Mechanisms Of Inflammatory Pain In The Temporomandibular Joint Disorder

Part1.Establish an inflammatory model of temporomandibular joint in rats and observe pain behaviorObjective:To observe the pain behaviors and detect the expression of c-fos and substance P(SP)in the spinal trigeminal nucleus of brainstem by establishing an inflammatory model of temporomandibular joint in rats.Methods:Rats were randomly divided into control group and experimental group.The temporomandibular joint cavity were respectively performed in saline and Complete Freund's adjuvant(CFA)injection,and then observed the pain behaviors in the observation box.Rats pain behaviors included orofacial friction,scratching and shrinkage head etc.Rats temporomandibular joint pain threshold and eating were measured lasts a week.Finally,immunohistochemistry(IHC)was used to detect the expression of c-fos and SP in the spinal trigeminal nucleus of brainstem.Results:Compared with the control group,the pain behaviors of rats in the experimental group are more in number and longer in time.After 24h intra-articular injection,the swelling of bilateral temporomandibular joint in the experimental group are more severe,the temporomandibular joint pain threshold and the food intake were significantly decreased.At the same time,the number of c-fos and SP positive cells in the spinal trigeminal nucleus was increased significantly.However,no significant change was observed in the control group.Conclusion:The rat model of temporomandibular joint inflammatory pain can be successfully induced by intra-articular injection with CFAPart2.Mechanisms of the synovitis in the pain of temporomandibular joint disorderObjective:To investigate the effect of expression of calcitonlin gene related peplide(CGRP)and SP in synovial tissue and trigeminal ganglion(TG),and infiltration of PGP9.5 nerve fibers in synovial tissue on rat TMJ inflammatory pain.Methods:TMJ pain threshold was measured at 1,2,3 and 4 weeks after injection.TMJ and TG of rat were harvested at corresponding time.TMJ was observed with hematoxylin eosin(HE)staining.The expression of CGRP,SP and PGP9.5 in synovial tissue and the expression of CGRP and SP in TG were detected by IHC.Finally,the gene expression level of CGRP,SP and PGP9.5 in synovial tissue were detected by RT-PCR.Results:Compared with the control group,the synovial tissue of the experimental group was significantly hyperplasia,and the articular cavity was full of inflammatory secretions.A large number of inflammatory cells and fat droplets can be seen in the proliferative synovial tissue.At 1 and 2 weeks experimental groups,the TMJ pain threshold was significantly lower than the control group.In the control group,synovial tissue shows no CGRP,SP and PGP9.5 expression,while in the experimental groups,the expression was significantly increased,especially at 1 and 2 weeks.At the same time,RT-PCR results showed that the CGRP,SP and PGP9.5 gene expression level was consistent with the protein expression.In the control group,there were a small number of neurons in the trigeminal ganglion to express CGRP and SP.However,at 1 and 2 weeks experimental group,the number of CGRP and SP positive neurons was significantly increased.Conclusion:the over expression of CGRP and SP in the synovial tissue and trigeminal ganglion,and the infiltration of PGP9.5 nerve fibers in synovial tissue were involved in TMJ inflammatory pain.Part3.Mechanisms of the osteoarthritis in the pain of temporomandibular joint disorderObjective:To investigate the effect of condylar cartilage degeneration and subchondral bone abnormal remolding on the temporomandibular joint osteoarthritis(TMJOA)pain.Methods:TMJ and TG samples were taken at 1,2,3 and 4 weeks after injection.Histological observation of condylar cartilage and subchondral bone was performed by HE,toluidine blue(TB),Safranin O(S.O),Masson trichrome staining and Micro-CT.Osteoclast and osteoblast activity were analyzed by tartrate-resistant acid phosphatase(TRAP)staining and osteocalcin(OCN)immunohistochemical staining respectively.The expression of NF-?B receptor activator ligand(RANKL)and osteoprotegerin(OPG)in condylar cartilage and subchondral bone were detected by IHC and the ratio of RANKL/OPG was evaluated.The expression of SP and PGP9.5 in TG was detected by immunofluorescence(IF)double staining.Results:Compared with the control group,the thickness of the condylar cartilage was reduced,the number of chondrocytes decreased and the cartilage matrix proteoglycan was decreased.Especially at 2 weeks experimental group,the osteoclast activity in subchondral bone was significantly enhanced and the number of osteoclasts was increased to the most associated with subchondral bone marrow lesions enlarged.At the same time,the ratio of RANKL/OPG in cartilage and subchondral bone was the highest.At 4 weeks experimental group,unmineralized new bones with irregular trabecular bone structure,up-regulated OCN expression,and decreased bone mineral density(BMD)were found in subchondral bone.Compared with the control group,the number of SP and PGP9.5 co-expressed neurons in the TG was significantly increased in the experimental group.Conclusion:The degeneration of condylar cartilage and the abnormal subchondral bone remodeling were associated with the decreased TMJ pain threshold and SP over-expression in the TG,indicating that the condylar cartilage matrix degradation,chondrocyte apoptosis,subchondral bone osteoclast activity and bone marrow edema lesions were involved in TMJOA pain.