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.

Association between the peri-implant bone structure and stress distribution around the mandibular canal: a three-dimensional finite element analysis.

The aim of this study was to elucidate the association between the bone structure at implant insertion sites and stress distribution around the mandibular canal by means of three-dimensional finite element (3D FE) analysis. Four FE models were created with slice data using micro-computed tomography (micro-CT), and 3D FE analysis was performed. Mechanical analysis showed that the load reached the mandibular canal via the trabecular structure in all FE models. High levels of stress were generated around the mandibular canal when the distance between the mandibular canal and the implant decreased. High stress levels were also observed when cortical bone thickness and bone volume/total volume (BV/TV) were low. Our findings suggest that load is transmitted to the mandibular canal regardless of differences in the thickness of cortical bone or cancellous bone structure, but excessive load may be generated in bone with thin cortical and coarse cancellous structures.

Consideration of shear modulus in biomechanical analysis of peri-implant jaw bone: accuracy verification using image-based multi-scale simulation.

The aim of this study was to clarify the influence of shear modulus on the analytical accuracy in peri-implant jaw bone simulation. A 3D finite element (FE) model was prepared based on micro-CT data obtained from images of a jawbone containing implants. A precise model that closely reproduced the trabecular architecture, and equivalent models that gave shear modulus values taking the trabecular architecture into account, were prepared. Displacement norms during loading were calculated, and the displacement error was evaluated. The model that gave shear modulus values taking the trabecular architecture into account showed an analytical error of around 10-20% in the cancellous bone region, while in the model that used incorrect shear modulus, the analytical error exceeded 40% in certain regions. The shear modulus should be evaluated precisely in addition to the Young modulus when considering the mechanics of peri-implant trabecular bone structure.

Evaluation of cortical bone thickness and root proximity at maxillary interradicular sites for mini-implant placement.

OBJECTIVES: The purpose of this study was to evaluate the interradicular cortical bone thickness, alveolar process width and root proximity for planning mini-implant placement in the maxillary alveolar process. MATERIAL AND METHODS: Eighty maxillae (right and left sides) of 40 Japanese adult skulls were examined. The samples were imaged and measured using a micro-CT system. Buccal and palatal interradicular cortical bone thickness, alveolar process width, and root proximity were measured in six interradicular sites from distal of central incisor to mesial of second molar. Buccal and palatal interradicular cortical bone thickness and alveolar process width were measured at 10 different vertical levels. Root proximity was measured at four different vertical levels. RESULTS: Buccal and palatal interradicular cortical bone thickness and alveolar process width tended to increase from crest to base of alveolar process. The buccal interradicular cortical bone thickness between canine and first premolar or between first premolar and second premolar was the greatest, and between central incisor and lateral incisor was the least. The palatal interradicular cortical bone was significantly thicker than the buccal. The root proximity between second premolar and first molar or first premolar and second premolar was the widest and between central incisor and lateral incisor it was the narrowest. CONCLUSIONS: The findings of this study suggest that recommendations when low dose 3D multislice CT or low dose cone beam imaging is not available, the results of this research may be useful in providing indicators for selecting the design of the placement site.

Variations of vascular distribution in the mandibular anterior lingual region: a high risk of vascular injury during implant surgery.

PURPOSE: To clarify variations of vascular distribution around the mandibular anterior tooth lingual region, an area in which vascular injuries have often been reported during dental implant surgery. The reasons for such injuries in this region are discussed from an anatomical perspective. MATERIALS AND METHODS: Anatomical dissections were performed on 100 sides of 50 cadavers used for anatomy education. Ten sides of 5 cadavers were injected intravascularly with methyl methacrylate, and penetration of the mandible was closely evaluated. RESULTS: In the mandibular anterior tooth lingual region, both the sublingual and submental arteries showed various distribution patterns. Distal branches basically penetrated the bone. CONCLUSIONS: In the mandibular anterior tooth lingual region, the sublingual and submental arteries traveled from the vicinity of the mylohyoid muscle attachment along the bone surface in an anterosuperior direction. Many of these blood vessels penetrated the alveolar mucosa in the anterior tooth region, and many distal branches of the vessels also finally penetrated the bone. This seems to explain why many vascular injuries are encountered around the mandibular anterior tooth lingual region during implant surgery.

Gene and protein expressions of vimentin and desmin during embryonic development of the mylohyoid muscle.

Meckel's cartilage is known to be involved in formation of the prenatal mandible. However, the relationship between Meckel's cartilage and the embryonic mylohyoid muscle during growth and development has been investigated only rarely. This study examined the expression of intermediate filaments in Meckel's cartilage and the embryonic mylohyoid muscle in fetal mice during morphological development. Specimens of E12-16 ICR mice sectioned in the frontal direction were subjected to immunohistochemistry for vimentin and desmin. Hematoxylin and eosin sections showed that the immature mylohyoid muscle began to grow along Meckel's cartilage during fetal development. Weak vimentin expression was detected in the mylohyoid muscle and surrounding tissues at E12. Desmin expression was detected specifically in the mylohyoid, and strong expression was evident after E13, and increased with age. It was inferred that the mylohyoid muscle is one the tissues developing from Meckel's cartilage, the latter exerting a continuous influence on the growth of the former. In the early stage, the surrounding mesenchymal tissues expressing vimentin formed a scaffold for the developing mylohyoid muscle. Muscle attachment at E13 showed steady desmin expression, which continued until maturity. This study suggested the possibility that Meckel's cartilage has an influence not only on the mandibular bone, but also on the development of the mylohyoid muscle attached to the mandibular bone. Furthermore, it revealed a stage of the developmental process of the mylohyoid muscle in which the expression of vimentin, which is a common protein in the surrounding tissue such as muscle and bone, induces the morphological formation of the mylohyoid muscle, cooperating with the surrounding structures.

Alignment of biological apatite crystallites at first molar in human mandible cortical bone.

The aim of this study was to quantitatively clarify the c-axis alignment of biological apatite (BAp) crystallites (hereafter referred to as BAp alignment) in the cortical bone of the human mandible first molar. Six mandible specimens were collected from the cadavers of six dentulous Japanese adults (mean age, 63.0 +/- 12.1 years) held at the Department of Anatomy, Tokyo Dental College. A microbeam x-ray diffraction system was used to determine BAp alignment in the mesiodistal direction. Bone mineral density (BMD) was also measured using 3-dimensional trabecular structure measurement software. The results showed that the degree of BAp alignment in the mesiodistal direction was low in the alveolar area and high at the base of the mandible, suggesting that BAp alignment in the alveolar area is affected by occlusal force. Moreover, it was observed that the correlation between BAp alignment and BMD was small, indicating that BAp alignment and BMD could be independent factors. Therefore, determining BAp alignment was important in the evaluation of bone quality, including bone strength.

Evaluation of the palatal bone for placement of orthodontic mini-implants in Japanese adults.

Mini-implants are increasingly being used for orthodontic anchorage in the palate. The anatomical structure of the jaw must be properly evaluated prior to use; however, there are a few research reports providing basic data regarding the palate. Bone thickness was measured and bone morphology evaluated in the palates of Japanese people. The palates of five Japanese adult cadavers and 15 skulls were examined. The samples were imaged and measured using the micro-CT system. In the mid-palatine suture region, the cortical bone had a complex mesh-like structure and was thicker than surrounding areas. Cortical bone thickness varied depending on the site. The mid-palatine suture region is an ideal site for mini-implant insertion; however, because bone and cortical bone thickness markedly decrease in the lateral region, careful attention should be paid when inserting mini-implants in the mid-palatine suture.

Site-dependent difference in the density of sympathetic nerve fibers in muscle-innervating nerves: a histologic study using human cadavers

The autonomic nerve supply of skeletal muscle has become a focus of interest because it is closely related to the adaptation of energy metabolism with aging. However, there is no comprehensive information concerning the sympathetic nerves present in muscle-innervating nerves (muscle-nerve). At the point of entry of muscle-nerves into 8 striated muscles (the soleus, extensor carpi radialis, infraspinatus, genioglossus, extra- ocular medial rectus, temporalis, lateral pterygoid, and digastricus anterior belly) in 15 cadavers of elderly people, we counted both tyrosine hydroxylasepositive nerve fibers (TH- fibers) and motor nerve fibers to estimate the ratio of THfiber/ motor fibers. The 3 limb muscles were found to have a high ratio (soleus, 58%; infraspinatus; 45%; extensor, 36%), whereas the 4 head muscles (digastricus, 23%; genioglossus, 15%; temporalis, 10%; lateral pterygoid, 6%; medial rectus, 1%) had relatively low ratios. The site- dependent characteristics of the THfibers seemed to reflect their commitment to muscle activity. However, some discrepant characteristics were noted: 1) In spite of the tonic and continuous activity required for both the genioglossus and infraspinatus, the proportions of TH-fibers were quite different between the tongue and the shoulder muscles; 2) Likewise, the soleus and extra-ocular rectus showed a considerable difference, even though rapid and phasic contraction is essential for both muscles. Rather than reflecting the influence of postnatal functional demand, these site- dependent characteristics might develop as a result of differences in sympathetic innervation of the striated muscles during fetal development, i.e., a short course along the arteries feeding the head muscles, or a long course along the muscle-nerves to the limb muscles (AU)

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Nerve terminal distribution in the human tongue intrinsic muscles: An immunohistochemical study using midterm fetuses.

Intrinsic tongue muscles, especially the transverse and vertical (T&V) muscles, regulate the shape of the tongue. However, little information is available on the nerve distribution pattern in human T&V muscles. Using S100 protein immunohistochemistry for paraffin-embedded histology, we investigated semiserial sagittal or frontal sections of eight human fetal tongues (180-240 mm crown-rump length: CRL). The height of the T&V muscle bundle showed a threefold difference between specimens with a small and a large CRL. Thus, the T&V muscles were still growing at the stages examined. In the intrinsic longitudinal muscles and all extrinsic tongue muscles, we observed the typical motor endplate band. In lower-magnification views, the T&V muscles also appeared to carry the band in the lateral part of the tongue, where the genioglossus muscle fibers did not cross these muscles. However, in higher magnification views, the nerve terminal distribution in the T&V muscles showed a unique rule: the nerve terminal for the transverse muscle bundle was located distantly from that of the adjacent vertical muscle bundle. This pattern seemed to be established during the stages examined. To provide such "distantly separated nerve terminals," thin nerve twigs took a highly curved course oblique to the T&V muscle bundles. We hypothesize that the unique nerve course and terminal distribution in the T&V muscles are a result of sorting to provide a good functional match between the nerve fiber and the muscle bundle. After sorting, the T&V muscle cells may initiate proliferation to increase the muscle bundle.

CD34-positive developing vessels and other structures in human fetuses: an immunohistochemical study.

CD34 is a well-known marker of progenitor cells of blood vessels and stromal tissues. Thus, CD34-positive cells have recently been used clinically in the field of vascular and orthopedic biotechnology because of their capacity to assist regeneration of injured tissues. However, to our knowledge, the in situ distribution of CD34-positive cells has not yet been described in the human fetus, with the exception of a few organs. In the present study, we conducted immunohistochemistry for CD34 using 12 human fetuses (9-15 weeks of gestation). CD34-positive structures showed a vessel-like appearance and were regularly arrayed in the viscera, nerves and lymph nodes, whereas in the body wall and extremities, they were distributed diffusely as fibrous tissues, such as the fascia and perimysium. The myocardium was also divided and bundled by CD34-positive fibrous tissues. In striated muscles, limited examples of CD34 expression were found in the tongue and extraocular muscles in which only vessels were positive. Lymphatic vessels were negative for CD34. In addition to their contribution to vascular development in any part of the body, CD34-positive mesenchymal tissues seem to play a critical role in the pattern formation of skeletal muscle, synovial tissue and other muscle/tendon-associated tissues in human fetuses.

Three-dimensional analysis of mesiobuccal root canal of Japanese maxillary first molar using Micro-CT.

The objective of this study was to three-dimensionally observe the morphological characteristics of mesiobuccal root canals of Japanese maxillary first molars using microcomputed tomography (Micro-CT) and classify root canal variations. This study used 90 maxillary first molars. Three-dimensional reconstruction was performed using data obtained by Micro-CT, and cross-sections of the root canals were observed. Moreover, the root canal morphology was classified by the configuration and root canal diameter, and was evaluated for occurrence using the classification by Weine et al. (1969) as a reference. Overall, single root canals were observed in 44.4%, incomplete separation root canals in 22.3%, and completely separate root canals (upper and lower separation root canals) in 33.3%. Mesiobuccal root canals often had intricate configurations, and accessory root canals (lateral canals and apical ramifications) were observed in most of the mesiobuccal root canals (76.7%), irrespective of whether there were ramifications of the main root canals. While there were no marked differences in the incidence of root canal ramifications between this study and earlier reports, the incidence of accessory root canals was higher in this study. This result may be explained by the far more superior visualization ability of Micro-CT than conventional methods, which allowed the detection of microscopic apical ramifications previously difficult to observe.

Immunohistochemical distribution of desmin in the human fetal heart.

Desmin is a member of the intermediate filaments, which play crucial roles in the maturation, maintenance and recovery of muscle fibers. Its expression has been examined in human cardiac muscle, rat and chicken, but its spatial distribution in the human fetal heart has not been described. The present study investigated desmin expression in the human fetal heart and associated great vessels in 14 mid-term fetuses from 9 to 18 weeks of gestation. Immunoreactivity for myosin heavy chain (MHC) and alpha smooth muscle actin (α-SMA), as well as neuron-specific enolase (NSE), was also examined. Increased expression of desmin from 9 to 18 weeks was clearly localized in the atrial wall, the proximal portions of the pulmonary vein and vena cava, and around the atrioventricular node. Desmin-positive structures were also positive for MHC. Meanwhile, the great vessels were also positive for α-SMA. The distribution of desmin exhibited a pattern quite different from that described in previous studies of rat and chicken. Thus, desmin in the human fetal heart does not seem to play a general role in myocardial differentiation but rather a specific role closely related to the maturation of the α-isozyme of MHC. Desmin expression in the developing fetal heart also appeared to be induced by mechanical stress due to the involvement of venous walls against the atrium.

A comparative study of myostatin, follistatin and decorin expression in muscle of different origin.

Muscle regeneration supports muscle function in aging, and plays a role in the functional impairment caused by progressive neuromuscular diseases. Major substances controlling this process are growth factors and the extracellular matrix (ECM). Thus, follistatin is known to antagonize the function of several members of the TGF-ß family of secreted signaling factors, including myostatin-the most powerful inhibitor of muscle growth characterized to date. Decorin-a small leucine-rich proteoglycan-traps myostatin and modulates its activity towards myogenic cells in the ECM. In addition, there are few reports concerning the regenerative muscle process of masseter muscles, which are of branchial arch origin, in mdx mice. Thus, in order to clarify the muscle regenerative process of masseter muscle, gene and protein expression of myostatin, follistatin and decorin were examined using the tibialis anterior (TA)muscle as a positive control. In both muscles, a gradual increase in mRNA myostatin, follistatin and decorin expression was detected, with the increase being greater in TA muscle than in masseter muscle. At 2 weeks, both muscles exhibited normal skeletal muscle cells. At 3 weeks, masseter muscle demonstrated scant areas of necrosis, whereas large necrotic zones were seen in TA muscle. At 4 weeks, the formation of necrotic tissue and presence of follistatin protein was observed clearly in masseter muscle. This result indicates that follistatin production is stimulated in the presence of necrosis. Interestingly, both muscles showed the same process of muscular formation, but with different time frames, which could be related to muscle origin.

Early fetal development of the rotator interval region of the shoulder with special reference to topographical relationships among related tendons and ligaments.

PURPOSE: There is a little information on the early fetal development of the rotator interval region of the shoulder, particularly with regard to whether topographical relationships among the ligaments and tendons change during development. METHODS: We examined the histological sections (transverse or frontal) of right or left shoulder in 20 mid-term human fetuses (7-15 weeks of gestation). RESULTS: The biceps tendon had an accompanying bursa-like cavity before the joint cavitation. The bursa for the tendon remained open to the joint cavity until 12 weeks. When reaching the glenoid, the biceps tendon involved and carried mesenchymal tissue around the coracoid process (the future coracohumeral ligament) toward the infraspinatus tendon. Until 10 weeks, the primitive glenohumeral ligament was established as simple collateral ligaments on the inner or humeral side of the rotator cuff tendons and the biceps long tendon. However, the subscapularis tendon crossed, attached to, and reformed the upper structure of the superior glenohumeral ligament. CONCLUSIONS: The early development of the coracohumeral ligament suggests that it is a primitive and basic structure. However, we hypothesize that mechanical demands from the subscapularis tendon and biceps long tendon are likely to change the primitive form of the rotator interval to the adult morphology, including the superior glenohumeral ligament. The significant modification evident during early fetal development suggests that anatomical reconstruction after rotator cuff tears should not be based on the "ideal" anatomy, especially that of the superior glenohumeral ligament, but on individual requirements.

Comparative histological study of levels 1-3 supportive tissues using pelvic floor semiserial sections from elderly nulliparous and multiparous women.

AIM: The connective tissue located between the uterine cervix and sacrospinous ligament (the uterospinous connective tissue; USCT) has recently been noted as the level 1 supportive tissue instead of the classical uterosacral ligament. We examined whether or not the USCT changes its histological architecture by vaginal delivery in correlation with the levels 2 and 3 supportive tissues. METHODS: In the pelvic floors of 17 female cadavers (9 nuliparous and 8 multiparous), we compared histological architectures among the USCT, arcus tendineus fasciae pelvis (ATFP) and perineal membrane (PM). RESULTS: The USCT was evident as a string-like tissue structure in multiparous women or a thick mesh in nuliparous women. It consistently contained fewer elastic and smooth muscle fibers than other levels. In contrast, the ATFP usually contained abundant elastic fibers and smooth muscle. Likewise, the PM also displayed a constant morphology. CONCLUSION: Although all three sites were likely to be injured by delivery, the USCT seemed to be more severely damaged and/or more difficult to be recovered than the ATFP and PM.

Dens evaginatus on occlusal surface of maxillary second molar: a case report.

Dens evaginatus is a rare dental anomaly characterized by the development of a tubercle on the occlusal surface of the tooth and can cause pulpitis, pulp necrosis, and periapical periodontitis due to tubercular fracture or attrition. Unlike with caries, pain caused by dens evaginatus may manifest itself in a distant location. Therefore, diagnosing the cause of that pain may prove problematic. Dens evaginatus usually occurs in the mandibular premolars. We report a successfully treated case in which dens evaginatus was difficult to diagnose due to distant radiation of pulpitis-induced pain. This pain occurred as a result of fracture of a tubercle located on the occlusal surface of the maxillary second molar, which is very rare.

Biomechanical role of peri-implant cancellous bone architecture.

PURPOSE: The aim of this study was to investigate the biomechanical role of trabecular bone around dental implants in the mandible. MATERIALS AND METHODS: The model in this study was made using micro-computed tomography data taken from a cadaver in whom endosseous implants had been in place for 15 years prior to death. Morphologic analysis and three-dimensional (3D) finite element analysis were performed to calculate the peri-implant loading path of the model in which the trabecular structure was accurately simulated. RESULTS: As seen through multiscale analysis using the homogenization method, the trabecular bone architecture around implants was isotropic for the most part. Also, 3D finite element analysis showed that compressive stresses oblique to the implant axis were transmitted to the lower constrained surface; tensile stresses oblique to the implant axis were transmitted to the upper constrained surface, and they intersected each other with vertical loading. The highest stress in cancellous bone was observed on perpendicular loading, and stress produced in trabeculae decreased approaching horizontal loading. CONCLUSION: Cancellous bone architecture around the implant was generally isotropic. 3D finite element analysis showed that cancellous bone trabeculae around implants dispersed stress by forming load transfer paths. The results suggest that trabecular bone plays a major role in supporting functional pressure exerted via the implant.

Three-dimensional observation of decrease in pulp cavity volume using micro-CT: age-related change.

We investigated three-dimensional decrease in the volume of the pulp chamber caused by age-related secondary dentin formation using micro-CT and evaluated the applicability of the results to evaluation of age taking into account sex, age and tooth type. Decrease was slightly higher in females than in males, and a higher correlation between decrease and aging was observed in females. A comparison between age-groups revealed that decrease progressed between the fifties and sixties in males, and the forties and fifties in females. A stronger correlation between aging and decrease was observed in the mandibular central incisors than in the mandibular second premolars. This correlation was higher than other correlations between sexes and age-groups.

Expression of intermediate filaments at muscle insertions in human fetuses.

Desmin and vimentin are intermediate filaments that play crucial roles in the maturation, maintenance and recovery of muscle fibers and mesenchymal cells. The expression of these proteins has not been investigated extensively in human fetuses. In the present study, we examined the immunohistochemical expression of intermediate filaments in skeletal muscles of the head, neck and thorax in 12 mid-term human fetuses at 9-18 weeks of gestation. We also used immunohistochemistry to localize the expression of the myosin heavy chain and silver impregnation to identify the fetal endomysium. Expression of desmin and vimentin was already detectable in intercostal muscle at 9 weeks, especially at sites of muscle attachment to the perichondrium. At this stage, myosin heavy chain was expressed throughout the muscle fibers and the endomysium had already developed. Beginning with punctate expression, the positive areas became diffusely distributed in the muscle fibers. At 15-18 weeks, intermediate filament proteins were extensively expressed in all of the muscles examined. Expression at the bone-muscle interface was continuous with expression along the intramuscular tendon fibres. These results suggest that the development of intermediate filaments begins in areas of mechanical stress due to early muscle contraction. Their initially punctate distribution, as observed here, probably corresponds to the earliest stage of fetal enthesis formation.