Effects Of Resvertrol On The Proliferation Of Human Melanoma A375 Cells And The Relationship With The PI3K/Akt/mTOR Signaling Pathway

Background: Malignant melanoma(melanoma)is one of the most aggressive skin tumors.Its annual growth rate of 3%-7% makes it the fastest growing tumor.At present,there are many treatments for melanoma.Surgical resection is the main treatment.Other treatments include radiotherapy,chemotherapy,molecular targeted therapy,immunotherapy and so on.However,there are many problems such as large side effects,high drug resistance and poor effect on the metastases.Therefore,it is particularly important to seek effective drugs and methods for the treatment of this disease.Some natural plant extracts have wide application prospects in the treatment of tumors because of their wide distribution,easy access to raw materials,little toxic and side effects,low cost and unique pharmacological effects.Resveratrol is a natural extract with many biological activities.Its biological activities include anti-tumor,anti-inflammatory,anti oxidant,anticoagulant,antiviral,anti angiogenesis and immunomodulation.A large number of experiments showed that resveratrol could inhibit proliferation and induce apoptosis of murine melanoma,hepatocellular carcinoma,colon cancer,lung cancer,breast cancer and gastric cancer etc in vitro and vivo.But for human melanoma cells,the mechanism of resveratrol is rare.Further exploration of its possible mechanism is crucial for its clinical application.Phosphatidylinositol 3-kinase / protein kinase B/ mammalian target of rapamycin(PI3K/AKT/m TOR)as an important intracellular signaling pathways,which is shown in many human tumors such as malignant melanoma,gastric cancer,leukemia and so on.The abnormal activation of this pathway leads to the mutation of some components in the pathway,which leads to downstream regulation of cell cycle,cell proliferation,apoptosis and transformation,and the imbalance of related proteins,which leads to the occurrence and development of tumors.At present,there are many targeted drugs for this pathway,but most of them are still in the stage of study in vitro.Their effectiveness and toxic and side effects still need to be further explored.Lei Junrong has reported that resveratrol can through the PI3K/Akt/m TOR signaling pathway reduces myocardial cells in mice with cerebral ischemia reperfusion injury,Jiang H and other studies have shown that resveratrol inhibits the proliferation of glioma by downregulating the PI3K/Akt/m TOR signaling pathway.We think there are association between resveratrol and melanoma and PI3K/Akt/m TOR signaling pathway.The aim of this study is to investigate the effect of resveratrol on the proliferation of human malignant melanoma A375 cell line and its relationship with PI3K/Akt/m TOR signaling pathway,so as to provide a basis for clinical application of resveratrol in melanoma.Objective: Resveratrol is a polyphenol compound,which is mainly found in grapes and is considered to have antitumor effects.Objective to study the effect of resveratrol on the proliferation of A375 cells and its relationship with PI3K/Akt/m TOR signaling pathway,so as to provide experimental evidence for resveratrol treatment of melanoma.Methods:Respectively at the concentration of 0?mol/l(control group),25?mol/l,50?mol/l,100?mol/l and 200?mol/l of resveratrol treatment of human melanoma A375 cells after 24 hours,then cell morphology was observed under light microscope,changes of cell cycle were observed with flow cytometry using Annexin PI double labeling method,and changes of PI3K/Akt/m TOR relative protein PI3 K,Akt,p-Akt,p-m TOR and Changes of cyclin D1 and cell cycle regulator P21 proteins were detected by Western blot.Results: After treatment of A375 cells with different concentrations of resvertrol 24 h,With the increase of drug concentration,the inhibition rate of cell proliferation gradually increased.The cells in G0/G1 phase increased with the increase of resvertrol concentration;and the cells in S and G2/M show the opposite trend.The proportion of G2/M phase of A375 cells in the blank control group(0 ?mol/l)accounted for 23.18±2.14 %,the G2/M phase of the experimental group(25?mol/l?50?mol/l?100?mol/l ? 200?mol/l)was 19.73±1.92%,15.47±2.01%,11.61±1.82%,5.63±0.89%,respectively,P<0.05,differences are statistically significant.Western blot showed that the PI3K/Akt/m TOR relative protein PI3 K,p-Akt,p-m TOR protein levels to increase as drug concentration decreased,but there is no same trend in Akt protein;the protein level of cyclin D1 gradually decreases with the increase of drug concentration,however,the P21 was Contrary.Conclusion: Resvertrol can obviously block the cell cycle in G0/G1 phase and inhibit the proliferation of A375 cells,one of the possible mechanisms is related to the inhibition of the PI3K/Akt/m TOR pathway and is related to the regulation of the expression of cyclin inhibitor P21 and cyclin D1 protein.