The Effect of LIPUS on TMJ Arthritis in Juvenile Rats

• Author / Creator

  Crossman, Jacqueline

• Juvenile idiopathic arthritis (JIA) is an inflammatory disease that can affect the temporomandibular joint (TMJ). TMJ-JIA can cause mandibular growth disturbances. Treatment options include medications targeting inflammation, orthodontic intervention, and surgeries; however, these medications are ineffective in many patients, none of these options stimulate mandibular growth, and surgery is risky, painful, and costly. This thesis aimed to investigate the effect of low intensity pulsed ultrasound (LIPUS) on TMJ-JIA in juvenile rats. LIPUS has been shown to have stimulatory effects on osteoblasts, chondrocytes, and synovial cells, and it has been demonstrated to have an anti-inflammatory effect on arthritis in animal models and more importantly LIPUS can stimulate mandibular growth.
  We first aimed to validate the collagen-induced arthritis (CIA) juvenile rat model of TMJ arthritis. Twenty-seven 3-week-old male Wistar rats were used. After in vivo micro-computed tomography (MicroCT) scanning, we divided them into 3 groups (n=9): the CIA group, the Saline group, and the Healthy group. The CIA group received CIA-TMJ injections (right and left) with an emulsion of type II collagen (Col-II) and Complete Freund’s Adjuvant, the Saline group received saline TMJ injections, and the Healthy group remained untreated. After 4 weeks, the rats were euthanized and ex vivo MicroCT scanned, and the TMJ tissues were prepared for analysis. Our results demonstrated that CIA rats had significantly less mandibular growth (p < 0.01), their synovial tissues were inflamed, there was an increased expression of interleukin (IL)-1β, matrix metalloproteinase (MMP)-13 and tumor necrosis factor (TNF)-α (p < 0.05), there was acute cartilage thickening, and Col-II expression was decreased. These results demonstrate the degenerative changes in TMJ cartilage due to CIA in the TMJ.
  Next, we studied the effect of LIPUS on CIA-induced TMJ arthritis. Seventy-two 3-week-old male Wistar rats underwent in-vivo MicroCT scans and were divided into 8 groups (n=9). Group 1 received CIA injections into the TMJs and immediate LIPUS treatment to the TMJs (20 minutes/day, 4 weeks); Group 2 received saline TMJ injections and immediate LIPUS treatment; Group 3 received CIA-TMJ injections and no immediate LIPUS (isoflurane anesthesia only, 20 minutes/day, 4 weeks); and Group 4 received saline TMJ injections and no LIPUS treatment. Group 5 received CIA-TMJ injections and after 4 weeks, received LIPUS treatment (delayed LIPUS application); Group 6 received saline TMJ injections and delayed LIPUS; Group 7 received CIA-TMJ injections and no delayed LIPUS (isoflurane only, 20 minutes/day, 4 weeks); and Group 8 received saline TMJ injections and no delayed LIPUS. The animals were euthanized, then ex vivo MicroCT scans were completed and tissues were prepared for analysis. We histologically analyzed H&E-stained TMJs, measured cartilage cell layer thicknesses and performed toluidine blue staining. Immunohistochemistry was completed to analyze expression of Col-II, Col-X, MMP-13, receptor activator of NF-κB ligand (RANKL), vascular endothelial growth factor (VEGF), transforming growth factor (TGF)-β1, IL-1β, IL-17, and TNF-α. ELISAs quantified levels of TNF-α and IL-1β in the blood serum, TGF-β1 (active/inactive forms) in the plasma, and IL-6 and IL-1β in the articular disc. We calculated mandibular and condylar growth using MicroCT analysis.
  In general, synovitis and condylar bone erosion were observed in the CIA TMJs, but less severe synovitis and minimal erosion was present in the LIPUS-treated TMJs. Immediate LIPUS treatment prevented acute cartilage thickening due to CIA (p < 0.05); increased proteoglycan production and Col-II, Col-X, and TGF-β1 expression; decreased MMP-13 and VEGF expression; and stimulated mandibular growth in CIA TMJs (p < 0.005). Delayed LIPUS treatment increased Col-II and TGF-β1 expression, but it did not affect cartilage thickness nor stimulate mandibular growth.
  This study validated the CIA juvenile rat model of TMJ arthritis. Our results show that LIPUS prevented mandibular growth disturbances and acute cartilage changes caused by CIA. The inflammatory effects from CIA didn’t appear to persist in the rats that received delayed LIPUS, so we observed minimal effects of CIA and LIPUS in these older rats. This study’s results have increased the understanding of this animal model of arthritis, its uses and limitations, as well as providing a greater understanding of LIPUS and its effects on the TMJ and mandibular growth in CIA-induced TMJ arthritic rats. These results will help to design future studies to further investigate LIPUS and TMJ arthritis, towards developing a new treatment for children TMJ-JIA.

• Subjects / Keywords

  • Arthritis
  • Therapeutic ultrasound
  • Animal model
  • Temporomandibular joint

• Graduation date

  Spring 2022

• Type of Item

  Thesis

• Degree

  Doctor of Philosophy

• DOI

  https://doi.org/10.7939/r3-cmyv-bf29

• License

  This thesis is made available by the University of Alberta Libraries with permission of the copyright owner solely for non-commercial purposes. This thesis, or any portion thereof, may not otherwise be copied or reproduced without the written consent of the copyright owner, except to the extent permitted by Canadian copyright law.