Berberine Promotes Mouse Breast Cancer Metastasis to Lung via Inducing Epithelial–mesenchymal Transition / 肿瘤防治研究

Objective To study the effect and mechanism of berberine (BBR) on the lung metastasis of mouse breast cancer via epithelial-mesenchymal transition (EMT). Methods CCK-8 and Transwell migration assays were utilized to investigate the proliferation and migration properties of breast cancer 4T1 cells after BBR treatment.Mouse 4T1-Luc cells were injected into mice under the fourth mammary fat pad, and the mice were then randomly divided into the control and BBR groups.The mice in the BBR group received daily intraperitoneal injections of BBR working solution and those in the control group were continuously intraperitoneally injected with the same volume of the solvent used to dissolve BBR powder.Tumor metastasis in the lungs of living mice was detected by using an in vivo imaging system.After 42 days of administration, lung metastasis was measured via microscopy and HE staining.Western blot analysis was used to examine the effects of BBR on the expression of EMT-related proteins (Vimentin and Snail) as well as the activation of the Akt and ERK signaling pathways. Results BBR significantly promoted 4T1 cell migration (P < 0.05).In vivo experiments showed that the number of lung metastases in the BBR group had significantly increased compared with that in control group (P < 0.05) as observed under microcopy and histological staining.Compared with the control group, BBR upregulated the expression levels of Vimentin and Snail as well as the phosphorylated levels of p-Akt and p-ERK (P < 0.05). Conclusion BBR may promote EMT and lung metastasis of breast cancer 4T1 cells by activating the expression of proteins in the p-Akt and p-ERK pathways.

Carcinosinumproduces phenotypic changes of 4T1 tumor cells in vitro

Considering that there are few published studies that specifically address the exclusive use of Carcinosinumin different potencies and, most of them focused on genotypic and clinical effects, the present study was proposed to identify possible phenotypic changes, including viability, expression of HER-2 and metastatic abilities, using 4T1 cells in vitroas a model. Carcinosinum was tested in different homeopathic potencies (12cH; 30cH; 200cH) mechanically prepared using sterile pure water. The time space between preparing the potencies and using them was 24 hours.The final dilutions were inserted into the culture medium in a volume equal to 10%, at the time of cell seeding. The same succussed vehicle used to prepare the drugs (70% ethanol) diluted 1:100 in sterile pure water was used as control. All treated cells were cultured in 25 mL flasks, with cell density of 5 x 105cells/mL. After 24 hours of treatment, cells were analyzed for apoptosis index using Annexin V kit and the Countess® system. The morphology of 4T1 cells was monitored by staining cell smears with hematoxylin-eosin and Giemsa methods. HER-2 expression was assessed by immunocytochemistry and metalloproteinase activity was assessed by zymography. The determination of the cytokine profile was performed using Cytometric Bead Array (CBA). The samples were evaluated in quadruplicate and the data were analyzed by one-way ANOVA. Carcinosinum30cH presented the highest apoptotic index and reduction of MMP-9-Pro expression; Carcinosinum200cH produced the highest positivity for HER-2 and no specific effect was seen after the treatment with Carcinosinum12cH. No change in cytokine expression was seen among treatments. We conclude that Carcinosinum30cH and 200cH can change phenotypic features important totumor development in vitro. The clinical meaning of these data deserves further investigation.

A hybrid bacterium with tumor-associated macrophage polarization for enhanced photothermal-immunotherapy

Remodeling the tumor microenvironment through reprogramming tumor-associated macrophages (TAMs) and increasing the immunogenicity of tumors via immunogenic cell death (ICD) have been emerging as promising anticancer immunotherapy strategies. However, the heterogeneous distribution of TAMs in tumor tissues and the heterogeneity of the tumor cells make the immune activation challenging. To overcome these dilemmas, a hybrid bacterium with tumor targeting and penetration, TAM polarization, and photothermal conversion capabilities is developed for improving antitumor immunotherapy in vivo. The hybrid bacteria (B.b@QDs) are prepared by loading Ag2S quantum dots (QDs) on the Bifidobacterium bifidum (B.b) through electrostatic interactions. The hybrid bacteria with hypoxia targeting ability can effectively accumulate and penetrate the tumor tissues, enabling the B.b to fully contact with the TAMs and mediate their polarization toward M1 phenotype to reverse the immunosuppressive tumor microenvironment. It also enables to overcome the intratumoral heterogeneity and obtain abundant tumor-associated antigens by coupling tumor penetration of the B.b with photothermal effect of the QDs, resulting in an enhanced immune effect. This strategy that combines B.b-triggered TAM polarization and QD-induced ICD achieved a remarkable inhibition of tumor growth in orthotopic breast cancer.

Effects of combination of pargyline and doxorubicin on proliferation and migration of tripple negative breast cancer 4T-1 cells / 中国药理学通报

Aim To study the combination of lysinespecifc demethylase 1 (lysine-specifc demethylase 1, LSD1) inhibitor pargyline and the chemotherapy drug doxorubicin on the proliferation, migration and invasion of murine triple negative breast cancer 4T-1 cells. Methods In vitro, the effect on the proliferation, invasion and migration of 4T-1 cells of the combination of these two drugs were detected with CCK-8 method, lactate dehydrogenase release test, Chou-Talay method, Scratch test, Transwell assay, Western blot and etc. Tumor-bearing mice were used to investigate the combined effect of these two drugs on the proliferation of 4T-1 cells in vivo. Results The combination of pargyline and doxorubicin effectively inhibited the proliferation, migration and invasion of 4T-1 cells. Compared with single drug group, the combination of these two drugs could significantly inhibit the proliferation of breast cancer and prolong the survival time of mice with triple negative breast cancer. Conclusions The combined application of pargyline and doxorubicin has a synergistic inhibitory effect on the proliferation, migration and invasion of mouse breast cancer 4T-1 cells, and has potential value for clinical treatment on triplenegative breast cancer.

Investigation of Mechanism of Tryptanthrin Inhibiting Breast Cancer in Mice Based on Proteomics / 中国实验方剂学杂志

Objective:A systematical study on the anti-breast cancer mechanism of tryptanthrin in breast cancer-bearing mice was done by Label-free proteomics. Method:UPLC-MS was used to detect the expressed-proteins of tryptanthrin inhibiting breast cancer in mice， chromatographic separation was achieved on the Ionoptics nano UPLC C18 column （0.075 mm×250 mm， 1.6 μm）， and gradient elution was performed with 0.1% formic acid aqueous solution-0.1% formic acid acetonitrile solution as mobile phase. Data acquisition was carried out in electrospray ionization （ESI） under the positive ion mode， the scanning range was m/z 100-1 700， MaxQuant 1.6.5.0 was used for database retrieval. Label-free proteomics with high resolution mass spectrometry was used to screen differentially expressed proteins between the model group of 4T1 breast cancer mice and oral administration group of tryptanthrin （100 mg·kg-1）. The proteomics of tryptanthrin against breast cancer was carried out. Result:A total of 3 997 proteins were identified in this proteomics research， and 2 911 proteins were quantifiable. A total of 750 differentially expressed proteins were identified between the model group and the tryptanthrin group， 286 proteins were up-regulated and 464 proteins were down-regulated. Gene ontology analysis showed that these differentially expressed proteins were mainly involved in biological processes of proliferation， cell migration， apoptosis， immunity， angiogenesis， inflammatory regulation， etc. Kyoto encyclopedia of genes and genomes pathway analysis further indicated that these proteins were mainly concentrated in T cell receptors， B cell receptors， Toll-like receptors， nuclear transcription factor-κB （NF-κB）， Ras proteins， interleukin-17， tumor necrosis factor， phosphatidylinositol 3-kinase/protein kinase B （PI3K-Akt）， mitogen-activated protein kinase （MAPK） and other signaling pathways. Conclusion:The differentially expressed proteins closely related to anti-breast cancer effect of tryptanthrin on 4T1 breast cancer mice are effectively screened out， including up-regulating proteins of leukocyte differentiation antigen 14 （CD14）， prostaglandin G/H synthase 2 （PTGS2）， E3 ubiquitin-protein ligase and down-regulating proteins of CD44， heat shock 70 kDa protein 1A （HSPA1A）， macrophage migration inhibitory factor （MIF）， NF-κB， ribosomal protein S6 kinase alpha-4 （RPS6KA4） and high mobility group protein B1 （HMGB1）. These findings suggest that tryptanthrin can inhibit breast cancer in mice mainly through regulating tumor inflammatory microenvironment.

Promotion effect of IL-4 and estradiol on growth of breast cancer cells in mice and its mechanism / 吉林大学学报(医学版)

Objective: To investigate the effect of interleukin-4 (IL-4) and estradiol on the biological behavior of breast cancer 4T1 cells of the mice, and to elucidate its mechanism. Methods: The 4T1 cells were cultured in vitro and added with different concentrations (0. 12. 5 . 25.0 . 50.0 and 100.0 fig • L 1 ) of IL-4 or estradiol (0. 6. 25. 12. 50. 25. 00 and 50.00 nmol • L ' ). The proliferation rate of the breast cancer 4T1 cells was measured by MTT method after treated for 72 h. The breast cancer 4T1 cells were divided into control group (without any treatment). IL-4 group (treated with 50. 0/ig • L 1 IL-4). estradiol group (treated with 12. 50 nmol • L 1 estradiol) and combination group (treated with 50. 0/ig • L 1 IL-4 + 12.50 nmol • L ' estradiol). MTT method was used to detect the proliferation rates of the breast cancer 4T1 cells in various groups, and flow cytometry was used to detect the percentages of the breast cancer 4T1 cells in different cell cycles in various groups, and Western blotting method was used to detect the expression levels of STAT6. p-ST AT 6. ERa. Erk. p-Erk. P70S6K. p-P70S6K. $6. and p-S6 in the breast cancer 4T1 cells in various groups. Results: Compared with 0 fig • L 1 IL-4 group, the proliferation rates of the breast cancer 4T1 cells in 25. 0» 50. 0 and 100. 0/ig • L 1 IL-4 groups were increased ( P< 0.05); compared with 0 nmol • L 1 estradiol groups, the proliferation rates of the breast cancer 4T1 cells in 12.50. 25. 00 and 50.00 nmol • L 1 estradiol groups were increased ( P<0.05). Compared with control group, the proliferation rate of the breast cancer 4T1 cells in IL-4 group was increased ( P-'CO. 05); compared with control group, the proliferation rate of the breast cancer 4T1 cells in estradiol group was increased ( P<0. 05); compared with IL-4 group or estradiol group, the proliferation rate of the breast cancer 4T1 cells in combination group was increased (P<0. 05). Compared with control group, the percentages of the breast cancer 4T1 cells at S phase and G/M phase in IL-4 group were increased (P∗C0. 05). and the percentage of the breast cancer 4T1 cells at G and Gi phases were decreased (P°-C0. 05); compared with control group, the percentage of the breast cancer 4T1 cells at S phase in estradiol group was increased ( P<0. 05). and the percentages of the breast cancer 4T1 cells at G and Gi phases were decreased (P<0.05). Compared with control group, the expression levels of ERa. p-Erk. p-P70S6K. and p-$6 in the breast cancer 4T1 cells in IL-4 group were increased ( P<0. 05). while the expression levels of p-$TAT6. ERa. p-Erk. p-P70$6K. $6. and p-$6 in the breast cancer 4T1 cells in estradiol group were increased (P<0.05); the expression levels of STAT6. p-$TAT6. ERa. p-ERK. p-P70$6K. and p-$6. in the breast cancer 4T1 cells in combination group were increased (P-C0. 05). Conclusion: The combination of IL-4 and estradiol can increase the expressions of IL-4 receptor (IL-4R) and estrogen receptor ( ER). and enhance the activation of Erkl. p70$6K kinase and phosphorylation of downstream $6 protein in the breast cancer 4T1 cells.

Preparation of sheddable PEG modified miRNA-complexing nanoparticles and its preliminary in vitro and in vivo study / 中国药科大学学报

@#This study was aimed to prepare sheddable PEG modified miRNA-complexing nanoparticles and investigate in vitro cellular uptake effect and in vivo distribution profile. The sheddable PEG material was synthesized through condensation. The sheddable PEG modified miRNA-complexing nanoparticles were successfully prepared by electrostatic interaction between gene vector and miRNA, and then ibuprofen was added to deshield PEG layer. The in vitro cellular uptake effect and in vivo distribution profile of nanoparticles were investigated on 4T1 model cells. As a result, the particle size of nanoparticles was 107. 7 nm and Zeta potential was 15. 8 mV. Compared to unsheddable PEG group, the cellular uptake effect by 4T1 tumor cells as well as the concentration on tumor regions was significantly improved in the sheddable PEG group. Results showed that this systen has a great potential application in the field of tumor treatment.

Establishment of mice breast cancer model in BALB/c mice with 4T1 cells / 中国免疫学杂志

Objective:To establish a model of mice breast cancer of 4T1 cells in BALB/c mice and choose the best model making method.Methods:Ninety mice were divided into three groups randomly ,with 30 in each group injected by 4T1 cells suspension of 1×106 ml-1 ,1×107 ml-1 ,1×108 ml-1 respectively.Each group of mice were randomly divided into two groups which were inoculated on the chest wall and lateral abdominal wall respectively.Tumor formation time ,tumor growth rate and the 8 week survival rate in each group were compared ,and pathological character was observed by C-erbB-2 immunohistochemistry staining.Results:Tumor growth rate in cells suspension of 1×107 ml-1 was high and grow steadily.Tumor growth rate wasn′t correlative with 4T1 cells injection in different parts.The result of C-erbB-2 immunohistochemistry staining was positive.Conclusion: Injection of 4T1 cells suspension with 1 ×107 ml-1 is the best way to male tumor model in three suspension.

Extracts of red yolk from duck eggs inhibit metastasis of human mammary carcinoma (4T1) cells in mice

The aim of the present study was to investigate the in vivo anti-metastatic activity of the red pigments of red yolk eggs laid by the ducks dieting on Potamogeton cripus L on the mammary carcinoma (4T1). The pigments were extracted with petroleum ether and acetone (2:1, v/v). BALB/c mice were divided into three groups (n=6), fed with the extracts at 150 mg/kg body weight (BW)/day (DEYE-H) or at 50 mg/kg BW/day (DEYE-L) and identical buffer without the extract (control group). The extracts were administered for 34 days. The treatment significantly inhibited the growth of orthotopical 4T1 tumour (DEYE-H vs control, 1:2; DEYE-L vs control, 2:3) and reduced the metastasis of tumour in the lungs (DEYE-H vs control, 4:7; DEYE-L vs control, 5:7), without statistical difference of body weight among the three groups.