Enzyme histochemical adaptive responses of the medial pterygoid muscle and two heads of the lateral pterygoid muscle to long-term soft diet feeding in growing rabbits.

The aim of the present study was to investigate the histochemical effects of long-term soft diet in the medial pterygoid muscle as well as the two heads of the lateral pterygoid muscles in growing rabbits. Eleven young rabbits were divided into two groups as solid diet (control group; n = 6) or soft diet (soft-diet group; n = 5) groups. After 6 months, muscle fibers from the medial and the two heads of the lateral pterygoid muscles were histochemically defined. In the medial pterygoid muscle, the percentage of the type 1 fiber cross-sectional area to total area was 10.1 ± 2.4% in the control group and 8.3 ± 3.0% in the soft-diet group, respectively. In the soft-diet group, there was a trend toward an increase in the number of type 2A fibers, and toward a decrease in the numbers of type 2B fibers in comparison with the controls. In the two heads of the lateral pterygoid muscle, the percentage of the type 1 fiber cross-sectional area to total area was 8.4 ± 7.5 and 3.3 ± 2.7%, respectively. Compared to that of the control group, the two heads in the soft-diet group showed a trend toward a decrease in the number of type 2A fibers. In addition, type 2B had a tendency to decrease in the number in the inferior head. In conclusion, this study suggests that long-term soft diet leads to adaptations of the pterygoid muscles. Two heads of the lateral pterygoid muscle revealed different adaptation from jaw-closing muscles under soft-diet conditions.

The role of lateral pterygoid muscle in the traumatic temporomandibular joint ankylosis: A gene chip based analysis.

Traumatic temporomandibular joint ankylosis (TMJA) is a common disease and disorder of the temporomandibular joint (TMJ); however, its pathogenesis has yet to be completely elucidated. In the authors' previous studies, the lateral pterygoid muscle (LPM) was confirmed to exert a function in distraction osteogenesis (DO) during the healing of a condylar fracture, which resulted in the formation of excess bone. The aim of the present study was to investigate alterations in the expression of any associated genes via an Affymetrix GeneChip method. The traumatic TMJA model was fabricated by a condylar fracture in the TMJ area of sheep with either a dissected LPM (LPD) or normal (LPN). The untreated sheep served as a control. At 4­ and 12 weeks post­surgery, the condylar zone was isolated to perform the gene chip analysis, which was performed according to a standard Affymetrix protocol. The validated genes were further evaluated by reverse transcription­quantitative polymerase chain reaction (RT­qPCR). The gene chip analysis indicated that the LPN gene expression pattern was similar compared with the DO process, while LPD was similar to that of normal bone fracture healing. The validated genes were collagen type II α1 chain, C­type lectin domain family 3 member A, interleukin 1A, cartilage oligomeric matrix protein, chondromodulin (LECT1), calcitonin receptor (CALCR), transforming growth factor (TGF)­ß1, Fos proto­oncogene (FOS), bone γ­carboxyglutamate protein and bone morphogenic protein (BMP)7, among which, BMP7, LECT1, CALCR and FOS were confirmed by RT­qPCR. In conclusion, the present study demonstrated that LPM exerts a DO effect during the pathogenesis of traumatic TMJA, which may provide a novel target for preventing TMJA.

Ultrastructural and biochemical changes of the medial pterygoid muscle induced by unilateral exodontia.

The aim of the present study was to investigate the histological, biochemical and ultrastructural effects of occlusal alteration induced by unilateral exodontia on medial pterygoid muscle in guinea pigs, Cavia porcellus. Thirty (n=30) male guinea pigs (450g) were divided into two groups: experimental-animals submitted to exodontia of the left upper molars, and sham-operated were used as control. The duration of the experimental period was 60 days. Medial pterygoid muscles from ipsilateral and contralateral side were analyzed by histological (n=10), histochemical (n=10), and ultrastructural (n=10) methods. The data were submitted to statistical analysis. When the ipsilateral side was compared to the control group, it showed a significantly shorter neuromuscular spindle length (P<0.05), lower oxidative metabolic activity, and microvessel constriction, in spite of the capillary volume and surface density were not significantly different (P>0.05). In the contralateral side, the neuromuscular spindles showed significantly shorter length (P<0.05), the fibers reflected a higher oxidative capacity, the blood capillaries showed endothelial cell emitting slender sprouting along the pre-existing capillary, and significantly higher blood capillary surface density, and volume density (V(v)=89% Mann-Whitney test, P<0.05). This finding indicated a complex morphological and functional medial pterygoid muscle adaptation to occlusal alteration in this experimental model. Considering that neuromuscular spindles are responsible for the control of mandibular positioning and movements, the professional should consider if these changes interfere in the success of clinical procedures in medical field involving stomatognathic structures.

Morphological and histological effects on the medial pterygoid muscle after unilateral exodontia in gerbils.

Morphological effects on the medial pterygoid muscle were evaluated in 20 male gerbils (average weight, 55 g) after occlusal alterations induced by extraction of left side molar teeth. Controls were only submitted to surgical stress. Sixty days after surgery, the groups were divided into two subgroups for the following studies: (1) observation of macroscopic morphology and vessels distribution (n=10); (2) light microscopy histological analysis (n=10). Group results were statistically compared using the Wilcoxon and Mann-Whitney tests, with a significant value of p<0.05. Statistical differences in biometric data were found between the left and right sides of the experimental group (p=0.043), and between the left side of the control group when compared to the same side in the experimental group (p=0.044). Vessels supplied by bundles of the external carotid artery in the medial pterygoid muscle did not show distribution differences in group comparisons. Histological alterations were found in the ipsilateral side of the experimental group, with a central localization of the nucleus and degenerative aspect of the fibers, usually near to internal aponeurosis. Fiber diameters seemed reduced and the neuromuscular spindles were localized near internal aponeurosis and had a modified appearance. It is concluded that the medial pterygoid muscle in the gerbil is sensitive to alterations of the masticatory movements.

A Cadaveric Study of Ultrasound-Guided Maxillary Nerve Block Via the Pterygopalatine Fossa: A Novel Technique Using the Lateral Pterygoid Plate Approach.

BACKGROUND AND OBJECTIVES: This study aimed to describe and assess the accuracy and feasibility of a novel technique for ultrasound-guided maxillary nerve block using the lateral pterygoid plate (LPP) approach via the pterygopalatine fossa (PPF) in a soft cadaveric model. METHODS: Ten soft cadavers were studied. The curved array ultrasound transducer probe was applied over 1 side of the face of the cadavers in the open-mouth posture. It was placed transversely below the zygomatic arch for identifying the border of the maxillary tuberosity and the LPP. We tilted the curve probe from the caudal to the cranial direction until the uppermost part of the PPF was identified. The in-plane needle approach was used from the anterior-to-posterior and lateral-to-medial directions through the fossa, and 3 mL of methylene blue dye was injected. RESULTS: The spread of injectate after ultrasound-guided maxillary nerve block using the LPP approach was successfully performed in all cadavers as demonstrated by visualized moderate to marked traces of methylene blue within the PPF. No accidental injections in the maxillary arteries or facial nerves were observed. CONCLUSIONS: This cadaveric study suggests that ultrasound-guided maxillary nerve block using the LPP approach via the PPF has a high degree of accuracy and feasibility. Further studies are required to confirm its efficacy and safety for clinical application.

Metabolic and vascular pattern in medial pterygoid muscle is altered by chronic stress in an animal model of hypodontia.

Psychological stress is an important perpetuating, worsening and risk factor for temporomandibular disorders of muscular or articular origin. Occlusion instability, by the way, is considered a risk factor of this pathology and can be reproduced in some experimental animal models. The exact physiologic mechanism underlying these relations however, remains unclear. Our purpose was to test the hypothesis that chronic stress and unilateral exodontia induce metabolic and vascular changes in the medial pterygoid muscle of rats. Adult Wistar rats were submitted to chronic unpredictable stress and/or unilateral exodontia and their plasma and medial pterygoid muscle were removed for analysis. The parameters evaluated included plasma levels of corticosterone, metabolic activity by succinate dehydrogenase, oxidative capacity by nicotinamide adenine dinucleotide diaphorase, capillary density by laminin and alfa-CD staining and reactive oxidative species production. Chronic unpredictable stress as an isolated factor, increased oxidative metabolism, capillary density and reactive oxygen species production at medial pterygoid muscle. Conversely, exodontia has a main effect in metabolism, promoting glycolytic transformation of muscle fibers. Association of both factors induced a major glycolytic pattern in muscle and vascular changes. Our findings provide insights into the mechanisms, possibly inducing metabolic and vascular alterations on medial pterygoid muscle of rats, by which chronic stress and occlusal instabilities might be involved as risk factors in the pathophysiology of temporomandibular disorders with muscular components.

Expression of MyHC isoforms mRNA transcripts in different regions of the masseter and medial pterygoid muscles in chimpanzees.

OBJECTIVE: The aim of this study is to examine the expression pattern of the different myosin heavy chain (MyHC) isoforms in the masseter and medial pterygoid muscles by real time quantitative polymerase chain reaction (RT-qPCR) to obtain information at molecular level which can be related to the functional characteristics of these two muscles. DESIGN: The masseter, deep and superficial portion, and medial pterygoid muscles of five adult Pan troglodytes were dissected in order to obtain samples of the anterior and posterior regions of each portion of the masseter and of the medial pterygoid. The expression of MyHC isoforms mRNA transcripts was analyzed by RT-qPCR. RESULTS: No significant differences in expression of MyHC isoforms between the masseter and the medial pterygoid were found. In contrast, when comparing the superficial and the deep portion of the masseter, we found that the MyHC-IIM isoform was expressed at a significantly higher level in the superficial portion. CONCLUSIONS: The superficial portion of the masseter and the medial pterygoid muscle have the same expression pattern regarding the different MyHC isoforms. On the other hand, the deep portion of the masseter, which is activated mainly during lateral and repositioning movements of the mandible, has a lower MyHC-IIM isoform expression than the superficial portion. Our findings provide new data on functional aspects of the masseter and medial pterygoid that can complement results obtained by other techniques.

Expression of myosin heavy chain isoform mRNA transcripts in the masseter and medial pterygoid muscles / Expresión de transcripciones de ARNm de isoforma de cadena pesada de miosina en los músculos masetero y pterigoideo medial

SUMMARY: Both the masseter and medial pterygoid muscles elevate the mandible, raising the lower jaw by acting simultaneously on the lateral and medial surfaces of the mandibular ramus. Nevertheless, electromyographic studies indicate that these muscles, as well as the superficial and deep heads of the masseter, act in a different way during mastication. We have analyzed by real time quantitative polymerase chain reaction (RT-qPCR) the expression of myosin heavy chain (MHC) isoforms in the masseter and medial pterygoid muscles in humans in order to identify possible differences in the expression patterns that may be related to functional differences identified with electromyography. Our findings indicate that the expression pattern of MHC isoforms in the two muscles is characteristic of fast and powerful phasic muscles. We have also observed a high percentage of expression of the MHC-IIx isoform and the expression of the MHC-M isoform at the mRNA level in both muscles, an isoform that does not translate into protein in the masticatory muscles of humans. The high percentage of expression of the MHC-IIx isoform in humans can be related to a high contractile speed of the masseter and medial pterygoid in humans. On the other hand, the low percentage of expression of the MHC-M isoform at the mRNA level in both muscles can be related to the complex evolutionary process that has reduced the size and force of the masticatory muscles in humans.

RESUMEN: Los músculos masetero y pterigoideo medial elevan la mandíbula actuando de forma simultánea sobre las caras lateral y medial de su rama. Sin embargo, los estudios electromiográficos indican que estos dos músculos actúan de forma diferente durante la masticación, de la misma forma que lo hacen las porciones superficial y profunda del músculo masetero. En el presente estudio hemos analizado mediante PCR en tiempo real la expresión de las isoformas de la cadena pesada de la miosina o myosin heavy chain (MHC) en los músculos masetero y pterigoideo medial en humanos, con la finalidad de identificar diferencias en los patrones de expresión que se puedan relacionar con las diferencias funcionales identificadas con la electromiografía. Nuestros resultados indican que el patrón de expresión de las isoformas de la MHC en los dos músculos es la característica de los músculos rápidos y potentes. También hemos observado un elevado porcentaje de expresión de la isoforma MHC-IIx y la expresión a nivel de ARNm de la isoforma MHC-M en los dos músculos, una isoforma que no se detecta a nivel de proteína en los músculos masticadores humanos. El elevado porcentaje de expresión de la isoforma MHC-IIx que hemos observado se puede relacionar con una elevada velocidad de contracción de los músculos masetero y pterigoideo medial en los humanos. Por otro lado, el bajo porcentaje de expresión de la isoforma MHC-M a nivel de ARNm en ambos músculos se puede relacionar con los procesos evolutivos complejos que han reducido el tamaño y la fuerza de los músculos masticadores en los humanos.

Chronic stress effects in contralateral medial pterygoid muscle of rats with occlusion alteration.

Temporomandibular disorder (TMD) has a high prevalence in our society, characterized by a severe pain condition of the masticatory muscles and temporomandibular joint. Despite the indication of multiple factor initiators of TMD, there is still controversy about its etiology and its pathophysiology is poorly understood. Using rats as experimental animals we investigated the effect of unpredictable chronic stress with or without unilateral molar extraction on the contralateral medial pterygoid muscle. Our hypothesis is that these two factors induce changes in morphology, oxidative metabolism and oxidative stress of muscle fibers. Young adult male Wistar rats (±200g) were divided into four groups: a group with extraction and unpredictable chronic stress (E+US); with extraction and without stress (E+C); without extraction and with unpredictable chronic stress (NO+US); and a control group without either extraction or stress (NO+C). The animals were subjected to unilateral extraction of the upper left molars, under intraperitoneal anesthesia with 4% Xylazine (10mg/kg) and 10% Ketamine (80mg/kg) on day zero. The rats of groups E+US and NO+US were submitted to different protocols of stress, from the 14th day after the extraction. The protocols were different every day for five consecutive days, which were repeated from the 6th day for five days more. Contralateral medial pterygoid muscles were obtained on the 24th day after the start of the experiment for morphological, metabolic, capillary density, and oxidative stress analysis. The data from capillary density showed a decrease of capillaries in animals subjected to dental extraction, compared with those without extraction and an increase of laminin expression in the group submitted to the unpredictable chronic stress when compared to the unexposed to stress. SDH test revealed a decrease of light fibers in the group submitted to unilateral extraction of molars, compared with this area in the control group. In E+US and NO+US groups, the deeply stained fibers increased compared to NO+C.·The exodontia factor was able to increase the ROS activity in muscle, whereas the stress factor does not significantly alter ROS in this tissue. It was concluded that both unpredictable chronic stress and the extraction induce metabolic and density of capillary changes in the contralateral medial pterygoid muscle to extraction, suggesting that these factors for a longer period of this experiment could induce muscle damage related to TMD.

Expression of unique and developmental myosin heavy chain isoforms in adult human digastric muscle.

Digastric muscle (DGM) is a powerful jaw-opening muscle that participates in chewing, swallowing, breathing, and speech. For better understanding of its contractile properties, five pairs of adult human DGMs were obtained from autopsies and processed with immunocytochemistry and/or immunoblotting. Monoclonal antibodies against alpha-cardiac, slow tonic, neonatal, and embryonic myosin heavy chain (MHC) isoforms were employed to determine whether the DGM fibers contain these MHC isoforms, which have previously been demonstrated in restricted specialized craniocervical skeletal muscles but have not been reported in normal adult human trunk and limb muscles. The results showed expression of all these MHC isoforms in adult human DGMs. About half of the fibers reacted positively to the antibody specific for the alpha-cardiac MHC isoform in DGMs, and the number of these fibers decreased with age. Slow tonic MHC isoform containing fibers accounted for 19% of the total fiber population. Both the alpha-cardiac and slow tonic MHC isoforms were found to coexist mainly with the slow twitch MHC isoform in a fiber. A few DGM fibers expressed the embryonic or neonatal MHC isoform. The findings suggest that human DGM fibers may be specialized to facilitate performance of complex motor behaviors in the upper airway and digestive tract.

Differential expression of myosin heavy-chain mRNA in muscles of mastication during functional advancement of the mandible in pigs.

Surgical and orthodontic treatment of retrognathia aims to improve orofacial function by adaptation and training of muscle capacity, which is connected with a change in muscle fibre-type proportions. The aim here was to analyse the proportion of myosin-heavy chain (MyHC) gene expression in type I (slow twitch/ST) and type IIb (fast twitch/FT) fibres during sagittal advancement of the mandible by reverse transcriptase-polymerase chain reaction (RT-PCR). The experiments were carried out on 10-week-old pigs (six test animals, six controls) over a 28-day period. Six pigs were fitted with acrylic bite blocks for sagittal advancement of the mandible. Tissue was taken from seven different regions of the masseter, temporal, medial pterygoid, and geniohyoid muscles. The 84 samples were used for histological fibre differentiation with ATPase staining and for isolation of total RNA. To measure the two MyHC isoforms, RT-PCR (in a single tube reaction with MyHC I, MyHC IIb, and GAPDH primers) was used. A significant increase was registered in the percentage of ST fibres and in mRNA from MyHC I in the anterior region of the masseter and in the posterior region of the temporal muscle of the treated animals. The proportion of ST fibres to FT fibres was increased by up to 12% after functional advancement of the mandible. The histological findings corresponded with the data for fibre mRNA generated by RT-PCR.

Desmin and nerve terminal expression during embryonic development of the lateral pterygoid muscle in mice.

OBJECTIVE: In adults, the lateral pterygoid muscle (LPM) is usually divided into the upper and lower head, between which the buccal nerve passes. Recent investigations have demonstrated foetal developmental changes in the topographical relationship between the human LPM and buccal nerve. However, as few studies have investigated this issue, we clarified the expression of desmin and nerve terminal distribution during embryonic development of the LPM in mice. METHODS: We utilized immunohistochemical staining and reverse transcription chain reaction (RT-PCR) to clarify the expression of desmin and nerve terminal distribution. RESULTS: We observed weak expression of desmin in the LPM at embryonic day (ED) 11, followed by an increase in expression from embryonic days 12-15. In addition, starting at ED 12, we observed preferential accumulation of desmin in the vicinity of the myotendinous junction, a trend that did not change up to ED 15. Nerve terminal first appeared at ED 13 and formed regularly spaced linear arrays at the centre of the muscle fibre by ED 15. The results of immunohistochemical staining agreed with those of RT-PCR analysis. CONCLUSION: We found that desmin accumulated in the vicinity of the myotendinous junction starting at ED 12, prior to the onset of jaw movement. We speculate that the accumulation of desmin is due to factors other than mechanical stress experienced during early muscle contraction. Meanwhile, the time point at which nerve terminals first appeared roughly coincided with the onset of jaw movement.

[An immunohistochemical study of distribution of insulin in lateral pterygoid muscle after functional mandibular protrusion in rats].

OBJECTIVE: The aim of this study is to investigate the effects of insulin on the lateral pterygoid muscle of growing rats after the functional mandibular protrusion. METHODS: An immunohistochemical (the ABC method) method was used to detect the location and distribution of insulin in the lateral pterygoid muscle of rats. RESULTS: 1. Insulin located in the cell membrane of the lateral pterygoid muscle. 2. The distribution of insulin in the lateral pterygoid muscle of rats was enhanced after the functional mandibular protrusion. CONCLUSION: Insulin may play an important role in adaptive remodeling of the lateral pterygoid muscle after the treatment of functional mandibular protrusion.

[Characteristic changes of acetylcholine receptor in rat lateral pterygoid muscles after functional mandibule advancement].

OBJECTIVE: The purpose of this study was to evaluate the maximum binding(Bmax) and affinity(Kd value) changes of acetylcholine receptor (n-AchR) in rat lateral pterygoid muscles after functional mandibule advancement. METHODS: 40 five-weeks-old male Sprague-Dawley rats were equally divided into experimental and control group. The mimic functional appliances were used in experiment group and the rats were killed after 1, 3, 7, 14 days. Radio-ligand binding assay (RBA) was applied to determine the maximum binding(Bmax) and Kd value of n-AchR of lateral pterygoid muscle. RESULTS: The Bmax of n-AchR in experimental group was higher than that in control group and the Kd value always kept in high level. CONCLUSION: The functional orthopedics can increase the Bmax of n-AchR in rapid growing rat lateral pterygoid muscles.

Distribution of substance P-like immunoreactive nerve fibers in temporomandibular joint soft tissues of monkey.

The distribution of substance P-immunoreactive and silver impregnated nerve fibers in the temporomandibular joint soft tissues of the Macaca fascicularis monkey was investigated in frozen sections. The pattern of substance P-immunoreactive structures in the soft tissues and periosteum of the temporomandibular joint was compared with the distribution of silver impregnated nerve fibers within these tissues. Presence of substance P-immunoreactive fibers was demonstrated in the temporomandibular joint capsule, disc attachments, fascia, adjacent periosteum and within the interfascicular connective tissue of the lateral pterygoid muscle. The overall distribution corresponded to that of silver impregnated nerve fibers. Substance P-immunoreactive nerve fibers were found in the adventitia of arteries in all vascularized temporomandibular joint soft tissues but could not be found in the adventitia of veins. No substance P-immunoreactive or silver impregnated nerve fibers were seen in the dense collagenous tissue forming the disc. Substance P is suggested to influence the major features of inflammation and to play a role in acute and chronic pain conditions.

An experimental orthopedic treatment of the rat mandible using a functional appliance alters the fibre and myosin types in masticatory muscles.

Masticatory muscles mediate the action of functional orthopedic appliances on mandible growth. When young rats were treated for four weeks with a postural hyperpropulsor of the mandible, an appliance designed to increase condylar cartilage growth rate, the proportion of fast non-fatigable fibres in the lateral pterygoid muscle increased significantly. Concomitantly, the amount of slow-myosin light chains increased in fibre extracts. This slow myosin originated from IIA fibres. By functional orthopedic treatment, the lateral pterygoid muscle was enriched in less fatigable fibres; the changes observed in the lateral pterygoid muscle were close to changes observed in other muscles after training.

Metabolic transitions in rat jaw muscles during postnatal development.

The program of acquisition of adult metabolic phenotypes was studied in three jaw muscles in order to determine the link between muscle metabolism and functional development. During early postnatal stages, there were similar transitions in the masseter, anterior digastric, and internal pterygoid muscles with respect to fiber growth, fiber type composition, and whole muscle energy metabolism. Oxidative capacity, as judged by the activities of the enzymes succinate dehydrogenase (SDH), malate dehydrogenase (MDH), and beta-hydroxyacyl CoA dehydrogenase (beta OAC), rose sharply after birth to reach near maximal levels by 3 weeks. The capacities for glycolytic metabolism represented by lactate dehydrogenase (LDH), and for high-energy phosphate metabolism represented by adenylokinase (AK) and creatine kinase (CK) activities, rose gradually, not reaching peak values until 6 weeks or later. Thus, the maturation of oxidative metabolism preceded that of glycolytic metabolism in the developing jaw muscles. This was documented for individual fibers in the masseter muscle. Differential metabolic maturation among the jaw muscles was evident beyond 3 weeks. All three jaw muscles attained their specific adult fiber-type profile by about 6 weeks. This maturation program differed from that of hindlimb muscles [Nemeth et al., J Neurosci 9:2336-2343, 1989] and diaphragm muscle [Kelly et al., J Neurosci 11:1231-1242, 1991], reflecting their differential energy demands for contractile performance.

[A radioimmunologic assay of insulin in lateral pterygoid muscle after functional mandibular protrusion in rats].

OBJECTIVE: To investigate the effect of insulin on the lateral pterygoid muscle after functional mandibular protrusion in rats. METHODS: Radioimmunoassay was used to examine the level of insulin in lateral pterygoid muscle quantitatively after functional mandibular protrusion in young growing rats. RESULTS: The amount of insulin in lateral pterygoid muscle in young growing rats enhanced after functional mandibular protrusion. CONCLUSION: Insulin plays an important role in adaptive remodeling of lateral pterygoid muscle after functional mandibular protrusion.

Comparison of phosphocreatine concentration in the human masseter and medial pterygoid muscles by 31P-CSI.

The aim of this study is to compare phosphocreatine (PCr) concentrations of human masseter and medial pterygoid muscles by a recently developed localized magnetic resonance spectroscopy (MRS) method, chemical shift imaging (CSI). The characteristic spectra of phosphorous metabolites including PCr and beta-ATP from the superficial part of the masseter (SM) and the deep part of the masseter (DM) and the medial pterygoid muscles (MPt) from 11 volunteers, 20-27-year-old were obtained. The study clearly demonstrated higher PCr/beta-ATP in the SM and MPt than in the DM both in mean values (P < 0.01) and in individual subjects. The results indicate that SM and MPt are power producers. There were no significant differences in the mean values of the PCr/beta-ATP ratios in SM and MPt, however, the PCr/beta-ATP ratios varied individually and the subjects could be divided into three distinct groups: values of MPt higher than SM (group A, 4 subjects); values of MPt almost equal to SM (group B, 3 subjects); and values of MPt lower than SM (group C, 4 subjects). There appears to be a close relationship between the PCr content as determined in the groups here and occlusal guidance.