Teza Adela 9.04 [304926]
ȘCOALA DOCTORALĂ
DOCTORAL THESIS
Ultrasonographic evaluation of the orofacial soft tissues
PhD Student: [anonimizat], Dudea D, Dudea S. Evaluation of periodontal tissues using 40 Mhz Ultrasonography. MedUltrasono 2013;4(2-3):17-22. ISI Factor de impact-1.265 (Study included in chapter 4)
Zimbran A, Dudea D, Gasparik C, Dudea S. Ultrasonographic evaluation of periodontal changes during orthodontic tooth movement. (study included in chapter 5)
TABLE OF CONTENTS
ABBREVIETIONS
INTRODUCTION
Diagnosis in the field of dentistry is the most important phase in establishing the right treatment plan. A [anonimizat].
[anonimizat], [anonimizat]'s complexity. Thus, microbiologic, histologic, serologic and imagistic examinations have become indispensable tools in precise diagnosis of clinical cases.
Periodontal imaging is a [anonimizat]. Currently, the only used periodontal imaging techniques are X-[anonimizat], do not permit observation of the superficial periodontium on the buccal and lingual surfaces.
[anonimizat] X-rays, [anonimizat]-maxillary system: [anonimizat], [anonimizat]. Cone beam computed tomography (CBCT) [anonimizat] a three-[anonimizat], yet its high price limits the indication of its use. [anonimizat]. [anonimizat].
These shortcomings are meant to be overcome with the help of a [anonimizat]-invasive, [anonimizat]: [anonimizat]: ophtalmology, [anonimizat], cardiology, orthopedics, [anonimizat], yet it is absent from the context of clinical dentistry. [anonimizat], [anonimizat].
[anonimizat] ‘Current phase of knowledge’, an overview of the scientific research in the filed of periodontal and muscular evaluation is thoroughly explained. There are matters that are not fully understood regarding applicability of ultrasound in the field of dentistry and future studies should focus on this issue.
Deriving from this premiss, within the current thesis, the following clinical and experimental studies have been conducted:
1. A feasibility study to test the possibility to use high frequency ultrasonography (40 MHz) on human peridontal tissues.
2. A reproducibility investigation conducted on 40 periodontal surfaces of pig jaw mandible.
3. An inovative research, which demonstrates the applicability of ultrasound imaging in the evaluation of periodontal, changes during orthodontic tooth movement.
4. A study to establish the correlation between the vertical facial pattern and the ultrasound-determined thickness of the maseter and orbicularis oris muscles.
5. A new approach to mandibular retrognatia: the influence of suprahyoid muscular thickness evaluated with the aid of ultrasounography.
The Ph.D. thesis entitled „The ultrasonographic evaluation of orofacial soft tissue” aims to demonstrate the application of ultrasound imaging within an experimental context as well as the clinical context of medical dentistry.
CURRENT STATE OF KNOWLEDGE
1. Anatomy
1. 1. Periodontal structures
The periodontium consists of the tissues that surround, envelop or embed the tooth: gingiva, periodontal ligament, cementum and alveolar bone. (fig.1) It has been divided into two parts: the gingiva, which covers the alveolar process and protects the underlying tissues, and the attachment apparatus, composed of the periodontal ligament, cementum and alveolar bone. [1,2]
1.1.1. Gingiva
Gingiva is the part of oral mucosa covered by keratinized epithelium (fig. 1). It has three main parts: free gingiva, interdental papilla and attached gingiva. All types of gingiva are specifically structured to function appropriately against microbial and mechanical damage. [3) exhibiting considerable variations in thickness, histology and differentiation. [1]
Free gingiva is the terminal border of the gingiva surrounding the teeth to form a collar of tissue. The colour of the free gingiva is usually light pink and averages between 0,5 and 2 mm in depth. [4]
The gingival sulcus is the shallow space bounded by the tooth surface on one side and the inner part of the free gingival margin (epithelium) on the other side. It extends from the free gingival margin to the junctional epithelium (0,69 mm average in depth)(5), which is a band of tissue (1 mm average wide) that attaches the gingiva to the tooth. Underneath the junctional epithelium, there is 1 to 1,5 mm connective tissue attachment to the root coronal to the crest of bone.
The interdental gingiva (interproximal papilla) represents the part of the free gingiva that occupies the gingival embrasure, which is the space between two adjacent teeth, beneath their area of contact.
Attached gingiva continues the free gingiva. It has a firm, resilient consistency and is tightly bound to the periostium of the alveolar bone. The width of the attached gingiva is is the distance between mucogingival junction and the free gingiva, and its dimensions decrease from the anterior part of the maxilla and mandible (3,5-4,5, respectively 3,3-3,9 mm) to the posterior segment (1,9 mm and 1,8 mm). (6)
1.1.2. Periodontal ligament
The periodontal ligament is a connective and vascular tissue that surrounds the roots of the teeth and connects them to the alveolar bone. Above the alveolar crest, the ligament is continuous with the connective tissue of the gingiva, and explains the development of periodontitis from gingivitis; at the apical foramen, it is continuous with the dental pulp so that the inflammation from the dental tissue can spread to the periodontal ligament. (7) In the literature, the average width of the periodontal ligament space is documented to be aproximately 0,2 mm, nonetheless, considerable variation exists. (1)
The principal fibers are the most important elements of the periodontal ligament. They are collagenous and arranged in bundles. The terminal portion of the principal fibers that are inserted into cementum and bone are called Sharpey’s fibers. Collagen is a protein composed of different amino acids, being responsible for maintenance of the framework and tone of tissue. (8)
The principal fibers of the periodontal ligament are arranged in six groups: the transseptal, alveolar crest, horizontal, oblique, apical and interradicular fibers. (1)
Transseptal group extends interproximally over the alveolar bone crest and is embedded in the cementum of adjacent teeth. These fibers may be considered as part of the gingiva since they do not have osseous attachment.
Alveolar crest fibers link the cementum to the alveolar crest. These fibers prevent the extrusion of the tooth (9) and resist lateral tooth movements.
Horizontal fibers extend at right angles to the long axis of the tooth from the cementum to the alveolar bone.
Oblique fibers represent the largest group in the periodontal ligament, bearing the brunt of vertical masticatory stress and transform them into tension on the alveolar bone.
Apical fibers radiate from the cementum to the alveolar bone, at the apical region. They appear only in completely formed roots.
Interradicular fibers extend from the cementum to the furcation areas of teeth with multiple roots. [1]
1.1.3. Alveolar Bone
The part of the maxillary and mandibular bones, which protects and supports the teeth, is known as alveolar process. It is a tooth-dependent bony structure, since it develops and undergoes remodelling with the tooth formation and eruption. (10) Therefore the shape, size, function and location of the teeth determine their morphology. A certain degree of repositioning of the teeth can be accomplished in response to orthodontic mechanics and occlusal forces that rely on the adaptability of the alveolar bone and associated periodontal tissues.
The alveolar bone is composed of: the external plate of the cortical bone, the inner socket wall of compact bone (alveolar bone proper or lamina dura on radiographs), and cancellous trabeculae between the two compact plates. The interdental septum consists of cancellous supporting bone enclosed within a compact border.
Remodeling is the main process of bony changes in shape, repair of wounds, resistance to forces. Bone resorption and bone formation constitutes one of the fundamental principles by which bone is necessarily remodelled throughout its life. Bone remodeling involves the activity of osteoblasts and osteoclasts, which form and resorb the mineralized connective structures of the bone. (11)
1.1.4. Cementum
Cementum is the avascular, calcified mesenchymal tissue that covers the anatomical root. The two types of cementum are primary (acellular) and secondary (cellular) (12), and they both consist of collagen fibrils and calcified matrix. (1)
The organic matrix of cementum is composed of type I and type III collagens. The inorganic material consists of 45-50 % Hydroxyapatite.
Acellular cementum covers the cervical third of the root and develops before the tooth reaches the occlusal plane. It does not contain cells and it has a thickness of 30 to 230 μm. (13) Sharpey fibers are the embedded portion of the principal fibers of the periodontal ligament (14) and make up most of the structure of the acellular cementum. They are inserted at right angles into the cementum and have an important role in tooth support. Their number, size and distribution increase with function. (1)
The cellular cementum contains cementocytes, in individual spaces, called lacunae, which communicate with each other through a system of canaliculi. The secondary cementum is more irregular and less calcified than the primary type.
Cementoenamel junction (the contact area between the cementum and enamel) has three possibilities: cementum overlaps the enamel (60-65% of cases); edge-on-edge butt joint (30%); cementum and enamel fail to meet (5%
1.2. Muscles of the dentomaxillary system
Muscles of the dentomaxillary system are classified according to their function: elevators of the mandible (masseter, temporal, internal pterygoid muscles); lowering muscles of the mandible (suprahyoid muscles); protrusive movement (internal pterygoid) and muscles of facial expression. The present thesis focuses on the superficial muscles, which can be easily identified with the use of ecography: masseter, suprahyoid muscles and orbicularis oris.
1. 2.1. Masseter muscle
Masseter is a powerful elevator muscle of the mandible, next to temporal and medial pterygoid. It is quadrangular in shape and is anchored above to the zygomatic arch and below to most of the lateral surface of the ramus of the mandible.
The superficial part of the masseter originates from the maxillary process of the zygomatic bone and the anterior two-thirds of the zygomatic process of the maxilla. It inserts into the angle of the mandible and the posterior part of the lateral surface of the ramus of the mandible.
The deep part of the masseter originates from the medial aspect of the zygomatic arch and posterior part of its inferior margin and inserts into the central and upper part of the ramus of the mandible as high as the coronoid process. (15)
1.2.2. Orbicularis Oris muscle
Orbicularis oris is a complex muscle consisting of deep and superficial fibers that completely encircle the mouth. The deep fibers are continuous with buccinator, while the superficial fibers are derived from other muscles. Its function is apparent when pursing the lips, as occurs during whistling. Some of the fibers originate near the midline from the maxilla superiorly and the mandible inferiorly, whereas other fibers are derived from both the buccinator, in the cheek, and the numerous other muscles acting on the lips. It inserts into the skin and mucous membrane of the lips, and into itself. Contraction of the orbicularis oris narrows the mouth and closes the lips. (15)
Orbicularis oris produces movements of the lips as in “puckering” or whistling. It has an important role in speech by changing the shape of the mouth and lips to help form recognizable sounds. It is also important in mastication as its contraction against the teeth helps to push food back between teeth when chewing. (16)
1.2.3. Suprahyoid muscles
Also called muscles of the floor of the mouth, they are located superiorly to the hyoid bone and include:
Milohyoid
Geniohyoid
Digastric
Stylohiyoid
These muscles pass in a superior direction from the hyoid bone to the skull or mandible and raise the hyoid, as occurs during swallowing. There is a finely tuned dynamic balance among all of the head and neck muscles
1.2.3.1 Digastric
The digastric muscle consists of an anterior and posterior belly connected by an intermediate tendon, which is attached to the body of hyoid bone. (17) The posterior belly originates from the mastoid notch; its fibers run downward inward and forward to the intermediate tendon. (18) The anterior belly of the digastric muscle is attached to the digastric fossa on the internal border of the mandible and runs downwards and backwards to the same digastric tendon as the posterior belly. It depresses and retrudes the mandible, and is also involved in stabilising the position of the hyoid bone and in elevation of the hyoid during swallowing.(17) When the hyoid bone is fixed, the right and left digastrics contract, the mandible is depressed and pulled backwards. When the mandible is stabilized, the digastric muscles with the suprahyoid and infrahyoid muscles elevate the hyoid bone. (18)
1.2.3.2 Mylohyoid muscle
It is superior to the anterior belly of the digastric and forms a muscular diafragm or the floor of the mouth. It originates from the mylohyoid line on the medial surface of the body of the mandible and blends with the mylohyoid muscle from the opposite side. The anterior and middle fibres run downwards and medially towards the midline to insert into a median fibrous raphe, which extends from the symphysis menti to the upper surface of the body of the hyoid bone.(15,16) Only the posterior fibers which originate at the level of the third molar, attach directly on the body of the hyoid.(19) It supports and elevates the floor of the mouth and elevates the hyoid bone.
1.2.3.3 Geniohyoid muscle
It is a strap-shaped muscle, made up of almost parallel fibers that run directly from the origin to insertion. (19) The geniohyoid arises from the inferior mental spine of the mandible and passes backward and downward to insert on the body of the hyoid bone. It is located superficially to the mylohyoid.
Depending upon which end of the muscle is fixed, geniohyoid can elevate the hyoid bone or depress the mandible. When the hyoid is pulled upwards and forwards, the floor of the mouth is shortened and the pharynx is widened ready to receive food.
Electromyography is the study of muscle function through the inquiry of the electrical signal the muscles emanate. (46) It is considered the most reliable and objective method for assessing changes in the electrical activity of the masticatory muscles.
Surface electromyography (sEMG) is currently used in children and adults, due to its advantages over intramuscular electromyography (needle and fine wires), because it is less invasive and easier to handle. {47)
Different sEMG studies of healthy, normo-occlusive adults have found a higher masseter activity in males compared with females during clenching, whereas, at rest position, no sex differences have been reported. (48-50 Comparing the subjects from different sagittal malocclusions, Miralles (51) reported that resting activity of the masticatory muscles was lower in Class I and II patients, than in subjects with Class III. Another study conducted by Antonin (52) demonstrated a significant difference between the masticatory muscles activity during mastication and swallowing, Class II division 2 and Class III.
The various movements of the jaw are produced by cooperative activity of several muscles bilaterally or unilaterally. Mandibular elevation is performed by temporalis, masseter and medial pterygoids and depression by the lateral pterygoids and digastric. The latter show their greatest activity in forceful opening of the mouth at the limit of depression of the mandible. Also, a high level of activity in the milohyoid muscle was reported, during functions of the dentomaxillary system, such as swallowing and suction. (52)
Electromyography of orbicularis oris muscles showed only slight myoelectric activity during rest position, although during aberrant oral activity, such as infant deglutition and thumb sucking, the activity was intense. (53)
3. Ultrasonography
3. 1. General overview
3.1.1 Definition and applicability
Ultrasonographic imaging is an evaluation method of the soft tissues that has been very used in recent decades in the medical domain: ophthalmology, obstetrics, neurosurgery, cardiology, orthopedics, rehabilitation medicine, pediatrics and oncology.
In the domain of medical application, the wavelength emission is generated with the help of electromechanical transductors, which use piezoelectric materials. (54) Ultrasound wavelengths have a frequency higher than 20kHZ, corresponding to the upper limit of human hearing.
3.1.2. Mechanism
Ultrasound (US) is linearly propagated through biological tissue, bearing resemblance to light beams. It is subject to transmission, reflection, refraction and diffraction, acoustic enhancement, absorption and diffusion. When ultrasound encounters an interface, part of the fascicle's energy is reflected back, generated the echo that represents the base of ultrasonographic diagnosis. The acoustic energy that is being reflected depends on the depharence between the acoustic impedance of the two layers forming the interface. (55) For example, the interface between soft tissue and bone will reflect approximately 40% of the fascicle's energy. Air and bone do not permit ultrasound transmission and thus make ultrasonographic investigation impossible. (54)
The fundamental principle of obtaining ultrasonographic information: the transductor functions alternatively as a generator and receptor of US. It regularly generates ultrasound impulses with the duration of 1 millisecond and then acts as a receptor of echo for 99 milliseconds, resulting in a pulse-echo cycle with the duration of 100 millisecond – pulsating characteristic. (55)
Each echo generates an individual electrical impulse. The corresponding electrical impulses can be visually represented in multiple ways. In dentistry, the most used are mode A and B.
Mode A (amplitude modulation) is the visual representation of a single line of spatial information. The horizontal line represents echo amplitude, while the vertical represents the depth from which the echo was received.
Mode B (brightness modulation) is the base upon which two-dimensional images are obtained. By moving the transductor, thousands of such lines of information are obtained side by side, generating the two-dimensional ultrasonographic image of a limited region of the organism, obtained in the plane that within the transductor's movement. (55)
Sound travels faster through solid surfaces, intermediary fast through liquid and slowly through gas. Most liquids behave as an ideal liquid, which means that energy transmission happens in longitudinal wavelengths. Propagation speed for ultrasound wavelengths in a liquid environment depend on particle density and resistance to compression Soft tissue can be considered such as a viscous fluid. Because density and compression module of most soft tissue is similar to that of water at 37° C, a propagation speed of approximately 1540 m/s is associated with mode B of pulse-echo. Variation in sound speed, due to heterogeneous distribution of soft tissue, or differences in local temperature, can determine errors in measuring distances or can cause image distortions. More complex issues can arrise when ultrasound interferes with the tooth or the alveolar processes – which represents a limiting factor for mode B. (56)
3.1.3. Advantages and disadvantages
The disadvantages of ultrasonographic imaging are limited to the possibility of causing tissue damage and teratogen effects in the case of lengthy exposer due to heat and acoustic cavitation. Still, ultrasound is used as a diagnostic method, and at a low intensity and pressure level, is proven to be minimally invasive.
Among the advantages of ultrasound are: relatively low cost, real-time diagnostic ability, evaluation of the mechanical tissue properties, ergonomic and non-invasive method. Despite these advantages, ultrasound is not being used in dentistry, and scientific literature holds few studies regarding the evaluation of oral structures.
Panoramic and retroalveolar X-ray and CT are conventional diagnostic methods in dentistry, yet their greatest disadvantage is the generation of ionizing radiation, which can be detrimental for patients and can not be used abusively. CT has a high cost, x-rays do not show morphological information such as ultrasound does.
3. 2. Periodontal ultrasound
The first studies were undergone by Baum in 1963, which used a 15 MHz transductor to visualize the internal structure of the tooth, yet the quality and clarity of the RG signal were not satisfactory. (57) Throughout time, and more-so within the last decade, ultrasonographic investigations have been used to detect cavity lesions, fissures and fractures of the tooth, soft tissue lesions, fractures of the viscerocranium, periodontal structural defects, measuring gum thickness, diagnosing pathology of the temporomandibular joint, and in oral implantology. (54)
The acoustic properties of enamel, dentine and soft tissue have been determined and thus studies can be undergone on models simulating dental structure.
Table I. Tissue acoustic properties (58)
Periodontal ultrasonographic evaluation dates from the year 1971, when Spranger tried to evaluate the height of alveolar growth in patients with periodontal disease. He reached the conclusion that this technique can offer important additional data to retroalveolar X-ray, if technical difficulties were to be overcome. (59)
In ideal experimental conditions (wavelengths propagating perpendicularly on smooth surfaces, perfectly aligned), Lost et al. (1988) managed to determine the diameter of periodontal ligaments with the help of type A uni-dimensional scans on treated pig mandibles. (60) In 1989, the team led by Lost managed to obtain one-dimensional ultrasonographic images of buccal/oral alveolar growth on pig mandible, succeeding in reproducing every measured mark. They also concluded that the 20 MHz transductor offers more accurate images than the 10 MHz, due to heightened resolution. (61)
The study lead by Eger (1996) proved the existence of a strong correlation between the measurements obtained with a 5 MHz transductor and those obtained with an endotonic instrument and calipers. Muller used the same transductor in the years 1998, 1999 and 2000 in order to determine the gum thickness before and after mucogingival surgery. (62)
Ghorayeb et al. (63) elaborates a study regarding diagnosing periodontal disease Due to the complexity of the periodontal structure and the small acoustic impedance at the interface between the gum and the ligaments, he concluded that it is still very difficult to evaluate these tissues ultrasonographically.
These technical problems have been overcome in the scientific research of Chifor et al. by fixing necessary markers in horizontal monitoring of bone resorption with the help of 20Mhz ultrasonography. The transductor had a longitudinal placement on the lateral area of the alveolar mandibular bone, and the results were compared the CT measurements and microscopic evaluation. Examination was undergone on the lingual surface of 4 pig mandibles and the following were identified: the enamel-cement junction, dental root, periodontal space at the emergence of the alveolar process, the coronary portion of the cortical bone (in order to evaluate horizontal bone resorption), as well as the distance between the enamel-cement junction and the coronary limit of the cortical bone. A statistically significant correlation was found between the measurements made with the 3 techniques. The authors concluded the accuracy of the measurements to be very high, the ultrasonographic method being of use in monitoring periodontal pathology. Because of the high frequency of ultrasound, access in the interdental area is very small and thus the authors have proposed using a specially adapted transductor for the oral cavity. (64)
Mahmoud et al. investigated the possibility of using a high frequency ultrasound system to reconstruct 3D images of the periodontal defects on the vestibular surface of human mandibles, in less than 30 seconds. The anatomic markers obtained on the 3D image corresponded to the mandibular anatomical structures. The authors of this study suggest the possibility of using high frequency ultrasonography in early diagnosis of severe periodontal disease. (65)
Tsiolis et al. (66) used 20MHz ultrasound in a study that attempted the evaluation of periodontal structures on pig mandibles, which have been froze beforehand. After gradual thawing, the transducer was placed at the level of the free gingival margin. All measurements were made from the level of the vestibular surface, due to accessibility. One marker was placed in the middle of the probe and then aligned to the top of the targeted tooth's cusp. Statistical analysis demonstrated that ultrasonography offered satisfying results regarding reproducibility and accuracy.
Evaluating gingival thickness with the help of a noninvasive diagnostic method such as ultrasound would help establishing the diagnosis and treatment plan in various clinical settings. Thus, many studies tried to develop a precise ultrasound device, adapted do the oral cavity. (67,68)
Müller and Könönen evaluated gingival thickness on 33 patients. The transductor, having a 4 mm diameter, was applied with moderate pressure, at the middle of the vestibular surface of every tooth. By measuring the recording time of echoes, gingival tissues was determined in the course of 2-3 seconds, being at least 0.5 mm, and the obtained image having a 0.1 mm resolution. The probing depth and gingival attachment were also evaluated, demonstrating the accuracy of ultrasound in the evaluation of periodontal structures. (69)
Another study, lead by Savitha and Vandana, on 32 patients in 338 areas, used module A of graphic representation of ultrasound wavelengths. They compared the measurements through the imagistic method with the traditional transgingival probing method. On the vestibular surface, the clinically evaluated mean thickness was 1.08, while the ultrasonographic mean was 0.86, and the thickness and the interdental papilla was 1.26 and respectively 0.77. The difference between the two methods was significant in the evaluation of the thickness of interdental papilla and not significant at the middle of the vestibular surface, especially in the canine area. (70)
Using the same ultrasound device (SDM, KRUPP Corporation, Essen, Germany; range of measurement 0.3-8.0 mm; 0.1 mm resolution; 5 MHz frequency; 3.0 sensor diameter mm), a more recent study lead by Cha evaluated the thickness of the fixed vestibular and palatine mucosa regarding the insertion of mini-implants with orthodontic purpose. The research was undergone on 61 patients aged between 19-35 years, of which 28 men and 33 women, the evaluated areas being: the proximity of the mucogingival line, at a distance of 4, respectively 8 mm from the free palatine gingival mucosa. The results demonstrated a greater gingival thickness in the maxillary area in men than in women, and a similar one in the mandibular and palatine region. The greatest gingival thickness was found in the anterior maxillary and posterior mandibular area and 4 mm in the anterior palatine area and 8 mm in the posterior area. (71) Thereby, the study demonstrates the applicability of ultrasound imaging in orthodontic practice, regarding the area selection for mini-implant insertion.
Due to technical limitations in the design of oral transductions and the lack of knowledge regarding the acoustic properties of human gingival tissue, ultrasonographic studies continue to be a challenge to researchers.
3.3 Ultrasonographic evaluation of masticatory muscles
Another use of ultrasound upon the cephalic extremity is the reproducible and effortless evaluation of muscle parameters pertaining to the head and neck complex. (72,73) This technique represents an improvement compared to conventional methods, regarding cost and speed of evaluating the thickness of the masseter muscle.
Radsheer et al. (74) in several studies, demonstrated that ultrasonography is an accurate and reproducible method for measuring the thickness of the masseter in vivo, due to its superficial position, and the changes that appear during growth in relation to its function.
In the study lead by Serra (75), the authors demonstrate higher reproducibility of ultrasound on the relaxed muscle than on the contracted muscle. Most studies were undergone on the masseter and temporal muscles, and the results were related to: age, sex, and muscle surface, force of mastication, weight, stereotype of mastication and pathology of the temporomadibular joint. The authors preferred ultrasound method, to CT and MRI, because of the priorly mentioned benefits. This technique is uncomplicated, and represents a considerable improvement relative to conventional methods for assessing masseter thickness, particular in terms of clinical availability and cost.
Recent studies demonstrated negative correlation between the thickness of the masseter muscle and the vertical growth pattern: hyperdivergent subjects proved a minimal thickness during contraction as well as during relaxation, compared to the normal and hypodivergent groups. (76) The study conducted by Lione, on 60 prebuberal subjects, demonstrated the accuracy of ultrasound imaging in evaluating muscles of mastication in vivo in children, as it allows a two-dimensional (2D) and a 3D analysis of the whole masseter. (77)
Frequencies between 5 to 13 MHz of the ultrasound transductors were used in different studies Kiliaridis (78); 1994; Emshoff, Bertram (7.5 MHz) (73), Trawitzki et al. (7.5 MHz) (79), and Naser-ud-Din et al. (5-13MHz) (80), conclusions were very similar: muscle thickness can be successfully measured using ultrasonography.
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Personal
Contribution
1. General hypothesis/objectives
The main objective of the present thesis is to introduce a new, reliable and accurate diagnostic tool in the field of dentistry. The research focused on two main directions: the evaluation of the anatomy and physiology of the periodontium and the characteristics of the head and neck muscles in relation to the development of bone and tooth structures.
The periodontal tissues are assessed mainly through clinical examination and conventional radiology, which do not provide enough information regarding the complex dynamic changes during masticatory and orthodontic forces.
The influence of each component of the dentomaxillary system over the development of musculoskeletal structures is not well understood. Multiple studies show divergent ideas regarding the etiology of dentomaxillary anomalies, yet precise knowledge lacks from the literature. Ultrasonography, with its simple, non-invasive and accurate characteristics, should be part of the clinical assessment of the head and neck soft tissues.
On the basis of the above-mentioned premises, the specific objectives of the present thesis were:
1. To determine whether high resolution Ultrasonography (40MHz) is a suitable method for healthy periodontal evaluation and if so, which are the structures that could be imagined on the ultrasound image.
2. To assess the reproducibility of ultrasonography on pig jaw periodontium, comparing the data obtained by two-trained radiologist.
3. To analyse the correlation between ultrasound and clinical measurements in respect to sulcus depth.
4. To determine if the changes that appear during orthodontic tooth movement, in the anatomical structures of the periodontium, could be monitored using ultrasonography
5. To demonstrate the influence of the orofacial muscles over facial morphology and tooth position, by means of ultrasound.
6. To investigate the influence of suprahyoid muscles on the position of the mandible in class 1 and 2 patients. The parameter assessed was the muscular thickness with the aid of ultrasonography.
2. Study 1. Evaluation of periodontal tissues using 40 MHz Ultrasonography- Feasibility Study
2.1. Introduction
The evaluation of periodontal tissues can be done through clinical examination (such as probing depth, assessment of gingival recession, tooth mobility) and complementary methods (radiological examination, blood tests and microbiological analysis). (1) Clinical examinations are time-consuming, user dependent and have disputable reliability. (81) X-ray examination (periapical, bitewing, panoramic) is not cost effective but the exposure to ionizing radiation and the lack of information about bone resorption from the buccal and lingual surfaces of the teeth represent important drawbacks of the method. (1)
Periodontal changes can be assessed more accurately using new imaging techniques like: cone-beam computed tomography; optical coherence tomography; optical spectroscopy and ultrasonography. (82-84)
Very high-resolution ultrasonography has been used mainly for the exploration of the skin or the anterior chamber of the eye. With the advent of new, easy to use transducers, novel applications emerged such as the exploration of the oral cavity or small laboratory animals (mice). (85) These complementary methods are attractive because they are noninvasive and comfortable for the patient.
Recent studies showed the validity and reliability of ultrasonography in the measurement not only of gingival thickness (86) but also of other periodontal structures, which cannot be assessed through inspection and palpation. (66) Very high resolution (20 MHz) in vitro US studies of periodontal structures using a dermatology-dedicated scanner further improved the knowledge on periodontal US appearance. (64, 87)
Xiang et al. demonstrates that ultrasonography can identify periodontal and oral tissue in vivo or ex vivo without the need to process or color and fix the samples, procedures necessary to other techniques (infrared spectroscopy and OCT). In addition, they conclude that ultrasonography represents a fast, noninvasive method, easy to use and reproducible in monitorizing periodontal disease. (84)
2.2. Objectives and Research Hypotheses
The aim of the present study is to assess the feasibility of in vivo transgenial assessment of the periodontal structures in humans, with the use of a 40MHz transducer. The null hypothesis of the research consists of
2.3. Materials and methods
Four volunteers with clinical healthy periodontal tissues were included in the study. Ethical approval and written consent were obtained.
In all volunteers, the 4 lower bicuspids were first evaluated clinically, through inspection and palpation. The endooral examination of the periodontal structures showed no signs of inflammation, resorption, tooth mobility or bleeding on probing. The patients did not complain of any symptoms regarding the periodontal tissues. The gingival sulcus depth (C1) (Fig .6 ) and the length of the clinical crown (C2) were measured with a periodontal probe.
After clinical assessment, the premolars were imaged ultrasonographically from the buccal incidence, with a percutaneous transgenial approach. A commercially available ultrasound scanner (Ultrasonix SonoTouch) with a linear 1.5 cm footprint, wideband 8-40 MHz transducer, operating at 40 MHz, was used (Fig. 7).
A fixed landmark (no.20 gutta-percha point) placed in the gingival sulcus, was used as reference. The images were obtained by positioning the transducer in a coronal plane in the lateral area of mandible alveolar bone.
The 40 MHz image revealed the cortical bone, the buccal surface of the tooth, the gingival sulcus with the reference point and the periodontal space in its most coronal position.
On the ultrasound image, the following micrometric level measurements were performed: (Fig. 8-10
):
D1 gingival sulcus depth
D2 free gingival thickness
D3 width of the periodontal space in the most coronal position
D4 distance between marginal gingiva and alveolar crest
D5 height of the clinical crown and
D6 height of the anatomic crown
Statistical differences between clinical and ultrasound measurements (sulcus depth and clinical crown) were evaluated using Wilcoxon Signed Rank Test, z= -1.221.
Fig 8. Coronal US image of the dento-periodontal structures at 40 MHz. Tick marks on the left side of the image indicate 2 mm.
Fig 9. Dental and periodontal US anatomy – 40 MHz image (top) and anatomic sketch (bottom – inspired after ref. [11]): 1 – dentin; 2 – enamel; 3- cementum; 4 – cemento-enamel junction; 5 – supra crestal fibers; 6 – gingival epithelium; 7 – periodontal ligament; 8 – crest of alveolar bone; 9 – gingival sulcus.
Table II Ultrasound measurements of the periodontal structures (millimiters)
2.4. Results
Fig 10. Measurements performed on the US image.
The 40MHz ultrasound image revealed the cortical bone, the anatomical and clinical crown and root, attached mucosa and the gingival sulcus.
The findings for D1 varied between 1.2-1.86 mm and for D2 between 0.65-1.34mm. The smallest variation of the values was found for D3: 0.21-0.39. The results of the measurements are presented in table II.
No statistical differences were found between clinical and imagistic measurements in respect to sulcus depth (Wilcoxon Signed Rank test, z = -1.221 based on positive ranks).
2.5. Discussions
The possibility to assess periodontal tissues with a dedicated 20 MHz dermatology scanner (Dermascan) was well documented by Chifor and coworkers, in an in vitro study. (64) The results demonstrated a high correlation between ultrasonography, Cone Beam computed tomography and direct microscopic section measurements as a reference standard, therefore eliciting the possibility to use 20 MHz ultrasonography in monitoring periodontal tissues.
In 1989, the team led by Lost managed to obtain one-dimensional ultrasonographic images of buccal/oral alveolar growth on pig mandible, succeeding in reproducing every measured mark. They also concluded that the 20 MHz transductor offers more accurate images than the 10 MHz, due to heightened resolution. (61)
To the best of our knowledge this is the first report on the use of very high-resolution ultrasound (40 MHz) in the assessment of the periodontal structures. The aim of this pilot study was to demonstrate the possibility to use high-resolution ultrasonography in vivo, for periodontal evaluation. Using the gutta-percha point as a landmark in the gingival sulcus of the buccal surface of the tooth, it was possible to determine the clinical and anatomical crown, the root with the periodontal space, the width of the free gingival margin and the distance between the alveolar crest and free gingiva.
This pilot study compared clinical data (probing depth) with ultrasound measurement of the sulcus depth and found a strong correlation between the two. However, the main drawback of the research is the absence of a gold standard technique to validate the findings. Future studies will focus on the accuracy and reproducibility of the ultrasound method related to the periodontal anatomy.
The ultrasound method has two main advantages: the absence of exposure to ionizing radiation, and a wider range of information. The present study confirmed the feasibility of external, transgenial approach for very high resolution US assessment of the periodontal structures. Further, in vitro and in vivo research is required and being performed, for the complete understanding of the US appearance of normal and diseased periodontal structures.
2.6. Conclusions
1. To the best of our knowledge this is the first report of the use of very high-resolution (40 MHz) ultrasound in the assessment of periodontal structures.
2. Data regarding gingival sulcus depth, free gingival thickness, and width of periodontal space, distance between marginal gingiva and alveolar crest, height of clinical and anatomical crown were obtained. No statistical differences between clinical and imagistic US measurement were obtained, in respect to probing depth.
3. Ultrasonography provides a non-invasive technique for periodontal assessment but further studies are mandatory to define a potential clinical role of the method.
3. Study 2. Reproducibility of high-resolution ultrasonography for measuring the periodontal structures.
3.1 Introduction
Ultrasonography is currently used to assess soft tissues in obstetrics, neurosurgery, cardiology, orthopedics, rehabilitation medicine, pediatrics and oncology.
Considering the many advantages of ultrasound: ergonomic, non-invasive, cost-effective and accurate, it would be beneficial to use it as a diagnosis tool in the field of dentistry, especially in periodontology.
Nowadays, periodontal structures are assessed mainly through clinical examination (inspection and probing depth) and conventional radiology (periapical radiography and bite-wing). (1) Unfortunately, clinical examinations are user-dependent, time-consuming and not very reliable (inter- and intrauser reliability is rather low). (81) While clinical attachment level can be assessed through periodontal probing, bone morphology and level can only be truly evaluated through radiographic evaluation, despite its known shortcomings (radiation exposure and lack of information on the lingual and buccal surfaces of the teeth).
Even though MRI and CT methods could overcome these disadvantages, a real time, non-invasive, low cost imaging technique, such as ultrasonography, would be preferable.
Tsiolis et al. (66) used 20MHz ultrasound in a study that attempted the evaluation of periodontal structures on pig mandibles, which have been froze beforehand. Statistical analysis demonstrated that ultrasonography offered satisfying results regarding reproducibility and accuracy. The authors speak of progress in ultrasonographic periodontal evaluation, of interest to the future of dentistry, if transductors with a frequency higher than 20MHz were to be used.
Therefore, a recent study lead by Zimbran et al. (89) was undergone in vivo with the use of 40MHz ultrasound transducer. The result suggested that this diagnostic method could be used in the future as a routine investigation in the evaluation and monitoring of periodontal tissues.
3.2. Objective and Research Hypothesis
The aim of this study was to investigate whether the ultrasound method can supply credible information regarding the structure of the periodontal tissues, comparing to probing depth and if high-resolution ultrasonography is a reproducible technique in the field of periodontal assessment. The null hypotheses were: 1. There is no statistical agreement between the probing depth technique and ultrasound measurement of the sulcus 2. Significant differences exist between the results of the two operators.
3.3. Materials and Methods
Pig jaws were selected as the experimental model, due to the similarities with the human periodontium and ease of obtaining the US scans. Twenty teeth, on 4 pig jaw mandibles were evaluated on their lingual and buccal surfaces using two methods: clinical probing depth and ultrasound imaging. In order to obtain reproducible and similar findings of the same site, a glued gutta-percha point was used as a landmark. Clinical probing depth (s) was performed with a manual conventional probe (Williams’s style), with the markings at 1, 2, 3, 5, 8, 9 and 10 mm. The evaluation site was the buccal and lingual surfaces of the lateral teeth, in contact with the gutta percha point. (fig. 11)
Ultrasound imaging involved the assessment of the periodontium using 8-40 MHz transducer with a linear 1.5 cm footprint. In order to test the technique’s reproducibility, two trained radiologists performed the ultrasound examination.
The pig mandibles were examined, not later than 3 hours after the sacrifice of the animal. The area of interest was between the dental crown near the bonded gutta-percha-landmarks and the mucogingival line. (fig. 12) The transducer was positioned in a longitudinal plane on the lateral alveolar bone, both lingually and buccaly.
The following measurements were recorded: width of the periodontal space in the most coronal position (D1), gingival sulcus depth (D2), free gingival thickness (D3), length of the suprachrestal fiber (D4) and height of the anatomic crown (D5). (fig. 13)
Recorded data were analyzed graphically and with the Shapiro-Wilk test for the relevance of normal distribution. Intra-class correlation coefficients were calculated for D1-D5 measurements in order to assess the reproducibility of the US method when two different raters performed the measurements.
Bland-Altman plots were computed to assess the agreement between the Perio probing and US measurement methods.
Fig.12 The gutta-percha (arrow) placed in the gingival sulcus
Fig. 13 Dental and periodontal area of assessment
3.4 Results
Intra-class correlation coefficients (ICC) and 95% confidence intervals for each of the five micrometric measurements are presented in Table III.
Mean values and standard deviations of the ultrasound measurements of the periodontium are presented in Table IV
No statistically significant differences were found between the two measurement methods for s (probing depth) and D2 (gingival sulcus depth) data sets (p>0.05), the first null hypothesis being rejected. Analyzing the Bland-Altman plot it could be observed that the mean difference between the s and D2 data sets was very close to 0 (-0.11) following a horizontal linear trend, suggesting a good agreement between the two measurement methods.
3.4 Discussions
The ICC is a general measurement of agreement between two or more different raters who perform measurements on the same sets of subjects. ICC is interpreted as follows: poor agreement for values between 0-0.2; fair agreement for values between 0.3-0.4; moderate agreement for values between 0.5-0.6; strong agreement for values between 0.7-0.8; almost perfect agreement for values >0.8.
In our study, for all of the areas evaluated, except D5, the ICC showed values very close to 1, suggesting a good agreement between the raters, and thus a very good reproducibility of the method.
The evaluation of the ultrasound image is considered to be subjective and depended upon the experience and knowledge of the radiologist. (55) In our study, two trained-radiologists from the same University Center performed the scanning of the pig jaw mandible, one our apart. The mechanical configuration of the ultrasonic instrument and the ability of the operator are both important factors in determining the reproducibility of the method. (90)
The two operators obtained very similar values for the width of the periodontal space in the most coronal position (D1), gingival sulcus depth (D2), free gingival thickness (D3) and length of the suprachrestal fiber (D4), therefore the null hypothesis for D1, D2, D3 and D4 was rejected. All these measurements had the same reference point for calculating the periodontal dimensions, thus explaining good agreement between the findings.
However, when referring to and length of the anatomical crown (D5), significant differences between the two operators were found. The variability in the position of the CEJ and the location of the tip of the tooth may have influenced the interpretation of the observers.
3.5 Conclusions
Ultrasonography represents a reliable and reproducible method for the assessment of periodontal structures, such as: free gingival thickness, gingival sulcus depth, and length of the suprachrestal fibers and width of the periodontal space in the most coronal position. Future studies using microscopic measurements and x-ray of the periodontal structures will be made in order to test the accuracy of the method.
Study 4. Ultrasonographic evaluation of periodontal changes during orthodontic tooth movement
4.1. Introduction
Tooth movement by orthodontic force application is characterized by remodelling changes in periodontal tissues, including, periodontal ligament (PDL), alveolar bone, and gingiva. (92)
The response to applied orthodontic forces is bone formation (due to osteoblasts) on the tension side and bone resorption on the compression side (due to osteoclasts) of the lamina dura. (93-95)
However, this represents an oversimplification of the three dimensional changes that appear in the microarchitecture of the periodontium. Several studies used histological staining and light microscopy, scanning electron microscopy (96), finite element models (97,98), genetic modifications (100) in order to demonstrate the complexity of periodontal transformations. These methods need sophisticated procedures and animal sacrifice, and cannot be used to evaluate longitudinal changes of periodontium due to orthodontic forces. (101)
A new technique aiming to observe real time, in vivo modifications induced by OTM in the morphology of PDL, alveolar bone and cementum would be beneficial in order to better conduct the treatment plan and to evaluate the tissue response to orthodontic forces.
Ultrasonography represents a new, accurate and noninvasive technique aimed to evaluate periodontal tissues. Several studies (102, 103) demonstrated that ultrasonography could be successfully used in assessing the cortical bone, sulcular depth, periodontal space, and length of the anatomical crown and the characteristics of the gingiva.
4.2 Objectives and research hypothesis
The aim of the present study was to assess whether changes that appear during orthodontic tooth movement in the anatomical structures of the periodontium can be monitored using ultrasonography. To the best of our knowledge, this is the first attempt to understand the modifications that appear during OTM using ultrasonic measurements of periodontal tissues, especially the gingival characteristics.
The null hypothesis of the present study was that by using ultrasonic measurements, anatomic features of periodontal structures (gingival sulcus depth; free gingival thickness; distance between marginal gingiva and alveolar crest; and width of the periodontal space in most coronal position) of teeth subjected to OTM would indicate the same results, overtime.
4.2 Materials and methods
The study was conducted on 5 patients, aged 14-25. In every case, maxillary canines were subjected to orthodontic retraction, as part of the orthodontic treatment plan; 8 teeth were measured as a result, all with typical dental anatomy (average root sizes, bone insertion, and shape). The institutional Ethical Committee approved the research protocol, and an informed consent was obtained from the subjects.
All patients were fitted with Alexander’s prescription preadjusted edgewise appliance of .018 slot (MiniMaster American Orthodontics). Upper first premolars were extracted due to severe crowding (8 upper bicuspids – 5 first right premolars and 3 first left premolars). (Fig. 15)
After levelling and aligning, the canines were distalized on 016 SS archwire, using elastomeric memory chain (AO) with a net force of approximately 100cN. The elastomeric chain delivering the force was measured with a dynamometer.
The average time for canine retraction was 5 months, which varied depending on the size of the initial space and bone density.
Fig. 15 Treatment mechanics after extraction of the premolars
A commercially available ultrasound scanner (Ultrasonix SonoTouch) with a linear 1.5 cm footprint, 40MHz -transducer was used, in order to evaluate the periodontal structures of the canines. Ultrasonographic scans were performed in three distinct areas of the canine’s buccal surface: mesial, middle and distal, with a percutaneous transgenial appropach.
The reference point was the bracket, placed in the center of the canine, which appeared hiperechoic on the US scan. The images were obtained by positioning the transducer in a longitudinal plane in the lateral area of the maxillary alveolar process. (Fig. 16)
The US scan revealed the cortical bone, the buccal surface of the tooth with the bracket placed in the center, the gingival sulcus and the periodontal space in the most coronal position. (Fig 17) On the 40 MHz image, the following distances were measured at a micrometric level: D1- gingival sulcus depth; D2- free gingival thickness; D3-distance between marginal gingiva and alveolar crest; D4-width of the periodontal space in the most coronal position.
All scans were performed by a single trained radiologist, in different moments of the orthodontic treatment: The US scans were performed in three different moments of OTM: before distalization (moment 1), after 2 days of 100 cN force delivery, when tooth displacement was expected (moment 2) and after 30 days to 60 days or the postlag stage (moment 3).
Paired sample t-test and Pearson’s correlation test were used to compare recorded data between different moments of the OTM.
4.3 Results
Average and standard deviations (SD) of US measurements (mm) (D1, D2, D3, D4) on the Mesial (MES), Middle (MID), Distal (DIS) side of the upper right canine, in three distinctive moments (moment 1, 2, 3) of OTM are presented in Table V.
Table V- Average and SD of US measurements (mm)
An increase of D1 (sulcus depth) was observed in all three areas of the periodontium, immediately after force delivery, whereas at the end of tooth movement the distance decreased considerably. However, significant differences were observed only for the mesial (p=0.031) and middle areas (p=0.048). A strong correlation was also noted for D1 before and immediately after force delivery (r=0.945, p<0.05).
D2 (free gingival thickness) was the most stable measurement during OTM, the free gingival thickness slightly decreased from the initial moment, yet no statistically significant difference being observed between the measurements (p>0.05).
The distance between marginal gingiva and alveolar crest (D3) was strongly correlated before and immediately after force delivery only for the mesial area (r=0.828, p<0.05). Furthermore, a very strong inverse correlation was observed for the middle area between moment 2 and 3 (r=-0.998, p<0.05).
The width of the periodontal space (D4) increased immediately after force delivery on the mesial side, whereas on the distal area the value decreased. In the last stage, D4 had a tendency to return to its initial size. Nonetheless, no significant difference was found between the measurements, in none of the three moments evaluated (p>0.05).
Charts representing the evolution of periodontal structures for D1 (sulcus depth), D2 (free gingival thickness), D3 (distance between marginal gingiva and alveolar crest), D4 (width of the periodontal space in the most coronal position), on the mesial area (blue line), middle area (red line), distal area (green line), during OTM, are presented as it follows:
4.4 Discussions
The definition of orthodontic tooth movement by Proffit is the result of a biologic response to interference in the physiologic equilibrium of the dentofacial complex by an externally applied force. (104)
Two main mechanisms are considered to be responsible for tooth movement—the application of pressure and tension to the PDL (105-107), and bending of the alveolar bone. (108,109)
However, few studies focused on the real time changes induced by OTM in the area of marginal periodontium. In this study, we propose a novel approach for the clinical evaluation of the free gingiva, gingival sulcus, suprachrestal fibers and position of the bone chrest. High-resolution ultrasonography (40 MHz) was used, for a better visualization of the superficial periodontium.
In the present study, a standardized orthodontic force of approximately 100cN was applied to the canine. The period of time between the scans were deliberately chosen, according to the lag stages found in the literature:
1. Initial stage (24-48h) represented by tooth displacement in the periodontal ligament space.
2. Lag stage lasts 20–30 days and is characterized by the formation of necrosis and hyalinization. In this lag stage there is little or no tooth movement.
3. Postlag stage, characterised by tooth movement mediated by bone remodelling through the agency of osteoclasts and osteoblasts on a background of neoangiogenesis. (110)
The null hypothesis was rejected since significant changes in the periodontium were measured with US method after orthodontic tooth movements. The distance between marginal gingiva and alveolar crest (D3) was strongly correlated before and immediately after force delivery only for the mesial area (r=0.828, p<0.05). Furthermore, a very strong inverse correlation was observed for the middle area between moment 2 and 3 (r=-0.998, p<0.05).
An increase of D1 (sulcus depth) was observed in all three areas of the periodontium, immediately after force delivery, whereas at the end of tooth movement the distance decreased considerably. However, significant differences were observed only for the mesial (p=0.031) and middle areas (p=0.048), on the opposite side of that of force delivery. A recent study demonstrated that gingival inflammation and dental plaque increase during orthoodntic treatment (111), which may explain the values obtained in our study.
The thickness of human PDL is reported to be around 0.1– 0.3 mm. (112) In our study, measurements of periodontal space in the most coronal position (D4) varied between 0,2 and 0,52, with an average value of 0,28 in the initial stage. The width of the periodontal space increased immediately after force delivery on the mesial side, whereas on the distal area the measurement decreased. In the last stage, D4 had a tendency to return to its initial size. Nonetheless, no significant difference was found between the measurements, in none of the three moments evaluated (p>0.05).
Although D2 (free gingival thickness) was the most stable measurement during OTM, no statistically significant difference was observed between the measurements (p>0.05). The prevalence, extent, and severity of gingival recession were correlated with past orthodontic treatment. (113) Future studies will be carried out to see if there is a significant change in the thickness of the gingiva, in the last stage.
The limit of the study was the relative small sample of subjects (5 patients- with 8 canines to be distalized), one of them was lost during the research. Further assessment of the modifications in the width, thickness and height of the marginal peridontium is needed, on a larger group of patients.
4.5 Conclusions
1. Ultrasonographic measurements, with high resolution, detected changes in the anatomic landmark of periodontal during orthodontic tooth movement.
2. Significant changes occurred immediately after force delivery on the middle and mesial area of the canine, for sulcus depth measurement and distance between marginal gingiva and alveolar crest.
.
5. Study 4. Ultrasonographic evaluation of masseter and orbicularis oris muscles in different in different vertical skeletal patterns.
5.1. Introduction
The interaction between masticatory muscle function and craniofacial growth was demonstrated in previous studies. (114-117) The bones that form the maxillofacial region are membranous bones, being more susceptible to the environmental factors (stimulating influence of muscles, bad habits, parafunctions) as compared to the long bones, which are formed by cartilaginous ossification. (118) The effects of muscle thickness on bone morphology can be explained by a theory that is recognized in the field of biodynamics as Wolff ’s law*, which correlates the internal structure and the shape of the bone to the function. (119)
The masseter is a superficial, bulky, powerful muscle, which elevates the mandible. (120) Its relation to vertical growth pattern was demonstrated in different studies, in children and adults. (121-123)
The position of the teeth is also influenced by orofacial muscles: the orbicularis oris muscle is a concentric, sphincter-like muscle around the mouth that closes, withdraws, and protrudes the lips. (121) Studies have shown that the upper and lower lips tend to move posteriorly following an orthodontic treatment with extractions and anterior teeth retraction. (122) However, changes in the soft facial profile also seem to be related to other variables, such as lip strain, structure, and thickness, together with incisor retraction. (123,124)
Radsheer (74,125) et al. and Bakke (126) et al. demonstrated that ultrasonography is an accurate and reproducible method for measuring the thickness of the masseter in vivo, due to its superficial position, and the changes that appear during growth in relation to its function. Ultrasonography has several advantages over MRI and computerized tomography because it is a fast, inexpensive and noninvasive technique, the equipment can be easily handled and transported. (126)
However, there are few studies that demonstrate the influence of the orofacial muscles over facial morphology and tooth position. (127)
The aim of the study was to evaluate the characteristics of the masseter and orbicularis oris muscles in patients who undergo orthodontic treatment, using 5-13MHz ultrasonography and to determine if these measurements are connected to certain dentofacial morphology.
5.2 Materials and Methods
Thirty patients (13 men and 17 women; age range 10-25 years) who underwent orthodontic treatment were selected to participate in this study. The subjects, with a normal physical development, no marked jaw asymmetries or craniofacial disorders, no congenital or developmental anomalies of the lips, mouth or face and no history of orthognathic tretment, were divided in three equal groups of 10, according to their anterior cranial base and mandibular plane angle (SNMP) and FMA: (Fig. 19)
Hypodivergent
Normodivergent
Hyperdivergent.
Cephalometric measurements Conventional skeletal landmarks were traced and 2 linear and 9 angular measurements were analysed in order to define the vertical growth pattern and position of the upper and lower teeth of the subjects (Table VI).
Table VI Measurements on the lateral cephalometric radiographs.
Ultrasonographic measurements: To analyze the orofacial muscles, a broadband linear transducer (Hitachi EUB 8500) with 5-7 MHz operating frequency was used. Water –based gel was applied to the probe before the imaging procedure.
For each subject, US scans of the masseter and orbicularis oris muscles were performed by a single trained radiologist, at the same real time scanner.
The patients were examined in a supine position for the assessment of the masseter and orbicularis oris muscles, only the orientation of the transducer varied:
1. For the masseter muscle, the transducer was oriented 30° to the Frankfort plane, perpendicular to the ramus of the mandible. (125,128) The scans were performed bilaterally during muscular relaxation (rest position, lips in gentle contact, presence of freeway space) and contraction (dental arches in maximum intercuspation).
Muscular thickness was measured as the linear distance between the most superficial and deepest points of the muscle, halfway between the origin and insertion (125) in two different conditions: in a relaxed position of the muscles, when the teeth were occluding gently and during maximal clenching, with the masseter muscle contracted. The measurements were made directly from the image at the time of scanning. (Fig. 21)
2. For the orbicularis oris muscle, the transducer was oriented longitudinally on the upper and lower lip of the patient, using a silicon pad for a better visualization. (g. 22)
Fig. 22 Position of the transducer for the assessment of orbicularis oris muscles
Fig. 23 US images of the superior and inferior orbicularis ori of the three vertical pattern a. hypodivergent b. normodivergent c. hyperdivergent
Statistical methods: Histograms and Shapiro-Wilk test were used to assess the normality of the data. Multivariate analysis of variance was conducted to test the hypothesis that muscular thickness does not vary among vertical skeletal patterns. Muscular thickness (masseter and orbicularis oris), FMA, SNMP, IMPA and U1SN were considered dependent variables, while vertical pattern, age and gender were set as independent variables. Multiple comparisons between variables were adjusted by the Bonferroni method at a significance level of α=0.05. The Pearson’s Correlation Coefficient was used to analyze the relationship between muscular thickness and vertical skeletal patterns.
5.3 Results
Thirty subjects (14 males, 16 females) were included in the present study: 9 subjects were normo- (1), 12 subjects were hipo- (2), and 9 subjects were hiper- (3). The age distribution was as follows: group 1: 10-15 years – 11 subjects, group 2:16-20 years – 7 subjects, group 3:21-25 years – 11 subjects, group 4:26-30 years – 1 subject.
Table VII Distribution of subjects into groups according to the vertical facial pattern.
All recorded data followed a normal distribution; hence parametric tests were further used to test the null hypothesis.
The results of the multivariate tests showed a significant interaction effect between cephalometric measurements (SNMP, Jarabak ratio and Sum angle) and: vertical skeletal pattern, age and gender (p<0.05). No significant effect was observed between muscular thickness of masseter and vertical skeletal pattern, age, or gender (p>0.05).
The masseter muscle thickness varied among the three vertical dentofacial patterns. Pairwise comparisons for SNMP showed significant differences between normo- and hypodivergent, hypo- and hyperdivergent (p<0.05). The thickness of the masster muscle in the hypodivergent group was significantly greater than in the hyper- and normo- divergent groups. During contraction, the masseter muscle thickness of the low angle individuals was also significantly higher than that of the hyper and normodivergent subjects. The difference between the contracted and relaxed state was important for all of the three vertical patterns: during contraction, the muscle thickness increased by 10 % when compared to the relaxed state. The right and left sides showed no significant differences in the dimension of the masseter muscles (p>0.05).
Gender also influenced SNMP values, females showing significantly higher values than males (p<0.05). Significant differences were observed between age groups as well: group 1 showed higher values than group 2(p<0.05).
Correlations between muscular thickness of masseter, SNMP and FMA are presented in Table VIII.
Table VIII- Correlation between muscular thickness and SNMP and FMA
When analyzing the influence of vertical skeletal pattern, age and gender upon thickness of orbicularis oris, IMPA and U1SN, multivariate test showed a significant interaction effect between orbicularis oris superior and gender, males showing slightly higher values than females (p<0.05). An increased thickness of the orbicularis oris muscle was found in patients who exhibit hypodivergent and normodivergent pattern, however the correlation was not significant (p>0.05).
The thickness of the orbicularis oris seemed to have an influence on the inclination of the upper and lower incisors (U1SN and IMPA): the larger the thickness of orbicularis oris, the greater the retrusion of the upper incisors, and vice-versa. Nonetheless, no significant correlations could be demonstrated between orbicularis oris superior and U1SN, and between orbicularis oris inferior and IMPA.
5.4 Discussions
The aim of the present study was to determine the characteristics of masseter and orbicularis oris muscles in subjects with different vertical growth patterns by means of a noninvasive method, with no biological effect on living tissue. (125) Ultrasonography of superficial muscles is an accurate, reproducible, easy, and rapid method, as long as the operator respects a certain protocol. (115, 129)
For the assessment of the masseter muscle the transducer was oriented 30° to the Frankfort plane, perpendicular to the ramus of the mandible, following previous recommendations (125,128) and in a supine position with a silicon pad over the lips, for a better visualization of the orbiculari oris muscle.
The type of skeletal muscles is influenced by various factors: physical activity (130), genetics (131) and sexual hormones level. (132)
Several studies demonstrated that hyperdivergent vertical pattern is associated with a decrease in masseter’s thickness, as compared to the normo and hypodivergent individuals. (77, 126 128) We obtained similar values for the masseter muscle: the hypodivergent patient had the maximum thickness both during relaxation and contraction, followed by the normodivergent; the thickness was minimum in the hyperdivergent patient. The explanation, according to Proffit and Fields (133), could be that in hyperdivergent subjects, the bite force is weak, allowing a greater eruption of the posterior teeth, thus leading to a backward rotation of the mandible; a decreased function could be correlated further with a less developed muscle.
The results of the present study show a large variation in masseter muscle thickness among males and females, during both relaxation and contraction, confirming the findings from other researches. (126-128)
There are certain studies, which proved the influence of perioral muscle forces, especially the tongue and lips, on the bucco-lingual positioning of the teeth in the arch. (134) Rasheed, Munshi (135) observed that open-bite subjects presented a thinner upper orbicularis oris, while deep-bite subjects presented a thicker lower orbicularis oris muscle and higher electromyographic activity compared to other types of occlusion. In our present study, the results demonstrate similar findings: the hypodivergent patient exhibited thinner orbicularis oris than the hyper and normodivergent patients.
In the study conducted by Jung et al. (137), the lip closing force indicated a great influence on the angulation of the maxillary incisors. Moreover, lip incompetence has been associated with a large ANB angle and a large overjet, which is related to greater activity of the mentalis muscle to reach an anterior oral seal. Our present study was focused on the influence of orbicularis oris muscles, both superior and inferior, over the saggital position of the upper and lower incisor. Although no significant correlation was found, a greater thickness of the inferior orbicularis oris appeared in subjects with retruded upper incisors. Future studies conducted on a larger sample of patients, might demonstrate the correlation from a statistical point of view.
However, to the best of our knowledge, the correlation between the thickness of orofacial muscles and a certain dentomaxillary anomaly: incisor protrusion in hyperdivergent pattern and retrusion determined by hypodivergent growth, it has not been investigated yet.. An increased thickness of the orbicularis oris and masseter muscles were found in patients who exhibit a hypodivergent and normodivergent pattern, whereas the hyperdivergent had the smallest volume of the orofacial muscles.
5.5 Conclusion
1. The results of this study showed important discrepancies in masseter dimension between the three vertical patterns: the hypodivergent patient had both of the muscles thicker than those of the hyper and normodivergent pattern.
2. The patient with retruded upper and lower incisors presented an increased thickness of the lower orbicularis oris muscle on the US images, yet no significant correlation was found
3. Based on our observation, muscular thickness (both masseter and orbicularis oris) seemed to influence dento-maxillary morphology both vertically (hypo-, normo-, hyperdivergent) and sagittally (retruded or protruded incisors). Further research on a larger sample of patients is necessary.
6. Study 5. The influence of suprahyoid muscles on the development of class I and II. An ultrasound approach.
6.1 Introduction
The correlation between form and function represents a subject of a long-standing debate regarding the extent to which muscular activity affects the size, orientation, shape and pattern of growth. Does function influence form, or is the other way round?
Clinical studies have shown that the direction and amount of mandibular growth is dependent not only on genetic factors but also on environmental ones. (137-140) Graber explained the adaptation of muscle function to morphogenetic pattern but also stated that a change in muscle function can initiate morphologic variation in the normal configuration of the teeth and supporting bone. (141) Experimental studies on animal subjects have demonstrated that morphological structures of the mandible diminish in size, after extirpation of certain muscles. (142,143)
According to Angle’s classification in 1890s, class II division 1 is defined as the distal position of the lower canine and molar compared to the upper ones. (144) This represents an oversimplification of dentomaxillary anomalies because it does not refer to vertical, sagittal and transversal relations of the bone structure.
Fisk and Graber, were the first who indicated that, the essential nature of Class 2 malocclusion is in a posterior position of the mandible. (145,146) Harris has confirmed their theory and also found that the mandible is, on the average, smaller in persons with Class II malocclusion than in Class I subjects. (147) Recent studies (148-151), which focused on the relation between mandibular size and growth pattern, could not draw a precise conclusion referring to the cause of mandibular retrognatia in class II patients.
Among the possible etiological factors, suprahyoid muscles seem to play an important role in the growth pattern of the mandible in patients with habitual mandibular retrognatia. (152,153) The study conducted by Spyropoulus (154) suggested that the removal of the suprahyoid musculature might be associated with changes in the development of the mandible.
6.2 Objectives and research hypothesis
On the basis of the previous theories and findings, the purpose of this study was to investigate whether the thickness of suprahyoid muscles determined with the use of ultrasound imaging, correlates with the position of the mandible, in class 1 and 2 patients. The null hypotheses were: 1. Thickness of suprahyoid muscles is the same regardless of gender. 2. There is no statistical correlation between suprahyoid muscular thickness and dentomaxillary class. 3. Muscular thickness does not influence the development and growth of the mandible.
6.3 Materials and methods
The sample consisted of 27 patients (males and females; average age 18.5) who sought orthodontic treatment in a private dental office. Among the samples, based on their ANB angle and Wits appraisal, 11 were Class 1 and 16 were Class 2. A written consent was obtained from each subject, and the study was approved by the institutional review board.
The inclusion criteria of the study were: permanent or mixed dentition, no previous orthodontic treatment or orthognathic surgery history, no signs of temporo-mandibular disorders and no severe facial asymmetry (<4 mm).
The sagittal differences between the maxillary and mandibular skeletal bases were measured with ANB angle and Wits appraisal (AoBo), which applies a vertical correction according to the occlusal plane. (Table IX)
To analyse the suprahyoid muscles, a broadband linear transducer (Hitachi EUB 8500) with 5-7 MHz operating frequency was used. The trials were conducted in a darkened room by a single trained radiologist, who performed the ultrasound scan once. The patient was asked to sit relaxed, in a supine position and to slightly lift the mandible for a better access. Water –based gel was applied to the probe before the imaging procedure and the transducer was positioned perpendicular to the direction of the muscle fibers and moved until optimized images had been provided. (Fig. 24)
Fig. 25 Coronal scan of the suprahyoid muscles: the mylohyoid (1), right and left geniohyoid muscles (2), the anterior belly of the digastric muscle right and left, the body of the mandible.
Histograms and Shapiro-Wilk test were used to assess the normality of the data. Multivariate analysis of variance was conducted to test the hypothesis that muscular thickness of the suprahyoid muscles does not vary among Class I and Class II patients. Muscular thickness (suprahyoid muscles), SNB, ANB and AOBO were considered dependent variables, while dentoskeletal anomalies (Class I and Class II), age and gender were set as independent variables. Multiple comparisons between variables were adjusted by the Bonferroni method at a significance level of α=0.05. The Pearson’s Correlation Coefficient was used to analyze the relationship between muscular thickness and SNB, ANB and AOBO.
6.4 Results
Twenty-seven subjects (13 males, 14 females) were included in the present study: 11 subjects were Class I, and 16 subjects were Class II. The age distribution was as follows: group 1: 10-15 years – 11 subjects, group 2: >16 years – 16 subjects.
All recorded data followed a normal distribution; hence parametric tests were further used to test the null hypothesis.
The results of the multivariate tests showed a significant interaction effect between dentomaxillary class (Class I and II) and AOBO (p<0.05). Significant effect was observed between muscular thickness of digastric and gender (p<0.05). However, a significant statistical interaction effect could not been observed between dentomaxillary class and muscular thickness of suprahyoid muscles (p>0.05).
Pairwise comparisons for muscular thickness of digastric muscle and dentomaxillary class showed no significant differences (p=0.078). However, we did not find enough evidence to suggest that the null hypothesis is false at the 95% confidence level (the observed power computed using α=0.05 was 0.425), possibly because of the reduced number of subjects included in the study.
Gender also influenced muscular thickness of digastric muscle, males showing significantly higher values than females (p<0.05).
Correlations between muscular thickness of digastric, SNA, ANB, AOBO are presented in Table X.
A good correlation between muscular thickness of digastric and AOBO was observed for Class II subjects (r=0.403, p=0.041, N=27).
No significant correlation could be demonstrated between muscular thickness of geniohyoid and mylohyoid muscles and AOBO, SNB, ANB. (Table XI)
6.5 Discussions
According to other studies mylohyoid, digastric and geniohyoid muscles can be explored ultrasonographically. (155) In our study, we were able to obtain high quality images of the suprahyoid muscles, with the aid of 40 MHz ultrasound transducer. (Fig. 25) Hence, the thickness of the right and left digastric and geniohyoid muscles were measured on the US scans. (Table X)
Table X Average and SD values of muscular thickness
Wits appraisal was the cephalometric linear measurement to divide the patients into dentomaxillary classes. The method entails drawing perpendiculars from points A and B onto the occlusal plane (AO and BO). The ANB angle was used as an additional, not as a reference measurement, because it does not take into account the rotation of the jaws and length of the cranial base. (156,157)
As expected, a significant interaction effect between dentomaxillary class (Class I and II) and AOBO (p<0.05) was obtained. When comparing gender and thickness of the suprahyoid muscles, a significant correlation was observed, males showing higher values than females (p<0.05), thus the first null hypothesis being rejected.
Anterior face height was reported to show a significant negative correlation with digastric muscle activity (158), enabling us to extend our past research on the vertical skeletal pattern.
Furthermore, Carlson et al. (159) documented a greater tendency towards relapse in mandibular advancement cases due to the stretching of the suprahyoid complex. According to other studies, the removal of the suprahyoid musculature might change the development of the mandible in a sagittal and vertical direction. (154)
Although, no significant differences (p=0.078) were obtained when comparing thickness of suprahyoid muscles for class I and class II anomalies, we could not reject the second null hypothesis (the observed power computed using α=0.05 was 0.425). An increased number of subjects included in the study could demonstrate that the null hypothesis is false.
The results from our research are important, since a good correlation between muscular thickness of digastric and AOBO (r=0.403, p=0.041, N=27) was obtained for class II subjects. Therefore a possible influence of anterior belly of the digastric over the severity of mandibular retrognatia could be documented: the thicker the digastric muscle, the severe the class II anomaly.
A thorough investigation of class II etiology, regarding the length of the body of the mandible, SNB angle, rotation of the mandible and muscular thickness is imperative in future studies.
However, when comparing geniohyoid and mylohyoid muscular thickness with AOBO, ANB angles no statistical correlation was found.
6.6 Conclusions
1. The mylohyoid, geniohyoid and anterior belly of the digastric muscles can be explored with the aid of 40MHz ultrasonography.
2. A positive correlation between muscular thickness of digastric and AOBO was observed for Class II subjects.
3. A more thorough research on a bigger number of subjects is necessary to investigate whether digastric muscular thickness influences the development of mandibular retrognatia.
8. General conclusions
1. Ultrasound imaging represents a reliable, uncomplicated and noninvasive technique, which can be successfully used in many fields of dentistry, such as periodontology and orthodontics.
2. Compared to other conventional methods for assessing periodontal and muscular structure, ultrasonography has many advantages, among which: clinical availability, cost and minimum radiance exposure.
3. The high-resolution transducer (40 MHz) was able to detect the gingival sulcus depth, free gingival thickness, and width of periodontal space, distance between marginal gingiva and alveolar crest, height of clinical and anatomical crown.
4. Ultraound measurements of the gingival sulcus depth were very similar to those of the probing depth technique.
5. A small dimension transductor would be more suitable for the evaluation of the intraoral soft tissues.
5. The interobserver reproducibility of the ultrasound measurements on periodontal structures was very good.
6. The applicability of ultrasonography extends to the filed of orthodontic biomechanics, regarding the possibility to assess, in real time, periodontal modifications during orthodontic tooth movement.
7. The middle and mesial area of the buccal surface of the canine, during orthodontic distalisation, showed significant changes immediately after force delivery.
8. Important discrepancies in masseter dimension were found between the three vertical patterns, with the aid of ultrasonography; thus the hypodivergent patient had both of the muscles thicker than those of the hyper and normodivergent pattern.
9. The patient with retruded upper and lower incisors presented an increased thickness of the lower orbicularis oris muscle on the US images, yet no significant correlation was found.
10. The results indicated that the thickness of master and suprahyoid muscles is depended upon gender, vertical skeletal pattern and dentomaxillary anomaly class.
11. The 40MHz resolution ultrasonography was able to explore the suprahyoid muscles, among which: anterior belly of the digastric, geniohyoid and mylohyoid.
12. A positive correlation between muscular thickness of digastric and AOBO was observed for Class II subjects, therefore the severity of mandibular retrognatia might be influenced by above-mentioned muscle.
13. A more thorough research on a bigger number of subjects is necessary to investigate whether digastric muscular thickness influences the development of mandibular retrognatia.
9. Originality and Innovative Contributions of the Thesis
The originality of the present thesis stands in the interdisciplinary approach to medical imaging science, orthodontics and periodontology. Each of the five studies consists in a different and innovative perspective over the applicability of ultrasonography in periodontal and muscular assessment.
The chapters included in the current state of knowledge offer a thorough and up to date synthesis of clinical and imaging methods for the dentomaxillary structures, which can benefit from the ultrasound evaluation.
The feasibility study represents the first attempt, in the literature, to use really high-resolution ultrasonography (40 MHz) on healthy human periodontal structures. Data regarding gingival sulcus depth, free gingival thickness, width of periodontal space, distance between marginal gingiva and alveolar crest and height of anatomical crown are visible and easy to measure on the US scan.
Although many studies demonstrated the possibility to use ultrasonography as a diagnostic tool in the field of dentistry, in the present, it is not part of the daily clinical practice.
Another original aspect of the Ph.D. resides in the study conducted on pig jaw mandibles, which confirms our previous findings regarding sulcus depth evaluation and demonstrates the reproducibility of the ultrasound method.
Orthodontic tooth movement induces many changes in the periodontal structures, yet clinicians do not have a reliable, accurate and non-invasive method to understand and predict them. According to our study, ultrasound imaging could be used to observe real time modifications after force delivery and help the orthodontist to predict and control treatment biomechanics.
Furthermore, two of the studies focused on muscular thickness assessment with the aid of ultrasonography and their implication in dentomaxillary anomalies. The influence of digastric muscle over the mandibular position in a sagittal direction could be considered groundbreaking, since no other study investigated it. The results showed a significant positive correlation between class II anomaly and digastric thickness.
Additionally, the masseter muscle is involved in the development of the vertical skeletal pattern, the mandibular divergency and muscular thickness being inversely proportional. Also, our research showed a possible correlation between the lower orbicularis oris muscle and the anterior-posterior position of the upper incisors.
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