| Objective: Matrix metalloproteinases (MMPs), a family of zinc-and calcium-dependent enzymes, mediate the invasion and metastasis properties of most malignant rumors through their ability to degrade most ECM components. Previous studies revealed that MMP-2 (Mr 72,000 gelatinase A) and MMP-9 (Mr 92,000 gelatinase B) are two important in MMPs, which belong to gelatinases, are critically associated with ovarian carcinomas. The induction and regulation of MMPs become the hot point in the science research in last few years. Substantial evidence has been accumulated that cytokines and growth factor play a critical role in MMPs expression. Studies indicate that the expression of gelatinases are regulated by EGFs bFGPN TGF et al. However, relative little is known about the regulation of MMP-2 MMP-9 by IGFs. Today there is no still report regard to the gelatinases regulated by the insulin-like growth factor-II (IGF-II). To further investigate the effect of cytokines and growth factor on the expression of MMPs, especially including MMP-2 and MMP-9 in local invasion and distant spread of ovarian cancer, we first measure thechange of gelatinases in three ovarian cancer cell lines treated with IGF-II.Method: Cell culture seeded on certain cell density was maintained in 24-well cell culture plates for 48 hours with 37 , and incubated 18 hours in serum-free medium following treatment with various concentrations of IGF-II. Condition media were collected , cleared by centrifugation, and performed gelatin zymography to assess MMP-2 and MMP-9 proteinase activity. Total RNA was extracted from the cells for reverse transcriptase-polymerse chain reaction (RT-PCR). In addition, we evaluated the proliferative effect of IGF-II on ovarian cancer cell lines.Result: (1) By gelatin zymography, we found that SKOV3 secreted MMP-9, not MMP-2; human ovarian cancer cell AO alone produce MMP-2; Neither MMP-2 nor MMP-9 was detected in the medium of 3AO. (2) The expression of MMP-2 mRNA, MMP-9 mRNA was analyzed by RT-PCR. MMP-9 mRNA not MMP-2 mRNA was expressed in SKOV3; AO and 3AO expressed both MMP-2 mRNA and MMP-9 mRNA. (3) A representative gelatin zymography showed that SKOV3 cells secreted only gelatinases with estimated molecular size of 92 kDa. IGF-II (10ng/mK 50ng/ml 100ng/ml) dose-dependently increased the secreted activity of 92-kDa gelatinase. Densitometric analysis showed that stimulation by IGF-II at the concentration of 10ng/ml 50ng/ml, 100ng/ml caused47.83+2.54%, 80.17+2.52%, 74.03+4.21%, increase as compared to control(p<0.05). In addition, MMP-2 undetected in either presence or absence IGF-II. The gelatin zymography of AO indicated small MMP-2 was secreted from conditoned medium, and its protein sythesis was promoted by IGF-II. From the concentration-effect curve, IGF-II increased the expression of MMP-2 by 17.45+2.54%, 94.79+11.63%, 40.68+7.78% at corressponding concentration of 10ng/ml 50ng/ml 100ng/ml, as compared to control(p0.01). Cell AO did not display 92kDa proteolytic band induced by IGF-II. We undetected MMP-2 and MMP-9 secretion in 3AO cell line either before or after IGF-II treatment. Moreover, the active form of two gelatinases was not detectable in the all of condition medium. (4) To investigate the induction of MMP-2 mRNA,MMP-9 mRNA after treatment with IGF-II (lOng/mK 50ng/mK lOOng/ml) for 18 nous, we conducted semiquantitive RT-PCR analysis. The expression of MMP-9 mRNA was observed in SKOV3 either before or after treatment with IGF-II. This find is not in line with result detected by gelatin zymography. By RT-PCR, we also observed the expression of MMP-9 mRNA after induction by IGF-II in AO and 3AO cell lines. The finding showed that the expression of MMP-9 mRNA was not further promoted by IGF-II. In addition, the expression of MMP-2 mRNA in AO or 3AO was further detected neither before nor after co-culture with IGF-II. (5) We examined the effect of IGF-IIon cell proliferation at the concentration of ranges from 10 ng/ml to 100 ng/ml. The cell nu...
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