Effects Of Doxorubicin Loaded Mesoporous Silica Nanoparticles On Proliferation,apoptosis And Autophagy Of Human Cervical Carcinoma Cells

Objective: To investigate the effects of doxorubicin(DOX)loaded mesoporous silica nanoparticles(MSNs)on proliferation,apoptosis and autophagy of human cervical carcinoma cells.Materials and Methods: HeLa and SiHa cells were chosed as objects of this study in order to present different histogenesis.MTT was used to test the cytotoxicity on HeLa and SiHa cells with DOX concentrations of 0.5,0.75,1,1.25,1.5 ?g/mL and 2.5,5,7.5,10 ?g/mL to determine the optimum loaded concentrations of DOX respectively.Adopting 1:1,1:2,1:4 and 1:8 as carrying proportions of DOX and MSNs.MTT was used to investigate the effects on cell proliferation.The cell apoptosis was analyzed by flow cytometry.Hoechst 33342 and MDC staining were applied to observe the cell apoptosis and autophagy.The variation of related proteins was tested by Western blot.Results:(1)Loaded concentrations of DOX: Based on the results of MTT,chosing 1 ?g mL-1 and 5 ?gmL-1 as loaded concentrations of DOX for Hela and SiHa cells respectively.(2)Effects of DOX loaded MSNs on cell proliferation: MTT assay showed a significant cytotoxicity between DOX-loaded MSNs groups and control groups in Hela and SiHa cells(P<0.05),and the effects were in a MSNs dose dependent manner.Meanwhile,the effects of inhibition on HeLa cells are more obvious than SiHa cells.With the increasing proportion of MSNs,morphological changes were more remarkable.(3)Effects of DOX loaded MSNs on cell apoptosis and autophagy: The images of Hoechst staining showed the exist of cell apoptotic characteristics in both cells,like nuclear condensation,and fragmented chromatin.The results of flow cytometry showed apoptosis in HeLa and SiHa cells.SiHa cells mainly manifested as early stage apoptotic cells while HeLa cells were late stage apoptotic cells and necrotic cells.The images of MDC staining showed the exist of autophagosomes in both cells.(4)The molecular mechanisms of DOX-loaded MSNs leaded to cell apoptosis and autophagy: The levels of p-Akt/Akt decreased in HeLa and SiHa cells.The protein expression of Bax/Bcl-2 and caspase-3 increased in both cells.The levels of P62 decreased and LC3 B increased in HeLa cells.ATG5 was upregulated in experiment groups except DOX group in HeLa cells.ATG5 in 1:8 DOX-loaded MSNs group and LC3 B in experiment groups except DOX group were upregulated in SiHa cells.The level of P62 decreased in SiHa cells.Conclusion:(1)DOX-loaded MSNs could inhibit the proliferation of HeLa and SiHa cells,and in a MSNs dose dependent manner.(2)DOX-loaded MSNs could lead to apoptosis and autophagy of HeLa cells and SiHa cells,The mainly different was SiHa cells manifested as early stage apoptotic cells while HeLa cells were late stage apoptotic cells and necrotic cells.The results showed a more suitable for human cervical squamous cell(SiHa)of DOX-loaded MSNs.(3)The mechanism of apoptosis could be the inhibition of PI3K/AKT pathway and changing the expression of caspase-3 and Bax/Bcl-2.The results of autophagy may due to the changing of key proteins P62,ATG5 and LC3 B of autophagy.