The morphometrical and topographical evaluation of the superior gluteal nerve in the prenatal period.

INTRODUCTION: Advances in medical science are helping to break down the barriers to surgery. In the near future, neonatal or in utero operations will become the standard for the treatment of defects in the human motor system. In order to carry out such procedures properly, detailed knowledge of fetal anatomy is necessary. It must be presented in an attractive way not only for anatomists but also for potential clinicians who will use this knowledge in contact with young patients. This work responds to this demand and presents the anatomy of the superior gluteal nerve in human fetuses in an innovative way. The aim of this work is to determine the topography and morphometry of the superior gluteal nerve in the prenatal period. We chose the superior gluteal nerve as the object of our study because of its clinical significance-for the practice of planning and carrying out hip surgery and when performing intramuscular injections.
MATERIAL AND METHODS: The study was carried out on 40 human fetuses (20 females and 20 males) aged from 15 to 29 weeks (total body length v-pl from 130 to 345 mm). Following methods were used: anthropological, preparatory, image acquisition with a digital camera, computer measurement system Scion for Windows 4.0.3.2 Alpha and Image J (accuracy up to 0.01 mm without damaging the unique fetal material) and statistical methods.
RESULTS: The superior gluteal nerve innervates three physiologically significant muscles of the lower limb's girdle: gluteus medius muscle, gluteus minimus muscle and tensor fasciae latae muscle. In this study the width of the main trunk of the nerve supplying each of these three muscles was measured and the position of the nerve after leaving the suprapiriform foramen was observed. A unique typology of the distribution of branches of the examined nerve has been created. The bushy and tree forms were distinguished. There was no correlation between the occurrence of tree and bushy forms with the body side (p > 0.05), but it was shown that the frequency of the occurrence of the bushy form in male fetuses is significantly higher than in female fetuses (p < 0.01). Proportional and symmetrical nerve growth dynamics were confirmed and no statistically significant sexual dimorphism was demonstrated (p > 0.05).
CONCLUSIONS: The anatomy of the superior gluteal nerve during prenatal period has been determined. We have identified two morphological forms of it. We have observed no differences between right and left superior gluteal nerve and no sexual dimorphism. The demonstrated high variability of terminal branches of the examined nerve indicates the risk of neurological complications in the case of too deep intramuscular injections and limits the range of potential surgical interventions in the gluteal region. The above research may be of practical importance, for example for hip surgery.

Introduction

The superior gluteal nerve (nervus gluteus superior) is an important element of the sacral plexus. Its fibers come out of branches of the abdominal spinal nerves L4, L5 and S1 [1]. The nerve leaves the pelvis through the suprapiriform foramen, which is the upper part of the greater sciatic opening (foramen ischiadicum majus) [2]. It then runs laterally with the superior gluteal artery between the gluteal muscles to innervate the hip joint as well as the muscles: m. gluteus medius, m. gluteus minimus m. tensor fasciae latae [3, 4] and, as confirmed by recent studies, also m. piriformis [5]. The analysis of available literature showed a dynamically growing number of publications concerning the described nerve. The majority of the most recent papers focuses on the relationship between the more common surgical interventions within the hip joint (femoral neck fractures, arthroplastic procedures, hip reconstruction procedures) and damage to the nerves passing through the greater sciatic opening [6, 7]. Due to the nature of the surgical approach to the hip joint, the most frequently damaged structures are the superior gluteal nerve and the superior gluteal artery [1].

Injury of the superior gluteal nerve is a complication of hip surgery that must still be reckoned with. The variability of the course of this nerve, which was studied on dissection material in adults, has a major influence on the risk associated with it [1, 8]. Acute superior gluteal nerve injury following lateral hip arthroplasty, assessed by EMG occurs in 11% to 77% of patients, but is usually temporary and most of these patients’ electromyograms recover completely after 3 months. However, in 11% of patients it persists after 9 months, and in one in 12 patients even after a year. In patients who underwent hip arthroplasty with a modified lateral approach, 4 out of 72 had chronic damage of the superior gluteal nerve [8]. Electromyographic examination (EMG) together with electroneurography (ENG) allows for an objective assessment of the degree of nerves and muscles damage in the examined patients [9]. The conclusion of these tests was, inter alia, the suggestion of performing an ultrasound examination before starting the procedure [1]. These studies, apart from their clinical significance, are of course also scientific, providing a more adequate description of the anatomical course of this nerve. No similar research on fetuses to understand the development of this nerve has been performed so far.

The process of limb formation in the embryonic period is relatively well known. Recently, the factors responsible for the location of the limbs, their polarity and identity have been identified [10]. The Pitx1 gene has been found to be the main marker of symmetry and identity of hind limbs in animals [11]. However, there is no precise data related to the development of innervation of limb muscles. Moreover, there are no studies describing the division of peripheral nerve branches and explaining the cause of the variability of nerve branches observed in further stages of ontogenesis. Some authors point out the high variability of sartorius muscle in the prenatal period, which allows to assume that morphological variants are congenital [12].

It has been demonstrated that the development of the lower limb consists mainly of its growth [13-15].

When analyzing the variability of the distribution of lower limb nerve branches, there are many literature papers on the following nerves: sciatic, common fibular or various cutaneous nerves [13, 16–18]. On the other hand, there is no detailed description of gluteal nerves in the prenatal period in the available literature.

There have been more and more surgical interventions in the perinatal period in recent years, especially in the case of deformities or in the presence of neoplastic lesions involving the lower limb [18].

In the case of newborns or infants, there are practically no scientific papers assessing the morphology of nerves innervating the gluteal region. All clinical recommendations are an extrapolation of data obtained from studies conducted on deceased adults [19]. Precise data on the location and distribution of the branches of these nerves is clinically essential to reduce the number of iatrogenic complications associated with damage to the inferior gluteal, superior gluteal and sciatic nerves. Although the most common injection site in the neonatal period is the lateral surface of the thigh (vastus lateralis muscle), understanding the developmental anatomy of the superior gluteal nerve is of practical importance for planning an injection later in childhood and in adulthood. An injury to the superior gluteal nerve results in the weakening of the limb’s abduction movements in the hip joint, the so-called duckling gait or Trendelenburg gait [6, 20]. Their common trauma can cause complete limb paralysis, resulting in massive clinical consequences at childhood [21, 22]. The World Health Organization estimates that around 12 billion injections into the gluteal region are made in connection with the administration of vaccines each year. Nerve damage in this area, including damage to the gluteal nerves, is one of the most common complications of these procedures [19]. The knowledge of the anatomy of this nerve, its development in the prenatal period and anomalies disturbing nerve conduction is therefore not only of cognitive importance but also of very significant clinical importance. Therefore, it is essential to study the details of morphology and topography of superior gluteal in the fetal period.

The aim of this study is to determine the topography and morphometry of the superior gluteal nerve in the prenatal period.

Material and methods

The study was carried out on human fetuses originating from the collection of the local Department of Anatomy at Wrocław Medical University. The whole material included 40 fetuses (20 females, 20 males) at the age from 15 to 29 weeks of fetal life with a total body length (v-pl, vertex-plantare) ranging from 130 to 345 mm (Table 1).

Table 1

Basic characteristics of the analyzed material.

Measurement feature	Female fetuses N = 20	Male fetuses N = 20	p-value
Morphological age, t (weeks):
Mean ± SD	21.7 ± 3.4	21.8 ± 3.3	0.963a
Median (Q1 –Q3)	22 (20–24)	20 (19–24)
Total body length, v-pl (mm)
Mean ± SD	239 ± 57	240 ± 51	0.981a
Median (Q1 –Q3)	250 (213–280)	219 (205–276)
Crown-rump length, v-tub (mm)
Mean ± SD	169 ± 37	169 ± 34	0.989a
Median (Q1 –Q3)	175 (154–193)	154 (146–191)
Body mass, m (g)
Mean ± SD	326 ± 195	360 ± 239	0.628a
Median (Q1 –Q3)	315 (182–438)	241 (182–492)

Q1 –lower quartile, Q3 –upper quartile

a t–test for independent samples; SD–standard deviation

It was obtained from local obstetric clinics as a result of premature and early childbirths and miscarriages between 1960 and 1996. The fetuses were stored in an appropriate solid preservative solution, containing ethanol, glycerol and formaldehyde in constant proportions, in a room with a permanent temperature.

The manner in which the fetuses were stored has not changed throughout the entire storage period. We have not included in the study fetuses with visible developmental malformations (any malformation in general) and those that did not have complete clinical documentation.

The value and credibility of the fetal collection has been confirmed in many scientific papers [15, 16, 23–26]. The basic characteristics of the study material are presented in Table 1. There were no statistically significant differences in somatic features of fetuses of both sexes (p > 0.05).

The following methods were used: anthropological method, preparatory method, image acquisition with a digital camera, computerized measurement system Scion for Windows 4.0.3.2 Alpha and Image I and statistical methods. These methods were also used in earlier publications by the authors’ team based on research material from the same fetal collection [27, 28].

The anthropological method allowed to determine the biological age of the examined fetuses. The morphological age was assessed using measurable parameters such as total body length (v-pl), crown-rump length (CRL or v-tub) and body weight [29]). The preparation method was based on exposing the superior gluteal nerves on 80 limbs using standard sectional instruments.

Image acquisition was performed using a high-resolution digital camera. The next step was the measurements in Scion Image for Windows 4.0.3.2 Alpha and Image J (National Institute of Mental Health–NIMH; https://imagej.nih.gov). Contrast improvement, sharpening, spatial processing were used to improve image quality. Metrological tests were carried out on the basis of a millimeter scale attached to each photographed specimen.

In the study, the width of the main trunk of the nerve supplying each of these three muscles was measured and the direction of the nerve course was observed after leaving the suprapiriform foramen.

The results of the measurements were analyzed statistically on the basis of Statistica PL statistical package (Statsoft, Tulsa, USA). The statistical analysis was carried out by an experienced statistician with high mathematical skills documented and confirmed in previous analyses [23, 30].

Variables with a distribution consistent with the normal distribution were analyzed using parametric methods. Student’s t-test (t-test) was used to assess the significance of differences between mean values in two independent groups. For quantitative variables, mean values (M), standard deviations (SD), medians (Me) and the lower (Q1) and upper (Q3) quartiles, the lowest (Min) and highest (Max) values were measured. Nominal qualitative variables (sex and type of nerve branch distribution) in multidirectional tables are presented as numbers (n) and structure indices (%). Relationships between categorized variables were assessed using Fisher’s exact test. The relationship between quantitative variables was analyzed by estimating Pearson’s r correlation coefficients. The test probability at the level of p < 0.05 was assumed as significant, whereas p < 0.01 was assumed as highly significant. The measurement results were statistically analyzed with the use of a package of statistical program Statistica v. 13.3 (TIBCO Software Inc.). Continuous quantitative variables were assessed for the consistency of their distribution with the theoretical normal distribution. The test W of Shapiro Wilk was used.

The study was approved by the local bioethics committee (No. 495/2020). The authors declare no conflict of interest.

Results

The basic characteristics of the material were included in Table 1.

The preparation revealed that the superior gluteal nerve innervates three physiologically relevant muscles of the lower limb’s girdle: gluteus medius muscle, gluteus minimus muscle and tensor fasciae latae muscle.

The typology of the distribution of the branches of the examined nerve was outlined. The authors found two types of distribution of the branches of the superior gluteal nerve: a tree form (Figs 1–5) and a bushy form. Within the bushy form, a different arrangement of branches that reach the muscles was found: vertical (Figs 6–9), horizontal (Figs 10–12). An additional classification is related to the number of secondary branches, which are numerous, with a small diameter of 0.1 mm or sparse in trunks with larger diameters (0.2–0.3 mm). The variety of morphology of the superior gluteal nerve is surprising, from simple, classic forms of a tree, to the connection of two trunks in a bushy form, located vertically or horizontally (within the muscles) forming ladder-shaped formations or nets with various mesh shapes. The preparation difficulties associated with small, easily damaged branches should be emphasized.

Fig 1

Superior gluteal nerve, tree type, morphological age 157–23–VI, v–tub 182, v–pl 258, mass 450, female sex, magnification 20x, 1– gluteus medius muscle, 2– gluteus minimus muscle, red arrow–branch gluteus medius muscle, yellow arrow–branch to gluteus minimus muscle, blue arrow–branch to tensor fasiae latae muscle (branch to tensor fasiae latae muscle is better visible in Fig 2).

Fig 5

Superior gluteal nerve, tree type, morphological age150–22–VI, v–tub 182,v–pl 258, mass 140, female sex, magnification x30, 1– gluteus medius muscle, 2– gluteus minimus muscle, red arrow–branch to gluteus medius muscle, yellow arrow–branch to gluteus minimus muscle, blue arrow–branch to tensor fasiae latae muscle.

Fig 6

Superior gluteal nerve bushy type, vertical course morphological age 162–24–VI, v–tub183, v–pl 270,mass 544, male sex, magnification 45x.

1– gluteus medius muscle, 2– gluteus minimus muscle, red arrow–branch to gluteus medius muscle, yellow arrow–branch to gluteus minimus muscle, blue arrow–branch to tensor fasiae latae muscle.

Fig 9

Superior gluteal nerve bushy type, vertical coursemorphological age 164–24–VI,v–tub191, v–pl 269. mass 489, male sex, magnification 20x, 1– gluteus medius muscle, 2– gluteus minimus muscle, red arrow–branch to gluteus medius muscle, yellow arrow–branch to gluteus minimus muscle, blue arrow–branch to tensor fasiae latae muscle.

Fig 10

Superior gluteal nerve, bushy type, horizontal–oblique course, the net with variform meshes morphological age 169–25–VII,V–tub 200, V–pl 294,mass 690, male sex, magnification 20x.

1– gluteus medius muscle, 2– gluteus minimus muscle, red arrow–branch to gluteus medius muscle, yellow arrow–branch to gluteus minimus muscle, blue arrow–branch to tensor fasiae latae muscle.

Fig 12

Superior gluteal nerve, bushy type, horizontal course, morphological age 113–17–V, v–tub 110,V–pl 140,mass 120, male sex, magnification 50x 1–gluteus medius muscle, 2–gluteus minimus muscle, 3–tensor fasciae latae muscle, red arrow–branch to gluteus medius muscle, yellow arrow–branch to gluteus minimus muscle, blue arrow–branch to tensor fasiae latae muscle.

Superior gluteal nerve, tree type, morphological age 184–27–VII, V–tub 220,V–pl 331.

124–18–V, V–tub 130,V–pl 190, mass 627, male sex, magnification 40x, –1– gluteus medius muscle, 2– gluteus minimus muscle, red arrow–branch to gluteus medius muscle, yellow arrow–branch to gluteus minimus muscle, blue arrow–branch to tensor fasiae latae muscle.

Superior gluteal nerve, tree type, morphological age 124–18–V, V–tub 130,V–pl 190, mass 140, male sex, magnification 30x.

Fig 11

Superior gluteal nerve, bushy type, horizontal–oblique course, the net with variform meshes, morphological age 169–25–VII,V–tub 200, V–pl 294,mass 690, male sex, magnification 20x.

Figs 10 and 11 are taken from both sides of the same specimen and are evidence of symmetry in the type of innervation., 1– gluteus medius muscle, 2– gluteus minimus muscle, red arrow–branch to gluteus medius muscle, yellow arrow–branch to gluteus minimus muscle, blue arrow–branch to tensor fasiae latae muscle.

Based on these observations a unique typology of the distribution of the branches of the examined nerve was created. The tree (Fig 13) and bushy, predominantly male (Fig 14) forms have been identified.

Fig 13

Superior gluteal nerve, tree type–anatomical diagram of the nerve path (thanks to Julia Derkowska), 1–gluteus medius muscle, 2–gluteus minimus muscle, 3–tensor fasciae latae muscle.

Fig 14

Superior gluteal nerve, bushy type–anatomical diagram of the nerve path (thanks to Julia Derkowska), 1–gluteus medius muscle, 2–gluteus minimus muscle, 3–tensor fasciae latae muscle.

The frequency of occurrence of different types of nerve on the left and right side and in female and male fetuses was analyzed by comparing them in 2 x 2 contingency tables (Table 2). They show the number (percentage) of fetuses in groups differing in sex and nerve type, the results of the independence tests and the values of the odds ratios and their 95% confidence intervals.

Table 2

Prevalence of superior gluteal nerve patterns in respect to sex and laterality.

The bold and italic text shows a statistically significant difference.

Type	Gender		F vs. M p-value*	OR (95% CI)
	Female (F) N = 20	Male (M) N = 20
Nerve pattern on the left:
Tree	8 (40%)	2 (10%)	0.068	6.00 (1.08–33.3)
Bushy	12 (60%)	18 (90%)		1.00 (ref.)
Nerve pattern on the right:
Tree	8 (40%)	2 (10%)	0.068	6.00 (1.08–33.3)
Bushy	12 (60%)	18 (90%)		1.00 (ref.)
Nerve pattern on both sides:
Tree	16 (40%)	4 (10%)	0.005	6.00 (1.79–20.1)
Bushy	24 (60%)	36 (90%)		1.00 (ref.)

* Pearson’s Chi–squared test with Yates’ continuity correction

There was no correlation between the occurrence of tree and bushy forms with the body side (p > 0.05), whereas the frequency of occurrence of bushy forms in male fetuses is significantly higher than in female fetuses (p < 0.01). The chance of occurrence of bushy nerve form in male fetuses is six times higher than in female fetuses (OR = 6).

The dimensions of the examined anatomical structures on both sides are equal (Table 3) and there is no statistically significant sexual dimorphism noted (p > 0.05). The results of measurements on the left and right sides and the female and male sexes were combined during the analysis of correlation and regression of the dimensions of the examined structures with the age of the fetus. It was due to demonstrated lack of sexual dimorphism and asymmetry.

Table 3

Width of the superior gluteal nerve branches to the particular muscles in respect to sex and laterality.

Measurement feature	Gender		F vs. M p-value
	Female (F) N = 20	Male (M) N = 20
MP L [mm]–MP width, left side			0.995a
Mean ± SD	4.2 ± 1.3	4.2 ± 1.2
Median (Q1 –Q3)	4.1 (3.1–5.3)	4.0 (3.2–4.6)
MP R [mm]–MP width, right side			0.848a
Mean ± SD	4.2 ± 1.3	4.2 ± 1.2
Median (Q1—Q3)	4.1 (1.9–6.5)	4.0 (2.8–6.8)
MP L vs. MP R	p = 0.311b	p = 0.558b
TFL L [mm]–the width of the nerve branch to TFL, left side			0.428a
Mean ± SD	0.26 ± 0.03	0.27 ± 0.05
Median (Q1 –Q3)	4.1 (3.1–5.3)	4.0 (3.4–4.6)
TFL R [mm]–the width of the nerve branch to TFL, right side			0.998a
Mean ± SD	0.26 ± 0.05	0.26 ± 0.04
Median (Q1 –Q3)	0.25 (0.24–0.28)	0.26 (0.24–0.30)
TFL L vs. TFL R	p = 0.388b	p = 0.180b
MED L [mm]–width of main nerve branch to MED, left side			0.758a
Mean ± SD	0.25 ± 0.05	0.25 ± 0.06
Median (Q1 –Q3)	0.25 (0.20–0.29)	0.25 (0.20–0.30)
MED R [mm]–width of main nerve branch to MED, right side			0.691a
Mean ± SD	0.24 ± 0.05	0.25 ± 0.06
Median (Q1 –Q3)	0.24 (0.21–0.27)	0.25 (0.20–0.29)
MED L vs. MED R	p = 0.091b	p = 0.126b
MINI L [mm]–width of main nerve branch to MINI, left side			0.139a
Mean ± SD	0.22 ± 0.04	0.24 ± 0.05
Median (Q1 –Q3)	0.21 (0.19–0.25)	0.26 (0.19–0.29)
MINI R [mm]–width of main nerve branch to MINI, right side			0.193a
Mean ± SD	0.22 ± 0.05	0.24 ± 0.05
Median (Q1 –Q3)	0.22 (0.19–0.25)	0.24 (0.20–0.29)
MINI L vs. MINI R	p = 0.762b	p = 0.322b
MINI bis L [mm]–the average width of nerve branches exiting the nerve trunk on the left side			0.606a
Mean ± SD	0.10 ± 0.02	0.11 ± 0.02
Median (Q1 –Q3)	0.10 (0.09–0.12)	0.10 (0.10–0.12)
MINI bis R [mm]–the average width of nerve branches exiting the nerve trunk on the right side			0.958a
Mean ± SD	0.11 ± 0.02	0.11 ± 0.03
Median (Q1 –Q3)	0.10 (0.10–0.13)	0.12 (0.10–0.13)
MINI bis L vs. MINI bis R	p = 0.149b	p = 0.937b

Q1 –lower quartile, Q3 –upper quartile, a t–test for independent samples; b t–test for dependent samples.

a t–test for independent samples

b t–test for dependent samples. MP–piriformis muscle; TFL–tensor fasciae latae muscle; MED–gluteus medius muscle; MINI–gluteus minimus muscle.

The values of linear correlation coefficients are presented in Table 4. In the case of statistically significant correlation (p < 0.05), they are also presented in the correlation diagrams, which include regression models (Figs 15–18).

Table 4

The correlation coefficients of the analyzed parameters with the age of the fetus.

Measuring feature	Correlation coefficient	Level of significance
MP (mm)	r = 0.608	p < 0.001
TFL (mm)	r = 0.177	p = 0.117
MED (mm)	r = 0.330	p = 0.003
MINI (mm)	r = 0.108	p = 0.117
MINI bis (mm)	r = 0.077	p = 0.520

MP–width of piriformis muscle, TFL–width of the main branch to tensor fasciae latae muscle, MED–width of main branch to gluteus medius muscle, MINI–width of main nerve branch to gluteus minimus muscle, MINIbis–the average width of nerve branches exiting the nerve trunk of the nervus gluteus superior

Fig 15

Correlation diagrams for the width of the piriformis muscle (MP) and width of the main branch to the gluteus medius muscle (MED) with the age of the fetus.

Fig 18

Correlation diagrams for the width of the nerve branch to the tensor fasciae latae muscle (TFL) with the width of the piriformis muscle (MP), the width of the main branch to the gluteus medius muscle (MED), the average width of the branches emerging from the main trunk of the nerve to the gluteus minimus muscle (MINI) and the average total width of the branches departing from the main trunk of the examined nerve (MINI bis).

The width of the piriformis muscle in the analysed period increases progressively at a linear rate of 0.22 mm per week and the width of the main branch to the gluteus medius muscle increases at a rate of 0.01 mm per week.

Due to the high variability of the dimensions of the nerve branches running to tensor fasciae latae muscle (TFL), the correlation between the dimensions of the main nerve branch running to gluteus minimus muscle (MINI) and the average width of the branches running away from the trunk of the main nerve (MINI bis), resulting from a high relative measurement error, is not statistically significant.

The growth of the nerve is proportional as evidenced by a statistically significant positive correlation of TFL, MP, MED, MINI and MINI bis (Figs 17 and 18).

Fig 17

Correlation diagrams for the width of the nerve branch to the tensor fasciae latae muscle (TFL) with the width of the piriformis muscle (MP), the width of the main branch to the gluteus medius muscle (MED), the average width of the branches emerging from the main trunk of the nerve to the gluteus minimus muscle (MINI) and the average total width of the branches departing from the main trunk of the examined nerve (MINI bis).

Discussion

The course of the superior gluteal nerve and its importance for the safety of surgical access to the hip joint was studied on dissection material in adults [8]. The authors distinguished two types of the course of this nerve: the spray pattern course and the transverse neural trunk pattern course. The course of the superior gluteal nerve was also studied by Jacobs and Buxton, who described two types of the course of this nerve. According to them, the ends of all branches of the nerve assumed an arcuate shape [31]. Perez et al. (2004) examined the dissection material of 19 adults [32]. They found that the superior gluteal nerve was in 89.48% cases divided into two branches and in 10.52% cases into three. The course of the superior gluteal nerve in adults was also studied by Ray et al. (2013). Among other things, they found that the most frequent number of branches to the individual muscles innervated by the superior gluteal nerve ranged from 2 to 3 [1].

Most of the work analyzing the development of the fetus was conducted using in vivo techniques. Various anatomical structures were studied: biparietal diameter, head and body circumferences, transverse brain dimensions, abdomen circumferences, and limb lengths [33-37].

The majority of the literature surrounding the development of the fetus has focused on fetal growth of the vertebral column [38-41]. In one study, authors evaluated the development of the sacral bone using ultrasonography [40], confirming the importance of biparietal diameter measurement in the assessment of physiological pregnancy. Similarly, in this paper, biparietal diameter was also used in preparation analysis to confirm the age of examined fetuses.

Throughout the study period, the scope of information derived from the available literature is mainly related to the analysis of topography and metrology of the superior gluteal nerve in adults [1, 3, 6].

The course and topography of the superior gluteal nerve is of significant importance in developing safe surgical access procedures to the hip joint [42]. Many authors also emphasize the high frequency of nerve injuries during pelvic bone fractures and hip dislocations. The nerve may also be the object of stab wound injuries or even the point of iatrogenic injuries resulting from intramuscular injections [43, 44].

With regard to prenatal studies, the available literature is extremely scarce. The only available studies related to the evaluation of the gluteal region concern, in most cases, the assessment of the superior gluteal muscle and analysis of sciatic nerve variability [27, 45].

Neuromuscular platelets of the gluteal region were analyzed through microscopic examination, which were based on fetal preparations as well as neurovascular pedicles. Microscopic analyses confirmed the high variability of the terminal branches of the superior gluteal nerve in the current study [46, 47].

The dynamics of growth of the examined nerve is proportional and comparable to the dynamics of other nerves analyzed in separate studies [48, 49]. This research results revealed that the width of the piriformis muscle in the analyzed period increases linearly at a rate of 0.22 mm per week, and the width of the main branch to the gluteus medius muscle average at a rate of 0.01 mm per week.

Similar results in terms of developmental dynamics were obtained when comparing piriformis muscle with other analyzed muscles in the fetal period [50]. Attention was drawn to the symmetrical development of the width of the piriformis muscle and the superior gluteal nerve as well as the branches diverging from the examined nerve. The aforementioned structures were responsible for the innervation of selected external pelvic muscles. In addition, the lack of sex differences in the analyzed material was also observed. The same characteristics were found in the analysis of development of gluteus maximus muscle [27, 45].

The present study introduces an innovative typology of branch distribution of the superior gluteal nerve, implementing a comprehensive mathematical analysis of the obtained results. These results are of great cognitive importance and can also be used in future research in connection with the dynamically growing specialities of neonatal and intrauterine surgery [51, 52].

Furthermore, the clinical significance of the study of the superior gluteal nerve in the prenatal period is important for the surgical treatment of defects in the lower limbs in infants and possibly also in the fetal period in the future. For example, in the developmental dysplasia of the hip (DDH), which affects approximately one percent of newborns. If it is diagnosed after the child is six months old, permanent deformation of the tissues of the hip joint may develop. The older the child, the more likely it will be to openly reduce it, with osteotomy of the femur or acetabulum to stabilize reduction. Late open reduction score is significantly inferior to long-term hip function and increases the risk of developing coxarthrosis [53].

The management of developmental dysplasia of the hip depends on age. The sooner neonatal hip instability is diagnosed and treated, the better. Initially, the procedure is based on the use of a device that holds the hip in the abduction. In the event of ineffectiveness of such a procedure, a surgical reduction is indicated [54].

Limitations of the study were two: the quantitative limitation and the limitation resulting from the nature of the fetal material. The quantitative limitation of the study was due to the fact that fetal material is currently particularly difficult to obtain, and obtaining fetuses for anatomical research is a big challenge. The limitation resulting from the nature of the material was due to specific features of fetal tissues distinguishing them from adult cadavers’ tissues. This requires from the anatomist much more precision, and often special techniques for the preservation, storage, and examination of the fetuses.

Conclusions

The morphology of the superior gluteal nerve has been determined. The bushy and tree forms have been identified. No lateralisation of individual types was observed. No sexual dimorphism and asymmetry were found.

This study has a significant clinical importance for planning and performing hip surgery and intramuscular injections in the gluteal area. It could help avoid the damages made to the superior gluteal nerve and mitigate the risks related to such surgeries. They are now among the most frequently made damages because of the nature of surgical access to the hip joint.

What is more, the clinical importance of our study of the superior gluteal nerve in the prenatal period is of great importance for the surgical treatment of lower limb defects in infants, and possibly also in the fetal period in the future (for example for developmental dysplasia of the hip—DDH).

15 Mar 2022

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Mateusz Koziej, MD, PhD, DSc

Academic Editor

PLOS ONE

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NO

At this time, please address the following queries:

a) Please clarify the sources of funding (financial or material support) for your study. List the grants or organizations that supported your study, including funding received from your institution.

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d) If you did not receive any funding for this study, please state: “The authors received no specific funding for this work.”

Please include your amended statements within your cover letter; we will change the online submission form on your behalf.

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NO

Please complete your Competing Interests on the online submission form to state any Competing Interests. If you have no competing interests, please state "The authors have declared that no competing interests exist.", as detailed online in our guide for authors at http://journals.plos.org/plosone/s/submit-now

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Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Yes

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2. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

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3. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

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4. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: Yes

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5. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: Dear Authors,

Thank you very much for the opportunity to review this great study! I have a few suggestions for study improvement:

1) Overall:

- Please unify English spelling - sometimes fetus and other times foetus is used. Please double check for English grammar mistakes.

2) Abstract:

- Please provide 1-2 sentences regarding the rationale for choosing the superior gluteal nerve for your current study. Why is knowledge regarding this particular nerve important?

- First sentence of the conclusions - please rewrite adding information referring to the gestation. If you have partly divided the abstract into sections, please stick to it and present it as Introduction, materials and methods, results and conclusions.

- Please rewrite last two sentences of the conclusions - they are a repetition from the results. Instead provide information as to how to use your results in everyday medical practice.

3) Introduction:

- Paragraph two, second sentence - please specify what risk and add its estimated prevalence.

- Penultimate paragraph - the authors discuss the clinical implications associated with the superior gluteal nerve. However, information regarding the possible causes, mechanisms of injury or situations in which it can be damaged prenatally or in the neonatal period is scarce. Please add 1-2 sentences pertaining to this particular population, as those will be the patients that could potentially benefit from the study results. Please also reconsider the information regarding vaccinations into the gluteus muscles - the newborns are predominantly vaccinated (namely against hep. B and TB in some countries) by utilising the vastus lateralis muscle.

4) Material and methods:

- Please explain the "v-pl" abbreviation the first time it is used.

- The crown-rump length ought to be written down with a hyphen.

- Please provide the first and third quartiles instead of min-max (those can be described in the text), as the quartiles are more informative (the former can present outliers). The mean and median ought to be presented with one decimal place (SD with two) for mathematical accuracy.

- Please specify "visible developmental malformations" - did it involve only malformations of the abdomen, pelvis and lower limb or any malformation in general? How many foetuses were excluded?

- Please carefully review the paragraph describing the statistical methods used. Some parts have been submitted twice. Min and max values surely could not have been estimated but measured - please rewrite.

5) Results:

- The information regarding the measurement method from paragraph two ought to be presented in methods section, not results. Likewise, table 1 (characteristics of the analyzed material) with its calculated values ought to be presented in the results (calculations lead to results) with only a brief description in the materials section. Please move table 1 to the results section.

- Figure 14 and Figure 13 in paragraph 3 should not be referenced prior to figures 1-12. Good practice is to number the figures consecutively in the order they appear (as per journal guidelines).

- Sentence "it is divided into two types: tree form (...)" requires rewriting (especially its second part) - possibly split into two sentences.

- Please stick to the "tree type" - do not introduce another descriptor as "woody type" if it refers to the same entity. Same goes for Table 2.

- The 95%CI for the OR in Table 2 have to be corrected. Please rename the table (possibly into: "Table 2. Prevalence of superior gluteal nerve patterns in respect to sex and laterality.").

- Please unify the results as written with a dot (.) describing the decimal place of the estimates (not comas).

- Same as with table 1, please provide Q1 and Q3 values instead of min-max for table 3. Muscle abbreviations have to be explained in table footnotes. Please rename the table (possibly into: "Width of the superior gluteal nerve branches to the particular muscles in respect to sex and laterality.").

- Please rewrite "gluteal muscle medium" unerneath the table 4.

6) Discussion:

- The paragraph regarding the development of the lumbar spine is not relevant to the current study. Please delete.

- Clinical significance of the fetal superior gluteal nerve has to be expanded. Please relate it more to the fetal (possibly fetal lower limb surgery), neonatal and infant applications, as this is the population that could benefit from the current study. The embryological pattern of the nerve will be less relevant in an adult due to the growth of the axial skeleton, muscular variabilities and personal history (namely injuries) in the later life.

- There is no limitations section to the discussion. Please amend.

7) Conclusions:

- Please provide 1-2 sentences on how the results of the current study can specifically be used in everyday medical practice.

8) Please acknowledge the statisticians in the acknowledgement section, as well as the donors as per anatomical journals editors recommendations (https://onlinelibrary.wiley.com/doi/abs/10.1002/ca.23671).

9) Please change section headings into "Table legends" and "Figure legends." The tables were not submitted as supplementary files, but part of the main manuscript.

10) Figures:

- There is no number 3 (TFL) visible on the figure (though described in figure legends). It is also hard to see that the blue arrow points to the nerve to TFL. Please amend.

- Figure 2 - yellow arrow seems to aim more at gluteus medius than minimus.

- Please double check all figures against their footnotes - in many cases TFL is only listed, but not marked.

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6. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

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Reviewer #1: No

[NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment files.]

While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com/. PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Registration is free. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email PLOS at figures@plos.org. Please note that Supporting Information files do not need this step.

2 May 2022

Reviewer #1: We have incorporated all of your suggestions into our revision, except part of suggestion 4 concerning decimal place for mathematical accuracy ("The mean and median ought to be presented with one decimal place (SD with two) for mathematical accuracy.").

After discussing with our statistician, we ask you to accept our version of the decimal places for the mean and median, because the number of decimal places depends on the accuracy of the measurement of the physical quantity. When measuring the length of biological structures (in millimeters), the actual average should be recorded to one decimal place. A measure of variation, i.e. standard deviation (SD), is always given in the same units with the same accuracy, so one decimal place is also enough.

Thank you for your help.

Submitted filename: Response to Reviewers.doc

Click here for additional data file.

30 May 2022

12 Jun 2022

Dear Reviewer,

We have incorporated all of your suggestions into our revision.

We have added section "limitations of the study" as the last paragraph of the discussion.

Thank you for your help.

Kind regards

Wojciech Derkowski, MD, PhD

Submitted filename: Response to Reviewers.doc

Click here for additional data file.

11 Jul 2022

21 Jul 2022

Dear Reviewer,

We have incorporated all of your suggestions into our revision.

We have changed in table 2 Fisher exact test for chi2 with Yates correction.

Thank you for your help.

Kind regards

Wojciech Derkowski, MD, PhD

Submitted filename: Response to Reviewers.doc

Click here for additional data file.

9 Aug 2022

The morphometrical and topographical evaluation of the superior gluteal nerve in the prenatal period.

PONE-D-21-33849R3

Dear Dr. Derkowski,

We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements.

Within one week, you’ll receive an e-mail detailing the required amendments. When these have been addressed, you’ll receive a formal acceptance letter and your manuscript will be scheduled for publication.

An invoice for payment will follow shortly after the formal acceptance. To ensure an efficient process, please log into Editorial Manager at http://www.editorialmanager.com/pone/, click the 'Update My Information' link at the top of the page, and double check that your user information is up-to-date. If you have any billing related questions, please contact our Author Billing department directly at authorbilling@plos.org.

If your institution or institutions have a press office, please notify them about your upcoming paper to help maximize its impact. If they’ll be preparing press materials, please inform our press team as soon as possible -- no later than 48 hours after receiving the formal acceptance. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information, please contact onepress@plos.org.

Kind regards,

Mateusz Koziej, MD, PhD, DSc

Academic Editor

PLOS ONE

Additional Editor Comments (optional):

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation.

Reviewer #2: All comments have been addressed

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2. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #2: (No Response)

**********

3. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #2: (No Response)

**********

4. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #2: (No Response)

**********

5. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #2: (No Response)

**********

6. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #2: Authors have improved their manuscript. There is nothing more to improve. Congratulations. It was a pleasure to revise this manuscript.

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7. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #2: No

**********

16 Aug 2022

PONE-D-21-33849R3

The morphometrical and topographical evaluation of the superior gluteal nerve in the prenatal period.

Dear Dr. Derkowski:

I'm pleased to inform you that your manuscript has been deemed suitable for publication in PLOS ONE. Congratulations! Your manuscript is now with our production department.

If your institution or institutions have a press office, please let them know about your upcoming paper now to help maximize its impact. If they'll be preparing press materials, please inform our press team within the next 48 hours. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information please contact onepress@plos.org.

If we can help with anything else, please email us at plosone@plos.org.

Thank you for submitting your work to PLOS ONE and supporting open access.

Kind regards,

PLOS ONE Editorial Office Staff

on behalf of

Dr. Mateusz Koziej

Academic Editor

PLOS ONE