Biological Treatments for Temporomandibular Joint Disc Disorders: Strategies in Tissue Engineering.

The temporomandibular joint (TMJ) is an important structure for the masticatory system and the pathologies associated with it affect a large part of the population and impair people's lifestyle. It comprises an articular disc, that presents low regeneration capacities and the existing clinical options for repairing it are not effective. This way, it is imperative to achieve a permanent solution to guarantee a good quality of life for people who suffer from these pathologies. Complete knowledge of the unique characteristics of the disc will make it easier to achieve a successful tissue engineering (TE) construct. Thus, the search for an effective, safe and lasting solution has already started, including materials that replace the disc, is currently growing. The search for a solution based on TE approaches, which involve regenerating the disc. The present work revises the TMJ disc characteristics and its associated diseases. The different materials used for a total disc replacement are presented, highlighting the TE area. A special focus on future trends in the field and part of the solution for the TMJ problems described in this review will involve the development of a promising engineered disc approach through the use of decellularized extracellular matrices.

[Effects of staurosporine on the contraction of self-assembled constructs of goat temporomandibular joint disc cells].

OBJECTIVE: The effects of the staurosporine on contraction of self-assembled constructs and extracellular matrix syntheses of goat temporomandibular joint discs were investigated. METHODS: Goat temporomandibular joint disc cells were isolated and cultured to P3, and 5.5×106 cells were combined with different concentrations of staurosporine (0, 0.1, 1, 10, 100 nmol·L⁻¹) in agarose wells and cultured for one week. The samples were frozen and sectioned. Safranin-O,  Picro-sirius red and immunohistochemical staining were performed to observe the distributions of the extracellular matrix and the expression of alpha-smooth muscle actin (α-SMA). Enzyme linked immunosorbent assay (ELISA) and Blyscan kits were utilized to quan--titatively detect the contents of type Ⅰ collagen (ColⅠ) and glycosaminoglycans (GAGs). RESULTS: Each group of goat temporo-mandibular joint disc cells in the agarose wells were gathered to self-assemble into a disc-shaped base for 4 hours and then to gradually contract into a round shape. The Picro-sirius red staining was strong and indicated collagen distribution. The Safranin-O staining observed GAGs throughout the entire construct. The expression of ColⅠ was strongly posi-tive in the staurosporine groups; however, the expression of α-SMA was weak. ColⅠ and GAGs contents in the stau-rosporine groups were greater than that of the control group, especially in the 10 nmol·L⁻¹ group (P<0.01). CONCLUSIONS: Staurosporine has a certain effect on the shrinkage of self-assembled constructs; however, such effect is not prominent. Staurosporine contributes to the construction synthesis of extracellular matrix.

Evaluation of articular disc loading in the temporomandibular joints after prosthetic and pharmacological treatment in model studies.

BACKGROUND: Temporomandibular joint dysfunction is often related to excessive load in the stomatognathic system. OBJECTIVES: The objective of the model tests, using numeric calculations, was to assess the articular disc loads in the temporomandibular joints after prosthetic and pharmacological treatment of functional disorders of the masticatory organ. MATERIAL AND METHODS: The study involved 10 patients, aged 21-48 years, of both sexes, randomly selected from a group of 120 patients treated with relaxation occlusal splints (60 patients, group I) and intramuscular injection of botulinum toxin type A (60 patients, group II), suffering from temporomandibular joint dysfunction with the dominant muscle component. In all subjects, a specialized functional examination was carried out. Treatment groups: occlusal splint therapy (group I) and intramuscular injection of botulinum toxin type A (group II). An assessment of the loads of 4 disc zones of the temporomandibular joints was carried out based on the results of clinical studies (phase I of the study), and numeric model tests (phase II). In the representatives of the study groups (5 patients in each group), measurements of occlusal forces and an evaluation of tension of the masseter and temporalis muscle were performed. RESULTS: The results of the average load values for all evaluated zones of the right and left articular disc differ in a statistically significant way in favor of group II, with the exception of the external mid part of the discs. In the case of the anterior of the right disc, the load was lower in patients belonging to group I than in those obtained in group II. CONCLUSIONS: Botulinum toxin type A significantly reduces the loads within the temporomandibular joints, generated by masseter muscle hypertonia.

[Chondroprotectors in complex treatment of internal TMJ disorders].

The article presents a detailed analysis of the clinical studies on the use of chondroprotector (Alflutop) in patients with persistent displacement of the articular disk of the temporomandibular joint in order to prevent the development of secondary osteoarthrosis. The study showed that the inclusion in the therapy chondroprotective action agents allows achieving short-term positive outcomes in most cases, relief of inflammation and pain, as well as improving joint function and stabilization of the thickness of the articular cartilage that provides cushioning function.

Outcomes of arthrocentesis for the treatment of internal derangement pain: with or without corticosteroids?

PURPOSE: Arthrocentesis (AC) is an acceptable treatment modality in the treatment of internal derangement (ID) pain. This study evaluated outcomes of AC on pain relief of ID pain with or without corticosteroids. MATERIALS AND METHODS: This single-blind clinical trial was conducted on 60 patients with ID pain who were randomly assigned to 2 groups (30 in each group) for AC. The AC of the upper joint space was then performed by using Ringer lactate under local anesthesia in both groups. In the second group, the procedure was followed by the administration of a single-dose intra-articular dexamethasone (8 mg). Assessments were made at baseline (T0), 1 month (T1), and 6 months (T6) after AC; pain, maximum mouth opening, and joint sounds before and after treatment up to 6 months were evaluated. Pain severity was documented according to Visual Analogue Scale. Age, sex, and skeletal maxillomandibular relationship were considered as variable factors, and irrigation with or without corticosteroids was a predictive factor of the study. Pain, click, and maximum mouth opening were other assessment outcomes of the study. RESULTS: Comparison of age, sex, and skeletal relationship did not show any significant differences between the 2 groups. Results did not demonstrate any difference for click between the 2 groups. Comparison of pain severity in T0, T1, and T2 between the 2 groups did not show any significant differences (P < 0.05). The repeat measure test revealed a significant change in T0, T1, and T2 for both groups (P < 0.001). MMO significantly changed between T0 and T1 and T0 and T6 in the 2 groups without any significant differences between them. CONCLUSIONS: The AC is an effective procedure for a short-term reduction of pain in temporomandibular disorder cases. It seems that AC using Ringer solution with or without corticosteroids may have the same effect on pain relief.

Temporomandibular joint orthopedics with anterior repositioning appliance therapy and therapeutic injections.

TMD orthopedics is the assessment, diagnosis and management of orthopedic disorders of the temporomandibular joint (TMJ). Anterior repositioning appliance (ARA) therapy for TMJ internal derangements is successful in long-term recapturing of disks in reducing and nonreducing joints at a rate of 64 percent and in regenerating degenerated condyles in some cases. ARA therapy for TMJ internal derangements is subjectively successful in relieving symptoms in reducing and nonreducing disk displacement TMJs in this study at an average rate of 94.5 percent.

A comparative study on the impact of intra-articular injections of hyaluronic acid, tenoxicam and betametazon on the relief of temporomandibular joint disorder complaints.

OBJECTIVE: The aim of this study was to compare the efficacy of intra-articular injections of three different agents with well known anti-inflammatory properties. MATERIALS AND METHOD: Between April 2010 and January 2013 a total of 100 patients who were diagnosed as temporomandibular joint disorder in the Department of Otolaryngology at Bozok University School of Medicine were prospectively studied. Patients with symptoms of jaw pain, limited or painful jaw movement, clicking or grating within the joint, were evaluated with temporomandibular CT to investigate the presence of cartilage or capsule degeneration. In the study group there were 55 female and 45 male patients who were non-responders to conventional anti-inflammatory treatment for TMJ complaints. The patients were randomly divided into four groups consisting of a control group and three different groups who underwent intra-articular injection of one given anti-inflammatory agent for each group. We injected saline solution to intra-articular space in the control group. Of three anti-inflammatory agents including hyaluronic acid (HA, Hyalgan intra-articular injection, Sodium hyaluronate 10 mg/ml, 2 ml injection syringe, Bilim Pharmaceutical Company, Istanbul, Turkey); betamethasone (CS, Diprospan flacon, 7.0 mg betamethasone/1 ml, Schering-Plough Pharmaceutical Company, Istanbul, Turkey) and; tenoxicam (TX, Tilcotil flacon, 20 mg tenoxicam/ml, Roche Pharmaceutical Company, Istanbul, Turkey) were administered intra-articularly under, ultrasonographic guidance. Following the completion of injections the, changes in subjective symptoms were compared with visual analogue scales, (VAS) scores at 1st and 6th weeks' follow-up visits between four groups. RESULTS: The HA group did significantly better pain relief scores compared to the, other groups at 1st and 6th weeks (p < 0.05). TX and CS groups' pain scores were better than control group values (p < 0.05, for both agents). The pain relief effect of TX was noted to decrease significantly between the 1st and 6th week (p < 0.05) (Fig. 1). We did not observe the same pattern in HA, CS and control (saline) groups between 1st and 6th week (p > 0.05). CONCLUSION: We found that HA produced better pain relief scores when compared to the other anti-inflammatory agents studied. The main disadvantage of HA is its relatively higher cost. Additionally it does not have a reimbursement status by state or private health insurance systems in Turkey. Despite the lower VAS scores, intra-articular TX and CS may be assessed as more economic alternatives to intra-articular HA injections.

Transcriptional regulation of proteoglycan 4 by 17ß-estradiol in immortalized baboon temporomandibular joint disc cells.

Temporomandibular joint disorders (TMDs) affect a significant portion of the population of the USA, with the majority of those seeking treatment being women of childbearing age. Owing to this striking sexual dimorphism it has been postulated that sex hormones play a role in the maintenance of normal temporomandibular joint (TMJ) function. Proteoglycan 4 (PRG4) is a secreted lubricating molecule required for maintaining low frictional levels within articular joints; however, its role in the TMJ is not well characterized. In this study we describe the development of immortalized baboon cells isolated from specific regions of the TMJ disc and their use in the investigation of PRG4 expression and localization patterns in the TMJ. We identified conserved estrogen response elements within the 5' flanking region of the PRG4 gene of several species, and found that treatment of baboon TMJ disc cells with estrogen led to reduced PRG4 promoter activity and reduced expression of PRG4 mRNA in vitro. The observed negative regulation of PRG4 by estrogen could lead to increased friction and degradation of joint components over time. This study, for the first time, provides evidence of the regulatory potential of estrogen on PRG4 gene expression and suggests a novel etiology for the gender disparity observed among TMD patients.

Effect of simvastatin injections on temporomandibular joint inflammation in growing rats.

PURPOSE: Juvenile idiopathic arthritis often affects the temporomandibular joint (TMJ), resulting in facial deformities, and intra-articular injections of anti-inflammatory steroids used in treatment may inhibit bone growth in the developing condyle. The purpose of this pilot study was to evaluate the anti-inflammatory properties of simvastatin (SIM), a bone anabolic drug, compared with the common steroid triamcinolone hexacetonide (TH) in experimental TMJ arthritis of growing rats. METHODS: Joint inflammation was induced by injecting complete Freund's adjuvant (CFA) into the TMJs of 32 growing (4-week-old) Sprague-Dawley rats while simultaneously receiving 1) ethanol drug carrier, 2) 0.1 mg of SIM, 3) 0.5 mg of SIM, or 4) 0.15 mg of TH. Six rats had no treatment to the TMJ. Animals were euthanized 28 days later, and TMJs were decalcified and stained with hematoxylin-eosin. RESULTS: Histopathologic TMJ results showed that CFA injection along with drug carrier induced increased thickness of the articular layer on the head of the condyle and inflammation of the retrodiscal area (CFA and ethanol). Although both TH and SIM reduced the articular layer thickness, 0.5 mg of SIM was more effective at reducing subsynovial inflammation. CONCLUSIONS: Intra-articular simvastatin showed anti-inflammatory properties in this TMJ model, prompting its further study in the growing TMJ, where bone anabolic properties would be important.

Poly (glycerol sebacate): a novel scaffold material for temporomandibular joint disc engineering.

The preponderance of temporomandibular joint (TMJ) disorders involving TMJ disc injury inspires the need to further explore tissue engineering strategies. The objective of this study was to examine the potential of poly (glycerol sebacate) (PGS), a biocompatible, biodegradable elastomer, as a porous scaffold material for the TMJ disc. Goat fibrochondrocytes were seeded on PGS at three seeding densities (25, 50, 100 million cells/mL scaffold), respectively, and cultured for 24 h, 2 weeks, and 4 weeks. The resulting histological, biochemical, and biomechanical properties were determined. Histological staining revealed an abundance of both collagen and glycosaminoglycans (GAG) throughout the high seeding density scaffolds at 4 weeks. There was also a significant increase in the cellular content in all groups over the four-week period, showing that the scaffolds promoted cell attachment and proliferation. The PGS scaffolds supported the deposition of large quantities of extracellular matrix, with differences noted between seeding density groups. At 4 weeks, the medium and high seeding density groups had significantly more collagen per scaffold (181±46 µg and 218±24 µg, respectively) than the low seeding density group (105±28 µg) (p<0.001). At 4 weeks, the medium and high seeding density groups also had a significantly higher GAG content per scaffold (702±253 µg and 773±187 µg, respectively), than the low seeding density group (324±73 µg) (p<0.001). The compression tangent modulus was significantly greater at 4 weeks than 24 h (123.6±86 kPa and 26.2±5 kPa, respectively) (seeding density groups combined) (p<0.001), with no differences between seeding groups at each time point. After 4 weeks, the tangent modulus of the low seeding density group was in a similar range of the goat TMJ disc (180±127 kPa compared to 304±141 kPa, respectively). The results show that cell seeding density and culture time do have an effect on both the biochemical and biomechanical properties of PGS scaffolds. These findings demonstrate that PGS has great potential as a scaffold material for TMJ disc engineering.

Protection of articular cartilage by intra-articular injection of NEL-like molecule 1 in temporomandibular joint osteoarthritis.

Several studies have proven the ability of NEL-like molecule-1 (Nell-1) to induce chondrogenesis and make it as a potential candidate for articular cartilage repair. In the current study, the chondroprotective effect of Nell-1 on osteoarthritis (OA) of the temporomandibular joint (TMJ) was investigated by intra-articular injection. Bilateral partial discectomy was performed in 24 rabbits to induce TMJOA. Four weeks later, the right TMJ was treated with Nell-1 as the experimental group, whereas the left was treated with physiologic saline as the control group. Twelve rabbits each time were randomly killed at 12 and 24 weeks after injection, respectively. Histologic observation and metabolic analysis by reverse transcription-polymerase chain reaction were used for evaluation. All TMJs appeared as OA-like histologic changes after intra-articular injection. However, the degree of osteoarthritis in the experimental group was less severe than that in the control group during the experimental time. The expression of type II collagen and aggrecan messenger RNA was significantly higher than the control group at 12 weeks after injection. However, no difference in the expression of aggrecanase or interleukin 1 messenger RNA was observed. The results suggest that intra-articular injection of Nell-1 may be a good alternative for the treatment of cartilage degeneration in OA.

Intra-articular corticosteroid injections to the temporomandibular joints are safe and appear to be effective therapy in children with juvenile idiopathic arthritis.

PURPOSE: The purpose of this study was to evaluate the safety and efficacy of intra-articular corticosteroid injections (IACIs) of the temporomandibular joint (TMJ) in children with juvenile idiopathic arthritis (JIA) when administered by an oral and maxillofacial surgeon without imaging guidance. MATERIALS AND METHODS: This was a retrospective study of children with JIA, seen at a single center, who were selected based on having received IACIs of the TMJ. All subjects received the intervention, which consisted of referral to a single oral and maxillofacial surgeon for TMJ IACI with 5 to 10 mg triamcinolone hexacetonide, under general anesthesia. Primary outcomes assessed in all subjects were the safety of the procedure and efficacy as determined by the change in maximal incisal opening (MIO). In addition, a subset of 31 subjects underwent repeat magnetic resonance imaging of the TMJ, permitting analysis of the change in the acute and chronic findings of arthritis in those patients. RESULTS: Sixty-three patients (68% female) received 137 IACIs. The mean age for diagnosis of JIA was 8.5 years, and the mean age at presentation for TMJ injections was 10 years. The injections were well tolerated: only 1 patient developed the steroid complication of hypopigmentation, and none developed degeneration or ankylosis. In terms of efficacy, the mean MIO increased from 40.8 ± 0.93 to 43.5 ± 0.90 mm (P = .001); in addition, changing the unit of analysis to individual joints, in patients who underwent repeat magnetic resonance imaging examination, 51% of TMJs showed magnetic resonance imaging evidence of improvement of arthritic changes, of whom 18% had complete resolution of TMJ arthritis. CONCLUSIONS: The results indicate that IACI of the TMJ can be safely performed by experienced oral and maxillofacial surgeons without a requirement for computed tomographic guidance. In addition, these results show that IACI may be effective in the management of TMJ arthritis, although further studies are required.

Repeated sodium hyaluronate injections following multiple arthrocenteses in the treatment of early stage reducing disc displacement of the temporomandibular joint: a preliminary report.

PURPOSE: The aim of this study was to assess the safety and clinical utility of intraarticular injection of sodium hyaluronate for the treatment of symptoms associated with internal derangement of the temporomandibular joint (TMJ). PATIENTS AND METHODS: Twenty patients, who have early stage temporomandibular dysfunction (TMD), were treated with intraarticular sodium hyaluronate injection and arthrocentesis. The patients received sodium hyaluronate (15 mg/ml) (1 ml) injections two times a week first following arthrocentesis with 200 ml Ringer's Lactate and the second without. The procedure was repeated for three times at weekly intervals for 3 weeks. Pre- and postinjection pain intensity, the presence of joint sounds, and interincisial distance were recorded. These data were evaluated by visual analog scale (VAS). RESULTS: The follow-up period was 6 months. There was a statistically significant reduction of pain intensity (p < 0.001) and joint sound (p < 0.001) in all patients. Initial measurement of maximal mouth opening (MMO) was 33.40 ± 3.75 mm. At the end of the follow-up period, the same measurements were repeated and interincisal distance was 49.3 ± 3.74 mm. The difference between these measurements was statistically significant (p < 0.001). CONCLUSION: We suggest that repeated sodium hyaluronate injections following multiple arthrocenteses with Ringer's Lactate is an effective and safe method for the treatment of early stage reducing disc displacement of TMJ.

Preliminary evaluation of histological changes found in a mechanical arthropatic temporomandibular joint (TMJ) exposed to an intra-articular Hyaluronic acid (HA) injection, in a rat model.

OBJECTIVE: This study investigates the histological effects of Hyaluronic acid injections in the treatment of induced temporomandibular joint (TMJ) osteoarthritis in rats. STUDY DESIGN: Twenty-four male Wister rats were subjected to induced mechanical osteoarthritis by manual hypermobility for 10 successive days. Animals were then divided into two groups; group I (control) and group II (experimental). Ten days after the induction of hypermobility, the right TMJ of the experimental animals was injected with a dose of 0.12 mg HA intra-articularly and 0.12 mg saline was injected into the left joint; while animals in the control group were left without any treatment. Two rats from group I were killed at one, two and six weeks; while 6 animals from group II were killed at one, two and four weeks post injection. RESULTS: The disk of the right joints in the experimental animals was of normal thickness and there was an increase in the thickness of the fibrocartilagenous layer. In the left joint; ulcerative changes in the disk were evident where the fibres were not well oriented and scalloped areas in the temporal bone area were present denoting osteoclastic activity. CONCLUSIONS: Repeated intra-articular TMJ injection of Hyaluronic acid appears to be a safe and effective way of inhibiting the progression of osteoarthritic changes in the joint through development of articular cartilage and reducing fibrous tissue proliferation.

Protocol on induction of TMJ articular disc degeneration in rats by utilization of botulinum toxin.

OBJECTIVE: To develop a model of experimental degeneration in the articular disc of the TMJ of rats through the use of botulinum toxin that can be used in future studies of degenerative diseases on fibrocartilage. METHODS: Aiming at the above-mentioned objective, 12 Lewis male rats were used and divided into two groups: CG, control group and DG, group of animals to which the botulinum toxin was administered (6 units/kg). The morphological analysis was carried out utilizing histological cuts stained with hematoxyline-eosine, toluidine blue and Picrosirius; the biochemical analysis was made by SDS-polyacrylamide gel electrophoresis. RESULTS: The DG showed peculiar characteristics regarding a degeneration joint disk, compatible with those described in literature as: reduction of cells number, general disorganization of cells direction and extracellular matrix, increase in glycosaminoglycans content and degradation of the tissue collagen. CONCLUSIONS: Based on the morphological and biochemical results, it was concluded that the proposed degeneration model showed to be satisfactory for futures studies of injuries and fibrocartilage regeneration processes.

Sodium hyaluronate improves outcomes after arthroscopic lysis and lavage in patients with Wilkes stage III and IV disease.

PURPOSE: Among patients with Wilkes stage III and IV disease undergoing arthroscopic lysis and lavage, does the use of an intra-articular injection of sodium hyaluronate (SH), when compared with Ringer lavage, result in better postoperative pain control and temporomandibular joint (TMJ) function? PATIENTS AND METHODS: We designed and implemented a randomized, double-blind, pilot controlled clinical trial. The study sample was composed of patients with middle Wilkes stage (late stage III and early stage IV) disease. Subjects were randomized to 1 of 2 treatment limbs. The treatment group received Ringer lactate plus an injection of 1 mL of SH after arthroscopy, whereas the control group was given Ringer lactate during arthroscopy. The primary outcome variables were pain and TMJ function measured by use of visual analog scales. Appropriate descriptive and bivariate statistics were computed. A P value less than .05 was considered statistically significant. RESULTS: The study sample was composed of 40 patients with 20 subjects enrolled in both treatment groups. There were no statistically significant differences between the 2 groups in terms of demographics and preoperative variables. Postoperative analgesia was statistically significant in the treatment group with respect to the control group on the visits on days 14 and 84. No statistically significant differences were observed between the 2 groups in the maximum interincisal opening and tolerance. CONCLUSIONS: An intra-articular injection of SH after arthroscopic lysis and lavage is effective in reducing pain in patients with TMJ dysfunction, enhancing postsurgical recovery. The analgesic effect of treatment with SH is maintained in the long term.

Effect of botulinum toxin-A in myofascial pain patients with or without functional disc displacement.

PURPOSE: To evaluate the effects of botulinum toxin-A in the treatment of patients who have myofascial pain with or without functional disc displacement. PATIENTS AND METHODS: Twenty-four participants were randomly assigned to the study by using Research Diagnostic Criteria for Temporomandibular Disorders. All patients were informed about botulinum toxin-A, and were required to give informed consent. Before the injections, patients were asked to fill out a Biobehavioral Questionnaire to evaluate their pain and psychological status, and afterward, electromyography of the right and left masseter and anterior temporal muscles was recorded. Saline was injected into the masseter and anterior temporal muscles in the placebo group, and botulinum toxin-A was used in the study group. On days 14 and 28, patients were asked to fill out a Biobehavioral Questionnaire again, and electromyography of the right and left masseter and anterior temporal muscles was recorded again. RESULTS: The study group showed improvement in pain and psychological status. Although a decrease in the action potentials of the masseter muscles on day 14 was followed by an increase on day 28, the reduction of pain scores and improvement in psychological status continued on day 28. CONCLUSIONS: The injection of botulinum toxin-A decreases the muscle action potential in 14 days. The patients also show improvement in pain and psychological status.

Scaffold and growth factor selection in temporomandibular joint disc engineering.

Temporomandibular joint disc tissue-engineering studies commonly fail to produce significant matrix before construct contraction. We hypothesized that poly-L-lactic acid (PLLA) non-woven meshes would limit contraction, allow for comprehensive mechanical evaluation, and maintain viability relative to polyglycolic acid (PGA) non-woven mesh controls. Additionally, we proposed that growth factor stimulation, while limiting contraction, would increase construct properties relative to previous reports. After 4 wks, cell proliferation and matrix deposition were similar between the two meshes, but PGA constructs had contracted significantly. Furthermore, only PLLA constructs could be tested in tension and compression. Additional PLLA constructs were formed, then treated with insulin-like growth factor-1 (10 ng/mL), transforming growth factor-beta 1 (5 ng/mL), or transforming growth factor-beta 3 (5 ng/mL). Transforming growth factor-beta 1 yielded the most cells, collagen, and glycosaminoglycans at 6 wks; these constructs also demonstrated improved mechanics. Analysis of these data demonstrated significant temporomandibular joint disc-engineering potential for PLLA and transforming growth factor-beta 1.

The effects of protein-coated surfaces on passaged porcine TMJ disc cells.

OBJECTIVE: Implantation of synthetic temporomandibular joint (TMJ) disc replacements aimed to alleviate pain and restore functional losses caused by TMJ disorders. Unfortunately, these synthetic replacements have been largely unsuccessful and in some instances have incited severe immune responses. Tissue engineering, however, may provide viable TMJ disc replacements. Towards this end, we have studied TMJ disc gene expression as a measure of protein production potential. With passage, collagen type I and aggrecan gene expression decrease in TMJ disc cell cultures. We hypothesize that surfaces coated with TMJ disc proteins may rapidly recover the lost gene expression in passaged TMJ disc cells. DESIGN: To study these effects, passages 0, 1, and 2 TMJ disc cells were plated in wells coated with aggrecan, collagen type I, collagen type II, or decorin. Safranin O staining was conducted to visualize cell aggregation. RESULTS: At passage 0, cultures appeared similar on each surface; however, by passages 1 and 2, aggrecan-coated and decorin-coated surfaces appeared to have more cell aggregates. Gene expression data did not correspond to these visual changes. No treated surface offered a significant change in aggrecan, collagen type I, or decorin expression relative to untreated controls. Furthermore, aggrecan and collagen type I gene expression dropped relative to samples taken prior to plating. CONCLUSIONS: These results indicate that, despite visual changes described by cell aggregates, protein coatings have limited effects for recovering TMJ disc gene expression in monolayer cultures.

Improving culture conditions for temporomandibular joint disc tissue engineering.

BACKGROUND: The temporomandibular joint (TMJ) is extremely important for activities like eating and talking, which can become painful and difficult for patients with TMJ dysfunction. Tissue engineering is a potential alternative to current surgical interventions through replacement of diseased or injured tissue with a functional construct. Since research with TMJ disc cells began relatively recently, optimal culturing conditions must be determined. METHODS: Metabolic additives, L-glutamine, L-alanyl-L-glutamine, sodium pyruvate, and insulin, were examined for their effects on TMJ disc cells in monolayer. Effects of L-proline were examined in three-dimensional (3-D) culture at concentrations of 0, 25 and 100 mg/l. RESULTS: The combination of L-glutamine, sodium pyruvate, and insulin improved cell proliferation rates without affecting collagen production or gene expression. No differences were observed in mechanical properties of the engineered constructs; however, collagen and glycosaminoglycan quantities normalized to cell number decreased at the highest concentration of L-proline. CONCLUSION: This work identified supplements for 2-D monolayer expansion. Other supplements or culture conditions still need to be investigated for 3-D tissue production. This work improves upon porcine TMJ disc cell culturing conditions, taking us closer to being able to engineer the TMJ disc.