Studies On The Anti-tumor Effects Of The Two GPX Mimics And Their Action Mechanisms

Cancer is very harmful to people's health, and affects people's normal life. Recently, there is no effective way to prevent and cure cancer. In many researches of anti-tumor drugs and theories, the relationship between free radical biology and tumor prevention is one of the most important research field.GPX is a kind of selenium-containing protein which is first found in mammal. It plays its role in the form of sec, catalyzes GSH to break down the peroxide in human body. Molecular biology and genetics demonstrate that the occurrence and development of many diseases are correlated with ROS, such as diabetes, cataract, Keshan disease, angioocardiopathy, inflammation and cancer. At the same time, the GPX activity is declined to different extent. For these reasons, GPX is an expectable antioxidant drug with important biology meaning.Because native GPX has many disadvantages , such as large molecular weight , instability in vitro , limited availability ,short half life and difficult to express by gene engineering method , it is limited in medicinal application. So many scientists pay a great deal of attention to study artifical GPX mimics in order to make it exert similar biological function as native GPX .We construct the two artifical mimics of GPX, selenium-containing cyclodextrin (Se-CD) and selenium-containing 5-mer peptide (Se-5-peptide), which GPX activities are 7.4 U/μmol and 11.0 U/μmol, respectively, and study their biological function to inhibit caner. The experimental results indicate that they have a good ability to inhibit cancers. 1 Inhibition of the growth of mice H22 cells by two kinds of GPX mimics and its mechanism.The changes of H22 cell morphology and quantity were observed using inverted microscope. MTT method is used to examine the inhibition of H22 cell proliferation by the drug and measure the drug's effect on the culture, such as the content of MDA, and the GPX activity. The H22 cell cycle and apoptosis were observed by flow cytometry. RT-PCR method was used to detect the expression of apoptosis-related genes Bcl-2, bax, caspase-9, Caspase-3. We use electron microscope to observe the drug effect on the cell shape.The results show that the GPX mimics can obviously inhibit the growth of H22, and reduce the content of MDA in the cell culture, and increase the GPX activity of the cell culture. The mimics can also retard the cell cycle at G0/G1 phase and reduce the distribution of proliferating cell at S phase and the synthesis of DNA at S phase, inhibiting the proliferating activity of tumor cell. The mimics can also reduce the expression of Bcl-2 in the cell, increase the expression of Bax, accelerate the apoptosis of H22, increase the expression of Caspase-9 and Caspase-3, activate the cell apoptosis pathway. Therefore, the mimics can inhibit the growth of the cell. We can see the putrescence and cavitate of the cells by microscope.2 The anticancer effect of two kinds of GPX mimics on H22 bearing mice and its mechanism.We observe the effect of GPX mimics on H22 bearing mice, such as the animal behavior, skin, psychosis, appetite, drink and weight. And we also observe its effect on tumor weight, thymus exponent and spleen exponent. We observe the effect on spleen lymphopoiesis, on NK cell killing activity, and the effect of the mimics on MDA, NO and GPX activity in tumor bearing mice blood. The result has shown that GPX mimics can inhibit the multiplication of cancer cells in H22 bearing mice , decrease the content of MDA in the bearing mice blood , enhance the GPX activity in blood , increase the thymus index and spleen index of the mice , increase the swallow ability of the mice celiac TAMs (tumor-associated macrophages) and NO release in blood of TAMs, increase the multiplication activity of mice spleen T cell , increase the killing activity of mice NK cell , reduce the expression of apoptosis Bcl-2 gene in tumour cell , obviously increase the expression of Bax gene, increase the expression of caspase-9,caspase-3 in tumour cell.3 Inhibition of the growth of human SMMC-7721 cel by 2-SeCD and its mechanism.We use invert discrepancy microscope to observe smmc-7721 in vitro, flow cytometry to detect the influence of 2-SeCD on SMMC-7721, fluorescence microscope to observe the effect of 2-SeCD on SMMC-7721, electrophoresis to detect the DNA-ladder, RT-PCR to detect the c-myc expression.The results show that with the proper dose of 2-SeCD it can obviously inhibit the cell growth, accelerate the cancer apoptosis, and decrease the expression of the c-myc gene expression.4 Inhibition of the growth of human HT1080 cells by 2-SeCD and its mechanism.To study the effect of 2-SeCD on HT1080 cells and its mechanism , the 2-SeCD of different concentrations was added into the medium in which the cell was cultured. The changes of cell morphology and quantity were observed by phase contrast microscope. The inhibition of HT1080 cell growth by 2-SeCD was detected by MTT. The cell cycle and cell apoptosis were obsereved by flow cytometry. RT-PCR method was used to detect the expression of apoptosis-related genes Bcl-2 and Bax.The result shows that the growth of the HT1080 cells was significantly inhibited by 2-SeCD in a dose-dependent way. 2-SeCD can reduce the distribution of the proliferating cell at S phase and significantly up-regulate Bax gene expression in cell , but has no obviously influence on Bcl-2 gene expression.