| Purpose:Salivary adenoid cystic carcinoma is one of the relatively common malignantsalivary gland tumors. Such cancer is characterized by its moistness, the vagueboundary with its surrounding tissues, the relapse after operation, the intrusion intovessels and the metastasis to remote parts through blood circulation. The curve forsalivary adenoid cystic carcinoma is still unitary now and lack of theoreticalresearch foundation. So it is necessary to carry out further study on its occurrenceand development based on molecule biology, and to explore new and effective cure.Homeobox gene family is a big family of coding the protein of transcriptionfactors, expression and coding a specific kind of transcriptional regulatory factors inthe specific growing stage of organism and in specific tissues. The expressing resultcan adjust the split of the cells and control the synthesis of relevant informationconveying cells and various hormone to influence the information conveying amongthe nearby cells and remote conveying, so that the growth and mature of the embryo can be adjusted. The expressing result also plays the role of master control in theprocess of multification and split of adult tissue cells.The sudden change of the sequence of a certain DNA of Homeobox gene canresult in the deterioration of phenotype change of normal cells. The abnormalexpression will make the split and multiplication of cells out of control and preventthe cells from splitting and growing, resulting in tumor.The occurrence and growth of salivary adenoid cystic carcinoma is a complicatedprocess mingled with a variety of factors and steps, involving the change of manykinds of genes, and relating to the mutual adjustment between the structure andfunction of transcription factors. As a big family in transcription factors, Homeoboxgene plays vary important role in the process of occurrence and growth of tumor. Ifthe key homeobox gene involved in the occurrence of salivary adenoid cysticcarcinoma can be found, the study on attack mechanism and clinical treatment ofsalivary adenoid cystic carcinoma will have a new moment. By using genemicroarray, this research selected homeobox gene relevant to the attack of salivaryadenoid cystic carcinoma and also tested them through realtime-PCR to inducechemically the standard cells of human salivary adenoid cystic carcinoma-so as tochange the biological character of cancer cells. The purpose of testing the influenceon the expressing of target gene was to find the key genes that control theoccurrence and growth and split inducing of salivary adenoid cystic carcinoma.Method:(1) Six paired specimens including salivary adenoid cystic carcinoma and itssurrounding normal salivary gland were obtained from six patients. Meanwhile twopaired specimens including salivary adenoid cystic carcinoma cells and normalsalivary gland were got. Total RNA was extracted from all specimens by means of Trizol method, and then mRNA was purified. Two equal quantities of mRNAsamples were reversely transcripted into fluro-marker-cDNA complex andhybridized with customized Oligo microarray, which containing numerous probes of232 human homeobox genes. Eight hybridized microarray chips were scanned byAgilent Scanner. All data were processed with professional software and differentlyexpressed genes were sorted out. The purpose was to find out some homeobox geneswhich were related to the oncogenesis of the salivary adenoid cystic carcinoma.(2) Salivary adenoid cystic carcinoma cell ACC-2 and ACC-M were chosen as theexperimental groups, and normal salivary gland tissue was used as control. TotalRNA was extracted by Trizol means and reversely transcripted into cDNA. Bymeans of quantitative real-time PCR detection, the mRNA expression of 3 suspectedhomeobox genes (TGIF, EVX1 and PITX1) were tested in different samples. Thenthe threshold value was compared to find out the differently expressed homeoboxgenes.(3) EVX1 gene was chosen as the target gene, and plasmid pEGFP-EVX1 wasconstructed. ACC-M cell was transfected by the plasmid. Then real-time PCR wasused to test the result, and the sequence was analyzed by Nucleotide-nucleotideBLAST (blastn). After 24 hours, the transfected cells were observed byfluent-microscope. The cell growth curve was assignmented, and the cell proliferation wastested by MTT method. The migration ability of the cells was decided by their ability to pass thefilter of 8μm aperture.Results:(1) 67 up-regulated genes and 54 down-regulated homeobox genes were found in thetissue samples, and 12 up-regulated genes and 15 down-regulated homeobox geneswere found in the cell samples. TGIF, EVX1, IRX2, EN2, HOP, IPF1, MEIS2 and PHOX2B were differently expressed both in the tissue and cell samples.(2) There was no difference in the expression of PITX1 between normal salivarygland and ACC-2, ACC-M. There was no difference in the expression of TGIF,EVX1 between normal salivary gland and ACC-2. Statistical significance was foundin the expression ofTGIF, EVX1 between normal salivary gland and ACC-M.(3) After the enhanced expression of EVX1 in the ACC-M, the tell proliferation wasfound to decrease by thegrowth curve and MTT method. The cell migration abilitydecreased too because the expression of EVX1 in the ACC-M was enhanced.Conclusion(1) Many homeobox genes were related to oncogenesis of salivary adenoid cysticcarcinoma, and three highly suspected genes (TGIF, EVX1 and PITX1) werescreened.(2) By means of quantitative real-time PCR detection, EVX1 gene was clarified asthe most suspected homeobox gene related to the salivary adenoid cystic carcinoma.(3) EVX1 gene might restrain the growth and metastasis of salivary adenoid cysticcarcinoma through the regulation of its cellular cycle.
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