THE ANATOMICAL RECORD 266:167–176 (2002) DOI 10.1002/ar.10050 Morphologic Changes in the TMJ Following Splint Wear BETTY J. SINDELAR,1* STAN EDWARDS,2 AND SUSAN W. HERRING2 1 School of Physical Therapy, Grover Center, Ohio University, Athens, Ohio 2 Department of Orthodontics, University of Washington, Seattle, Washington ABSTRACT Intraoral splints are a commonly used dental treatment for a variety of conditions. Because such splints alter the condyle– disc–fossa relationship, they probably change the loading status of the temporomandibular joint (TMJ), including the TMJ disc. Collagen, a major constituent of the disc, acts to resist tensile loading, and it is presumed that the fiber orientations of the individual disc bands reflect their functional loading. Therefore, the purpose of this study was to examine effects of intraoral splint wear on TMJ morphology in general, and collagen orientation of the intra-articular disc in particular. Young adult, female miniature pigs were divided into three groups: open-bite splint, protrusive-bite splint, and unsplinted control. Splints were worn for 2 months, after which the TMJ discs were harvested for histological examination and stereological analysis, and the skulls were cleaned. Although the splints had no effect on skull dimensions, changes were seen in the TMJs. The discs of the protrusively-splinted group showed an increased thickness of the posterior band (P ⬍ 0.015) and minor changes in collagen orientation of the anterior band. The most striking change was the presence of a degenerative osseous defect on the medial side of the mandibular condyle in half of the splinted animals. These results indicate that prolonged splint wear can induce remodeling and even injury of TMJ tissues. Anat Rec 266:167–176, 2002. © 2002 Wiley-Liss, Inc. Key words: temporomandibular joint disc; porcine; morphology; intraoral splints; collagen Intraoral splints are used in dentistry to treat a variety of conditions, such as occlusal malalignments, temporomandibular joint (TMJ) dysfunctions, and sleep apnea (Bondemark, 1999; Carels and van der Linden, 1987; Gianelly et al., 1970; Grim, 1995; Johal and Battagel, 1999; Kimmel, 1994; Major and Nebbe, 1997; Nitzan, 1994; Wright et al., 1995). All splints change the occlusal relationship of the teeth and, hence, the spatial relationship of the TMJ components, although the precise effect varies with the splint (Ito et al., 1986). Additionally, splint wear must affect disc structure, as indicated by studies showing changes in shape (Ferrari and Herring, 1995) and glycosaminoglycan (GAG) content (Mao et al., 1998; Sindelar et al., 2000). However, detailed morphology has not previously been assessed. The TMJ disc is composed of approximately 80% water (Sindelar et al., 2000); the remaining 20% is mostly collagen. Prior studies on the human, bovine, dog, rabbit, monkey, sheep, and rat TMJ disc have shown that the fibers of the intermediate zone are oriented predominantly in the anterior-posterior (AP) direction, while the fibers of the anterior and posterior bands are more multidirectional or “basket-woven” (Berkovitz, 2000; Desai et al., 1996; Gillbe, 1973; Landesberg et al., 1996; Mills et al., 1988; © 2002 WILEY-LISS, INC. Mills et al., 1994b; Minarelli et al., 1997; Minarelli and Liberti, 1997; Strauss et al., 1960; Taguchi et al., 1980; Teng and Xu, 1991). The arrangement or structural organization of the collagen fibers is thought to correspond to the mechanical function of the disc with fibers paralleling the direction of applied tensile loading (Smith et al., 1981). Thus, if altered loading occurs, the disc may adapt by reorientation of the collagen fibers to accommodate the direction of the newly applied tensile stresses (Scapino and Mills, 1997). In previous studies, surgically induced anterior displacement of the rabbit TMJ disc did in fact lead to changes in the collagen fiber orientation of the anterior bands and intermediate zones (Mills et al., 1994a; Mills and Scapino, 1993). In particular, the fibers of the intermediate zone became more multidirectional, resem- Grant sponsor: NIH/NIDR; Grant number: 5 RO1 DE11236; Grant sponsor: NIH; Grant number: T35 DE07150. *Correspondence to: Betty Sindelar, Ph.D., School of Physical Therapy, Grover Center, Rm W295, Ohio University, Athens, OH 45701. Fax: (740) 593-0292. E-mail: email@example.com Received 29 November 2000; Accepted 10 December 2001 Published online 14 February 2002 168 SINDELAR ET AL. Fig. 1. Intraoral splints. Top: Chrome-cobalt ramp splints, occlusal view. Splints were cemented onto occlusal surface of premolars and molars of 8-month-old animals. Bottom: Protrusive splint (PS), lateral view. Anterior is to the left. bling the posterior band. However, it is always difficult to assess the effect of surgery on the involved tissues. Therefore, we proposed to alter the load on the TMJ disc in a nonsurgical way, by changing the fossa– disc– condyle relationships through the use of intraoral splints. The purpose of this study was to examine the effect of intraoral splint wear on the collagen fiber orientation of all disc bands. Two types of splints were used. One, which protruded the mandible, was assumed to bring the condyle to a more anterior functional position on the disc. The SPLINT-INDUCED TMJ MORPHOLOGIC CHANGES second was a simple opening splint that presumably left the joint components in their normal positions. The control group wore no intraoral splint. It was expected that in the protrusively splinted group the collagen fibers of the intermediate zone would become more multidirectional, while the fibers of the anterior band would become more unidirectional. We hypothesized that, because of loading in the altered position, the anterior band would functionally become the intermediate zone, and would therefore orient more of its fibers along the AP axis of the disc. The miniature pig was chosen as the animal model because like humans, pigs are omnivorous, have bunodont molars, and use a transverse chewing stroke (Bermejo et al., 1993; Herring, 1976; Weaver et al., 1962). The disc of the TMJ is quite similar to man in gross structure, histologic and biochemical composition, and response to mechanical loading (Berg, 1973; Christensen, 1975; Fontenot, 1985; Meister et al., 1973; Kopp, 1976; Sindelar et al., 2000). Because of the limited availability of fully mature miniature pigs, young adults were used, creating a possible confounding problem due to growth. Previous studies noted not only dental changes but facial growth alterations in monkeys and pigs as a result of intraoral bite block wear (Altuna and Woodside, 1985; Carlson and Schneiderman, 1983; Ferrari and Herring, 1995; Schneiderman, 1989). Therefore, an additional objective of this project was to examine the skull and mandible for bony growth or adaptive changes that could have occurred as the result of intraoral splint wear. Because the animals used in this study were post-pubertal and experiencing slow growth, we did not expect to find size differences among the groups. However, we did expect some evidence of injury to the TMJ tissues because most previous studies noted pathologic effects (whereas Ramfjord and Blankenship (1981) did not). MATERIALS AND METHODS Nineteen female Hanford miniature pigs were acquired in sibling sets at 7 months of age. The subjects were randomly divided into three groups: control (C), openingonly (control) splint (CS), and protrusive splint (PS). Impressions using custom-made forms and bite registrations were taken on all pigs. Each animal received surgical implantation of a small radio-opaque screw near the root of the left maxillary and mandibular canines. These screws were distant from the TMJ (approximately 14 cm) and jaw muscles, and did not interfere with chewing function (Sindelar, unpublished EMG data). They allowed the AP positioning of the mandible to be checked on videofluoroscopy during chewing. Prior to splint delivery, all pigs were examined with a baseline videofluoroscopy chewing session to determine the relative position of the maxilla and the mandible in the AP direction. At 8 months of age, the animals in the CS and PS groups were fitted with chrome-cobalt ramp splints that covered the occlusal surfaces of the maxillary and mandibular molars (Fig. 1). The CS and PS splints increased the height of the bite by 5 mm at the first molariform tooth while the PS splint also positioned the mandible 7 mm anteriorly. All splints were cemented onto the teeth and were worn continuously for 2 months. Splints were checked daily and, when necessary, replaced as quickly as possible. As stated previously, the C group received no intraoral appliances. Additional videofluoroscopy sessions occurred within 2 days of splint de- 169 Fig. 2. Superior view of disc. Portions marked “C” were used for this study. The center hatched area (11 mm wide) was used in another study. The “slice” section was used to measure disc thicknesses. livery, 1 month post-delivery, and 2 months post-delivery. These sessions were used to assess whether the animals were chewing into the splint position (i.e., the ramp portions were coming into contact during the closed portion of the chewing cycle). Qualitative comparisons of the relative positions of the implanted screws at the closed portion of the chewing cycle were made to confirm that the mandible was advanced in the PS group but not in the CS and C groups. Food and water were provided to all animals per breeder guidelines (Charles River, Wilmington, MA). After 2 months of splint wear, the animals were euthanized with an intracardiac injection of pentobarbital (10 cc; Guidelines of the Animal Care Committee, University of Washington, Seattle, WA). The left TMJ disc was dissected out and frozen. The heads were harvested and cleaned. Disc Left discs were thawed at room temperature. Approximately 40% of each disc was removed from the middle by two sagittal cuts (Fig. 2) and were used for a different study. The remaining medial and lateral portions were used for this study. A 1-mm sagittal slice was taken from the lateral edge of the medial section and coded to blind the investigators to treatment group. Images of these slices were captured using a videocamera and analyzed using NIH Image. The thinnest portion of the intermediate zone and the thickest portions of the anterior and posterior bands were measured three times and averaged. The slice was then fixed with ethanol and processed for paraffin vacuum-embedding using a standard protocol (Bancroft and Stevens, 1996). To assess for the effects of processing on shrinkage, these slices were then remeasured. Collagen Pattern The medial and lateral disc portions were fixed in ethanol, embedded in paraffin, and sectioned in the horizontal plane from the superior surface at 7.0 m (Bancroft and Stevens, 1996). Sections were stained with Direct Red, which identifies collagen almost exclusively (Junqueira et al., 1979; Sweat et al., 1964). Three separate sections taken from the disc core were analyzed for each region: lateral anterior, intermediate, and posterior bands; and medial anterior, intermediate, and posterior bands. The images were captured (Nikon T4, Kodak Ektachrome 160T color film) while being viewed under a microscope at 50⫻ (Nikon Eclipse 5100) (Fig. 3). These images were then converted to black and white bitmap 170 SINDELAR ET AL. Fig. 4. Average angle frequency distributions. C ⫽ control group, CS ⫽ control splint group, and PS ⫽ protrusive splint group. Angles of 0° and 180° indicate AP orientation, whereas 90° indicates mediolateral orientation. format for stereological analysis using a custom software program, MacAzimuth (written by Prof. John Rensberger, Geological Sciences, University of Washington). This software determines the orientation of all strings of six contiguous pixels relative to a fixed axis; further information about the software can be found in Teng et al. (1997) and Rensberger and Watabe (2000). The fixed axis was the AP axis of the disc. Output is given as a frequency distribution of angles and constitutes a direct measure of the orientation of the collagen fibers in the image (Fig. 4). For each image, average orientation (mean angle) was calculated. The standard deviation of the mean angle was taken as a measure of anisotropy, the nonrandom directionality of the collagen fibers. Skull Measures Fig. 3. Representative horizontal sections from the lateral portion of a control group disc. Top: Anterior band. Middle: Intermediate zone. Bottom: Posterior band. Direct Red staining. Anterior is to the top; medial is to the right. Prior studies (Altuna and Woodside, 1985; Carlson and Schneiderman, 1983; Ferrari and Herring, 1995) have indicated that intraoral splint wear alters the dimensions of the maxilla and mandible. Therefore, the following widths and lengths were measured on the maxilla and mandible of the dried skulls (Fig. 5): Right (RML) and left mandibular length (LML): infradentale to the most posterior point on the condyle. 171 SPLINT-INDUCED TMJ MORPHOLOGIC CHANGES Fig. 5. Upper left: Lateral view of pig skull. RML ⫽ right mandibular length. Upper right: Posterior view of mandible. MW ⫽ maximum mandibular width, CNW ⫽ condylar neck width, and CW ⫽ coronoid width. Lower left: Superior view of mandible. MA ⫽ mandibular arch width. Lower right: Palatal view of skull. MXA ⫽ maxillary arch width and PL ⫽ palate length. Maximum mandibular width (MW): taken near the mandibular angles. Width at condylar neck (CNW): taken immediately below the scar from the attachment of the posterior capsule. Width at coronoid processes (CW): taken at the superior-most point. Mandibular arch (MA) and maxillary arch (MXA) width: taken between the lingual surfaces of the first molars at the posterior cemento-enamel junction on the mandible and maxilla, respectively. Palate length (PL): hard palate length. Two different standardization measures were employed: 1) cube root of weight in kilograms, and 2) basicranial axis length (Radinsky, 1984). The basicranial axis length was considered preferable to total skull length, which would have been affected by changes in upper jaw dimensions. All measurements were taken three times and averaged for final analysis. Articular Surfaces The articular surfaces of the temporal fossa and the mandibular condyle were examined under the dissecting microscope for signs of degenerative changes. Images of the condylar articular surfaces were captured using a videocamera and analyzed using NIH Image. All images were blinded as to treatment group. The medial-lateral width and AP length of the articular surface were measured, and surface areas were calculated using NIH Image. Measurements were taken three times, averaged for final analysis, and standardized to appropriate power of weight. For all measurements, a nested analysis of variance (ANOVA) was used. The individual pigs were randomly assigned to treatment group, so “pig” was the random effect whereas “treatment group” was the fixed effect. A Fisher’s exact test was used to examine the degenerative changes in the condylar articular surface. RESULTS The fluoroscopy sessions showed that the animals fully occluded into their splints in almost all masticatory cycles (85% or more). Examination of the screw position at occlusion verified that the PS group did have protruded mandibular positions. Overall, average disc dimensions were 26.8 ⫾ 2.2 mm for the medial-lateral width and 14.5 ⫾ 1.5 mm for the AP length, with no difference among groups. Regardless of group, the posterior band was always the thickest and the 172 SINDELAR ET AL. intermediate zone was the thinnest (P ⬍ 0.01) (Table 1). Additionally, a significant treatment/band interaction was present for the posterior band, which was thicker in the PS than in the other groups (P ⬍ 0.015). Histologic preparation of the disc led to a linear shrinkage in the tissues of 21.5%. The mean angles of the collagen fiber orientation within the intermediate zones of all groups were aligned around the AP axis or 0° in our system (Table 2). Orientation of the anterior and posterior bands ranged from 81° to 96° in all groups. Within each treatment group, the orientation of the intermediate zone was significantly different from the other bands (P ⬍ 0.001), whereas the anterior and posterior bands could not be distinguished. A band-location interaction was present in the C and CS groups (P ⬍ 0.001) (Fig. 6). In the anterior bands, the medial locations were inclined slightly posterolaterally, while the lateral locations were inclined slightly posteromedially (Fig. 7). In the posterior bands, the medial locations showed collagen to be almost perfectly mediolateral (90°), but the lateral locations sloped slightly anteromedially. In contrast, the PS group showed no significant band-location interaction. In the posterior band, the medial-lateral difference was reduced, and in the anterior band both locations were oriented very closely to the mediolateral axis. Visual examination of the intermediate zones showed that the collagen fibers displayed a marked crimping (Fig. 3B), similar to the pattern found in ligament and tendon. Much less crimping was seen in the anterior and posterior bands. The magnitude of the waviness in the intermediate zone made the analysis of parallelness impossible because the program measured the direction of the crimps rather than the direction of the fibers. Therefore, the analysis of fiber direction was only performed on the anterior and posterior bands, both medial and lateral components. Results of this analysis indicated that no significant differences were present relative to band, location, or treatment (Table 3). The splints had no effect on skeletal dimensions, including those of the condyle with or without either standard- ization technique (Table 4). However, there was a trend for the condylar measurements (width, perimeter, and area) to be smaller in the PS group. Furthermore, visual TABLE 1. Mean disc thickness measurements ⴞ S.D. (mm) Groups Anterior band Intermediate zone Posterior band C CS PS 4.0 ⫾ 0.7 3.9 ⫾ 0.5 4.1 ⫾ 0.5 1.2 ⫾ 0.1 1.3 ⫾ 0.2 1.3 ⫾ 0.3 5.2 ⫾ 0.5 6.1 ⫾ 1.4 7.6 ⫾ 1.5* Within each treatment group, bands were significantly different (P ⱕ 0.001). In addition, the posterior band of the PS group was thicker than the other groups (P ⬍ 0.02). C, control (n ⫽ 6); CS, control splint (n ⫽ 6); PS, protrusive splint (n ⫽ 7). Fig. 6. Mean collagen fiber orientation (⫾ standard error). The C and CS groups showed significant mediolateral differences in the anterior and posterior bands (P ⬍ 0.001) but the PS group did not. TABLE 2. Mean angles (degrees) of collagen fibers relative to the AP axis ⴞ standard error Anterior band C CS PS Intermediate zone Posterior band Medial Lateral Medial Lateral Medial Lateral 85.2 ⫾ 2.0 87.5 ⫾ 2.0 88.9 ⫾ 1.9 94.4 ⫾ 2.0 96.2 ⫾ 2.0 88.4 ⫾ 1.8 ⫺0.5 ⫾ 2.0a 1.5 ⫾ 2.0a ⫺1.5 ⫾ 2.0a 0.7 ⫾ 2.0a 1.6 ⫾ 2.0a 1.6 ⫾ 2.0a 89.3 ⫾ 2.1 88.2 ⫾ 2.1 90.4 ⫾ 1.9 82.2 ⫾ 2.0 80.6 ⫾ 2.0 85.1 ⫾ 1.9 Indicates significance within treatment group (P ⬍ 0.001). C, control group (n ⫽ 6); CS, control splint group (n ⫽ 6); PS, protrusive splint group (n ⫽ 7). a 173 50.4 ⫾ 6.0 50.6 ⫾ 3.4 50.9 ⫾ 5.6 48.7 ⫾ 5.0 49.8 ⫾ 6.9 49.0 ⫾ 5.3 No significant differences noted. C, control group (n ⫽ 6); CS, control splint group (n ⫽ 6); PS, protrusive splint group (n ⫽ 7). Mean standard deviation of the mean angle ⫾ S.D. inspection of the condyles showed a defect (Fig. 8) present on the medial aspect of the articular surface in 53.8% of the splinted animals (26 condyles inspected) (Table 5). No defects of this type were found in any of the 10 mandibular condyles of the control group. No significant difference was noted between the splinted groups. When present, the defect was found bilaterally, appeared to be pre-mortem, and appeared to be indicative of degenerative joint disease. The cranial components of the joints appeared normal in all cases. 154.4 ⫾ 75.7 149.5 ⫾ 32.5 125.7 ⫾ 20.9 71.2 ⫾ 21.8 71.2 ⫾ 15.7 59.0 ⫾ 5.2 23.0 ⫾ 1.6 23.2 ⫾ 1.9 23.0 ⫾ 0.4 † P 37.6 ⫾ 1.8 38.1 ⫾ 1.9 38.7 ⫾ 2.1 36.6 ⫾ 1.4 34.7 ⫾ 2.2 36.1 ⫾ 2.1 155.7 ⫾ 3.8 155.3 ⫾ 4.7 157.4 ⫾ 5.7 † 76.4 ⫾ 4.2 76.5 ⫾ 3.2 77.5 ⫾ 4.1 PL MXA MA CW CNW Numbers given are the raw values of the measurements in mm or mm2. RX ⫽ treatment group where C ⫽ control (n ⫽ 6 except for † where n ⫽ 5), CS ⫽ control splint (n ⫽ 6), and PS ⫽ protrusive splint (n ⫽ 7). W ⫽ medial-lateral width of the articular surface of the condylar head. P ⫽ perimeter of the articular surface of the condylar head. A ⫽ surface area of the condylar head. For all other abbreviations, see text. No medial-lateral differences were noted in the condylar articular surface measurements, so side measures were averaged for this table. No significant differences were noted. 47.8 ⫾ 2.4 46.2 ⫾ 7.9 50.5 ⫾ 3.4 95.0 ⫾ 2.9 98.5 ⫾ 3.0 98.1 ⫾ 4.0 51.7 ⫾ 7.6 48.7 ⫾ 4.0 48.8 ⫾ 6.1 104.5 ⫾ 4.4 106.0 ⫾ 3.4 104.8 ⫾ 4.0 Lateral 204.8 ⫾ 5.8 203.0 ⫾ 5.7 208.3 ⫾ 4.0 Medial 205.4 ⫾ 5.1 202.3 ⫾ 5.3 207.3 ⫾ 4.6 Lateral C CS PS Medial MW C CS PS Posterior band LML Anterior band RML TABLE 3. Measure of randomness RX Fig. 7. Schematic representation of the mean angle orientation of the collagen fibers of the left disc. Solid squares represent areas examined for fiber orientation. Thin solid lines across the disc surface represent 90° orientation. Anterior is to the top. TABLE 4. Measurements of the skull and mandible (mean ⴞ S.D.) † W A SPLINT-INDUCED TMJ MORPHOLOGIC CHANGES 174 SINDELAR ET AL. Fig. 8. Example of the observed defects. Superior surface of the right condyle from a PS animal. Arrow points to the defect present on the medial articular surface in 55% of the TMJs of the splinted animals. Bar length ⫽ 1 cm. TABLE 5. Presence of defect on the medial aspect of the articular surface of the mandibular condyle* Defect presence Yes No Total Control a 0 10 10 Control splint Protrusive splint Total 6 6 12 8 6 14 14 22 36 *See Fig. 8 for a typical example. Different from other groups (P ⱕ 0.005). Mandibular condyles from one of the control animals were unavailable for observation. a DISCUSSION The splints altered the occlusal relationship by changing the position of the mandible, not by affecting growth. The lack of change in the measured skeletal parameters among the treatment groups is undoubtedly related to the age of the subjects. When the splints were delivered, the animals were 8 months of age, which is past puberty. Therefore, although these splints were similar to those used in other studies (Altuna and Woodside, 1985; Carlson and Schneiderman, 1983; Ferrari and Herring, 1995; Schneiderman, 1989), they did not induce bony growth adaptations. However, the splints apparently had an adverse impact on the TMJ structures, as indicated by the defect on the medial condylar articular surface and the trend of the PS to have smaller condylar articular surfaces. That the changes resulted from splint wear rather than the other experimental procedures (e.g., marker screw placement) is indicated by the absence of any changes in the otherwise identically treated control animals. The defects observed suggest that the splints altered the loading on the medial side of the joints in a way that the animal was unable to accommodate, resulting in extensive bony destruction. It is clear that the protrusive splint caused remodeling of the disc. Within the disc, the most obvious morphological change was the increased thickness of the PS posterior band. This finding is similar to that reported previously (Ferrari and Herring, 1995; Scapino and Mills, 1997), and suggests an adaptation to a larger posterior joint space created by the anterior displacement of the mandible via the splint. However, the internal structure of the posterior band was not altered by splinting and continued to feature multidirectional collagen fibers. Even with splints, the intermediate zone retained its strong alignment with the AP axis of the disc, which is consistent with previous reports of the collagen alignment of this zone in other species (Mills et al., 1988; Mills et al., 1994b; Strauss et al., 1960; Taguchi et al., 1980; Teng and Xu, 1991). It would be reasonable to assume that this zone sees a large tensile force in the AP direction during physiologic function with or without splints. Of interest is the fact that the intermediate zone also has a very large concentration of highly sulfated GAGs, indicating that this zone also sees a large compressive force during physiologic function (Sindelar et al., 2000). These extremes of compressive and tensile loading during function make this tissue unique. Our principal hypothesis, that the anterior band would develop an AP collagen orientation, was not borne out. SPLINT-INDUCED TMJ MORPHOLOGIC CHANGES Indeed, the CS and PS anterior bands showed fewer AP oriented collagen fibers than the controls (Figs. 3 and 7). That the PS anterior band had even more transverse fibers than normal is further indicated by the loss of the usual medial-lateral differences (Figs. 6 and 7). Although these alterations are subtle, they suggest that functional loading did change, albeit not in the expected way. Other studies have shown that when a disc band or even the posterior discal attachment becomes loaded like the intermediate zone, that area begins to remodel itself into a pseudo-intermediate zone (Mills et al., 1994a; Scapino and Mills, 1997; Scapino, 1983). While expected, major orientation changes were not observed in the PS anterior band in this study. Given the ages of the experimental animals and the relatively short duration of the splint wear, there simply may not have been enough time for remodeling to occur. Additionally, since the splints only function when the teeth are in occlusion, the experimental animals did not spend the entire 2 months in the altered loading condition. It is important to note here that in addition to chewing, pigs spend a considerable amount of time bruxing. Yet there certainly was a large proportion of time with no tooth-to-tooth contact. Thus, a longer wear time or an experimental modification to control condyle position at all times might have resulted in more obvious histologic changes. Alternatively, in the absence of direct observation of the functioning joints, we cannot be certain that the PS splint actually forced the condyle to function against the anterior band rather than the intermediate zone. We emphasize at this point that the relationship of the loading among the condyle, disc, and fossa is not clear in the splinted positions, especially the PS state. Based on the results of this study, our initial assumption that the condyle would simply be translated anteriorly and the mechanics of movement would not be different from the normal state appears to be false. Perhaps additional splint wear time would clear up this matter, but we would also suggest that monitoring the condyle– disc–fossa relationship during in vivo movement is absolutely necessary to understand altered loading mechanics. In summary, the 2-month splint wear resulted in degenerative changes in the medial aspect of the mandibular condyle, an increase in the superior-inferior thickness of the posterior disc band, and a greater similarity in the angulation of medial vs. lateral collagen fibers in the anterior band. While the first result was found in both splinted groups, the latter two results were found only in the protruded group. ACKNOWLEDGMENTS We thank John Rensberger, Ph.D., for help with the MacAzimuth program, Ms. Patricia Emry for lab assistance, and Todd Alonzo, Ph.D., for help with the statistical analysis. LITERATURE CITED Altuna G, Woodside D. 1985. Response of the midface to treatment with increased vertical occlusal forces. Angle Orthod 55:251–263. Bancroft J, Stevens A. 1996. Theory and practice of histological techniques. 4th ed. New York: Churchill Livingstone. 766 p. Berg R. 1973. Contribution to the applied and topographical anatomy of the temporomandibular joint of some domestic mammals with particular reference to the partial resp. total resection of the articular disc. Folia Morphol (Praha) 21:202–204. 175 Berkovitz B. 2000. Crimping of collagen in the intra-articular disc of the temporomandibular joint: a comparative study. J Oral Rehab 27:608 – 613. Bermejo A, González O, González JM. 1993. The pig as an animal model for experimentation on the temporomandibular articular complex. Oral Surg Oral Med Oral Pathol 75:18 –23. Bondemark L. 1999. Does 2 years’ nocturnal treatment with a mandibular advancement splint in adult patients with snoring and OSAS cause a change in the posture of the mandible? Am J Orthod Dentofacial Orthop 116:621– 628. Carels C, van der Linden F. 1987. Concepts on functional appliances’ mode of action. Am J Orthod Dentofacial Orthop 92:162–168. Carlson DS, Schneiderman ED. 1983. Cephalometric analysis of adaptations after lengthening of the masseter muscle in adult rhesus monkeys, macaca mulatta. Arch Oral Biol 28:627– 637. Christensen LV. 1975. Elastic tissue in the temporomandibular disc of miniature swine. J Oral Rehab 2:373–377. Desai S, Johnson DL, Howes RI, Rohrer MD. 1996. Changes in the rabbit temporomandibular joint associated with posterior displacement of the mandible. Int J Prosthodont 9:46 –57. Ferrari CS, Herring SW. 1995. Use of a bite-opening appliance in the miniature pig: modification of craniofacial growth. Acta Anat 154: 205–215. Fontenot MG. 1985. Viscoelastic properties of human TMJ discs and disc replacement materials. J Dent Res 64:163. Gianelly AA, Ruben MP, Risinger R. 1970. Effect of experimentally altered occlusal vertical dimension on temporomandibular articulation. J Prosthet Dent 24:629 – 635. Gillbe GV. 1973. A comparison of the disc in the craniomandibular joint of three mammals. Acta Anat 86:394 – 409. Grim DL. 1995. Seeing the larger medical picture: airway enhancement for true orthodontic health. J Gen Orthod 6:5– 8. Herring SW. 1976. The dynamics of mastication in pigs. Arch Oral Biol 21:473– 480. Ito T, Gibbs C, Marguelles-Bonnet R, Lupkiewicz S, Young H, Lundeen H, Mahan P. 1986. Loading on the temporomandibular joints with five occlusal conditions. J Prosthet Dent 56:478 – 484. Johal A, Battagel J. 1999. An investigation into the changes in airway dimension and the efficacy of mandibular advancement appliances in subjects with obstructive sleep apnoea. Br J Orthod 26:205–210. Junqueira LCU, Bignolas G, Bretani RR. 1979. Picrosirius staining plus polarization microscopy, a specific method for collagen detection in tissue sections. Histochem J 11:447– 455. Kimmel SS. 1994. Temporomandibular disorders and occlusion: an appliance to treat occlusion generated symptoms of TMD in patients presenting with deficient anterior guidance. J Craniomandibular Pract 12:234 –240. Kopp S. 1976. Topographical distribution of sulphated glycosaminoglycans in human temporomandibular joint disks. J Oral Pathol 5:265–276. Landesberg R, Takeuchi E, Puzas JE. 1996. Cellular, biochemical and molecular characterization of the bovine temporomandibular joint disc. Arch Oral Biol 41:761–767. Major PW, Nebbe B. 1997. Use and effectiveness of splint appliance therapy: review of literature. J Craniomandibular Pract 15:159 – 166. Mao JJ, Rahemtulla F, Scott PG. 1998. Proteoglycan expression in the rat temporomandibular joint in response to unilateral bite raise. J Dent Res 77:1520 –1528. Meister R, Berg R, Berg P. 1973. Beiträge zur topographischen und angewandten anatomie des kiefergelenkes (articulatio temporomandibularis) einiger haussäugetiere unter besonderer berücksichtigung der resektionsmöglichkeiten des discus articularis. Z Exper Chirurg 6:437– 448. Mills DK, Daniel JC, Scapino R. 1988. Histological features and in-vitro proteoglycan synthesis in the rabbit craniomandibular joint disc. Arch Oral Biol 33:195–202. Mills DK, Scapino R. 1993. Extracellular matrix changes following disc displacement in the rabbit CMJ. J Dent Res 72:371. Mills DK, Daniel JC, Herzog S, Scapino RP. 1994a. An animal model for studying mechanisms in human temporomandibular joint disc derangement. J Oral Maxillofac Surg 52:1279 –1292. 176 SINDELAR ET AL. Mills DK, Fiandaca DJ, Scapino RP. 1994b. Morphologic, microscopic and immunohistochemical investigations into the function of the primate TMJ disc. J Orofac Pain 8:136 –154. Minarelli AM, Liberti EA. 1997. A microscopic survey of the human temporomandibular joint disc. J Oral Rehab 24:835– 840. Minarelli AM, Del Santo M, Liberti E. 1997. The structure of the human temporomandibular joint disc: a scanning electron microscopy study. J Orofac Pain 11:95–100. Nitzan DW. 1994. Intraarticular pressure in the functioning human temporomandibular joint and its alteration by uniform elevation of the occlusal plane. J Oral Maxillofac Surg 52:671– 679. Radinsky L. 1984. Basicranial axis length v. skull length in analysis of carnivore skull shape. Biol J Linn Soc 22:31– 41. Ramfjord S, Blankenship J. 1981. Increased occlusal vertical dimension in adult monkeys. J Prosthet Dent 45:74 – 83. Rensberger JM, Watabe M. 2000. Fine structure of bone in dinosaurs, birds and mammals. Nature 406:619 – 622. Scapino RP. 1983. Histopathology associated with malposition of the human temporomandibular joint disc. Oral Surg Oral Med Oral Pathol 55:382–397. Scapino RP, Mills DK. 1997. Disc displacement internal derangements. In: McNeil C, editor. Science and practice of occlusion. Chicago: Quintessence Publishing Company. p 220 –234. Schneiderman E. 1989. A longitudinal cephalometric study of incisor supra-eruption in young and adult rhesus monkeys (macaca mulatta). Arch Oral Biol 34:137–141. Sindelar B, Evanko S, Alonzo T, Herring S, Wight T. 2000. Effects of intraoral splint wear on proteoglycans in the temporomandibular joint disc. Arch Biochem Biophys 379:64 –70. Smith JFH, Canham PB, Starkey J. 1981. Orientation of collagen in the tunica adventitia of the human cerebral artery measured with polarized light and the universal stage. J Ultras Res 77:133–145. Strauss F, Christen A, Weber W. 1960. The architecture of the disk of the human temporomandibular joint. Helv Odontol Acta 4:1– 4. Sweat F, Puchtler H, Rosenthal SI. 1964. Sirius red F3BA as a stain for connective tissue. Arch Pathol 78:69 –72. Taguchi N, Nakata S, Oka T. 1980. Three-dimensional observation of the temporomandibular joint disk in the rhesus monkey. J Oral Surg 38:11–15. Teng S, Xu Y. 1991. Biomechanical properties and collagen fiber orientation of TMJ discs in dogs. I. Gross anatomy and collage fiber orientation of the discs. J Craniomandib Disord 5:28 –34. Teng S, Choi I, Herring S, Rensberger J. 1997. Stereological analysis of bone architecture in the pig zygomatic arch. Anat Rec 248:205– 213. Weaver ME, Sorenson FM, Jump EB. 1962. The miniature pig as an experimental animal in dental research. Arch Oral Biol 7:17– 24. Wright E, Anderson G, Schulte J. 1995. A randomized clinical trial of intraoral soft splints and palliative treatment for masticatory muscle pain. J Orofac Pain 9:192–199.