Expressions Of Transforming Growth Factor β1and Matrix Metalloproteinase-9in Experimental Tympanosclerosis

Tympanosclerosis (TS) is the end-stage process of continued inflammation in the middle ear cleft, which is seen after recurrent or chronic otitis media(OM), and its mainly performances is progressive hearing loss, and it often induces conductive deaf. Its etiology is unknown, but it is widely thought to be an excessive healing response to infection or trauma, for instance, acute and chronic suppurative otitis media and secretory otitis media etc. As we known, streptococcus pneumoniae and staphylococcus aureus are common pathogenic bacteria in OM. In recent years, studies found that various tissues and organs had same features in development of sclerosis:begin with damaging of parenchymal cells and, then induce extracellular matrix deposits and tissue remodeling eventually. Above two stages are still found in formation of TS. Transforming growth factor β1(TGFβ1), as one of the important factors to fibrosis, can promote ECM synthesis, inhibit ECM degradation and inhibit MMP-9synthesis directly. Literatures have reported that TGFβ1can not only inhibit immune responses, but also can reduce the body’s natural immune function through reducing secretion of sticky protein in SOM. In addition, TGF β1was known as one of important factors in promoting the formation of fibrosis., and studies have proved that the expression of it were significantly increased in different tissues and organs, such as liver, kidney and lung fibrosis. Matrix metalloproteinases (MMPs) is the main protein enzymes, which participate in degradation of extracellular matrix components. Matrix metalloproteinase-9(MMP-9), as a member of this family, can also degrade type4collagen fiber except extracellular matrix components. Turkish scholars have found that antagonist drugs of matrix metalloproteinases can effectively prevent the development of TS. Previous studies showed that, increased activity of MMP-2and MMP-9can degrade excessive sedimentary of extracellular matrix, destroy normal structure basement membrane and then initiate the occurrence of fibrosis in early period of liver fibrosis; and continued rising of MMP-2and MMP-9further promote degradation of normal matrix components, and finally form a vicious circle and jointly promote the fibrosis process at stage of cirrhosis. Other researches reported that a variety of inflammatory cells factor, hormones and growth factor (such as TGF (31, etc.) can induce expressions of MMPs, while TGFβ1can also inhibit matrix metalloproteinases synthesis and promote expressions of metal protease inhibitors to reduce degradation of the extracellular matrix. As so far, surgery is the main treatment to TS, and there is no effective drugs to treat and prevent the development of TS in the clinical practice. Through studying the functions of transforming growth factor beta and matrix metalloproteinases in various inflammation and fibrosis diseases, many scholars have put forward that targeted therapy may be a new direction to treat and prevent the development of these diseases. The worsening of TS will affect normal communications, and thereby reduce the patients’quality of living and working in different degrees. Therefore, the aim of this experiment is to study the expressions of transforming growth factor beta1and matrix metalloproteinase-9in formation of TS through establishing animal models of TS. Then on the basis of this work, we want to further explore the possible molecular mechanism of the two factors in development of TS and provide guiding significance for clinical researches and new targeted treatments and preventions of TS.Objectives:Our study was to investigate the expressions of TGFβ1and MMP-9in an experimental tympanosclerosis(TS) and explore their possible roles in the development of this disorder.Methods:70guinea pigs were utilized in this study, of which ten were chosen to serve as control and the other sixty were chosen to set up TS model by inoculation of type-3Streptococcus pneummoniae. Then the sixty animals were further divided into six subgroups on the basis of six time points, including1week、2weeks、3weeks、4weeks、6weeks and8weeks. Observation of myringosclerosis by Otomicroscopy, and findings of the morphological changes and calcium depositions by Hematoxylin-eosin and Von Kossa staining were performed to evaluate the animal model of TS. The expressions of TGF(31and MMP-9were detected by Western blot and immunohistochemistry.Results:Slight Sclerotic changes in tympanic membrane(TM) were observed from week2and extensive myringosclerosis(MS) was found from week6under Otomicroscopy. Pathology changes of TS and calcification were obviously appeared in tympanic membrane and middle ear mucous membrane from week6by HE and Von Kossa stain. The expressions of TGFβ1and MMP-9were found by Immunohistochemistry, and both were seen in the cytoplasm of fibroblast cells and inflammatory cells which were widely distributed in tympanic membrane and middle ear mucosa at week6. The expression of TGFβ1protein was detected significantly increase in the formation of tympanosclerosis. MMP-9increased from week1to week4, and then declined from week6. The expressions of TGFβ1and MMP-9were statistically significant compared with the corresponding control group (P<0.05).Conclusion:Our data indicate that the alteration in expressions of TGF-β1and MMP-9were involved in the development of TS, which may be an important mechanism underlying the pathogenesis of TS. Additionally, it maybe useful for further researches of pathogenesis、clinical treatment and prevention.