The Effects Of Dimercaptopropanesulfonic Acid(DMPS) On The Cellular Biomethylation Of Arsenic And The Effects Of Selenium On Arsenic And Platinum Induced In HepG2 Cells

Organic arsenic(iAs),as a kind of carcinogen,has a strong toxic effect.It is closely related to the occurrence and development of a variety of diseases,including cancer.For a long period,we use arsenic as anticarcinogen,it made a success in APL(Acute promyelocytic leukemia)therapy.However,its toxicity will damage the organism and bring physical sufferings to the patients.Therefore,the research of reducing the toxicity of arsenic has an important significance for alleviating the patients’ pain availably.At present,platinum-based antitumor drugs are the most commonly used in clinical for cancer treatment.Cisplatin has been used for more than three decades in standard chemotherapy regiments.However,it has been greatly hampered by drug resistance and severe side effects,like nephrotoxicity and neurotoxicity.Synergistic anticancer as a new method was proposed recently to solve the above problems,simultaneously improve the efficiency of cancer treatment.Selenium is an essential dietary element for humans and is regarded as a protective agent against cancer.Moderate selenium intake would reduce the risks of cancer.Although the mode of anticancer action of selenium is not fully understood yet,several mechanisms,such as antioxidant protection by selenoenzymes,specific inhibition of tumor cell growth by Se metabolites,modulation of cell cycle and apoptosis,and effect on DNA repair have all been proposed.Moreover,recent studies suggest that Se has a potential to be used not only in cancer prevention but also in cancer treatment where in combination with other anticancer drugs to increase efficacy of cancer therapy.Therefore,to exploit the effects of Se on treatment of cancer in combination with other drugs is of great significane for human health and cancer therapy.This thesis is mainly divided into the following three aspects on the basis of the work our research group,which is about the biological effects of arsenic and selenium compounds in HepG2 cells:1.Based on HepG2 cells,we used high performance liquid chromatography-inductively coupled plasma-mass spectrometry(HPLC-ICP-MS)to analysis the effects of DMPS on the content of As3+ and biomethylation of As3+ and MMA3+in HepG2 cells.Meanwhile,we investigated the expression of human arsenic(Ⅲ)methyltransferase(hAS3MT)by western-blot as well as the accumulation of cellular reactive oxygen species(ROS)and apoptosis by flow cytometry influenced by DMPS in HepG2 cells.The results suggested that DMPS at low concentrations dramatically decreased the content of arsenic in HepG2 cells and inhibited the cellular methylation of As3+.We also found that DMPS competitively coordinated with As3+ and MMA3+ to inhibit the up-regulation of arsenic on the expression of hAS3MT and block the enzymatic methylation.Moreover,DMPS eliminated arsenic-induced accumulation of ROS,which might contribute to the antidotal effects of DMPS on arsenic toxicity.2.We select HepG2 cell to explore the biological effects of selenium on arsenic in cells.At first,we detected the activity of HepG2 cells by MTT induced by selenium and arsenic.Then,we analyzed the apoptosis rate,level of reactive oxygen species(ROS)by flow cytometry.Finally,we made use of RT-PCR and Western-blot to investigate the apoptosis associated gene expression level(Hmox1、Bax、Bcl-2、Caspase-3、hAS3MT)and protein expression level(Bax、Bcl-2、Caspase-3、NF-κB).The results show that the effects of selenium and arsenic on HepG2 cells were concentration dependent and time dependent.Selenium could significantly enhanced apoptosis rate caused by arsenic at high concentration.In addition,selenium and arsenic could increase ROS accumulation to promote apoptosis.RT-PCR and Western-blot demonstrated that selenium down regulation NF-κB and inhibit Bcl-2 expression,as a result of cell death.3.To clarify the biological effects of selenium on cisplatin induced in HepG2 cells,we conducted MTT assay to analyze the activity of HepG2 cells primarily.Then cell apoptosis and ROS level were tested by flow cytometry.Meanwhile,we investigate the expression level of gene-associated with apoptosis including Hmoxl、Bax、Bcl-2、Caspase.Our study results prove that selenium and cisplatin could induce HepG2 cell apoptosis in concentration-and time-dependent manners.The apoptosis index induced by cisplatin is related to selenium concentration.At low concentration,they demonstrated antagonistic effects,while selenium played a synergism role with cisplatin at high concentration.On the other hand,selenium and cisplatin could make intracellular ROS rise and promote cell apoptosis.RT-PCR indicated Caspase-3 was down-regulated by selenium in HepG2 cells.