Structure-function relationships of TMJ lateral capsule-ligament complex

Abstract

The human temporomandibular joint (TMJ) lateral capsule ligament (LCL) complex is debated as a fibrous capsule with distinct ligaments or ligamentous thickening, necessitating further evaluation of the complex and its role in TMJ anatomy and mechanics. This study explores the ultrastructural arrangement, biomechanical tensile properties, and biochemical composition of the human LCL complex including region-specific differences to explore the presence of a distinct temporomandibular ligament and sex-specific differences to inform evaluations of potential etiological mechanisms. LCL complex ultrastructural arrangement, biomechanical properties, and biochemical composition were determined using cadaveric samples. Statistical modeling assessed sex- and region-specific effects on LCL complex tissue properties. Collagen fiber coherency, collagen fiber bundle size, and elastin fiber count did not differ between sexes, but females trended higher in elastin fiber count. LCL complex water and sGAG content did not differ between sexes or regions, but collagen content was higher in the anterior region (311.0 ± 185.6 μg/mg) compared to the posterior region (221.0 ± 124.9 μg/mg) (p = 0.045) across sexes and in males (339.6 ± 170.6 μg/mg) compared to females (204.5 ± 130.7 μg/mg) (p = 0.006) across regions. Anterior failure stress (1.1 ± 0.7 MPa) was larger than posterior failure stress (0.6 ± 0.4 MPa) (p = 0.024). Regional differences confirm the presence of a mechanically and compositionally distinct temporomandibular ligament. Baseline sex-specific differences are critical for etiological investigations of sex disparities in TMJ disorders. These results have important biomechanical and clinical ramifications, providing critical baseline tissue material properties, informing the development of TMJ musculoskeletal models, and identifying new areas for etiologic investigations for temporomandibular disorders.

Keywords: Extracellular matrix composition; Second harmonic generation microscopy; Temporomandibular joint disorders; Tissue biomechanics; Tissue ultrastructure.

Figure 1.

(A) Dissected intact human temporomandibular joint, with the LCL complex surface exposed. (B) For biochemical and biomechanical testing, LCL complex specimens were prepared from the anterior and posterior regions of the LCL complex. (C) Representative image of how bone-ligament-bone units were clamped for tensile biomechanical testing. Specimens containing the zygomatic arch, LCL complex, and mandibular condyle were prepared for biomechanical testing. Bone-ligament-bone units were clamped at the bony locations (zygomatic arch and mandibular condyle) of each unit for testing. (D) For biomechanical tensile testing, LCL complex specimens underwent sequential stress-relaxation tests at 10% strain and 20% strain, followed by a strain-to-failure (STF) experiment. Specimens were tested using a ramp strain input, with a strain rate of 1% strain-per-second. For each relaxation phase, specimen strain was held constant for 1800 seconds. Preceding each test phase, specimens underwent five cycles of preconditioning at 10% strain for the 10% strain stress-relaxation experiment, and 20% strain for the 20% strain stress-relaxation and strain-to-failure experiments. Between all tensile tests, specimens were allowed to rest at 0% strain for 300 seconds. STF data is truncated. (E) To characterize baseline sex-specific and region-specific differences in LCL complex biomechanics, specimen Elastic Modulus and Failure stress were determined. (F) Fung’s viscoelastic parameters were calculated for the loading and unloading stress response. Example stress relaxation curve and QLV curve fit shown.

Figure 2.

(A) Representative Second Harmonic Generation (SHG) microscopy image of the intact human temporomandibular LCL complex with bundle diameter measurements shown in yellow. In the human LCL complex, collagen fibers are irregularly arranged, with an anterosuperior to posteroinferior direction with principal fiber direction in the superior-inferior direction. (B) No statistically significant differences between males (n=7) and females (n=3) in collagen fiber coherency (left) or fiber diameter (right) were determined. Box plots show median and interquartile range of collagen fiber coherency and fiber diameter to display data variability. (C) Representative false color composite images of the underlying collagen ultrastructure (red) with the co-localized elastic fiber autofluorescence (green). (D) Distinct elastic fibers were counted, and differences between males and females were determined. There was no statistically significant difference between males and females in the total number of elastic fibers. Box plots show median and interquartile range of elastin fiber count to display data variability.

Figure 3.

Tensile biomechanical stress-relaxation tests were performed on bone-ligament-bone units at 10% strain, 20% strain and strain-to-failure (top) [n=11M, 11F]. Tensile elastic modulus and failure stress were calculated from resulting stress-strain curves during ramp loading, and sex- and region-specific differences in baseline LCL complex properties were determined. There were no significant sex-specific differences in LCL complex biomechanical properties, however a significant region-specific effect was determined between anterior and posterior regions of the human LCL complex. The anterior region was significantly stiffer and failed earlier than the corresponding posterior region (p=0.044) within the multivariate model (bottom). Tensile moduli at 10% strain were smaller than those at 20% strain (p<0.001) and strain-to-failure (p<0.001), with no significant difference between 20% and strain-to-failure (p=0.140). The anterior region was stiffer than the posterior region at 20% strain and strain-to-failure with no significant differences at 10% strain following Bonferroni correction. Box plots show median and interquartile range of tensile modulus and failure stress to display data variability. Significant differences between regions or strain conditions are indicated above plots (p<0.05).

Figure 4.

Biochemical assays determining sex and region-specific differences in LCL complex composition were performed for water content (left), total collagen content (second from left), hydroxyproline content (second from right), and total sGAG content (right) [n=11M, 11F]. There was no significant sex- or region-specific differences in water content or sGAG content. Significant effects of sex (p=0.006) and region (p=0.045) were observed in total collagen content and hydroxyproline content. Box plots show median and interquartile range of water, collagen, hydroxyproline, and sGAG content to display data variability. Significant regional and sex differences are indicated above plots (p<0.05).