Influence of the growth and epithelial-mesenchymal transformation of breast cancer mediated by integrin subunit α3 transfection with ultrasound targeted microbubble destruction
Objective To investigate the influence on the growth and epithelial-mesenchymal transformation(EMT)of breast cancer by integrin subunit α3(ITGA3)transfection mediated with ultrasound targeted microbubble destruction(UTMD).Methods The expression of ITGA3 in breast cancer samples from the TCGA database was analyzed based on the GEPIA platform,and survival curves were drawn to evaluate the impact of ITGA3 expression on the survive of breast cancer patients.Breast cancer BT-549 cells were divided into LIP-NC group(liposome mediated negative control plasmid),LIP-ITGA3 group(liposome mediated ITGA3 overexpression plasmid transfected cells),UTMD-NC group(UTMD-mediated negative control plasmid transfected cells)and UTMD-ITGA3 group(UTMD-mediated ITGA3 overexpressed plasmid transfected cells).Cells in UTMD-NC and UTMD-ITGA3 groups were accepted ultrasonic irradiation(frequency 1 MHz,sound intensity 0.75 W/cm2,duration 45 s)during transfection.Cell morphology of UTMD-NC and UTMD-ITGA3 groups was observed by scanning electron microscopy.Cell viability was detected by MTT assay.Cell migration ability was detected by scratch assay,and cell invasion ability was detected by Transwell assay.Western blot analysis was performed to quantify the expression of ITGA3 and EMT-and STAT3 pathway-related proteins in BT-549 cells.A xenograft tumor model of breast cancer was established by subcutaneously injecting tumor cells into nude mice.After 35 d of transplantation,the nude mice were sacrificed and the xenograft tumors were harvested and weighed.Immunohistochemistry was used to detect the expression of ITGA3,N-cadherin and PCNA of xenograft tumor tissues in each group.Results Bioinformatics analysis showed that the expression of ITGA3 in breast cancer samples was significantly lower than that in normal breast samples(P<0.05).The low expression of ITGA3 was associated with the poor prognosis in breast cancer patients(HR=0.81,P=0.000 72).In vitro experiment showed that cells in the LIP-NC group were spherical under scanning electron microscopy,with no obvious pits or pores on the surface of the cell membrane.The cells in the UTMD-NC group showed no change in morphology,while with rough areas and small pits on the cell membrane.The cell viability,migration ability and invasion ability of UTMD-ITGA3 and LIP-ITGA3 groups were decreased compared with the control groups(UTMD-NC group and LIP-NC group),and the differences were statistically significant(all P<0.05).The cell viability,migration ability and invasion ability of UTMD-ITGA3 group were lower than those of LIP-ITGA3 group(all P<0.05).The relative expression of ITGA3 protein in the LIP-ITGA3 group cells was higher than in the LIP-NC group.The relative expression of ITGA3 protein in the UTMD-ITGA3 group was higher than in the UTMD-NC and LIP-ITGA3 groups,with statistically significant differences(all P<0.01).The relative expression levels of Vimentin,N-Cadherin,p-STAT3,c-Myc,CyclinD1 and PCNA proteins in the LIP-ITGA3 group and UTMD-ITGA3 group were significantly lower than those in the UTMD-NC group and LIP-NC group,while the relative expression level of E-Cadherin protein was significantly higher than that in the UTMD-NC group and LIP-NC group(all P<0.01).Additionally,the relative expression levels of Vimentin,N-Cadherin,p-STAT3,c-Myc,CyclinD1 and PCNA proteins in the UTMD-ITGA3 group were significantly lower than those in the LIP-ITGA3 group,and the relative expression level of E-Cadherin protein was significantly higher than that in the LIP-ITGA3 group(all P<0.05).In vivo experiment showed that the weight of nude mice xenograft tumors in the UTMD-ITGA3 group and LIP-ITGA3 group was significantly decreased compared with the UTMD-NC and LIP-NC groups,respectively,with statistically significant differences(all P<0.01).Moreover,the weight of nude mice xenograft tumors in the UTMD-ITGA3 group was significantly decreased compared with the LIP-ITGA3 group(P<0.01).The relative expression level of ITGA3 protein in the xenograft tumor tissues of nude mice in the UTMD-ITGA3 group was significantly higher than that in the LIP-ITGA3 group(P<0.01).The relative expression levels of N-cadherin and PCNA proteins in the xenograft tumor tissues of nude mice in both the UTMD-ITGA3 and LIP-ITGA3 groups were significantly lower than those in the UTMD-NC and LIP-NC groups(all P<0.01).Additionally,the relative expression levels of N-cadherin and PCNA proteins in the UTMD-ITGA3 group were significantly lower than those in the LIP-ITGA3 group(all P<0.01).Conclusion UTMD mediated ITGA3 overexpression plasmid transfection can effectively inhibit the growth and EMT of breast cancer,providing a new strategy for breast cancer treatment.
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