The origin of squamous cell components in salivary gland tumor has been not yet clarified in detail. The squamous cell differentiation from adenocarcinoma has been reported in various carcinoma by HPV transfection in vitro. The adenocarcinoma cells adjacent to the squamous cell carcinoma components were positive for HPV. This is thought to indicate that after adenocarcinoma cells are transfected with HPV, they undergo morphological changes, and that squamous cell differentiation follows. The purpose of this study were to examine the effects of HPV-16 E6/E7 gene transfection into SGT cell line from human salivary gland adenocarcinoma, and to study the relation between the E6/E7 gene and squamous differentiation. Plasmid pBR322 containing HPV-16 was transfected into cultured SGT cell line using lipofectin method. Hygromycin was used as a selection marker. The presence of HPV E6/E7, transglutaminase 1, and involucrin mRNAs and protein in E6/E7 gene transfected cells was investigated by RT-PCR and immunoslot blot method. The apoptosis index was analysed by flow cytometry. The growth rate of E6/E7 gene transfected cells was reduced. E6/E7 transfected SGT cells increased apoptosis index. Involucrin and TGase I mRNAs by the squamous cell differentiation was most conspicuous in the E6/E7 gene transfected cell compared with non transfected cells. Squamous cell differentiation demonstrated in the transfectedSGT cell line, which expressed E6/E7 fusion gene mRNA.E6/E7 gene transfected cells showed squamous cell differentiation, expressing involucrin and TGase 1 protein by immunoslot blotting. The transfected SGT cell which expressed E6/E7 gene mRNA showed the squamous cell differentiation particularly clearly, and apoptosis was also demonstrated. It suggested that E6/E7 gene transfection into human salivary gland adenocarcinoma cells might induce clear squamous cell differentiation and contribute to study the pathogenesis of human salivary gland adenocarcinoma.

Urokinase-type plasminogen activator (uPA) and plasminogen activator type 1 (PAI-1) inhibitor contribute to the invasiveness of many carcinomas. It will be helpful to study clinical behavior of patients with malignant tumor by analysis of their expression. Expression of uPA and PAI-1 in human salivary gland tumors has been rarely reported in vitro. The purpose of this study were to investigate the protein expression of uPA and PAI-1 in SGT cell line compared to oral SCC and HeLa cell lines and to study migration and adhesion assay. All the cell lines were cultured under DMEM with 10% FBS at at 37oC in a 5% CO2 incubator. We studied a possible association between cytosolic uPA and PA-1 concentrations in SGT cell line compared to any other cell lines through cell migration and adhesion assay, and enzyme-linked immunoassay(ELISA). In migration assay SGT cell line was about 2 .5-4 folds higher than another cell lines. In adhesion assay SGT cell line was about 1.1-2 folds higher than another cell lines. uPA cytosolic concentrations of SGT cell line was about 3-10 folds, while PAI-1 was about 2.5-10 folds. Oral SCC cell lines were the lowest value. Both uPA and PAI-1 concentrations were correlated with migration and adhesion assay. High cytosolic concentrations of uPA and PAI-1 was correlated with migration and adhesion assay. It suggested that these markers might be specific marker for SGT cell line and would be contributed to treatment and prognosis of human salivary gland adenocarcinoma

The transcription factor nuclear factor-kB (NF-kB) plays an important role in regulating cell growth, apoptosis, and metastatic functions. Constitutive activation of NF-KB has been observed in various cancers; however, molecular mechanisms resulting in such activation remain elusive. Numerous evidences showed that over expression of TGase 2 might be linked with constitutive activation of NF-KB. To understand the pathways responsible for constitutive activation of NF-kB is important for rational design of NF-kB inhibitors for cancer therapy. Human salivary gland adenocarcinoma has been most aggressive solid tumors. Current therapy does not significantly improve survival rates. Thus, to investigate new therapeutic modalities against this adenocarcinoma is necessary. The purpose of this study was to study a constitutive activation of NF-kB with the expression of TG2 in SGT cell line origianted from human salivary gland adenocarcinoma by TGase 2 activity, RT-PCR and immunoslot blotting method. SGT cell line showed the highest TGase 2 activity and NF-kB activation than any other cell line. All the cell lines showed increased NF-kB mRNA activation after A231027 treatment than that of control. In immunoslot blotting, SGT cell line showed NF-kB activation correlated with TGase 2 expression after A231027 and BSA. It suggested that there might be a direct correlation between TGase 2 expression and NF-kB activation in SGT cell line.

Cyclosporine A (Cs A) which is a highly lipophilic cyclic undecapeptide mainly used for its immunosuppressive properties exert a wide spectrum of biological activities including fungicidal antiproliferactive, anti-inflammatory and chemotherapuetic effects. Human salivary gland adenocarcinoma is very aggressive characteristics, which is need to get the effective chemotherapuetic methods. Subconfluent SGT cell cultures have been treated with CsA at in vivo relevant concentrations for 24h. MTT assay for cellular proliferation of cultured SGT cell line has been performed and TGase 1 activity assay for cellular differentiation has been detected in the CsA-treated samples. It suggested that CsA could have an inhibitory effect in the proliferation of SGT cell line but no in the differentiation.

Taxol(paclitaxel) is used in chemotherapy against several cancer. Treatment of tumor cell lines with taxol induces apoptosis, but exact mechanism is not sufficiently understood. The purpose of this study was to investigate apoptosis by the inhibitory effect of paclitaxel on the motility properties of human salivary gland adenocarcinoma cell lines. Paclitaxel inhibited cell motility induced by soluble and immobilized attractant. It suggested that paclitaxel would be a potent inhibitor of salivary gland adenocarcinoma cell motility independent of its cytotoxic and apoptotic activity.

AJthough salivary gland adenocarcinoma accounts for third prevalence rate of all salivary gland tumors. it is one of the most aggressive solid tumors. Current therapy does not s ignificantly improve survival rates. Thus‘ investigating new therape utic modali t ies aga inst sali va ry gland adenocarcinoma is necessary. Manumycin A. a natural product o{ Streptα7Jyces parvuJus‘ inhi bits farn esy l- transferase by competition with farnesyl pyrophosphate groups . Manumycin has shown antitumor activity in several ex per‘ imental systems through identifying the regulatory pathway of apoptosis. The hi erarchical relationship of caspase-8 to caspase-3 and caspase-9 in the drug-induced a poptosis pathway in antitumol effect is not clear. The hi erarchical relations hip between cytochrome c and the caspases and provided evidence to support the hypothesis that the release of cytochrome c was upstream of caspase activation in the enhanced apoptosis induced by manumycin A Manumycin A has not been examined extensively in human salivary gland tumor and has not yet been clarified. The purpose of this study were to investigate mRNA and protein expression of Bc l- 2 、Bax, Cytochrome C‘ caspase- 3 , 一8 and -9 in SGT cell line by RT-PCR and immunoslot blotting, and to a pply its results to exami ne iLs chemoprevention for salivary gland adenocarcinoma. MTI assay showed about 50% cellular viability of SGT cell line treated by 50μM manunycin A Bcl-2. Bax‘ and caspase-8 mRNA expression in SGT cell line were unchangeable after 50μM manu nycin A Cytochrome C‘ caspase-3 and -9 showed about 1.5-5 folds higher mRNA expression in SGT cell line than that of control a nd DMSO- t reated group a fter 50M manunycin A. Bcl-2, Bax, and caspase-8 protein expression in SGT ce ll line were unchangcable after 50μM manunycin A. Cy Lochrorne C, caspase-3 and -9 showed about 2-7 fo lds higher protein express ion in SGT cell line than that of control and DMSO-treated group after 50μM manunycin A. mRNA expression was assoc iated with protein expression in SGT cell line after 50μM manunycin A. It suggested that manumycin A would induce poptotic effect on SGT cell line by caspase-3 and - 9 activation through cytochrorne c release. And man umycin A will be a useful chemoprevention drug for human salivary gland carcinoma in future.

Adenocarcinoma NOS of salivary glands is characterized by a high rate of local recurrences and metastasis. Long-term survival rate of Adenocarcinoma NOS lis not promising. Thus, different chemotherapeutical approaches had been proposed for this neoplasm, including apoptosis induction by drugs. The current treatment of choice of adenocarcinoma NOS is controversible, and an effective treatment for them is not yet available. Chemotherpeutic agents that can be inhibit or reverse the tumor growth by targeting apoptotic pathways will be new candidates for cancer prevention and therapy. The purpose of this study were to study the effect of Brefeldin A(BFA) as apoptotic inducing agent in SGT cell line from human submandibular adenocarcinoma NOS and apply these results to make a plan of treatment and prognosis of salivary gland tumors involving adenocarcinoma NOS. SGT cells were treated with a 300μM BFA solution in serum-free medium during 18 hours. SGT cells were grown in DMEM with 10% fetal bovine serum served as controls. The growth curve and MTT assay for succinyl dehydrogenase activity were performed. For apoptotic analysis, fragmentation of genomic DNA was confirmed with gel electrophoresis. Transmission electron microscopy was assessed for the effect of BFA on SGT cells phenotype. Apoptotic cell recognition and counting were carried out with Annexin-V, caspase 3 and APo2.7 antibody through flow cytometry. Growth of SGT cell line was abrutply decreased after 1 day of BFA treatment. MTT assay for succinyl dehydrogenase activity of the cells showed about 55% after 300μM BFA treatment. Destruction of cellular organells, numerous vacuolation in the cytoplasm & nucleus, chromatin margination, & fragments of nucleus were seen with TEM after 300μM BFA treatment. DNA fragmentation of SGT cell line was induced by 300μM BFA treatment and confirmed by gel electrophoresis from genomic DNA extraction. Late apoptosis of the cells through flow cytometric analysis of Annexin-V staining as induced by 300μM BFA treatment. Early apoptosis of the cells through flow cytometric analysis of caspase 3 and Apo 2.7 staining was induced by 300μM BFA treatment. It suggested that early and late apoptosis of SGT cell line would be induced by Brefeldin A treatment in vitro study. This work evaluated the efficacy of BFA, a potent apoptosis inducer, on SGT cultured cell line. And BFA as chemotherapeutic agent will be used as the treatment choice for adenocarcinoam NOS, and be need to apply BFA to in vivo study & clinical approach in future.

Established SGT cell line from human submandibular gland adenocarcinoma was used to study the TGase expression on a cellular level in vitro. Transglutaminase 2(TGase 2) is assoacitated with apoptosis, GTP binding protein, and cell marix interaction. The role of TGase 2 in salivary gland tumors is not clear yet. The pupose of this study were to examine the TGase expression of SGT cell line compared to other tumor cell lines, and to apply these results to the pathogenesis of salivary gland tumor. TGase enzyme assay of SGT, SCC-15, HN 4 and HeLa tumor cell line was 3 times repeated, and calculated. Immunoslot blot for semiquantitative protein analysis was done. The obtained results were as follows. 1. SGT cell line showed the highest TGase 2 enzyme activity(about 6-16 folds) irrespective of pre or postconfluency. 2. HN 4 cell line showed the highest TGase 1 enzyme activity(about 2-3 folds) irrespective of pre or postconfluency. 3. Under postconfluency TGase 1 induction was not induced, but slightly increased in all tumor cell lines. 4. TGase enzyme activity in all tumor cell lines was accompanied with TGase protein formation. From the aboving results, the higher TGase 2 expression of SGT cell line suggested that they would come from submandibular ductal cells and have a important role in the pathogensis of salivary gland tumors.