The importance of TNF-alpha in arthritis is well documented. It may be that TNF-alpha is also markedly involved in muscle inflammation (myositis). An animal model where this can be investigated is needed. A newly developed rabbit myositis model involving pronounced muscle overuse and local injections of substances having proinflammatory effects was therefore used in the present study. The aim was to investigate the patterns of TNF-alpha expression in the developing myositis and to evaluate the usefulness of this myositis model for further TNF-alpha research. Human rheumatoid arthritis (RA) synovial tissue was examined as a reference. TNF-alpha immunoexpression and TNF-alpha mRNA, visualized via in situ hybridization, were detected in cells in the inflammatory infiltrates of the affected muscle (soleus muscle). Coexistence of TNF-alpha and CD68 immunoreactions was noted, suggesting that the TNF-alpha reactive cells are macrophages. Expression of TNF-alpha mRNA was also noted in muscle fibers and blood vessel walls in areas with inflammation. These findings demonstrate that TNF-alpha is highly involved in the myositis process. The model can be used in further studies evaluating the importance of TNF-alpha in developing myositis.

TNF-alpha is known to be involved in muscle damage and inflammation (myositis). The relationships between the TNF-alpha system and muscle fiber necrosis/regeneration and the tachykinin system in this situation are unclear. We have an experimental rabbit model related to unilateral muscle overuse which leads to marked muscle derangement and myositis bilaterally. Using this model, staining for TNF-alpha, in parallel with staining for the substance P-preferred receptor (NK-1R) and desmin were performed. Desmin staining was used as a reference concerning identification of degeneration/regeneration and the soleus muscle was the muscle examined. It was observed that the inflammatory cells, as well as blood vessel walls in the myositis areas, expressed TNF-alpha mRNA. Muscle fibers that were interpreted to represent necrotic fibers expressed TNF-alpha mRNA reactions and showed NK-1R immunoreactions, the reactions being confined to white blood cells that had infiltrated into the fibers. Muscle fibers that were interpreted to be in a regenerative state expressed patchy/widespread TNF-alpha mRNA and point like NK-1R immunoreactions. Abnormal muscle fibers thus showed TNF-alpha mRNA as well as NK-1R immunoreactions. Normal muscle fibers never showed these reactions. Occurrence of inflammatory cell and muscle fiber TNF-alpha mRNA reactions was equally marked in the myositis areas of the contralateral side as in these areas of the ipsilateral experimental side. The observations show that the TNF-alpha system is much involved in the processes that occur in the muscle derangement/myositis processes. The involvement relates to effects in processes of both regeneration and muscle fiber necrosis. It may be that substance P via activation through the NK-1R influences the TNF-alpha expression. The findings of TNF-alpha upregulation also for the contralateral side show that the TNF-alpha system is involved both ipsi and contralaterally during the development of myosits/muscle affection in response to unilateral overuse.

BackgroundTNF-alpha is suggested to be involved in muscle damage and muscle inflammation (myositis). In order to evaluate whether TNF-alpha is involved in the myositis that occurs in response to muscle overuse, the aim was to examine the expression patterns of TNF receptors in this condition.MethodsA rabbit muscle overuse model leading to myositis in the soleus muscle was used. The expression patterns of the two TNF receptors Tumor Necrosis Factor Receptor type 1 (TNFR1) and Tumor Necrosis Factor Receptor type 2 (TNFR2) were investigated. In situ hybridization and immunofluorescence were utilized. Immunostainings for desmin, NK-1R and CD31 were made in parallel.ResultsImmunoreactions (IR) for TNF receptors were clearly observed in white blood cells, fibroblasts and vessel walls, and most interestingly also in muscle fibers and nerve fascicles in the myositis muscles. There were very restricted reactions for these in the muscles of controls. The upregulation of TNF receptors was for all types of structures seen for both the experimental side and the contralateral nonexperimental side. TNF receptor expressing muscle fibers were present in myositis muscles. They can be related to attempts for reparation/regeneration, as evidenced from results of parallel stainings. Necrotic muscle fibers displayed TNFR1 mRNA and TNFR2 immunoreaction (IR) in the invading white blood cells. In myositis muscles, TNFR1 IR was observed in both axons and Schwann cells while TNFR2 IR was observed in Schwann cells. Such observations were very rarely made for control animals.ConclusionsThe findings suggest that there is a pronounced involvement of TNF-alpha in the developing myositis process. Attempts for reparation of the muscle tissue seem to occur via both TNFR1 and TNFR2. As the myositis process also occurs in the nonexperimental side and as TNF receptors are confined to nerve fascicles bilaterally it can be asked whether TNF-alpha is involved in the spreading of the myositis process to the contralateral side via the nervous system. Taken together, the study shows that TNF-alpha is not only associated with the inflammation process but that both the muscular and nervous systems are affected and that this occurs both on experimental and nonexperimental sides.

Using a rabbit model leading to myositis in response to exercise-induced muscle overuse, we have previously observed that TNF-alpha is involved in the exercised muscle in early developing myositis as well as both ipsi-and contralaterally in the myositis which develops in response to a lengthened period of overuse. It is unknown if TNF-alpha can also be engaged contralaterally in early stages of myositis. The hypothesis was that this is the case. It was therefore evaluated whether the TNF-alpha system is early involved contralaterally. An experimental model of 1 week of overuse of the soleus muscle on one side leading to myositis was used, and in situ hybridization and immunohistochemistry were applied to study the expression patterns of TNF-alpha in the soleus muscle in the contralateral side. TNF-alpha was expressed in the myositis process which occurred contralaterally. There were thus TNF-alpha mRNA reactions in the cells of the inflammatory infiltrates, in blood vessel walls and in certain of the muscle fibers. Parts of the latter were necrotic fibers, whereas others were interpreted to be in a regenerative stage. TNF-alpha immunoreactions were seen for infiltrating white blood cells. The observations show that the TNF-alpha system is early involved in the cross-over effects that occur in response to unilateral muscle overuse leading to myositis bilaterally. TNF-alpha is likely to have pro-inflammatory and destructive effects but also to have effects in the muscle regenerative processes. The occurrence of an early involvement of the TNF-alpha system contralaterally to the injury side shows a new aspect of importance of this system in inflammation.