| Salivary gland adenoid cystic carcinoma, which comprized 27% of all salivary gland malignant tumors, is one of the most frequently encountered malignance. It is very common in submandibular gland and small-salivary gland and has a high metastatic potential. It is well known that hormone and hormone-like substances play important roles in controlling cell differentiation and proliferation. It has been reported that estrogen receptor (G2) exists not only in target cells of mammary gland and womb, but also in non-target organs such as liver, kidney, colon, pancreas, oral mucosa and skin. 17-B Estrogens(E2) is the most cancer promoting substance among 7 carcinogens that FDA declared. Since E2 is still widely used clinically and because E2 has carcinogenisis potential, the relationship of E2 and ER has attracted wide attention. Up to now, there is no report available about the effect of E2 on adenoid cystic carcinoma cell proliferation and metastasis.In this study, adenoid cystic carcinoma (Sacc 83) cells were treated with E2 at clinically used dosage to investigate the effects of E2 on the proliferation and metastasis of SAcc83 cells.The effects of E2 on the proliferation of Sacc83 cells were studied by MTT assay, cell counting, clonogenic assay and flow cytometry. The expression of proliferation cell nuclear antigen was examined with immunocytochemical method. The matrix metalloproteinase-9 (MMP-9) activity was tested with sodium dodecyl sulfide polyacrylamide gel electrophoresis (SDS-PAGE). The invasion potential of the cells was investigated with chicken embryo heart invasion assay. The in vivo growth and metastasis of SAcc83 cell induced tumor were studied in ovary-resected female nude mice. 106SAcc83 cells in 0.2ml of serum-free medium were injected subcuteneously into each of 4 nude mice. In the tested group, 0.2ml of 9.55 X 10-5mol/L E2 was given subcuteneously to each mice daily. The size of the subcuteneous neoplasm was measured and calculated daily untill day 15. Then the mice were sacrificed and the tumous were removed, measured and histologicaly examined. In vivo metastatic potential of the cells was examined with tail vein injection of 10 cells into 10 ovary-removed female nude mice. E2 was given to 5 tested animal with the schedule mentioned above. 42 days after injection, the mice were sacrificed, the metastasis foci on the lung surfaces were counted. Following results were obtained:1. MTT assay revealed that E2 at 10-5, 10-6, 10-7,10-8 and 10-9mol/L promoted the proliferation of Sacc83 cells, 10-7mol/L of E2 showed the strongest effect, with an increase in cell proliferation by 34.4% (P<0.01) .2. Cell growth curves were measured by cell counting method and the cell population doubling time (DT) was calculated. The cell DTs in groups of control, 10-7, 10-8 and 10-9 mol/L of E2 DT (h) were 52.8, 43.7, 50.9 and 47.1, respectively. The shortening rates (%) of cell DT in E2 groups were 17.2, 3.6 and 10.8, respectively.3. Cell ndivision index was increased by by 33.3%, 40.9% and 17.8%, respectively, when the cells were treated with E2 at 10-7mol/L for 1, 2 or 3 days.4. Clonning efficiency of the cells treated with 10-7 mol/ L of E2 in soft argarculture was increased by39.0% (P<0.01) .5. Flow cytometer analysis displayed that S phase cells was increased by \23%, 3.3% and 9.7%, respctively after 12, 18 and 24h treatment with E2 at 10-7mol/L.6. Cells treated with E2 at 10-7mol/L showed increased expression of PCNA.7. Chicken embryo heart invasion test displayed that the ratio of the tumor cell invasion area in 10-7 mol/L E2 group and control group was 0.640+0.079 and 0.291 ?0.051 respctively (P<0.01).8. Cells treated with E2 10-7mol/L displayed higher expression of MMP-9 when observed with SDS- PAGE.9. Neoplasm developed in all nude mice after transplantation of 106 cells (8/8 in each group). The weight doubling time of the neoplasia was 76.5h in the mice treated with E2, whereas that in the control group was 61.5 h. Nude mice were sacrificed 15 days...
|
PDF Full Text Request