Electrical Conductivity Method to Determine Sexual Dimorphisms in Human Temporomandibular Disc Fixed Charge Density.

To investigate potential mechanisms associated with the increased prevalence of temporomandibular joint (TMJ) disorders among women, the study objective was to determine sex-dependent and region-dependent differences in fixed charge density (FCD) using an electrical conductivity method. Seventeen TMJ discs were harvested from nine males (77 ± 4 years) and eight females (86 ± 4 years). Specimens were prepared from the anterior band, posterior band, intermediate zone, medial disc and lateral disc. FCD was determined using an electrical conductivity method, assessing differences among disc regions and between sexes. Statistical modeling showed significant effects for donor sex (p = 0.002), with cross-region FCD for male discs 0.051 ± 0.018 milliequivalent moles per gram (mEq/g) wet tissue and 0.043 ± 0.020 mEq/g wet tissue for female discs. FCD was significantly higher for male discs compared to female discs in the posterior band, with FCD 0.063 ± 0.015 mEq/g wet tissue for male discs and 0.032 ± 0.020 mEq/g wet tissue for female discs (p = 0.050). These results indicate FCD contributes approximately 20% towards TMJ disc compressive modulus, through osmotic swelling pressure regulation. Additionally, FCD regulates critical extracellular ionic/osmotic and nutrient environments. Sexual dimorphisms in TMJ disc FCD, and resulting differences in extracellular ionic/osmotic and nutrient environments, could result in altered mechano-electro-chemical environments between males and females and requires further study.

Locator retention: does variation in number and configuration matter?

This pilot study was designed to collect initial data on overdenture attachment retention in varying configurations of attachment location in an implant-retained mandibular overdenture. A clear acrylic model of a mandible with 6 numbered implants and Locator resilient abutments was used to simulate implant placement in a patient. A clear acrylic denture was fabricated with 6 Locator housings to match the implants in the model. Attachments were tested in 4 different configurations: 2 implants, 2 and 5 (T25); 4 implants, 2-5 (T2345); 4 implants, 1, 3, 4, and 6 (T1346); and 6 implants, 1-6 (T1-6). Clear nylon male inserts were used for each test. The mean overall retentive strength across all 20 pulls was 576.0 N for configuration T1-6, 354.9 N for configuration T1346, 350.7 N for configuration T2345, and 189.9 N for configuration T25. Mean retentive strength also stabilized after the 7th pull for all 4 configurations, resulting in nonsignificant declines in retentive strength within each specific configuration after 7 pulls. Configuration T1-6 exhibited the greatest retentive strength relative to all other configurations both initially and after repeated application of force. Configurations T1346 and T2345 had similar retentive strengths, and both had greater retentive strength than T25. However, despite these differences, all 4 configurations exhibited similar losses in retentive strength from the repeated application of force during the first 7 pulls until stabilization occurred shortly thereafter.

Tensile biomechanical properties of human temporomandibular joint disc: Effects of direction, region and sex.

Approximately 30% of temporomandibular joint (TMJ) disorders include degenerative changes to the articular disc, with sex-specific differences in prevalence and severity. Limited tensile biomechanical properties of human TMJ discs have been reported. Stress relaxation tests were conducted on TMJ disc specimens harvested bilaterally from six males and six females (68.9±7.9 years), with step-strain increments of 5%, 10%, 15%, 20% and 30%, at 1% strain-per-second. Stress versus strain plots were constructed, and Young׳s Modulus, Instantaneous Modulus and Relaxed Modulus were determined. The effects of direction, region, and sex were examined. Regional effects were significant (p<0.01) for Young׳s Modulus and Instantaneous Modulus. Anteroposteriorly, the central region was significantly stiffer than medial and lateral regions. Mediolaterally, the posterior region was significantly stiffer than central and anterior regions. In the central region, anteroposteriorly directed specimens were significantly stiffer compared to mediolateral specimens (p<0.04). TMJ disc stiffness, indicated by Young׳s Modulus and Instantaneous Modulus, was higher in directions corresponding to high fiber alignment. Additionally, human TMJ discs were stiffer for females compared to males, with higher Young׳s Modulus and Instantaneous Modulus, and female TMJ discs relaxed less. However, sex effects were not statistically significant. Using second-harmonic generation microscopy, regional collagen fiber organization was identified as a potentially significant factor in determining the biomechanical properties for any combination of direction and region. These findings establish structure-function relationships between collagen fiber direction and organization with biomechanical response to tensile loading, and may provide insights into the prevalence of TMJ disorders among women.

Comparison and evaluation of biomechanical, electrical, and biological methods for assessment of damage to tissue collagen.

In regard to evaluating tissue banking methods used to preserve or otherwise treat (process) soft allograft tissue, current tests may not be sufficiently sensitive to detect potential damage inflicted before, during, and after processing. Using controlled parameters, we aim to examine the sensitivity of specific biomechanical, electrical, and biological tests in detecting mild damage to collagen. Fresh porcine pulmonary heart valves were treated with an enzyme, collagenase, and incubated using various times. Controls received no incubation. All valves were cryopreserved and stored at -135 °C until being rewarmed for evaluation using biomechanical, permeability, and cell viability tests. Statistically significant time dependent changes in leaflet ultimate stress, (p = 0.006), permeability (p = 0.01), and viability (p ≤ 0.02, four different days of culture) were found between heart valves subjected to 0-15 min of collagenase treatment (ANOVA). However, no statistical significance was found between the tensile modulus of treated and untreated valves (p = 0.07). Furthermore, the trends of decreasing and increasing ultimate stress and viability, respectively, were somewhat inconsistent across treatment times. These results suggest that permeability tests may offer a sensitive, quantitative assay to complement traditional biomechanical and viability tests in evaluating processing methods used for soft tissue allografts, or when making changes to current validated methods. Multiple test evaluation may also offer insight into the mechanism of potential tissue damage such as, as is the case here, reduced collagen content and increased tissue porosity.

Decreased Mechanical Strength and Collagen Content in SPARC-Null Periodontal Ligament Is Reversed by Inhibition of Transglutaminase Activity.

The periodontal ligament (PDL) is a critical tissue that provides a physical link between the mineralized outer layer of the tooth and the alveolar bone. The PDL is composed primarily of nonmineralized fibrillar collagens. Expression of secreted protein acidic and rich in cysteine (SPARC/osteonectin), a collagen-binding matricellular protein, has been shown to be essential for collagen homeostasis in PDL. In the absence of SPARC, PDL collagen fibers are smaller and less dense than fibers that constitute WT PDL. The aim of this study was to identify cellular mechanisms by which SPARC affected collagen fiber assembly and morphology in PDL. Cross-linking of fibrillar collagens is one parameter that is known to affect insoluble collagen incorporation and fiber morphology. Herein, the reduction in collagen fiber size and quantity in the absence of SPARC expression was shown to result in a PDL with reduced molar extraction force in comparison to that of WT mice (C57Bl/6J). Furthermore, an increase in transglutaminase activity was found in SPARC-null PDL by biochemical analyses that was supported by immunohistochemical results. Specifically, collagen I was identified as a substrate for transglutaminase in PDL and transglutaminase activity on collagen I was found to be greater in SPARC-null tissues in comparison to WT. Strikingly, inhibition of transglutaminase activity in SPARC-null PDL resulted in increases in both collagen fiber thickness and in collagen content, whereas transglutaminase inhibitors injected into WT mice resulted in increases in collagen fiber thickness only. Furthermore, PDL treated with transglutaminase inhibitors exhibited increases in molar extraction force in WT and in SPARC-null mice. Thus, SPARC is proposed to act as a critical regulator of transglutaminase activity on collagen I with implications for mechanical strength of tissues.

Impact of storage solution formulation during refrigerated storage upon chondrocyte viability and cartilage matrix.

Various preservation solutions have been evaluated for longer hypothermic cartilage storage for tissue transplantation; however, the results are mixed. This research was carried out to determine whether phosphate-buffered saline (PBS) or organ preservation solutions would preserve both the extracellular matrix and chondrocytes of articular cartilage better than culture medium during refrigerated storage in the time frame that cartilage is stored for clinical use. Porcine cartilage plugs were stored, without the underlying bone, in culture medium with and without fetal bovine serum (FBS), PBS, Belzer's and Unisol solutions for 1 month at 4°C. Metabolic activity was tested using a resazurin reduction method, and matrix permeability was evaluated by measuring electrical conductivity. Storage in culture medium with 10% FBS was shown to provide good cartilage metabolic function for 7 days, decreasing to about 36% after 1 month of storage. There was no significant difference between samples stored in culture medium with and without FBS after 1 month of storage (p = 0.5005). Refrigerated storage of cartilage in PBS and two different solutions (Belzer's and Unisol) designed for optimal refrigerated tissue and organ storage results in loss of chondrocyte function and retention of matrix permeability. In contrast, the opposite, namely significantly better retention of chondrocyte function and loss of matrix permeability, was observed with culture medium. Future research should be focused on combining retention of chondrocyte function and matrix permeability by storage solution formulation.

Effect of mechanical strain on solute diffusion in human TMJ discs: an electrical conductivity study.

This study investigated the effect of mechanical strain on solute diffusion in human TMJ discs (mean cadaver age 77.8) using the electrical conductivity method. The electrical conductivity, as well as small ion diffusivity, of male and female TMJ discs was determined under three compressive strains. In the male group, the average disc electrical conductivity (mean ± SD) at 0% strain was 5.14 ± 0.97 mS/cm, decreased to 4.50 ± 0.91 mS/cm (-12.3%) at 10% strain, and 3.93 ± 0.81 mS/cm (-23.5%) at 20% compressive strain. Correspondingly, the average disc relative ion diffusivity at 0% strain was 0.44 ± 0.08, decreased to 0.40 ± 0.08 (-8.9%) at 10% strain, and 0.36 ± 0.08 (-16.7%) at 20% compressive strain. In the female group, the average disc electrical conductivity at 0% strain was 5.84 ± 0.59 mS/cm, decreased to 5.01 ± 0.50 mS/cm (-14.2%) at 10% strain, and 4.33 ± 0.46 mS/cm (-25.8%) at 20% compressive strain. Correspondingly, the average disc relative ion diffusivity at 0% strain was 0.49 ± 0.05, decreased to 0.43 ± 0.04 (-11.3%) at 10% strain, and 0.39 ± 0.04 (-19.9%) at 20% compressive strain. The results indicated that mechanical strain significantly impeded solute diffusion through the disc. This mechanical strain effect was larger in the female than in the male human TMJ disc. This study may provide new insights into TMJ pathophysiology.

Allogeneic heart valve storage above the glass transition at -80°C.

BACKGROUND: Cryopreserved allogeneic heart valves are usually stored and transported below -135°C; however, such methods require expensive equipment for both storage and transportation. METHODS: In this study, vitrified porcine aortic valves were stored on either side of the cryoprotectant formulation's glass transition temperature (-119°C) at -80°C and -135°C, using a newly formulated vitrification solution (VS83) consisting of a combination of 4.65M dimethyl sulfoxide, 4.65M formamide, and 3.30M 1,2-propanediol. Three groups of valves were studied: (1) fresh; (2) VS83-preserved, stored at -80°C; and (3) VS83-preserved, stored at -135°C. RESULTS: Using the VS83 cryoprotectant concentration formulation, cracking was not observed during valve storage. No ice-related events were detectable during 5°C rewarming by differential scanning calorimetry. All cryopreserved tissue samples demonstrated significantly less viability than fresh samples (p<0.01). No significant viability differences were observed between the VS83-preserved groups stored at -80°C and -135°C. Material testing did not reveal any significant differences among the three test groups. Multiphoton imaging of VS83-preserved heart valves stored at -80°C and -135°C demonstrated similar collagen and elastin structures. CONCLUSIONS: These results indicate that VS83-preserved heart valves can be stored and transported at temperatures in the vicinity of -80°C with retention of extracellular matrix integrity and material properties. The VS83 preservation of heart valves at -80°C without the need for liquid nitrogen should result in both decreased manufacturing costs and reduced employee safety hazards. Moreover, it is anticipated that low cell viability may result in less immunogenicity in vivo.

Impact of Hypothermia upon Chondrocyte Viability and Cartilage Matrix Permeability after 1 Month of Refrigerated Storage.

BACKGROUND: The purpose of this research was to assess the extracellular matrix and chondrocytes of articular cartilage during refrigerated storage and to determine whether changes could be detected in the time frame that cartilage is stored for clinical use. MATHODS: Porcine cartilage was stored as either bisected femoral heads with bone attached or plugs without the underlying bone in culture medium with fetal bovine serum for 1 month at 4 °C. Metabolic activity was tested using a resazurin reduction method on intact tissue and viable cell recovery after enzymatic tissue digestion at each time point. Cartilage plug permeability was evaluated by measuring electrical conductivity. RESULTS: Storage in culture medium provided good cartilage viability and metabolic function for 7 days; however, significant changes were observed in femoral heads (p < 0.05). All mean chondrocyte assessment values were <30% of fresh controls at 28 days. Cartilage plugs tended to perform better after 7 days of storage than the femoral heads and retained significantly higher metabolic activity (mean = 94.5% vs. 70.5%; p < 0.05). Cartilage plugs demonstrated consistent changes in electrical conductivity after 28 days of storage (p < 0.05). CONCLUSION: Refrigerated storage of cartilage results in both loss of chondrocyte viability and matrix permeability.