| Objectives To investigate the effect of Gra-Epo anti-tumor and the mechanism of reducing liver injury caused by drugs through graphe-loaded epothilone(GrapheneEpothilone,Gra-Epo).Intervened nude mouse xenograft model of triple negative breast cancer.Methods 1 Human triple-negative breast cancer MDA-MB-231 cells were routinely cultured and subcultured.MDA-MB-231 cells were injected to nude mice from it'subcutaneous,and the nude mouse models of triple negative breast cancer were made.The tumor tissues were taken to verify models by HE staining,Estrogen Receptor(ER),Progesterone Receptor(PR)and Human epidermalgrowth factor receptor-2(HER2)were detected by immunohistochemistry to further verify tumor models.2 Forty-eight tumorigenic nude mice were selected and randomly divided into four groups: control group,paclitaxel(PTX)group,Epothilone(Epo)group and graphene-loaded epothilone(Gra-Epo)group.The mice of each group were Injected drugs in intraperitoneally once every two days at a dose of 0.3ml for each mouse,and the concentrations of drugs were 8mg/kg,4mg/kg and 4mg/kg,respectively.Continue administration for 3 weeks.3 The tumor tissues were taken after the mice were killed,and the diameter and circumference of tumors were measured and recorded.The microscopic changes of tumor tissues were observed under the microscope after HE staining.4 The protein of Cysteinylaspartatespecificproteinase-3(Caspase-3)and Vascular Endothelial Growth Factor(VEGF)in tumor tissues were detected by Western Blot and immunohistochemistry.5 The expression of glutathione(GSH),superoxide dismutase(SOD)and malondialdehyde(MDA)in liver homogenate were detected by enzyme-linked immunosorbent assay(ELISA).6 The pathological changes of liver in each group were observed after HE staining.7 Ultrastructural pathological changes of hepatocytes were observed under scanning electron microscope.8 Serum enzymatic indicators: alanine aminotransferase(ALT),aminotransferase(AST)and alkaline phosphatase(ALP)were measured by ELISA,respectively.9 The experimental data of each group were analyzed by SPSS18.0 software,and these data were analyzed by one-way analysis of variance.P<0.05 was considered statistically significant.Results 1 The nude mice grew well after MDA-MB-231 cells were inoculated in subcutaneous.The nodules of the subcutaneous transplanted tumor gradually increased and highlighted the surface of the skin at about 8 days,they are hard by touch and mobility were poor.HE staining and immunohistochemical staining confirmed that the tumors were triple negative breast cancer.2 The tumor inhibition rates of epomycin(Epo)group,graphene-loaded epothilone(Gra-Epo)group and paclitaxel group were 29.18%,41.63% and 20.33%,respectively.The results showed that the tumor inhibition rates of the three drug groups were compared with that of the control group in pairs,and the difference was statistically significant(P<0.05).3 The tumor tissues were stained by HE and observed under microscope: the cells of normal saline control groups had disordered for cells arrangement,and the volume of the cells were larger than than normal's.The morphology of the cells were irregularity.The cell structures for the tumor tissues of Epo group,graepo group and purple PTX group were disordered,and necrotic.4 The protein expression of caspase-3 and VEGF in the tumor tissues were detected by western blot and immunohistochemical staining: Western Blot results showed that the expression of caspase-3 protein were increased and the expression of VEGF were decreased for the three treatment groups in the tumors compared with that in the normal saline group.The expressions of caspase-3 protein were observed in the cytoplasm under microscope through immunohistochemical staining.Compared with the control group,the expressions of caspase-3 protein in the tumors were increased in the three treatment groups,and statistical analysis showed significant difference(P<0.05).VEGF expressed in the cytoplasm.Compared with the normal saline group,the expressions of VEGF were decreased in the three groups,and statistical analysis showed significant difference(P< 0.05).5 The expressions of enzyme indicators ALT,AST and ALP in liver function showed that the expression of ALT,AST and ALP were increased for Epo group,gra-epo group and PTX group.The difference between the three groups were statistically significant(P<0.05).6.The expressions of MDA were increased in the Epo group,GraEpo group and PTX group,and the expressions of GSH and SOD were decreased in the three groups,statistical analysis showed significant difference(P<0.05).Compared with the PTX group,the expressions of MDA in the Epo group and Gra-Epo group were decreased by 36.21% and 26.25%,respectively.Compared with the Epo group and the Gra-Epo group,the expressions of GSH in the PTX group were increased by 56.31% and 26.21%,respectively.Compared with in the paclitaxel group,the expressions of SOD in Epo group and Gra-Epo group were increased by 25.49% and 40.16%,respectively.Statistical analysis showed that the difference was statistically significant(P<0.05).7 HE stained for liver tissues: the liver tissues of the control group had intact liver lobule and hepatic sinus physiological structure,and the morphology of liver cells was complete and order.Most of the hepatocytes in the PTX group were swollen with varying degrees of cytoplasm,and the hepatic lobule structure was destroyed to varying degrees.The degree and number of hepatocyte injury in the epomycin(Epo)group and the graphene-loaded epothilone(Gra-Epo)group were lower than those in the paclitaxel group.8 Electron microscopic observation showed that the nucleus of hepatocytes in the control group was normal,no morphological changes and the nuclear membrane was intact.A large number of mitochondria are distributed in the cytoplasm and are structurally ordered.In the paclitaxel group,hepatocytes showed morphological changes,the number of mitochondria in the cytoplasm decreased significantly and the structure was absent,and the endoplasmic reticulum was arranged in a disorderly dispersed manner.Compared with the paclitaxel group,the Epo group(Epo)group and the graphene-loaded epothilone(Gra-Epo)group had lighter nuclear structure and cytoplasmic organelle structure changes,and the nuclear membrane defect was lighter.The number of mitochondria is reduced to a lesser extent.Conclusions 1 Epothilone has significant inhibitory effect on the tumor in the nude mice model of human triple negative breast cancer MDA-MB-231.The antitumor effect of graphene as its carrier is enhanced,and its mechanism is related to the increase of caspase-3 and decrease of VEGF in triple negative breast cancer tissues.2 After loading with graphene as a carrier,ebomycin can maintain the stability of liver biochemical indicators and reduce drug-induced liver damage.The mechanism is related to the good bioavailability of graphene.Figure27;Table6;Reference 117... |
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