The Regulation And Its Mechanism Of AQP5Affected By MBD2in HSG Cells After Radiation Injury

Clinical practice shows that the salivary glands of the patients got oral andmaxillofacial malignancies (nasopharyngeal cancer, oral cancer and throat cancer) alwaysbe damaged after radiation therapy which would cause progressive hyposalivation, similarto Sj(o|¨)gren’s syndrome (SS), manifesting dry mouth. Sj(o|¨)gren’s syndrome (SS) is a class ofchronic autoimmune diseases. The study in etiology found that the exception of synthesisand expression within aquaporin5(AQP5) is closely related to SS. The reduction of AQP5plays an important part in SS salivation abnormal. The study has shown that DNAmethylation is involved in the expression inhibiting of the AQP5as a mechanism thatcannot be ignored. Demethylase(MBD2)is an important part of the DNA methylation. Thegene has the ability of methylation silencing, also with regulatory proteins of DNAmethylation which show the methylase activity, performances a significant role in theepigenetic regulation of a variety of tumors. Biology software testing analysis reflect thatMBD2(methyl-CpG binding domain2)can regulate methylation of AQP5as a part ofmethylation system, though the mechanism was unknown.Objective：To established the injured model of human submandibular gland(HSG)byradiation for study on the relationship between MBD2and AQP5. Increasing the AQP5directionally by the mediating of MBD2, and investigate the expression of AQP5in theinteraction of MBD2and its regulation mechanism, offering new perspectives for theclinical treatment of Sj(o|¨)gren’s syndrome.Method：Through the basis of the analysis on the related of AQP5and SS bybioinformatics test, the subject focused on the molecular and cellular level. First of all,establishing the injured cellular model of human submandibular gland(HSG)by radiation.Using immunofluorescence, RT-PCR and western-blot, detect the differentially expressedof AQP5in HSG before or after radiating. Then, overexpression vector of adenovirus wouldbe established and transfect to HSG. For proving the adjusting of MBD2in AQP5, RT-PCR and western-blot were used to testing the expression of AQP5.Result：1Established the injured cellular model of human submandibulargland(HSG)by radiation.2The expression of AQP5would be inhibited not only in mRNA,also in protein, by the60cobalt-γ-rayed radiating.3Expression vector of adenovirus hasbeen established.4MBD2, can promote the expression of AQP5in HSG cell lines.Within the using of immunofluorescence, RT-PCR and western-blot, we estimated theexpression of AQP5in a HSG model, and the relationship with MBD2. In subsequentexperiments, by means of double fluorescent promoter reporter system, EMSA andchromatin immunoprecipitation (CHIP) to detect the MBD2how to involved in AQP5expression patterns, also to investigate in depth the interaction of MBD2in AQP5and itsregulation mechanism. Furthermore, theoretical and experimental basis for SS treatmentthrough AQP5would be provided by the demethylation research.