Effects of live attenuated measles vaccine Hu191 strain on epithelial mesenchymal transition,proliferation and migration of 4T1 breast cancer cells / 中国生物制品学杂志

@#Objective To investigate the effects of live attenuated measles vaccine Hu191 strain(MV-Hu191)on epithelial mesenchymal transition(EMT),proliferation and migration of 4T1 breast cancer cells.MethodsCCK-8 and clone formation assay were used to analyze the effect of MV-Hu191 on the proliferation of 4T1 cells;The effect of MV-Hu191 on 4T1cell migration was analyzed by cell scratch test;The expression of EMT pathway proteins(MMP-2,MMP-9,E-cadherin)in4T1 cells was detected by Western blot;4T1 tumor-bearing mouse model was established in female BALB/c mice. The model mice were divided into control group(PBS),MV-Hu191(1 × 106TCID50)group and paclitaxel group(15 mg/kg),with 10 mice in each group,and injected into tumor at the dosage of 100 μL every 2 d for 5 times. At 28 d after administration,the effects of MV-Hu191 on survival time,tumorigenicity and metastasis in vivo were observed;The pathological characteristics of lung tissue and tumor tissue were observed by HE staining under microscope;The expression of EMT pathway proteins(MMP-2,MMP-9 and E-cadherin)in tumor tissue was detected by immunohistochemical staining.Results The results of in vitro experiment showed that,compared with the control group,MV-Hu191 inhibited the proliferation and migration of 4T1 cells(F = 2. 811 and 13. 535,P = 0. 001 and 0. 002,respectively),down regulated the expression of MMP-2 and MMP-9(F = 45. 433 and 9. 744,P = 0. 011 and 0. 038,respectively),and up regulated the expression of Ecadherin(F = 7. 001,P = 0. 032);The results of in vivo experiment showed that MV-Hu191 significantly prolonged the survival time of tumor-bearing mice,and decreased the tumor quality(F = 8. 301,P = 0. 003)and the number of pulmonary nodules metastasis compared with the control group(F = 33. 792,P = 0. 000);MV-Hu191 treated tumor tissue gap was small,the cells were round,and the alveolar contour was clearly visible;The expression of MMP-2 and MMP-9 in MVHu191 treated tumor tissue decreased significantly(F = 6. 705 and 9. 047,P = 0. 028 and 0. 023,respectively),while the expression of E-cadherin increased significantly(F = 3. 468,P = 0. 039).ConclusionMV-Hu191 signi-ficantly inhibits the proliferation and migration of 4T1 breast cancer cells,antagonizes the tumorigenicity and lung meta-stasis of 4T1 tumorbearing mice,and prolongs the survival time of mice. The possible mechanism of MV-Hu191 against breast cancer is closely related to the regulation of EMT pathway protein expression.

Possible mechanism of 191 measles attenuated live vaccine strains inhibiting the proliferation of triple negative breast cancer MDA-MB-231 cells based on proteomics technology / 中国肿瘤生物治疗杂志

@#[摘 要] 目的：基于蛋白质组学技术探讨麻疹减毒活疫苗191株（MV-Hu191）在体内外对三阴性乳腺癌MDA-MB-231、4T1细胞的影响及其作用机制。方法：采用CCK-8法检测MV-Hu191对MDA-MB-231和4T1细胞增殖的影响；液相色谱-质谱联用技术分析MV-Hu191处理对MDA-MB-231细胞中蛋白质谱的影响，多重数据库筛选蛋白质谱中的典型差异蛋白质并进行GO、KEGG、亚细胞定位与功能注释。瘤内注射1×106 TCID50 MV-Hu191干预4T1细胞移植瘤模型小鼠，流式细胞术检测小鼠脾组织中T细胞亚群，ELISA法检测小鼠血清TNF-α和IL-6含量。结果：体外实验结果表明，MV-Hu191具有抑制MDA-MB-231和4T1细胞增殖的作用，差异均具有统计学意义（P<0.01）。蛋白质组学分析结果显示，MV-Hu191作用MDA-MB-231细胞后明显上调蛋白质有38个、下调有12个；差异表达的蛋白质主要参与细胞黏附、信号受体激活、细胞代谢、应激反应等生物学过程，22个差异蛋白质亚细胞定位位于细胞外，KEGG功能分类显示与免疫调节功能相关的差异蛋白质最多且均为上调蛋白，包括C4A、C8B、SERPINF2、A2M、SERPINC1、CTSB、SERPING1、C5；PPI预测发现免疫相关差异蛋白与CD4、CD8、TNF-α及IL-6相互关联。体内实验结果显示，MV-Hu191干预组小鼠脾组织中CD4+ T细胞数量略高于对照组，但差异无统计学意义（P>0.05），CD4+/CD8+ T细胞比值明显高于对照组（P<0.05），血清TNF-α和IL-6含量显著上升（均P<0.01）。结论：MV-Hu191显著抑制MDA-MB-231、4T1细胞增殖及拮抗4T1细胞荷瘤小鼠成瘤性，其机制可能是MV-Hu191通过激活免疫效应分子实现抗肿瘤作用。

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.

The anti-tumor activity of andrographolide on breast cancer of mice and its mechanism / 中国药理学通报

Aim To evaluate the effects of androgra- pholide ( Andro) on suppressing tumor growth and improving mitochondrial function on mouse breast cancer and explore its mechanism.Methods MTT assay was performed to measure the effect of Andro on the growth capacity of mouse breast cancer cell line 4T1.Mice were treated with Andro, then tumor volume measured, organ index calculated and Hematoxylin - eosin ( HE ) stained to detect the inhibitory effect of Andro on tumors.Lactate assay kit was used to detect the lactate level in mouse serum to check glycolysis discrepancy.To illustrate the transformation in oxidative phosphorylation ( OXPHOS) , Acetyl coenzyme A ( Acetyl-CoA ) assay kit was used to ascertain Acetyl-CoA content level , Western blot was used to detect the protein expression of pyruvate dehydrogenase ( PDH) , and ATP assay kit was used to ascertain ATP content level.The mitochondria functions were analyzed: oxygen ( ()2 ) consumption was measured by Clark oxygen electrode, and mitochondrial membrane potential was detected by JC-1 staining.Results Andro could effectively inhibit the proliferation of 4T1 cells and the growth of tumors, and had no significant damage on normal organs.An- clro reduced serum lactic acid content, indicateing that Andro inhibited the process of glycolysis.The expression of PDH, content of acetyl-CoA and ATP content increased with the increase of Andro concentration.It showed that Andro could up-regulate oxidative phosphorylation.In addition, the ()2 consumption and mitochondrial membrane potential increased in 4T1 cells, indicating that Andro could recover mitochondrial function.Conclusions Andro can inhibit the growth of mouse breast cancer, and its mechanism may be related to the inhibition of glycolysis level, restoration of OXPHOS and improvement of mitochondrial function.

Immunomodulatory effect of adriamycin in mouse 4T1 model of breast cancer / 生物工程学报

To explore the immunomodulatory effect of adriamycin on 4T1 breast cancer. We used a tandem mass tag-based quantitative proteomic method to detect differential proteins in breast cancer tissues, and multiple bioinformatics databases to analyze the differentially expressed proteins in the proteome. Also, we used enzyme-linked immunosorbent assay to detect the effects of adriamycin on helper T cells 1 and 2 in breast cancer tissues, and flow cytometry to detect CD4+ T cells, CD8+ T cells and regulatory T cells. We discovered the immunomodulatory targets of adriamycin in differential proteins. In total 170 differential proteins were significantly up-regulated, whereas 58 were markedly down-regulated. In addition, 73 proteins were involved in immune regulation. Kyoto encyclopedia of genes and genomes enriched important protein pathways related to cytokines and factor receptors, interleukin 17 pathway and cancer transcriptional regulatory pathways. These pathways and important differential proteins related to immunomodulatory functions were ultimately regulated by adriamycin on CD4+ T cells, CD8+ T cells and regulatory T cells, thereby affecting the prognosis of breast cancer. Moreover, adriamycin significantly increased interleukin 2, CD4+ T and CD8+ T (P<0.01) and markedly reduced regulatory T cells (P<0.05). The function of adriamycin against triple-negative breast cancer was closely related to the immunoregulation process of the differential proteins Ighm, Igkc, S100A8, S100A9 and Tmsb4x. Adriamycin could regulate the content of helper T cells 1 cytokines, CD4+ T and CD8+ T lymphocytes in breast cancer and reduce the number of regulatory T cells to produce immunomodulatory effects.

Reactive oxygen species and senescence modulatory effects of rice bran extract on 4T1 and NIH-3T3 cells co-treatment with doxorubicin

Objective:To determine the effect of rice bran extract (RBE) in combination with doxorubicin on 4T1 triple-negative breast cancer cells and NIH-3T3 cells. Methods:RBE was obtained by maceration with n-hexane. The phytochemical profile of RBE was observed using high-performance liquid chromatography. Cytotoxic effect of RBE was evaluated through MTT assay. In addition, flow cytometry was used for cell cycle and apoptosis analysis. Cellular senescence was observed using SA-β-Gal assay and intracellular reactive oxygen species (ROS) levels were evaluated using DCFDA staining. The pro-oxidant property of RBE was also evaluated through 1-chloro-2,4-dinitrobenzene spectrophotometry and molecular docking. Results:RBE was obtained with a yield of 18.42％ w/w and contained tocotrienols as the major compound. RBE exerted no cytotoxic effect on 4T1 and NIH-3T3 cells. However, RBE in combination with doxorubicin decreased 4T1 cell viability synergistically (combination index<0.9) and induced apoptosis and senescence on 4T1 cells. RBE significantly decreased senescence in doxorubicin-treated NIH-3T3 cells. Additionally, RBE did not increase ROS levels in doxorubicin-treated 4T1 cells. Meanwhile, the combination of RBE and doxorubicin reduced ROS levels in NIH-3T3 cells. RBE significantly reduced glutathione-S-transferase activity and alpha-tocotrienol interacted with glutathione-S-transferase in the glutathione binding site. Conclusions:Rice bran may be used as a co-chemotherapeutic agent to improve the therapeutic effectiveness of doxorubicin while protecting against the cellular senescence effects of doxorubicin on healthy cells.

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.

Targeting peptide-decorated biomimetic lipoproteins improve deep penetration and cancer cells accessibility in solid tumor

The limited penetration of nanoparticles and their poor accessibility to cancer cell fractions in tumor remain essential challenges for effective anticancer therapy. Herein, we designed a targeting peptide-decorated biomimetic lipoprotein (termed as BL-RD) to enable their deep penetration and efficient accessibility to cancer cell fractions in a tumor, thereby improving the combinational chemo-photodynamic therapy of triple negative breast cancer. BL-RD was composed of phospholipids, apolipoprotein A1 mimetic peptide (PK22), targeting peptide-conjugated cytotoxic mertansine (RM) and photodynamic agents of DiIC18(5) (DiD). The counterpart biomimetic lipoprotein system without RM (termed as BL-D) was fabricated as control. Both BL-D and BL-RD were nanometer-sized particles with a mean diameter of less than 30 nm and could be efficiently internalized by cancer cells. After intravenous injection, they can be specifically accumulated at tumor sites. When comparing to the counterpart BL-D, BL-RD displayed superior capability to permeate across the tumor mass, extravasate from tumor vasculature to distant regions and efficiently access the cancer cell fractions in a solid tumor, thus producing noticeable depression of the tumor growth. Taken together, BL-RD can be a promising delivery nanoplatform with prominent tumor-penetrating and cancer cells-accessing capability for effective tumor therapy.

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.

Effect of Panax Notoginseng Saponins on Breast Cancer Cell Line 4T1 in Tumor-bearing Mice Through MEKK1/SEK1/JNK1/AP-1 Pathway / 中国实验方剂学杂志

Objective:To explore the potential mechanisms of Panax Notoginseng Saponins （PNS） on growth inhibition of breast cancer cell line 4T1 in tumor-bearing mice by investigating the mitogen-activated protein kinase kinase kinase 1 （MEKK1）/stress activated protein kinase （SAPK）/extracellular regulated protein kinases （Erk） Kinase （SEK1）/c-Jun N-terminal kinase 1 （JNK1）/activator protein-1 （AP-1） signaling pathways. Method:The 4T1 breast cancer mice model was established. Forty-eight mice with successful modeled and randomly divided into the low， medium and high-dose PNS groups （10， 20， 40 mg·kg-1） and the model control group （12 mice in each group）. The PNS groups received intraperitoneal injection with dosage of 10 mL·kg-1， while the controlled group was given the same dosage of saline. After administration with PNS for 28 days， tumor tissues were isolated， weighed， sliced and homogenized. Tumor cell apoptosis was detected by TdT mediated-dUTP nick end labeling （TUNEL） staining. The mRNA expressions of MEKK1， SEK1， JNK1 and AP-1 in tumor tissue were detected by Real-time polymerase chain reaction（Real-time PCR）. The protein expressions of MEKK1， SEK1， JNK1 and AP-1 in tumor tissue were detected by immunofluorescence staining and Western blot. Result:Compared with model group， the tumor weights of medium-dose and high-dose PNS groups were decreased significantly （P<0.05）. TUNEL staining showed that the number of apoptotic tumor cells increased with the rise of dosage of PNS （P<0.05）. The medium-dose and high-dose PNS groups showed a significant increase in the mRNA expressions of MEKK1， SEK1， JNK1 and AP-1 as well as the protein expressions of MEKK1， SEK1， JNK1 and AP-1 in tumor tissues （P<0.05）， with statistically significant differences （P<0.05）. Conclusion:PNS could inhibit the tumor growth of breast cancer cell line 4T1 in tumor-bearing mice， which may be related to the activation of MEKK1/SEK1/JNK1/AP-1 signaling pathways.

Avaliação dos efeitos da Suplementação do Ácido Graxo ômega 3 em modelo de tumor de mama murino e mucosite induzida por 5-Fluouracil / Evaluation of the effects of Omega 3 Fatty Acid Supplementation in a model of murine breast tumor and mucositis induced by 5-Fluouracil

O ômega 3 é um ácido graxo poliinsaturado utilizado como agente terapêutico no tratamento de diversas doenças. Evidências sugerem efeito anti-inflamatório com sua utilização. A escolha do tratamento eficiente que minimize os efeitos adversos do tratamento oncológico tem sido desafiadora para a prática clínica. Dessa forma, ele seria interessante na prevenção e / ou tratamento da mucosite, uma inflamação que acomete as mucosas da boca ao ânus, decorrente da utilização de quimioterápico e radioterápico. As consequências deste agravo vão desde a perda de peso, anorexia e odinofagia, ao aumento do risco de translocação bacteriana, em virtude do aumento da permeabilidade intestinal (PI). Tais complicações podem levar à interrupção do tratamento quimioterápico até que o paciente se restabeleça. Trabalhos anteriores do nosso grupos de pesquisa demonstraram que o ômega 3 foi capaz de atenuar a perda de peso e reduzir a PI pela regulação da apoptose de células intestinais. Entretanto, possíveis efeitos da suplementação do ômega 3 no tumor não foram investigados. O presente estudo avaliou o efeito deste composto na mucosite induzida por quimioterápico em modelo de tumor de mama murino. Para tanto, camundongos Balb/c foram divididos nos grupos controle (CTL), controle tumor (CTLTU), tumor ômega 3 (TUW3), tumor 5-fluouracil (TU5FU) e tumor 5-fluouracil ômega 3 (TU5FUW3). Nos animais dos grupos com tumor foi realizada a inoculação das células tumorais (dia 1) e, uma vez que os tumores já se encontravam palpáveis (dia10), os grupos TUW3 e TU5FUW3 iniciaram a suplementação com ômega 3 pela ração por 10 dias, enquanto os demais animais receberam ração controle. Ao final desse período, os animais dos grupos TU5FU e TU5FUW3 receberam injeção intraperitoneal de 5-FU (300mg/kg) para indução da mucosite, enquanto os demais animais receberam a injeção de solução salina. Após 72 horas, os animais foram eutanasiados para coleta de dados. A suplementação com ômega 3 não foi capaz de prevenir a perda de peso em decorrência da mucosite. Entretanto, os animais do grupo TU5FUW3 apresentaram redução da PI por meio da recuperação das junções firmes ZO-1 e ocludina, além da preservação da arquitetura dos vilos. Os animais do grupo CTLTU também apresentaram aumento da PI e redução da expressão das proteínas de junção, provavelmente pela liberação de citocinas TNF-α e IL-1ß, conforme dosagem destas no baço. Nos animais suplementados com ômega 3 foi observado redução do tamanho do tumor e do número de metástases tumorais, além de redução da proliferação de células tumorais e aumento da afinidade das células tumorais ao quimioterápico, levando à apoptose destas. Este processo de citotoxicidade seletiva foi observado pela redução da toxicidade hepática. Diante desses resultados, pode-se concluir que a suplementação de ômega 3, por período de dez dias, foi capaz de atenuar os efeitos da mucosite induzida por quimioterápico, sem causar prejuízo à sua atividade antitumoral.

Omega 3 is a polyunsaturated fatty acid used as a therapeutic agent in the treatment of various diseases. It is widely accepted that its use can exert an anti-inflammatory effect. Thus, it would be interesting in the prevention and treatment of mucositis, which is described as an inflammation that affects mucosal membranes from the mouth to the anus, due to the use of chemotherapy and radiotherapy. Patients who suffer from this outcome, usually presents weight loss, anorexia and odynophagia,also there is an increased risk of bacterial translocation, due to the increase of the intestinal permeability (IP). Such complications can lead to the interruption of chemotherapy treatment until the patient is recovered. Previous work has shown that omega 3 was able to reduce weight loss and reduce IP by reducing intestinal cell apoptosis. However, the effects of omega 3 supplementation from the diet on the tumor remains unknown. This study aimed to investigate the effect of this compound on chemotherapy-induced mucositis in the murine breast tumor model. For that, BALB/ c mice were divided into the control (CTL), tumor control (CTLTU), omega 3 tumor (TUW3), 5-fluouracil tumor (TU5FU) and 5-fluouracil omega 3 tumor (TU5FUW3) groups. Tumor cells were inoculated (day 1) and, since they were already palpable (day 10), groups TUW3 and TU5FUW3 started receiving the chow supplemented with omega 3 for 10 days, while the other animals received control chow. At the end of this period, animals from groups TU5FU and TU5FUW3 received an intraperitoneal injection of 5-FU (300mg / kg) to induce mucositis, while the other animals received an injection of saline. After 72 hours, all animals were euthanized for data collection. Omega 3 supplementation was not able to prevent weight loss due to mucositis. However, animals from TU5FUW3 group showed a reduction in IP by recovering the tight-junctions ZO-1 and occludin, and by preventing the mucosal damage. Animals from CTLTU group also showed increased PI and reduced expression of tight-junction proteins, probably due to the release of cytokines TNF-α and IL-1ß, according to their dosage in the spleen. In addition, our results have demonstrated that animals supplemented with omega 3, have shown a reduction in the size of the tumor and the number of metastasis in lungs. We have found that the reduction was mediated through a reduction in the proliferation of tumor cells and also by an increased affinity of tumor cells to the chemotherapeutic drug, leading to their apoptosis. This selective cytotoxicity process leads to the reduction of liver toxicity. Given these results, we can conclude that omega 3 supplementation, for ten days, was able to attenuate the effects of chemotherapy-induced mucositis, without causing damage to its antitumor activity.

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.

Total alkaloids of Coptidis Rhizoma combined with exercise inhibits tumor growth of orthotopically transplanted 4T1 breast cancer mice by blocking cell cycle G_1/S transformation / 中国中药杂志

Breast cancer is one of the leading causes for cancer-related death among women worldwide. Coptidis Rhizoma has antibacterial,anti-inflammatory,anti-tumor and other pharmacological activities,but whether exercise could synergistically promote the role of RC in the treatment of breast cancer has not been reported. In this experiment,the effects and mechanism of total alkaloids of Coptidis Rhizoma combined with exercise on the tumor growth of orthotopically transplanted 4 T1 breast cancer were systemically studied in mice. Balb/C mice transplanted with 4 T1 cells in situ were used as models. The total alkaloids of RC(145 mg·kg-1·d-1) alone or in combination with exercise(10 m·min-1,30 min/time,5 times/week) were given for 28 days,and then the changes in body weight and tumor volume,tumor weight,interleukin-1β(IL-1β),serum estradiol(E2) content,and expression levels of estrogen receptor α(ERα),cell cycle related proteins CDK4,CDK6,cyclin D1,CDK2,and cyclin E in tumor tissues. The results showed that total alkaloids of Coptidis Rhizoma could significantly inhibit the growth of 4 T1 breast cancer in mice(P< 0. 01),and exercise significantly promoted the anti-tumor activity of total alkaloids of Coptidis Rhizoma(P<0. 01),and reduced E2 and IL-1β levels in mice. Western blot and flow cytometry showed that the total alkaloids of Coptidis Rhizoma combined with exercise could down-regulate the protein expression levels of ERα,CDK4,CDK6,cyclin D1,CDK2 and cyclin E in cancer cells,block the transformation of G1/S in 4 T1 cell cycle,and inhibit DNA synthesis in breast cancer cells. The total alkaloids of Coptidis Rhizoma combined with exercise showed synergistic effect in inhibition of tumor growth in mice with orthotopically transplanted 4 T1 breast cancer.

Effects of polysaccharides from Ginkgo biloba on proliferation of 4T1 breast cancer cells and expression of GLUT family genes / 中国药理学通报

Aim To investigate the effects of polysac-charides from Ginkgo biloba on the proliferation, apop-tosis of mouse 4T1 breast cancer cells and the possible mechanism. Methods 4T1 cells in logarithmic growth phase were treated with polysaccharides from Ginkgo biloba of different concentrations. The effect of poly-saccharides from Ginkgo biloba on inhibition of prolif-eration and cytotoxicity of 4T1 cells was determined by MTT assay and trypan blue exclusion assay respective-ly. The apoptotic effect of polysaccharides from Ginkgo biloba on 4T1 cells was detected by DAPI staining. qRT-PCR experiments were carried out for the detec-tion of gene expressions of the glucose transporter fami-ly upon the treatment with the polysaccharides from Ginkgo biloba. Results Polysaccharides from either Ginkgo biloba leaf or Ginkgo biloba exocarp significant-ly inhibited the proliferation of 4T1 cells in a dose-and time-dependent manner. Moreover, with the increasing doses of polysaccharides, cell viability decreased, ac-companied by the increased cell cytotoxicity and apop-tosis. qRT-PCR results showed that polysaccharides from Ginkgo biloba significantly reduced glucose trans-porter 1 gene expression. Conclusions Polysaccha-rides from Ginkgo biloba can both inhibit 4T1 cell pro-liferation and induce cell apoptosis, and by regulating glucose transporter family gene expression, it interfered with cell energy metabolism, which infers that the effects of cell proliferation inhibition as well the apopto-sis induction might be due to the regulation of glucose transporter family gene expression.

In vivo and in vitro effects of Miao Medicine Indigofera stachyoides extracts on breast cancer 4T1 cells / 中草药

Objective To study the effects of Indigofera stachyoide extracts on the breast cancer cells (4T1) in vivo and in vitro. Methods MTT method was used to detect the antitumor activity of I. stachyoides extracts in 4T1cells in vitro, the inhibition rate of cell proliferation, and half inhibition concentration (IC50). The established mice model with 4T1 solid tumor were randomly divided into model, extracts of I. stachyoides (petroleum ether phase, ethyl acetate phase, n-butanol phase, water phase, and ethanol extracts) groups, and cisplatin group. After being administered for 15 d, mice body weight and victera index were measured; The observation of tumor pathology and the calculation of tumor inhibition rate were performed. Results IC50 of ethyl acetate phase, n-butanol phase, ethanol extracts of I. stachyoides on 4T1 cells in vitro reached 228.9, 323.4, and 322.6 μg/mL, respectively. The tumor inhibition rates of petroleum ether phase, ethyl acetate phase, n-butanol phase, water phase, and ethanol extract of I. Stachyoides, and cisplatin group on 4T1 mice were (55.88 ± 6.68)%, (66.67 ± 14.32)%, (65.71 ± 12.38)%, (53.81 ± 16.17)%, (43.73 ± 25.73)%, and (76.85 ± 11.38)%, respectively. In the different extraction parts of I. stachyoide, the petroleum ether group had the effects of reducing the spleen index, increasing the thymus index and IL-2 level, and the ethyl acetate part was the best partaccording to tumor volume and the tumor suppressor rate. HE staining showed that the tumor cells in petroleum ether extract group were less than that in the model group, the cell arrangement was loose, the pathological mitosis and tumor cell infiltration were less than those of model group, and there was a small amount of lymphocytes and macrophages infiltration. Conclusion The extracts of I. stachyoides can inhibit the growth of 4T1 tumor cells in vivo and in vitro, and its mechanism may enhance the body immunity, so as to inhibit the tumor growth.

Psychotria viridis: Chemical constituents from leaves and biological properties

ABSTRACT The phytochemical study of hexane, chloroform and methanol extracts from leaves of Psychotria viridis resulted in the identification of: the pentacyclic triterpenes, ursolic and oleanolic acid; the steroids, 24-methylene-cycloartanol, stigmasterol and β-sitosterol; the glycosylated steroids 3-O-β-D-glucosyl-β-sitosterol and 3-O-β-D-glucosyl-stigmasterol; a polyunsaturated triterpene, squalene; the esters of glycerol, 1-palmitoylglycerol and triacylglycerol; a mixture of long chain hydrocarbons; the aldehyde nonacosanal; the long chain fat acids hentriacontanoic, hexadecanoic and heptadenoic acid; the ester methyl heptadecanoate; the 4-methyl-epi-quinate and two indole alkaloids, N,N-dimethyltryptamine (DMT) and N-methyltryptamine. The chemical structures were determined by means of spectroscopic (IR, 1H and 13C NMR, HSQC, HMBC and NOESY) and spectrometric (CG-MS and LCMS-ESI-ITTOF) methods. The study of biologic properties of P. viridis consisted in the evaluation of the acetylcholinesterase inhibition and cytotoxic activities. The hexane, chloroform, ethyl acetate and methanol extracts, the substances 24-methylene-cycloartanol, DMT and a mixture of 3-O-β-D-glucosyl-β-sitosterol and 3-O-β-D-glucosyl-stigmasterol showed cholinesterase inhibiting activity. This activity induced by chloroform and ethyl acetate extracts was higher than 90%. The methanol and ethyl acetate extracts inhibit the growth and/or induce the death of the tumor cells strains B16F10 and 4T1, without damaging the integrity of the normal cells BHK and CHO. DMT also demonstrated a marked activity against tumor cell strains B16F10 and 4T1.

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.

Anti-breast cancer properties and toxicity of Dillenia suffruticosa root aqueous extract in BALB/c mice

Objective: To determine the anti-breast cancer activities and the safety oral consumption of Dillenia suffruticosa root aqueous extract (DRAE) in BALB/c mice. Methods: In the anti-breast cancer study, female BALB/c mice were divided into five groups (n = 12), which were (1) positive control (with breast cancer, untreated), (2) negative control (without breast cancer, untreated) and other three groups of mice with breast cancer treated with 1 000, 500 and 250 mg/kg of DRAE, respectively, by oral gavage for 28 days. All mice except from the negative control group were injected into the mammary fat pad with 4T1 cells (1 × 105 4T1 cells/0.1 mL of phosphate buffer solution). DRAE was administered orally on Day 11 after the tumor has developed. Results: The tumor volume of the 1 000 mg/kg of DRAE group reduced significantly compared to the positive control while treatment with 500 mg/kg of DRAE had signif-icantly inhibited metastasis to the heart. In the acute toxicity study, treatment with up to 5 000 mg/kg of DRAE was not toxic to the animals, indicating its safety when a large amount of this plant extract was ingested. Based on the sub-acute toxicity study, treatment of the highest dose of DRAE (1 000 mg/kg) had mild liver toxicity indicated by mild focal hemorrhage. Conclusions: DRAE possesses anti-breast cancer properties but at the same time it shows mild toxicity to the liver. The non observable adverse effect dose for DRAE is 500 mg/kg.