. For the distal enthesis of the clavicular head of pectoralis major on the humerus, New Zealand males represented the largest area (6.4 cm²), and Thai females the smallest (3.9 cm²). The sternohyoid distal enthesis on the manubrium was larger in males from both samples, ranging from 2.5 cm² (New Zealand females) to 4.4 cm² (Thai males).

3.3.3. Sex and population differences

For cranial landmark entheses, there was evidence for sex differences for only sternocleidomastoid and longissimus capitis (Table 3). In the New Zealand sample, males exhibited significantly larger standard area measurements than females for sternocleidomastoid (p = 0.009) and longissimus capitis (p = 0.015) landmark entheses. In the Thai sample, there were significant sex differences for the longissimus capitis landmark enthesis (p = 0.029) standard data, with males exhibiting larger values.

More variation between sexes was seen in landmark entheses on the clavicle, for both samples (Table 3). New Zealand males had significantly larger entheses than New Zealand females for the pectoralis major (clavicular head) (p = 0.006) and costoclavicular (p = 0.035) landmark entheses. When normalised for height, these differences were evident only for the pectoralis major landmark enthesis (p = 0.017). For the Thai group, males exhibited larger landmark enthesis areas for pectoralis major (p = 0.005), subclavius (p = 0.028) and the costoclavicular ligament (p = 0.004). With normalised data, only the costoclavicular ligament landmark enthesis showed a significant difference (p = 0.023).

For distal entheses (Table 4), there were few significant sex differences observed. For the New Zealand sample, males had larger enthesis areas (standard data) than females for the pectoralis major (clavicular head) enthesis at the humerus (p < 0.001). In the Thai sample, males had larger area measurements for the sternocleidomastoid clavicle enthesis, for both standard (p = 0.002) and normalised (p = 0.003) data.

For landmark entheses, no population differences were observed on the cranium, although several were apparent for the clavicle (Table 3). The New Zealand group exhibited significantly larger landmark enthesis areas on the clavicle (standard and normalised data) for the sternohyoid, the subclavius and the costoclavicular ligament. Evidence of a significant population difference was found for one distal enthesis (sternocleidomastoid clavicular enthesis) for normalised data, which was larger in the Thai sample (p = 0.046) (Table 4).

3.4. Intra‐observer reliability

All enthesis measurements had excellent intra‐observer reliability (Table 5), with ICCs higher than 0.9, with the exception of the upper trapezius landmark enthesis (ICC of 0.89).

TABLE 5.

Intra‐observer reliability for enthesis area measurements.

Muscle/ligament	Enthesis	N	Intraclass correlation coefficient	95% confidence interval
Upper trapezius	Cranial	6	0.89	0.21–0.99
	Clavicle	12	0.97	0.90–0.99
Semispinalis capitis	Cranial	12	0.95	0.83–0.99
Nuchal ligament	Cranial	12	0.99	0.92–0.99
Sternocleidomastoid	Cranial	12	0.99	0.94–0.99
	Clavicle	12	0.98	0.94–0.99
	Manubrium	12	0.98	0.77–0.99
Splenius capitis	Cranial	12	0.99	0.96–0.99
Longissimus capitis	Cranial	12	0.94	0.78–0.98
Pectoralis major (clavicular head)	Clavicle	12	0.99	0.99–0.99
	Humerus	12	0.98	0.91–0.99
Sternohyoid a	Clavicle	12	0.99	0.98–0.99
Subclavius	Clavicle	11	0.99	0.98–0.99
Costoclavicular ligament	Clavicle	12	0.99	0.97–0.99

Note: Bold values indicate excellent reliability. Poor = <0.5; Moderate = 0.5–0.75; Good = 0.75–0.9; Excellent = >0.9. Measurements taken by the first author.

Abbreviations: N, number of entheses.

^a Sternohyoid manubrium enthesis not included due to lack of data.

4. DISCUSSION

This study explored architecture, sexual dimorphism and population variation in muscles, ligaments and entheses associated with the nuchal crest, mastoid process and rhomboid fossa of the clavicle in New Zealand and Thai individuals. Overall, the architectural data for muscles were similar to previous research, although size was generally smaller (Bayoglu et al., 2017; Borst et al., 2011; Kamibayashi & Richmond, 1998; Van Ee et al., 2000) and new data are presented for ligament and enthesis architecture. Additionally, although few population differences were observed, the Thai sample was more sexually dimorphic than the New Zealand sample for muscle size, with a small number of sex differences observed for the enthesis area in both groups, but not for ligament mass. This research adds data to the existing literature on muscle architecture and sexual dimorphism of the head and neck, while also presenting new data in areas that are understudied in anatomy, including ligament and enthesis architecture and population variation.

4.1. Muscle, ligament and enthesis architecture

This research provides architectural data to supplement the existing literature on widely studied muscles, such as trapezius (Johnson et al., 1994) and sternocleidomastoid (Kennedy et al., 2017) and also presents novel data on under‐studied muscles, ligaments and entheses.

Soft tissue proximal and distal attachment sites were mostly consistent with those described in the literature, with some differences, including instances of anatomical variation in the upper trapezius (Bakkum & Miller, 2016), and are discussed below. Noting variation in attachments can be important for surgical applications, particularly in cases where knowledge of muscle locations and potential variation is key to performing surgery correctly and preventing injury (Clark et al., 2019; Kadri & Al‐Mefty, 2007; Lee et al., 2014).

For the upper trapezius, only 30% of individuals exhibited a proximal attachment at the occipital bone, with the remainder of the attachments solely at the nuchal ligament. These findings contrast with previous literature, where the upper trapezius is described to attach proximally to both the occipital bone and nuchal ligament (Abbott & Lucas, 1954; Allia & Gorniak, 2013; Bayoglu et al., 2017; Borst et al., 2011; Fielding et al., 1976; Giacomo et al., 2008; Houseman et al., 2000; Johnson et al., 1994; Kamibayashi & Richmond, 1998; Kawtharani & Hasan, 2018; Mercer & Bogduk, 2003; Phadnis & Bain, 2015; Rea, 2016; Van Ee et al., 2000; Vasavada et al., 2011; White et al., 2011). The lack of proximal skeletal attachments for the upper trapezius in this study differed from the expected outcome and may warrant further investigation of this attachment site in other populations. Limited skeletal attachments may affect the biomechanical function of the muscle, possibly providing decreased stability of the posterior neck, and could have implications for cervical injuries, such as whiplash (Au et al., 2016; Östh et al., 2017).

The nuchal ligament also exhibited some instances of unique proximal (C2) and distal (C5) attachments in the Thai sample. These contrast with previous descriptions, which identified proximal attachments at the cranium and C1 and distal attachments at C6 and C7 (Allia & Gorniak, 2013; Dean & Mitchell, 2002; Fielding et al., 1976; Humphreys et al., 2003; Johnson et al., 2000; Kadri & Al‐Mefty, 2007; Mercer & Bogduk, 2003; Mitchell et al., 1998; Nash et al., 2005; Ono et al., 2012; Takeshita et al., 2004; White et al., 2011). The presence of these variations in only Thai individuals may be evidence of some population variation in nuchal ligament attachment sites.

Both semispinalis capitis and longissimus capitis demonstrated variation in distal attachments. For semispinalis capitis, a number of individuals from both samples had attachments to the spinous process (six) and nuchal ligament (one), which varies from the standard attachment site of the transverse processes (Bayoglu et al., 2017; Borst et al., 2011; Rezasoltani et al., 1998; Vasavada et al., 2011; White et al., 2011). For longissimus capitis, attachments to only the cervical (nine individuals from both samples) or thoracic vertebrae (one Thai individual) were observed in this study, which contrasts with the expected attachment to both cervical and thoracic (Bayoglu et al., 2017; Borst et al., 2011; Elliott et al., 2018; Houseman et al., 2000; Kamibayashi & Richmond, 1998; Vasavada et al., 2011).

Mean muscle PCSA in this study was generally smaller than previously reported in dissection studies, which could be due to a number of factors such as age or preservation techniques. Of the eight muscles, the upper trapezius (1.9–3.2 cm²) and subclavius (0.3–0.5 cm²) demonstrated consistent muscle size with previous dissection data: the upper trapezius ranges between 2.0 and 3.5 cm² (Bayoglu et al., 2017; Borst et al., 2011; Johnson et al., 1994; Kamibayashi & Richmond, 1998) and subclavius has a reported value of 0.3 cm² (Bayoglu et al., 2017). The mean sizes for the remaining six muscles—semispinalis capitis, sternocleidomastoid, splenius capitis, longissimus capitis, clavicular head of pectoralis major and sternohyoid—were smaller than previous dissection studies. For example, the mean PCSAs of semispinalis capitis (1.4–2.8 cm²), is less than reported in four dissection studies: 3.1–5.5 cm² (Bayoglu et al., 2017; Borst et al., 2011; Kamibayashi & Richmond, 1998; Van Ee et al., 2000).

It is unclear why most muscles exhibited a smaller PCSA than those in previous dissection studies, particularly because the age ranges and methods were similar. While age of the cadaveric samples was similar across studies, age‐related changes, such as osteoarthritis and disc degeneration (Buikstra & Ubelaker, 1994; Ortner, 2003), were observed on most cervical vertebrae in both New Zealand and Thai samples and may be a contributing factor to the differences in muscle size. These degenerative changes can lead to reduced mobility in the neck (Frontera et al., 2000; Okada et al., 2011), and, combined with a general reduction in cervical range of motion with age (Pan et al., 2018), could be one explanation for decreased muscle size. Although Okada et al. (2011) found no relationship between muscle volume and disc degeneration, other muscle size variables, such as length, could be affected by degenerative changes in the cervical vertebrae. Neck pain is also related to reduced muscle size (Snodgrass et al., 2019; Uthaikhup et al., 2017) and this could have had additional influence, although it was not known whether any of the donors in this study had experienced neck pain while living, due to lack of medical history information.

Another potential reason that values may be smaller in this study is preservation techniques, as half of the individuals (those from New Zealand) were embalmed (Crosado et al., 2020); however, many previous dissection studies also used embalmed cadavers (Bayoglu et al., 2017; Borst et al., 2011; Johnson et al., 1994; Kamibayashi & Richmond, 1998; Kennedy et al., 2017). Additionally, the other half of the sample (from Thailand) was not embalmed, and fresh tissue can provide a more accurate representation of muscle morphology (Crosado et al., 2020). Therefore, it is not easy to identify the role that preservation techniques may have played in the differences in reported muscle size.

While findings for muscle PCSA can be compared with data from the existing literature, this is not possible for ligament mass and enthesis area data, given the lack of information on these architectural parameters. Although some studies provide costoclavicular ligament width/length measurements (Gu et al., 2020; Tubbs et al., 2009), measurement standards differ and, therefore, values are difficult to compare. Additionally, some papers present enthesis area data; however, due to varying methods, including measurements taken from dry bone (Noldner & Edgar, 2013) or using the triangular area of the enthesis (Heron's formula) (Lee et al., 2014), these data cannot be compared with the area measurements in this study.

4.2. Sex differences

In this study, sexual dimorphism was identified in muscles and entheses, but not ligaments, and there were fewer enthesis sex differences than originally anticipated, based on the knowledge that the skeletal morphology at these sites is sexually dimorphic (Buikstra & Ubelaker, 1994; Rogers et al., 2000; Walker, 2008).

For muscle data, sex differences appeared almost exclusively in the Thai sample (seven of eight muscles), but were only evident in one of the eight muscles in the New Zealand sample. The differences in muscular sexual dimorphism between the Thai and New Zealand groups could be due to several factors similar to those discussed above, such as age, population variation or preservation techniques.

Advanced age is a major factor that can impact data obtained from cadaveric samples and, although it is common for most studies of this nature (Kamibayashi & Richmond, 1998; Kennedy et al., 2017; Van Ee et al., 2000), the findings may not be applicable to younger individuals. As discussed above, muscle size typically decreases with older age for several reasons (Frontera et al., 2000), which is also true for the cervical muscles (Okada et al., 2011). Age is also a factor associated with sarcopenia, or age‐related muscle loss (Karlsson et al., 2015), meaning younger individuals typically have greater muscle mass and ligaments can also degrade with age (McCarthy & Hannafin, 2014). Although both samples consisted of older individuals, the Thai group was younger, with a mean age of 69 years, compared with 83 years for the New Zealand group. The older age of the New Zealand sample may have affected the data in this study, particularly compared with the Thai sample. Age and age‐related changes (Frontera et al., 2000; Okada et al., 2011; Pan et al., 2018; Uthaikhup et al., 2017) could have led to a decrease in muscle size for the New Zealand sample, which may have contributed to the limited sex differences, as males and females had similar muscle sizes. However, the females in the New Zealand sample were older (mean age of 85 years) than the males (mean age of 80 years), which would suggest that females should have noticeably smaller muscle sizes than males, but this was not the case. Furthermore, despite representing the older sample, the New Zealand individuals did not have the smallest muscles overall. In fact, the Thai females, who also represented the youngest group (mean age of 64 years), consistently exhibited the smallest muscle sizes. This likely indicates that sexual dimorphism is still observable in these data, which was expected. However, when examining male muscle sizes from both groups, age may have played a role in the differences seen between the Thai male (mean age of 73 years) and New Zealand male samples, as the Thai males consistently had larger muscle sizes. While advanced age is likely a contributing factor to variation in muscle size in this study, the data here demonstrate that its influence is not straightforward and that sexual dimorphism is still observable.

Another reason the muscular sexual dimorphism in the New Zealand sample differed from that in the Thai sample could be population variation. It has been demonstrated that Asian populations are more skeletally gracile than European populations (Garvin, 2012; İşcan & Steyn, 2013) and the same could be true for muscle size. However, while the female muscle data are consistent with skeletal trends (Garvin, 2012, İşcan & Steyn, 2013), as the New Zealand females were more muscularly robust than the Thai females, the Thai males were more robust than the New Zealand males. Additionally, although the Thai sample was more muscularly sexually dimorphic than the New Zealand sample, there were few significant differences found when testing population variation in muscle size. It is clear that population variation plays a role in skeletal research (Bidmos & Asala, 2004; de Paiva & Segre, 2003; Franklin et al., 2006; Orish et al., 2014; Suazo et al., 2009; Tallman, 2019; Walker, 2008) and it is likely also responsible for some of the differences seen between the New Zealand and Thai samples. However, in terms of population variation, muscle and skeletal robusticity trends appear to diverge, and this highlights the novel nature of this study, which presents population‐specific soft tissue and skeletal data for comparison. More research is needed in this space to better understand how muscle architecture may differ between populations.

The differences in preservation techniques between samples may have also played a role in the variable muscular sexual dimorphism observed in New Zealand (embalmed) and Thai (fresh frozen) samples. However, research has found there is limited tissue shrinkage with some embalming techniques (Cutts, 1988), particularly with the solution used for the New Zealand sample (Crosado et al., 2020). Nevertheless, since embalming can impact soft tissues through shrinkage (Kennedy et al., 2017) and drying (Crosado et al., 2020), it should always be considered as a possible factor in variation seen in data, especially when using samples with different preservation methods.

No sex differences were seen in ligament size for any of the groups in this study, which could mean that mass is not an appropriate variable for representing overall ligament size, when analysing soft tissue relationships. These findings could also mean that ligaments are simply not sexually dimorphic tissues and only act structurally and functionally (Sinnatamby & Last, 2011).

Entheses exhibited some sexual dimorphism (five of 15 entheses for both samples), particularly those associated with the mastoid process and rhomboid fossa. These significant sex differences may represent the skeletal sexual dimorphism seen in anthropological sex estimation methods (Buikstra & Ubelaker, 1994, Rogers et al., 2000, Walker, 2008), although age and population should be considered when interpreting these results.

It has been suggested that skeletal morphology may become more robust with increasing age (Buikstra & Ubelaker, 1994; Konigsberg & Hens, 1998; Meindl et al., 1985; Walker, 1995); however, previous research into the impact of age on skeletal sex estimation has shown varying results (Garvin, 2020; Garvin et al., 2014; Lesciotto & Doershuk, 2018; Rogers et al., 2000; Tallman, 2019; Walker, 2008). For example, Lesciotto and Doershuk (2018) found age had almost no influence on sex estimation from the cranium in their study on 272 Americans, aged 20 to 92 years, and Garvin (2020) suggested that the inclusion of age in skeletal sex estimation does not meaningfully improve outcomes, especially if age is also estimated. This could mean that cranial and clavicle sexual dimorphism is something that appears at puberty and does not change significantly over time, similar to other secondary sex characteristics, such as size or pelvic morphology (Foster et al., 2012; Schonfeld, 1943; Wells, 2007).

Sexual dimorphism in enthesis size was consistent across both populations but, compared with results for muscles, the Thai group expressed more sexual dimorphism in muscles than entheses, while the New Zealand sample had more in entheses than muscles. Based on these findings, it appears New Zealand males and females have similar muscle size, while they are skeletally different, suggesting more skeletal sexual dimorphism. In contrast, this indicated the Thai sample had more muscular sex differences but were less skeletally sexually dimorphic. These results are similar to those from Tallman (2019), who found that Thai males and females exhibited similar cranial morphology (Buikstra & Ubelaker, 1994) and less skeletal sexual dimorphism.

4.3. Population differences

There were no significant population differences in standard muscle size data, despite differences in muscular sexual dimorphism between the New Zealand and Thai samples. This suggests that the variation between the samples was representative of sex, rather than population differences. This also indicated that, although age and preservation methods may be confounding factors, they did not cause significant differences when comparing muscle size data between samples, since there was limited evidence of population variation.

When muscle data were normalised for height, some population differences became apparent, which may be due to body size variation; however, demographic data show that stature across populations was similar for each sex. Despite this, normalising these data using stature may not be best for removing the body size variable, as this study focuses on several neck muscles, and using the height of the cervical spine may be more effective (Jaric, 2002; Östh et al., 2017). Although this was beyond the scope of the study, evaluating muscle data by considering other body size factors may provide further insight into these results.

Some significant population differences were observed for entheses (three of 15 for standard data). Despite the small number, the presence of any significant differences between populations highlights the need for separate analyses of each sample. It also emphasises the importance of population‐specific studies representing diverse groups of people (Garvin et al., 2014; Tallman, 2019; Tallman & Go, 2018).

4.4. Limitations

The main limitations in this study were the advanced age of the individuals, the differences in preservation processes for the cadaveric material and the inability to conduct intra‐ and interobserver error analysis for soft tissue architectural parameters. Although imaging studies on living people would allow for broader age ranges, the types of analyses required for some aspects of this study, such as enthesis area measurement, would not have been possible and, therefore, the use of cadaveric material was necessary, despite the age limitation. Additionally, although differences in preservation processes across samples introduce limitations to the data, exploration of population variation was important to this research and, therefore, both samples were included. Finally, due to the destructive nature of the dissection process and shrinkage following the removal of tissues for measurement, it was not possible to assess soft tissue architectural parameters for intra‐observer reliability at different time points, or for interobserver reliability across different researchers. Therefore, reliability testing was limited to testing intra‐observerer error for entheseal area measurements.

5. CONCLUSION

Based on the dissection of 20 cadavers from New Zealand and Thailand, this research presents architectural data for muscles, ligaments, and entheses of the head and neck and demonstrated that muscular sex differences were more prominent in the Thai than the New Zealand sample. Both groups had similar sex differences for enthesis size and very few population differences existed for muscles and entheses. No significant sex or population differences were observed for ligament size. Overall findings indicate that sexual dimorphism and population differences can be observed in some muscles and entheses in New Zealand and Thai populations; however, the same variation is not present for ligaments. These architectural data could be applied to various areas of future research that explores cervical surgical applications, injury risk (such as whiplash) management, biomechanical modelling, and biological anthropology. Further anatomical research on these cervical tissues is encouraged to not only supplement these architectural data but to also expand knowledge relating to sex and population differences in anatomy.

AUTHOR CONTRIBUTIONS

Jade S. De La Paz: Study design, dissections, enthesis measurements, intra‐observer reliability, statistical analyses, drafting the manuscript, edits. Hallie R. Buckley: Supervising work, commenting on drafts and the final version of the manuscript. Siân E. Halcrow: Supervising work, commenting on drafts and the final version of the manuscript. Nawaporn Techataweewan: Advising on and organising work in Thailand, commenting on drafts and final version of the manuscript. Stephanie J. Woodley: Senior author, supervising work, dissection training and supervision, commenting on drafts and final version of the manuscript.

CONFLICT OF INTEREST STATEMENT

The authors declare that there are no conflicts of interest to report.

De La Paz, J.S. , Buckley, H.R. , Halcrow, S.E. , Techataweewan, N. & Woodley, S.J. (2023) Architecture of head and neck soft tissues and associated entheses: An exploration of sexual dimorphism in, and population differences between, New Zealand and Thai individuals. Journal of Anatomy, 243, 110–127. Available from: 10.1111/joa.13853

DATA AVAILABILITY STATEMENT

The data that support the findings of this study are available from the corresponding author upon reasonable request.