Statins Inhibit Cell Grouth Of NK/T-Cell Lymphoma

BackgroundMalignant lymphoma(ML) is a malignant tumor that derived from the lymph nodes and other lymph hematopoietic tissue, which can be divided into Hodgkin's lymphoma(HL) and non-Hodgkin's lymphoma(NHL).NK / T-cell lymphoma(NKTCL) is a type of NHL, and has high incidence of lymphoma.The distribution has certain geographical features, mainly in Asia. Generally speaking, the disease often involves tissues such as the nasal cavity, nasopharynx, tonsils, palate and other upper respiratory or digestive tract may also be involved, as well as the skin, testis, brain and bone marrow.Severe cases may appear Hemophagocytic syndrome. This type of lymphoma often invades blood vessels, and is capable of releasing perforin, so there will be a large number of perforated necrotic bone and soft tissues, and this could cause irreversible dysfunction of corresponding organ. The etiology of this disease is not clear.There are large numbers of studies indicating that it is closely associated with chronic EB virus infection. The disease has the characteristics of high degree of invasion, rapid progression, poor prognosis and short survival. Even if it achieved complete remission, it could easily recurrer in the latter period. Most prognosis in the early stage is well, but prognosis in the advanced period is poor. Currently, there is no standard first-line therapy, but in recent years, along with the progress of the study, the efficacy of DDGP chemotherapy that based on gemcitabine and pegaspargase toward NKTCL has been significantly improved. The NKTCL cells usually express the mRNA of drug resistance genes, and the product of this gene, such as P-glycoprotein can be combined with some chemotherapy drugs resistance.As a result, this may lead to unsatisfactory treatment effect, and reduced survival.Statins are reductase inhibitors of 3-hydroxy-3-methylglutaryl coenzyme A(HMG-CoA), which are capable of blocking the mevalonate pathway. The drug can not only reduce cholesterol level, but also protect the cardiovascular system. Mevalonate pathway is an important intracellular pathway to maintain cell metabolism activity, and the final products of this pathway include cholesterol and dolichol.Obstruction of this pathway can significantly affect the important biological functions of cells. In recent years, a number of laboratory and clinical data demonstrate that besides the effect of lowering cholesterol level, statins are also beneficial to the prevent and treat other diseases.For example, statins significantly improve the vascular endothelial function and stable the atherosclerotic plaque on vascular wall artery.It can also play against inflammatory and immunomodulatory, protect the nerve, reduce the progression of chronic kidney disease and prevent the cancer. The anti-cancer property of statins has become a hot research. Relevant researches on the anti-cancer property of statins exist in many tumor cells, such as breast cancer cells, liver cancer cells, lung cancer cells, prostate cancer cells and leukemia cells. Moreover, the existing animal experiments and prospective clinical trials have also confirmed that statins drugs can reduce the incidence of cancer, reduce tumor growth, slow its transfer speed, and reverse tumor resistance.In this study, NKTCL cell lines of NKL and YTS are regarded as research objects and different concentrations of simvastatin(SIM) and fluvastatin(FLU) are acted on the cell lines.CCK8 assay is used to test the growth and inhibition of cells, flow cytometry(FMC) is used to test the effect of drugs on cell cycle, and then AnnexinV-PI and Western blot methods are used to test the apoptosis of cells. In the following step, FLU and gemcitabine(GEM) will be combined.CCK8 and Western blot will be used to test the effects on cell proliferation and apoptosis to after the action of the two drugs. It is hoped that through this study, we can understand if statin drugs have the function of tumor suppression to NKTCL, and if it can be used together with clinical chemotherapeutics for antitumor, so as to find more effective methods of treating the disease. ObjectiveExplore the influence of fluvastatin and simvastatin on the inhibition of cell proliferation and induced apoptosis of NKTCL cell lines; in addition, discover the effect of combination therapy with gemcitabine. Materials and Methods1. Cell culture: YTS cultured in a 1640 medium containing 10% of fetal bovine serum, NKL in a medium containing 10% of fetal bovine serum and interleukin-2, both of which placed in a incubator with the temperature of 37 ?, containing 5 % of C02 for culture. Replace or add new medium every 2-3 days according to the cells condition.2. CCK8 assay: test the level of cell proliferation after the action of FLU and SIM with different concentrations, and the level of cell proliferation before and after the mixture of GEM and FLU.3. FMC test cell cycle: 48 hours of different concentrations of FLU(0,5,10,25,50uM) act on the two cell lines and the resistance of the cell cycle tested before and after the drug in according to the cell cycle kit.4. AnnexinV-PI Apoptosis detection: 48 hours of different concentrations of FLU(0,5,10,25,50uM) act on the two cell lines and the operation conducted in accordance with the relevant kit instructions.5. Western blot method to detect apoptosis-related proteins: the total cellular protein of both YTS and NKL cell lines was extracted after 48 hours action of SIM and FLU with the same dose, and then this method was used to detect the expression of Cleaved caspase-3 and Cleaved caspase-9.After 48 hours of combining FLU and GEM on YTS and NKL, this method was used to detect the expression of Cleaved caspase-3 and Cleaved caspase-9. Results1. SIM and FLU are capable of inhibiting the cell proliferation of YTS and NKL: by using CCK8 method, the growth of YTS and NKL cell lines is suppressed after the action of SIM and FLU, it could be found that different concentrations of SIM and FLU both have inhibiting effect on the two cell lines after 48 and 72 hours; in addition, NKL is sensitive to statins compared with YTS, FLU has stronger anti-tumor effect than SIM, and it is in a concentration-dependent manner.However, such drugs have no significant effect on cell viability of peripheral blood mononuclear cells of normal people.2. GEM and FLU combination has synergistic effect: the proliferation and inhibition levels of both YTS and NKL cell lines were significantly increased with the action of the combitation on YTS(49.81±2.01)% and on NKL(51.68±2.47)% compared with the same dose of GEM alone or FLU alone, and the inhibition rate tended to increase along with the increase of FLU concentration.3. FLU can lead to cell cycle arrest: after different concentrations of FLU(0,5,10,25,50uM) acting on the two cell lines, the proportion of cells in G0 / G1 phase was increased, while the proportion of cells in S phase was decreased, and the change was in a concentration-dependent manner.4. FLU is capable of inducing NKL and YTS apoptosis: after FLU(0,5uM, 25uM) acting on the two cell lines, cell apoptosis was gradually increased along with concentration increasing..5. Both FLU and SIM are capable of causing an increase in the expression of apoptosis-related protein, and have a synergistic effect with GEM: the expression of Cleaved caspase-3 and Cleaved caspase-9 was increased after FLU and SIM with the same dose act on both cell lines, and the cells were more sensitive to FLU than to SIM. Expression of apoptosis protein with the combination of FLU and GEM was more than any single drug. Conclusions:1. SIM and FLU inhibits the proliferation of NKTCL cell lines.2. FLU and GEM combination exerts a significant synergistic effect.