RT-PCR Examination of Inflammatory Mediators in TMJ Inflammation

Objectives: Previous results from this group demonstrated that adjuvant-induced inflammation of the TMJ produced a significant increase in inflammatory mediators within the tissues of the TMJ. However, changes at the gene level have not been determined. Therefore, the goal of this study was to use RT-PCR to examine changes in the expression of various neuropeptides and neurotrophins during inflammation of the TMJ. Methods: Ten adult (200-225 g) male Sprague Dawley rats were injected with complete Freund's adjuvant (CFA, 50 mg in 50 ml of paraffin oil) into the superior joint space of the TMJ to induce acute inflammation. The rats were sacrificed 48 hours later along with ten age-matched, uninjected controls. TMJ tissues (disc, retrodiscal, and capsule) were removed and homogenized. Subsequently, RNA was extracted and RT-PCR was run to examine changes in RNA expression for calcitonin gene-related peptide (CGRP), substance P (SP) and nerve growth factor (NGF). The RT-PCR band densities were quantified from three separate experiments via computer-assisted two-dimensional densitometric scanning using the public domain NIH Image Analysis Program 1.59b4f. Results: Densitometric analysis of the RT-PCR results indicate that CGRP mRNA was increased in the inflamed TMJ tissues by 56% (p<0.01) relative to the uninjected controls. SP mRNA was also significantly increased by 48% (p<0.01), while NGF mRNA was increased only slightly (23%, p<0.05) relative to the controls. Conclusions: This study demonstrates that adjuvant-induced inflammation of the TMJ resulted in an upregulation of mRNA expression of the neuropeptides CGRP and SP, and the neurotrophin NGF, during the acute stage of inflammation of the TMJ, providing further evidence of their role and importance in this process. This study was supported in part by NIH/NIDCR Traineeship T35 DE07188 and the Baylor Oral Health Foundation.