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pp.49-57
Salivary gland adenocarcinoma(AdCa NOS) is one of the major causes of mortality among malignant salivary gland tumors. New therapeutic measure are needed to improve the outcome for patients with AdCa NOS because current therapy does not significantly improve survival rates. Transglutaminase 2(TGase 2) was implicated in forming cross-linked protein polymer, apoptosis and matrix interaction. And also TGase 2 expression is up-regulated in proliferation, migration, invasion, and metastasis of cancer cells. shRNA which has emerged as an effective method to target specific genes for silencing has provided new opportunities for cancer therapy. But there has been rarely reported using shRNA-TGase 2 transfection in AdCa NOS. The purpose of this study were to examine the specific inhibition of TGase 2 mRNA and protein expression by siRNA transfection of TGase 2 through RT-PCR and immunoslot blotting, and to study proliferation, migration and invasion assay of SGT cell line from AdCa NOS. Cell cycle analysis showed that the downregulation of shRNA-TGase 2 caused the accumulation of cells in the sub-G0/G1 phase. In migration assay, suppressing shRNA-TGase 2 inhibited the capacity of the cells to migrate compared to parental cells. In invasion assay, cells transfected with shRNA-TGase 2 decreased in invasion when compared to SGT and vector transfected cells. shRNA-TGase 2 expressing plasmids efficiently downregulated TGase 2 mRNA and TGase 2 protein expression. It suggested that the shRNA-TGase 2 targeting system against TGase 2 could have a therapeutic potentiality for malignant salivary gland tumors, especially in inhibiting and/or preventing cancer cell proliferation, migration and invasion.
Ⅰ. INTRODUCTION
Ⅱ. MATERIALS and METHODS
1. Cell Culture, siRNA-expressing PlasmidConstruction and Transfections
2. Cell Cycle Analysis
3. Cell Migration Assay
4. Cell Invasion Assay
5. RT-PCR
6. Sample Preparation for TGase assay
7. Conditions for TGase Assay
8. Immunoslot blot
Ⅲ. RESULTS
Ⅳ. DISCUSSION
REFERENCES
