Immunohistochemical study of interleukin-1beta and interleukin-1 receptor antagonist in an antigen-induced arthritis of the rabbit temporomandibular joint.

BACKGROUND: In temporomandibular joint (TMJ) arthritis, there is limited knowledge of the relationship between interleukin-1beta (IL-1beta) and interleukin-1 receptor antagonist (IL-1ra), as well as the source of these cytokines. We investigated the development of an antigen-induced arthritis in the rabbit TMJ immunohistochemically. METHODS: Unilateral TMJ arthritis was induced in 32 adult New Zealand White rabbits. From 6 h to 12 weeks after induction of arthritis, topology of IL-1beta and IL-1ra were observed. RESULT: The acute stage of induced arthritis lasted for one week after induction, thereafter it became chronic. In the early phase of the acute stage, infiltrating inflammatory cells, as well as synovial cells, produced IL-1beta and IL-1ra. In the late phase of the acute stage, the main source of these cytokines was subsynovial fibroblasts. In this phase of arthritis, IL-1beta and IL-1ra did not appear to be produced by synovial cells. From the early to intermediate phase of the chronic stage, proliferating synovial cells produced IL-1beta and IL-1ra. In this phase of the arthritis, these cytokines were also observed in a cluster formation in chondrocytes. CONCLUSION: This arthritis model shows a staging of the joint inflammation process with time. IL-1beta and IL-1ra are produced by a certain kind of cells depending on the stage of inflammation.     

Relationship between TNF-α and TUNEL-positive chondrocytes in antigen-induced arthritis of the rabbit temporomandibular joint

BACKGROUND: Terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate biotin nick end-labeling (TUNEL) staining is a widely accepted method for the detection of DNA fragmentation in nuclei of apoptotic cells. Tumor necrosis factor (TNF)-alpha is closely associated with changes in condylar cartilage and modulates apoptosis in various tissues including cartilage. The aim of this study was to investigate the relationship between apoptotic chondrocytes and TNF-alpha in a rabbit model of arthritis. METHOD: Unilateral temporomandibular joint (TMJ) arthritis was induced in 20 adult New Zealand White rabbits. From 1 day to 6 weeks after the induction of arthritis, immunohistochemical analysis for TNF-alpha and TUNEL was performed. RESULTS: In condylar cartilage, TNF-alpha-positive cells and TUNEL-positive cells were localized together. TNF-alpha-positive chondrocytes seemed to precede TUNEL-positive cells. CONCLUSIONS: The results of the present study suggest that TNF-alpha may be involved in apoptosis and/or apoptotic necrosis of chondrocytes as TMJ arthritis progresses from the acute to chronic stage.     

Interleukin-1beta in antigen-induced arthritis of the rabbit temporomandibular joint

The aim was to investigate joint perfusate levels of interleukin-1beta (IL-1beta) in antigen-induced monoarthritis of the rabbit temporomandibular (TMJ) and knee joints. Twenty-four adult male New Zealand White rabbits were divided into three groups: a control group as well as TMJ arthritis and knee joint arthritis groups. After sensitization, unilateral arthritis was induced by intra-articular injection with ovalbumin and the contralateral joint was injected with saline 3 weeks after induction of arthritis. Joints were then perfused continuously with saline and samples were collected at 10-min intervals over a 50-min period. The IL-1beta concentrations in the samples were then analyzed. After killing the animals, the joints were examined histologically. The IL-1beta concentrations in the samples from the arthritic TMJs and knee joints were significantly higher than in the saline-injected and the control joints. Histological signs of chronic arthritis of similar severity were found in both joints. The IL-1beta levels in the samples from the arthritic TM and knee joints correlated with the histological severity of the arthritis, including pannus formation. In conclusion, this study shows that IL-1beta is released in the synovium of rabbit TMJs and knee joints during antigen-induced arthritis, and that high IL-1beta levels in synovial fluid are associated with histological signs of inflammation including, pannus tissue formation.     

间接性颞下颌关节损伤后IL-8的表达及其意义

目的：通过观察羊颞下颌关节（TMJ）间接创伤后不同时期髁突软骨中IL-8的表达，探讨IL-8在TMJ创伤后导致TMJ骨关节病过程中的作用。方法：用自制撞击装置造成山羊双侧颞颌关节间接性创伤，分别于伤后2h,7d,1月，3月取材，并以正常TMJ作为对照，用免疫组化法进行观察。结果：TMJ创伤后髁突软骨中IL-8的表达明显强于正常对照组，其着色程度越靠近结构破坏区越明显。结论：IL-8在TMJ创伤后导致TMJ骨关节病过程中起重要作用。     

Inflammatory cell and cytokine patterns in patients with chronic polyarthritis and temporomandibular joint involvement

OBJECTIVE: To investigate the occurrence of selected markers for inflammatory cells and cytokines in patients with chronic polyarthritis (CPA) and temporomandibular joint (TMJ) involvement. MATERIAL AND METHODS: Eleven patients (11 joints) with CPA and TMJ disorder were included in the study. Synovial specimens were obtained during TMJ open surgery and these were subjected to immunohistochemistry on frozen sections post-fixed with paraformaldehyde and with the cell membranes permeabilized by saponin. In all patients, the cytokines IL-1alpha, IL-1beta, IL-1ra, TNFalpha, IFNgamma, IL2, and TGFbeta were investigated using specific antibodies. The occurrence of macrophages and T-lymphocytes was investigated using immunohistochemistry with monoclonal antibodies against antigens CD68 and CD45RO, respectively. In addition, PCNA was used as a marker for cell proliferation. RESULTS: Staining of IL-1alpha, IL-1, and TGF was seen in all 11 specimens, IFN? in 1, TNFalpha in 4, and IL-2 in none. CD45RO-positive T cells were detected in 7 specimens, CD68-positive macrophages in 6, and cell proliferation seen with PCNA was noted in 8. CONCLUSIONS: The predominant cytokines of TMJ CPA were IL-1alpha, IL-1beta, and TGFbeta, and there appeared to be no differences between the subgroups (rheumatoid arthritis, psoriatic arthritis, and ankylosing spondylitis) involved. Moreover, the cytokine pattern of TMJ CPA patients seemed to differ from patients with osteoarthritis, as shown in our previous study. The main difference was the absence of IFNgamma and TNFalpha in TMJ CPA patients and a stronger TGFbeta and IL-1alpha expression.     

Porphyromonas gingivalis survives within KB cells and modulates inflammatory response

BACKGROUND/AIMS: The purpose of the study was to investigate the intracellular survival of Porphyromonas gingivalis as a possible mechanism for maintaining periodontitis. METHODS: P. gingivalis strains, the strain ATCC 33277 and seven clinical isolates, were co-cultured with KB cells. The number of intracellular bacteria was determined up to 3 days after infection. In addition, the numbers of KB cells per well, the concentrations of the cytokines interleukin-1beta (IL-1beta), IL-6, IL-8 and tumour necrosis factor-alpha (TNF-alpha) and the arginine-specific amidolytic activity were measured. The 16S rRNA of P. gingivalis and the mRNA expression of IL-1beta, IL-6, IL-8, TNF-alpha and rgpA were also determined. RESULTS: All the P. gingivalis strains studied were able to survive within KB cells. In contrast to the reduced values of colony-forming units at day 3, equal and higher levels of 16S rRNA were seen in comparison to day 0. Arginine-specific amidolytic activity declined in all samples during infection. Expression of mRNA for rgpA was not found after infection of KB cells by P. gingivalis strains. IL-8 was detectable in all samples 2 days after infection with P. gingivalis strains. Principal components analysis underlined a correlation between the arginine-specific amidolytic activity 1 h after infection and both the released IL-8 and the mRNA expression of IL-8. Associations were found between the cultivable numbers of intracellular P. gingivalis and the mRNAs of IL-1, IL-6 and TNF-alpha at the day of infection. CONCLUSION: The results indicate survival of P. gingivalis within epithelial cells, possibly in a non-cultivable stage. Invasion into cells modulates the virulence properties of P. gingivalis as well as the inflammatory response of the cells.     

Temporal changes in inflammatory mediator concentrations in an adjuvant model of temporomandibular joint inflammation

AIMS: To determine temporal changes in the concentrations found in the temporomandibular joint (TMJ) and trigeminal ganglion of 3 specific classes of inflammatory mediators commonly linked with conditions of joint inflammation. The intent was to determine whether concentrations of the neuropeptide calcitonin gene-related peptide (CGRP), the neurotrophin nerve growth factor (NGF), and the proinflammatory cytokines interleukin-1beta (IL1beta) and tumor necrosis factor-alpha (TNF-alpha) are altered in the trigeminal ganglion and TMJ tissues during various stages of adjuvant-induced inflammation of the rat TMJ. METHODS: Adult male rats received bilateral TMJ injection of complete Freund's adjuvant (CFA), while control rats did not receive CFA treatment. The trigeminal ganglion and TMJ tissues were collected at 2 days, and 2, 4, and 6 weeks postinjection and analyzed using either radioimmunoassay or enzyme-linked immunosorbent assay. RESULTS: In the trigeminal ganglion, both CGRP and NGF concentrations were significantly elevated in comparison to controls from 2 days to 4 weeks; however, the patterns of increase differed. Concentrations of each inflammatory mediator were significantly elevated in the TMJ tissues of CFA-injected animals at 2 days and continued to be significantly elevated throughout the 6-week period. CGRP content remained at peak levels from 2 days through 6 weeks, while peak content for NGF, IL-1beta, and TNF-alpha was found at 2 days through 2 weeks. CONCLUSION: The results suggest that the development of CFA-induced inflammation of the TMJ was accompanied by a variable increase in the concentration of different classes of inflammatory mediators in both the trigeminal ganglion and TMJ tissues, which implies that each class of inflammatory mediator may play a significant role during different stages in the onset and exacerbation of the inflammatory process.     

Protective effects of Tacrolimus, a calcineurin inhibitor, in experimental periodontitis in rats

OBJECTIVE: Periodontitis is a well-appreciated example of leukocyte-mediated bone loss and inflammation with pathogenic features similar to those observed in other inflammatory diseases, such as arthritis. Since Tacrolimus, is an immunomodulatory drug used for the treatment of some cases of arthritis, we hypothesized that it may modulate periodontal disease. DESIGN: Using a murine model of ligature-induced periodontal disease, we assessed the effects of daily administrations of Tacrolimus (1mg/kg body weight) on bone loss, enzymatic (myeloperoxidase) analysis, differential white blood cells counts, airpouch exudate and cytokine expression for 5-30 days. RESULTS: Radiographic, enzymatic (myeloperoxidase) and histological analysis revealed that Tacrolimus reduced the severity of periodontitis. More specifically, Tacrolimus suppressed the expression of serum interleukin (IL-1beta), tumour necrosis factor (TNF-alpha), IL-6, airpouch exudate PGE(2) and leukocytosis usually observed after the induction of periodontitis. Tacrolimus treatment in periodontitis-induced rats conferred protection against the inflammation-induced tissue and bone loss associated with periodontitis, through a mechanism involving IL-1beta, TNF-alpha and IL-6. CONCLUSIONS: The effects of Tacrolimus on periodontal disease pathogenesis may provide clues to a novel approach to host modulation therapy in destructive periodontal disease.     

Tumour necrosis factor-alpha and apoptosis in the rat temporomandibular joint

The purpose of this investigation was to investigate the roles that tumour necrosis factor-alpha (TNF-alpha) and apoptosis play during acute inflammation of the temporomandibular joint (TMJ). Adult male Sprague-Dawley rats were injected with complete Freund's adjuvant (CFA) into the TMJ or kept as uninjected controls. The TMJ tissues were removed 2 days post-injection to mimic conditions of acute inflammation and analysed for changes in expression of TNF-alpha, the receptor TNF-R1, caspase-3 and -8, and apoptosis. Concentrations of TNF-alpha, TNF-R1, caspase-3 and -8, and apoptosis were significantly elevated in CFA-injected animals compared to uninjected controls. Tissue incubation with TNF-alpha caused a significant increase in caspase-3 and -8. Also, levels of apoptosis were significantly increased during inflammation, which could be inhibited by the addition of either anti-TNF-alpha neutralising antibody or caspase inhibitors. TNF-alpha may play a significant role in the onset of acute CFA-induced TMJ inflammation, and activation of apoptosis signalling pathways may be involved.     

Clinical course of an antigen induced arthritis model in the rabbit temporomandibular joint

The aim of this study was to investigate clinical features of an antigen-induced arthritis model in the rabbit temporomandibular joint (TMJ) and compare them to those in knee joint (KJ) arthritis. Eighteen adult male New Zealand White rabbits were divided into a TMJ arthritis group and a KJ arthritis group. Monoarthritis was induced unilaterally with ovalbumin and clinical observations, including joint swelling, skin surface temperature (SST) over the joints, and the withdrawal reflex threshold (WRT) to noxious pressure, were performed over a 3 week period. The joints were then evaluated histologically. The WRT was decreased during the 3 weeks after induction of TMJ arthritis, together with mild but significant joint swelling. The arthritic KJ also showed significant swelling and reduced WRT during the 3 week period. A significant increase of SST over the arthritic TMJ was present during the first week, while changes in SST over the KJ were inconsistent. The histological evaluation showed chronic arthritic features in all arthritic joints of both groups and no difference in the severity of arthritis was found between the TMJ and the KJ.     

Tumor necrosis factor-α increases chemokine gene expression and production in synovial fibroblasts from human temporomandibular joint

BACKGROUND: Synovitis, which is characterized by infiltration of inflammatory cells, often accompanies progression of clinical symptoms of the temporomandibular joint (TMJ). Synovial fibroblasts of the TMJ are believed to play important roles in progression of synovitis. The purpose of this study was to examine production and gene expression of chemokines by synovial fibroblasts stimulated by tumor necrosis factor-alpha (TNF-alpha). METHODS: Protein levels of chemokines were measured by enzyme-linked immunosorbent assay (ELISA). Gene expression of chemokines was analyzed by real-time polymerase chain reaction (PCR). RESULTS: Production of interleukin (IL)-8, growth-related oncogene (GRO)-alpha, monocyte chemoattractant protein (MCP)-1, and regulated upon activation normal T-cell expressed and secreted (RANTES) protein by synovial fibroblasts was increased by TNF-alpha. In contrast, stromal cell-derived factor (SDF)-1alpha, macrophage inflammatory protein (MIP)-1alpha and -1beta were not detectable in conditioned media of synovial fibroblasts, with or without TNF-alpha treatment. Increases in gene expression of IL-8, GRO-alpha, MCP-1, and RANTES in response to TNF-alpha treatment were detected. CONCLUSIONS: Increased protein production and gene expression of chemokines by synovial fibroblasts in response to TNF-alpha treatment appears to play an important role in recruitment of inflammatory cells into synovium and the progression of synovitis in the TMJ.     

Reactive oxygen species participation in experimentally induced arthritis of the temporomandibular joint in rats

In the temporomandibular joint (TMJ), it has been hypothesized that mechanical stresses lead to the oxidative stress of articular tissues. It has also been postulated that cells pertinent to arthritis-including endothelial cells and synovial cells-when stimulated by mechanical stresses and/or pro-inflammatory cytokines, promote oxidative damage. To determine the involvement of reactive oxygen species (ROS) in the diseased joint, we studied the generation of ROS in synovial fluid (SF) from interleukin-1alpha (IL-1alpha)-induced TMJ arthritis by electron spin resonance (ESR) spectroscopy, using the spin trap 5,5-dimethyl-1-pyrroline N-oxide (DMPO). The TMJ arthritis was experimentally induced in rats by the injection of human recombinant IL-1alpha into the TMJ; control rats were treated with normal saline solution. We found that the detected radicals in the collected SF were identified as a 1:2:2:1 quartet, characteristic of the hydroxyl radical-DMPO spin adduct. The ESR signal intensity of the hydroxyl radical-DMPO spin adduct in the SF from IL-1-treated rats was significantly higher than that from the control rats (P < 0.01). The results of ESR study also showed that hydroxyl radical (HO*) was increased in a time-dependent fashion in the presence of superoxide anion radical (O2*-) scavenger superoxide dismutase (SOD); the formation of DMPO-HO* was strongly inhibited by the iron chelater deferoxamine. We could measure higher levels of free iron (Fe2- and Fe3-) in the SF from TMJ arthritis than in that from controls (P < 0.05). Analysis of the data obtained from the present study suggests that the HO* radical detected in SF from IL-1-induced TMJ arthritis is generated via a modified Haber-Weiss reaction (biological Fenton reaction) in which O2*- can subsequently result in the production of H2O2 through dismutation reaction by SOD. Thus, HO* may be generated from the reaction of resultant H2O2 with free iron ions. The results presented here provide the first evidence of involvement of ROS in IL-1-induced TMJ arthritis.     

Antigen-induced arthritis of the temporomandibular joint via repeated injections of bovine serum albumin in domestic pigs

Rheumatoid arthritis (RA) is a chronic inflammatory disease and the temporomandibular joint (TMJ) is affected in up to 50%, resulting in pain, limited mouth opening and dental malocclusion. The outcome of conservative and surgical therapies is unsatisfying in many cases. The purpose of this study was to establish a large animal model of antigen-induced arthritis (AIA) of the TMJ that enables the investigation of the pathogenesis of RA and the evaluation of new therapies. In five domestic pigs, systemic immunization was performed via consecutive intramuscular injections of bovine serum albumin (BSA). Then, AIA was induced via the application of BSA into the TMJ. Injection with saline served as the control. After ten weeks, the joints and adjacent tissues were harvested for histological analysis and cytokine quantification. The changes observed in the AIA specimens included severe synovial inflammation, cartilage-specific glycosaminoglycan content loss, and cartilage surface and discus alterations as well as the formation of chondrocyte clusters. Protein analyses of the synovia showed enhanced levels of IL-1β, IL-6, TNFα and VEGF. A porcine model of immunologic arthritis of the TMJ was successfully established. This model may be used in future studies to investigate the underlying pathogenesis of RA and new therapeutic strategies.     

颌面枪弹伤软组织中前炎症细胞因子的表达

目的 观察颌面枪弹伤局部软组织中白细胞介素（ｉｎｔｅｒｌｅｕｋｉｎ,ＩＬ）６、８（ＩＬ－６、ＩＬ－８）、肿瘤坏死因子（ｔｕｍｏｒ ｎｅｃｒｏｓｉｓ ｆａｃｔｏｒ－ａｌｐｈａ,ＴＮＦ－α）的表达情况,分析前炎症细胞因子对伤口愈合的影响。方法应用免疫组织化学方法检测３种前炎症细胞因子在狗下颌枪弹伤模型伤口局部软组织中的表达,结合伤口组织病理学变化分析创伤愈合过程中前炎症细胞因子的作用机理。结果 枪弹     

颞下颌关节盘摘除术关节区组织病理学变化的实验研究

目的 建立兔颞下颌关节盘摘除术实验动物模型,研究关节盘摘除术早期,关节区组织形态学变化.方法 用10只新西兰大白兔,实验组8只行双侧关节盘摘除术;2只为正常对照组.术后1周、2周、4周、10周各处死2只,切取关节组织,进行组织病理学观察.结果 髁突及关节结节关节软骨连续性破坏,功能区关节软骨下骨组织直接暴露于关节腔内,非功能区则软骨细胞各层增生明显,表面纤维层增生变厚,呈现出纤维性粘连样改变.暴露在关节腔部分的骨组织表面致密,髓腔内的骨小梁吸收,伴微小囊肿形成.髁突关节软骨、骨组织及滑膜出现早期骨关节炎样改变.结论 兔关节盘摘除术后早期,关节区组织表现为骨关节炎样改变,不是适应性改变.     

Interleukin-1β induces interleukin-6 mRNA expression and protein production in synovial cells from human temporomandibular joint

BACKGROUND: Interleukin (IL)-1 beta and IL-6 were found to be elevated in fluid from the temporomandibular joint (TMJ), although the source of these cytokines was not elucidated. There is little known about the function and response of synovial cells in the TMJ. The purpose of this study was to prepare cultured human synovial cells (HTS cells) from the TMJ and to investigate IL-6 production in HTS cells incubated with IL-1 beta. METHODS: HTS cells were isolated from temporomandibular joint synovial tissue using an outgrowth method and then cultured in Ham's F12 medium containing 10% fetal bovine serum. The HTS cells were treated with or without IL-1 beta for 3, 6, 9 and 24 h. IL-6 and soluble IL-6 receptor (sIL-6R) levels in cultured supernatant were measured by ELISA. IL-6 mRNA expression was investigated using immunocytochemistry and RT-PCR. RESULTS: HTS cells were morphologically heterogeneous. IL-1 beta increased IL-6 production in HTS cells. In those treated with IL-1 beta, several cells were strongly stained in the cytoplasm around the nucleus, while several cells were weakly stained in this area. IL-1 beta also stimulated IL-6 mRNA expression. In contrast, sIL-6R could not be detected in cells treated with or without IL-1 beta. CONCLUSIONS: IL-1 beta increased IL-6 production in synovial cells resulting from an increase in IL-6 mRNA expression. Enhanced production of IL-6, which is associated with bone resorption and inflammatory response, seems to be related to the progression of TMJ disorders.     

rhIL-1Ra抑制大鼠颞下颌关节骨关节炎软骨破坏的研究

目的:探讨关节腔内注射重组人IL-1Ra对实验性大鼠颞下颌关节炎软骨关节修复的影响.方法:取SD大鼠24只,用关节腔内注射Ⅱ型胶原酶法建立双侧颞下颌关节骨关节炎模型,1周后,右侧(实验侧)关节腔内一次性注射重组人IL-1Ra5 μg(稀释于0.05 ml生理盐水),左侧(对照侧)注射等量生理盐水.建模后2周、4周各处死12只动物,取颞下颌关节标本进行HE染色、免疫组化染色及RT-PCR检测,用Mankin评分方法评估TMJ组织病理变化程度.另取1只SD大鼠用作空白对照,2周后处死.结果:2周时双侧颞下颌关节软骨均呈现不同程度的关节病损,实验侧和对照侧Mankin计分分别为1.33±0.52和2.00±6.63(P ＞0.05).4周时,实验侧改良和对照侧Mankin评分分别为3.00±0.63和6.50 ±0.84(P＜0.05).AD-AMTS-4、5的蛋白及mRNA表达也低于对照侧(P＜0.05).结论:颞下颌关节腔内注射重组人IL-1Ra能缓解骨关节炎导致的软骨病损,其治疗机制可能是通过抑制ADAMTS-4、5的表达来实现的.     

Orthodontic tooth movement and de novo synthesis of proinflammatory cytokines.

Interleukin-1beta (IL-1beta), interleukin-6 (IL-6), and tumor necrosis factor-alpha (TNF-alpha) are proinflammatory cytokines that are thought to play a role in bone remodeling, bone resorption, and new bone deposition. In the present work, in situ hybridization was performed to measure the messenger RNA expression of IL-1beta, IL-6, and TNF-alpha at 3, 7, and 10 days after the application of orthodontic force on the maxillary first molars of 12 rats. The contralateral side and 3 untreated rats served as controls. Measurements of the messenger RNA expression were selected as the means to investigate the role of orthodontic force in de novo synthesis of proinflammatory cytokines. After the application of force, the induction of IL-1beta and IL-6 was observed to reach a maximum on day 3 and to decline thereafter. No messenger RNA induction of either cytokine was measured in the control teeth. The messenger RNA expression of TNF-alpha was not detected at any time point of this study in the experimental or contralateral sides or in the control animals. Our data support the hypothesis that these proinflammatory cytokines may play important roles in bone resorption after the application of orthodontic force.     

Reduced mandibular growth in experimental arthritis in the temporomandibular joint treated with intra-articular corticosteroid

The aim of this investigation was to study the effect of intra-articular (i.a.) corticosteroid injections (IACIs) in the temporomandibular joint (TMJ) on mandibular development in antigen-induced TMJ arthritis. Ten-week-old female New Zealand white rabbits (n = 42) were randomly divided into four groups: group A, control (no injections); group B, placebo (repeated i.a. TMJ saline injections); group C, untreated arthritis (repeated induction of TMJ arthritis); and group D, steroid (repeated induction of TMJ arthritis + IACI). All animals had two tantalum implants inserted in the right side of the mandible serving as stable landmarks for later growth analysis. One implant was inserted close to the symphysis and one in the molar region. Computerized tomographic (CT) full-head scans were carried out at 14 (T1) and 26 (T2) weeks of age. (Dropout of animals at T2; group C, n = 7, and group D, n = 3.) Absolute and relative intra- and inter-group growth variations were evaluated during the growth period by comparison of CT scans. One-way analysis of variance was used for T1 statistical analysis, and absolute intra-group and relative inter-group growth differences between T1 and T2 were evaluated by Student's t-tests. At T2, the animals in the group A had greater sagittal and vertical mandibular growth compared with the other three groups. TMJ arthritis caused diminished mandibular growth. However, relative mandibular growth was significantly less in group D. The findings of this study do not indicate a positive long-term effect in the use of IACI in the TMJ as an early treatment intervention against TMJ inflammation in growing individuals.