| Lymphoma is a malignant clonal disease that originates from hematopoietic stem cells.This study has shown that realgar NPs can inhibit a variety of tumor cells,especially leukaemia.However,realgar has disadvantages of high dose and low bioavailability in clinical practice.The clinical treatment of lymphoma with realgar has been reported frequently,but the research on its mechanism is still unclear.Objective In our previous study,we found that realgar NPs has a strong inhibitory effect on B-cell lymphoma Raji cells at the cell level,and the dosage is obviously lower than that of crude realgar.But the research on the mechanism of action is not in-depth.This study with the same dose of crude realgar as control,aims to further explore the molecular mechanisims of realgar NPs on the apoptosis and autophagy of Raji cells.It provides a theoretical basis for the clinical application of realgar NPs in the treatment of lymphoma.Methods In this experiment,the cells were divided into blank group and drug administration group?crude realgar and realgar NPs?.An optical microscope was used to observe the growth of Raji cells under the action of crude realgar and realgar NPs.Cell activity was detected by CCK-8 experiment.The morphology and nucleus of Raji cells were observed by fluorescence microscopy.Flow cytometry was used to detect the apoptosis of Raji cells.Western blot was used to study the apoptosis and autophagy of Raji cells from the perspective of molecular biology.Results From the results of the CCK-8 experiment,compared with the blank group,the proliferation ability of Raji cells was inhibited for 24h by using crude realgar and realgar NPs at the concentrations of 1?g·m L-1and 5?g·m L-1,and the inhibitory effect depended on the concentration.The inhibitory effect of 5?g·m L-1realgar NPs group was more significant.The Fluorescence microscopy results showed that the Raji cells in the blank group were round.Compared with the blank group,the number of Raji cells in crude realgar group was not significantly different from that of the blank group.Compared with the blank group and the crude realgar group,Raji cells in the realgar NPs group were inhibited in cell proliferation,the number of Raji cells was reduced,and nuclear shrinkage occurred.The central part of the nucleus was significantly highlighted.With the increase of drug concentration,the stronger the inhibition of cell proliferation,the central part of the nucleus was more prominent.The results of flow cytometry showed that crude realgar and realgar NPs can induce apoptosis phenomenon in Raji cells,its ability to induce the apoptosis is concentration-dependent.In the case of drug effect time is 24 h,with a concentration of 1?g·m L-1realgar,the total apoptotic rate of Raji cells treated with the crude realgar group was 6.7%,the rate of apoptosis of Raji cells treated with realgar NPs group was 8.6%;Raji cells were treated with 5?g·m L-1realgar,the total apoptotic rate of Raji cells treated with the crude realgar group was 7.5%,the rate of apoptosis of Raji cells treated with realgar NPs group was 22.6%.The total apoptosis rate of blank cells was 5.0%?the realgar NPs vs.the crude realgar,P?0.01?.Western blot results showed that after the Raji cells were treated with crude realgar and realgar NPs for 24hours,compared with the blank group,the expression of pro-apoptotic protein Bax increased gradually and and the expression of anti-apoptotic protein Bcl-2 increased.Compared with the blank group,the expression content of autophagy-related protein Beclin1 increased gradually with the increase of drug concentration,while the expression of P62 protein showed the opposite trend.Conclusion Realgar NPs can inhibit the proliferation of Raji cells in a concentration and time-dependent manner.Realgar NPs can induce apoptosis of Raji cells,which may be achieved by mitochondrial pathway.At the same time,realgar NPs can also induce autophagy in Raji cells by up-regulating Beclin-1 protein expression and down-regulating P62 protein expression. |
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