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@#Objective To establish and validate a method for the determination of the interesting protein expression level of recombinant adeno-associated virus(rAAV)infected cells,so as to monitor the product quality in different stages of rAAV9production process.Methods After incubation of serial diluted rAAV samples with infection enhancer Envirus-AAV,the human malignant glioblastoma cells(U87-MG)pretreated with hydroxyurea(HU)were infected.Using rAAV9 reference as the standard,the expression level of glutaryl-CoA dehydrogenase(GCDH)was detected by ELISA,and the specificity,accuracy,precision,linear range,limit of quantitation(LOQ)and durability of the method were verified.Eight batches of rAAV9 samples were detected by the established method.Results The A_(450)-A_(630) value of the sample buffer was 0.3,which was slightly lower than the lowest dilution point(1 ng/mL)of the four-parameter standard curve for protein quantification.The average recoveries of samples with 150%,100% and 50% theoretical relative titer levels were in the range of 100.0%-107.3%.The RSDs of the target protein expression level of the samples with three theoretical relative titer levels detected by the same experimenter three times and different experimenters were all less than 25%.There was a good linear relationship between rAAV9 samples and the target protein expression levels in the range of 50%-150% theoretical relative titer levels,and the linear regression equation was y = 1.077 x-0.022,R~2= 0.984.The LOQ of the method was 0.59,namely 6.0×10~(12) vg/mL.After U87-MG cells were incubated with HU for different time(18,21,24 h),and the culture supernatant was stored under different conditions(room temperature for 0.5 h,below-60 ℃ for 12 h,below-60 ℃ for 24 h).The RSDs of target protein expression levels were all less than 25%.The target protein expression levels of 1-8 batches of rAAV9 samples were 111%,121%,72%,65%,86%,75%,102% and 91%,respectively.Conclusion The established method for the determination of the target protein expression level after rAAV infection has good specificity,accuracy,precision and durability,and can be used for the quality control of products in different stages of rAAV9 production.
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Glioblastoma is aggressive brain tumour with poor prognosis with conventional chemotherapy, hence there is need to find alternative targets for developing newer treatment. Advent of new treatment methods involving medicinal plants have shown to reduced Cancer mortalities and prevents development of drug resistance for chemotherapy. Present study aimed at investigates the anti-proliferating activity of two promising medicinal plants, Ocimum sanctum and Centella asiatica. We studied the effect of their plant extract on U87MG Glioblastoma cells proliferation, survival effect and apoptosis. Cytotoxic activity was assessed, after the plant extract treatment on U87MG using MTT assay with dose of 1 mg/mL to 25mg/mL and apoptosis assess was done using Annexin V assay with the three dose (1.5 mg/mL, 2 mg/mL and 2.5 mg/mL). Survivin gene expression was studied using QRT-PCR (Rotar gene Q, Qiagene) has a marker of proliferation. Ocimum sanctum and Centella asiatica treatment of U87MG cells with dosage of 1.5 mg/mL, 2.0 mg/mL, 2.5 mg/mL showed increase in mean apoptotic cells 2.8 %, 4.9%, 10 % and 3.1%, 5.8% and 7.2%, respectively, compared to untreated U87MG cells. Survivin gene analysis of U87MG cells showed down-regulation in gene expression and differences was significant in comparison to untreated control group with both the plant extract, Centella asiatica showed more down-regulation (97% with 2.5 mg/mL) than Ocimum sanctum. Ocimum sanctum and Centella asiatica exhibited promising anti-proliferating activity and induces apoptosis by down regulation of survivin gene expression
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【Objective】 To explore the establishment methods of transgenic human umbilical cord mesenchymal stem cells (hUC-MSCs) overexpressing tumor necrosis factor(TNF)-related apoptosis-inducing ligand (TRAIL) based on the transposons, and attempt to apply it on the nude mice mode with glioma. 【Methods】 PiggyBac transposon system specially designed by us was used to prepare non-targeting and Her2-targeting hUC-MSCs that can stably express TRAIL through puromycin screening. The glioma cells expressing firefly luciferase (U87MG-LUC) were injected into the skull of the immunodeficient mice (BALB/c-nu/nu) with 1×106 cells per mouse. After 7 days of injection, the mice transplanted with U87MG were detected with a small animal living imager to determine the size and location of the tumors in skull. Then we injected the glioma-transplantation nude mouse with two kinds of transgenic hUC-MSCs expressing TRAIL (named as untarget-TRAIL and target-TRAIL, respectively), or the non-transgenic hUC-MSCs (all 1×106 cells per mouse) or PBS (named as WT-MSCs and PBS for negative control) respectively, and then monitored the changes of tumor signals by a small animal living imager every week for 3~4 weeks. 【Results】 After six passages to expand the cells, the both transgenic cell lines can stably express TRAIL gene. Their ratio of green fluorescent protein (GFP) positive cells can reach 93%-97%, and the positive ratio of their MSC-specific surface markers still maintained normal (CD34+, CD45+, and HLA-DR+ all <0.1%, CD90>99%, CD73>88%, and CD105 >60%). The median survival time (d) of U87MG-transplanted nude mice in the groups of untarget-TRAIL, target-TRAIL, WT-MSCs, and PBS was 41 vs 39 vs 24 vs 23(P<0.05). 【Conclusion】 The transgenic hUC-MSCs overexpressing TRAIL gene can significantly prolong the survival time of nude mice with brain glioma.
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@#[Abstract] Objective: To explore the inhibitive effect of asiatic acid (AA) on paclitaxel (PTX)-resistant glioma cells and its possible mechanism. Methods: The effects of AA on the proliferation and apoptosis of glioblastoma U87MG cells were detected by CCK-8 assay, Real-time quantitative polymerase chain reaction (qPCR) and Western blotting. The drug-resistant glioma cell line PR-U87MG was established by culturing the cells in concentration-increasing PTX. With U87MG cells as control, the PTX-resistance of PR-U87MG cells was confirmed using CCK-8 assay, and the mRNA and protein levels of MDR1 and LRP were measured with qPCR and western blotting. PR-U87MG cells were treated with AA, PTX or AA+PTX, and then the cell viability and apoptosis of each group were measured with CCK-8 assay, qPCR and Western blotting. Results: PTX-resistant PR-U87MG cell line was successfully established. AA inhibited the viability of U87MG and PR-U87MG cells in a dose-dependent manner (P<0.01) and significantly promoted their apoptosis (P<0.01). Compared with the group treated with AA or PTX alone, the group treated with the combination of AA and PTX had significantly decreased protein levels of PARP1 (P<0.01), drug-resistant related proteins (Pgp-1 and LRP [lung resistance protein], all P< 0.01), and markedly increased caspase 3 (P<0.01). Conclusion: AA could effectively enhance the sensitivity of U87MG cells to PTX, and the mechanism may be related to the suppressed expression of drug efflux-associated proteins Pgp-1 and LRP.
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Aim To establish human U87-MG glioma model in nude mice brain and to observe the characteristics of the tumor growth. Methods Human U87-MG glioma cells were cultured in vitro. 5 μL of cell suspension containing 3.0 ×1010·L-1, 4.0×1010·L-1and 5.0×1010·L-1respectively was inocula-ted into the right caudate nucleus of 18 male nude mice brain un-der the guidance of stereotaxic apparatus, separately, whereas another 6 nude mice as the control group, were inoculated into the same volume of Hanks solution. The moving and survival state of rats with gliomas were observed. The examinations of the tumors formation, volumes, metastasis and histopathology were performed and the obtained brain samples were stained with HE and immunohistochemistry. Results All the tested rats of dif-ferent inoculation doses developed brain tumors without extracra-nial metastasis. The mean survival time of three groups was (46.50 ± 3.27) d,(38.50 ± 3.28) d and (30.67 ± 3.51) d,respectively. The tumors showed the similar morphological fea-tures and immunophenotype to human glioma. There was positive expression of GFAP and S-100 in the tumors. Conclusions The orthotopic implantation model of human U87-MG glioma, by in-oculating quantitative U87-MG cells stereotaxically into the brains of the nude mice, is successfully established with 100 yield of intracranial tumor and no extracranial growth extension. It resembles the histopathological and morphological features of human glioma,which can be used as a reliable animal model for the study of the tumorigenesis, pathogenesis, biological charac-teristics and therapy of glioma.
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To explore the inhibitory effect of timosaponin AⅢ on the proliferation of human glioblastoma cell line U87MG and investigate its related mechanism. As compared with the model group, the tumor weight was significantly reduced in timosaponin AⅢ-treated group. Timosaponin AⅢinhibited the proliferation of U87MG cell line in a dose-dependent manner. It up-regulated the gene and protein expression levels of p21, meanwhile inhibited the protein expression levels of β-Catenin, Cyclin D1 and Bcl-2. It also inhibited the translocation of β-Catenin into nucleus, suppressed the phosphorylation expression of ERK, but increased the phosphorylation expression of p38 and JNK. Combined use of JNK inhibitor SP600125 and p38 inhibitor SB203580 could decrease p21 and increase β-Catenin protein expressions. Timosaponin AⅢ inhibited the proliferation of human glioblastoma cell line U87MG partly by intervening MAPK and Wnt/β-Catenin signal pathways.
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AIM: To explore the effects of chromodomain protein 8 (CBX8) on the proliferation and apoptosis of human glioma cells.METHODS: The expression of CBX8 in the tissues and cells was detected by Western blot and RT-qPCR.The overexpression (Flag-CBX8) and silencing (sh-CBX8) vectors of CBX8 were constructed and transfected into glioma T98G cells and U87MG cells.The cell proliferation was detected by MTT assay and BrdU staining.The cell apoptosis was analyzed by flow cytometry.The protein expression of Rb/E2F1 was detected by Western blot.RESULTS: Compared with normal brain tissues and astrocytes, the expression of CBX8 was increased in the glioma tissues and glioma cells.Overexpression of CBX8 promoted the cell proliferation, inhibited the cell apoptosis, and upregulated the protein levels of Rb/E2F1.On the contrary, silencing of CBX8 inhibited the cell proliferation, promoted the cell apoptosis, and decreased the protein levels of Rb/E2F1 in the T98G cells and U87MG cells.Moreover, the expression of cyclin D1 and Bcl-2/Bax ratio were reduced after transfection with sh-E2F1 in the T98G cells and U87MG cells.CONCLUSION: CBX8 may regulate the proliferation and apoptosis of glioma cells through Rb/E2F1 pathway.
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Objective To investigate the effects of salidroside on proliferation and invasive ability of glioma U87-MG cells; To discuss the its mechanism to induce apoptosis of U87-MG cells. Methods U87-MG cells were cultured in vitro for 24 h under different concentrations of salidroside and camptothecin. The proliferation of U87-MG cells was detected by MTT assay. The apoptosis rate of U87-MG cells was detected by flow cytometry. Transwell assay was used to detect the invasive ability of U87-MG cells. ROS was detected by indirect fluorescent labeling. The expressions of Caspase-3, Bcl-2, Bax, E-cadherin, N-cadherin and matrix metalloproteinase-9 (MMP-9) in U87-MG cells were detected by Western blot. Results Compared with the blank control group, U87-MG cells had significant inhibitory effect on the growth of U87-MG cells in each administration group, and the invasive ability of U87-MG cells was significantly reduced after 10, 50, 100 μg/mL salidroside was intervened, and 10, 50, 100 μg/mL salidroside for 48 h for U87-MG cells could induce apoptosis of the cells; the level of ROS was positively correlated with the concentration of salidroside; 10, 50, 100 μg/mL salidroside up-regulated the expressions of Caspase-3, Bax and E-cadherin, and down-regulated the expressions of Bcl-2, N-cadherin and MMP-9. Conclusion Salidroside can induce apoptosis of U87-MG cells and inhibit the invasive ability of U87-MG cells.
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Moléculas orgânicas fluorescentes são uma importante ferramenta para biologia celular. Compostos ideais para esta aplicação devem ter alto brilho (produto do coeficiente de atenuação molar e do rendimento quântico de fluorescência), ser fotoestáveis e internalizáveis, não comprometer a viabilidade celular e interagir com biomoléculas com algum grau de especificidade. Nesta Tese de Doutorado é apresentado o estudo do uso de cBeet120, uma betalaína cumarínica artificial, e células de glioma humano da linhagem U87-MG. Betalaínas são pigmentos de plantas que apresentam alta biocompatibilidade que servem como material de partida para o desenvolvimento de derivados funcionais. A sonda se acumula principalmente no núcleo das células U87- MG e marca principalmente nucléolos via interação com proteínas. A presença de DNAse ou RNAase elimina a marcação nuclear, sem afetar a fraca marcação citoplasmática de fundo. Estudos de inibição de transporte sugerem que cBeet120 é internalizada por transportadores de L-glutamato da família de transportadores de amino ácidos excitatórios (EAAT). O uso de artemisinina para inibição Ca2+-ATPases aumenta a velocidade de internalização de cBeet120 em células U87-MG. Quando irradiada com luz de cor ciano, cBeet120 no interior do núcleo de células vivas é fotoativada, resultando em um aumento da intensidade de fluorescência com o tempo (monitorado por 90 min) e o deslocamento hipsocrômico do máximo de emissão. Em células fixadas com paraformaldeído, o padrão de marcação da célula se torna mais difuso e a sonda emite fluorescência sem fotoativação. Medidas de tempo de vida de fluorescência em solução e imageamento por microscopia de tempo de vida de fluorescência permitem inferir a ocorrência da formação de um complexo proteína-cBeet120 ou um produto de transiminação que pode estar sujeito a isomerização cis/trans
Fluorescent organic molecules are an important tool for cell biology. Ideal compounds for this application must have high brightness (product of the molar attenuation coefficient and fluorescence quantum yield), be photostable and internalizable by cells, do not compromise cellular viability and interact with biomolecules with some degree of specificity. In this Doctorate Thesis, we describe the interaction of cBeet120, an artificial coumarinic betalain, and human glioma cells of line U87-MG. Betalains are plant pigments that exhibit high biocompatibility that serve as starting material for the development of functional derivatives. The probe accumulates mainly in the nucleus of the U87-MG cells and mainly marks nucleoli via interaction with proteins. The presence of DNAse or RNAase eliminates nuclear labeling, without affecting the poor background cytoplasmic labeling. Transport inhibition studies suggest that cBeet120 is internalized by L-glutamate transporters from the excitatory amino acid transporter (EAAT) family. The use of artemisinin for inhibition Ca2+-ATPases increases the rate of cBeet120 internalization in U87-MG cells. When irradiated with cyan colored light, cBeet120 within the nucleus of living cells is photoactivated, resulting in an increase in fluorescence intensity over time (monitored for 90 min) and the hypochromic shift of the emission maximum. In cells fixed with paraformaldehyde, the labeling pattern of the cell becomes more diffuse and the probe emits fluorescence without photoactivation. Fluorescence life-time measurements in solution and fluorescence life-time imaging microscopy allows to infer the occurrence of the formation of a protein-cBeet120 complex or the formation of a transimination product that may be subject to cis/trans isomerization
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Coumarines/analyse , Beta vulgaris/métabolisme , Mécanismes moléculaires de l'action pharmacologique , Gliome/complications , Glioblastome/complications , Bétalaïnes , FluorescenceRésumé
OBJECTIVE@#To study the effects of Transient receptor potential cation channel, subfamily V, member 1 (TRPV1) combined with lidocaine on status and apoptosis of U87-MG glioma cell line, and explore whether local anesthetic produces neurotoxicity by TRPV1.@*METHODS@#U87-MG cells were divided into control group, gene silencing group, empty vector group and TRPV gene up-regulation group. For cells in each group, flow cytometry was employed to detect the intracellular calcium ion concentration and mitochondrial membrane potential at different time point from cellular perspective. Cell apoptosis of U87-MG was assayed by flow cytometry and MTT from a holistic perspective.@*RESULTS@#Calcium ion concentration increased along with time. The concentration in TRPV1 gene up-regulation group was significantly higher than those in other groups at each time point (P < 0.05). After adding lidocaine, mitochondrial membrane potential in U87-MG significantly increased (P < 0.05). This increasing trend in TRPV1 gene up-regulation group was more significant than other groups (P < 0.05), while in TRPV1 gene silencing group, the trend significantly decreased (P < 0.05). Flow cytometry result and MTT result both showed that cell apoptosis in each group significantly increased after lidocaine was added (P < 0.05). This increasing trend in TRPV1 gene up-regulation group was more significant than other groups (P < 0.05), while in TRPV1 gene silencing group, the trend significantly decreased (P < 0.05). Moreover, apoptosis was more severe along with the increasing concentration of lidocaine (P < 0.05).@*CONCLUSIONS@#In this study, it was proved that lidocaine could dose-dependently induce the increase of intracellular calcium ion concentration, mitochondrial membrane potential and apoptosis in U87-MG glioma cell line. The up-regulation of TRPV1 enhanced cytotoxicity of lidocaine, which revealed the correlations between them. Lidocaine might have increased intracellular calcium ion concentration by activating TRPV1 gene and induced apoptosis of U87-GM glioma cell line by up-regulating mitochondrial membrane potential.
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Objective: To study the effects of Transient receptor potential cation channel, subfamily V, member 1 (TRPV1) combined with lidocaine on status and apoptosis of U87-MG glioma cell line, and explore whether local anesthetic produces neurotoxicity by TRPV1. Methods: U87-MG cells were divided into control group, gene silencing group, empty vector group and TRPV gene up-regulation group. For cells in each group, flow cytometry was employed to detect the intracellular calcium ion concentration and mitochondrial membrane potential at different time point from cellular perspective. Cell apoptosis of U87-MG was assayed by flow cytometry and MTT from a holistic perspective. Results: Calcium ion concentration increased along with time. The concentration in TRPV1 gene up-regulation group was significantly higher than those in other groups at each time point (P < 0.05). After adding lidocaine, mitochondrial membrane potential in U87-MG significantly increased (P < 0.05). This increasing trend in TRPV1 gene up-regulation group was more significant than other groups (P < 0.05), while in TRPV1 gene silencing group, the trend significantly decreased (P < 0.05). Flow cytometry result and MTT result both showed that cell apoptosis in each group significantly increased after lidocaine was added (P < 0.05). This increasing trend in TRPV1 gene up-regulation group was more significant than other groups (P < 0.05), while in TRPV1 gene silencing group, the trend significantly decreased (P < 0.05). Moreover, apoptosis was more severe along with the increasing concentration of lidocaine (P < 0.05). Conclusions: In this study, it was proved that lidocaine could dose-dependently induce the increase of intracellular calcium ion concentration, mitochondrial membrane potential and apoptosis in U87-MG glioma cell line. The up-regulation of TRPV1 enhanced cytotoxicity of lidocaine, which revealed the correlations between them. Lidocaine might have increased intracellular calcium ion concentration by activating TRPV1 gene and induced apoptosis of U87-GM glioma cell line by up-regulating mitochondrial membrane potential.
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Os gliomas são tumores cerebrais definidos patologicamente pela presença de células com características histológicas e imuno-histoquímicas que evidenciam diferenciação glial. Dentre eles, os astrocitomas são os mais frequentes em adultos. Estes tumores normalmente apresentam infiltração difusa no tecido adjacente, são resistentes aos tratamentos e têm uma tendência natural para a progressão maligna. O tratamento padrão atual consiste na realização de ressecção cirúrgica do tecido tumoral seguida de radio e quimioterapia concomitantes, porém o prognóstico permanece extremamente pobre. O quimioterápico padrão-ouro no tratamento de GBM é o agente alquilante de DNA temozolamida (TMZ). Entretanto, os danos induzidos pela TMZ podem ser revertidos pela ação da maquinaria de reparo de DNA, impedindo a morte celular e levando à resistência do GBM ao tratamento. No presente estudo correlacionamos a expressão dos genes ATM, BRCA2, BRIP1, EXO1, NEIL3, RAD54L e XRCC2, envolvidos em reparo de DNA e sabidamente superexpressos em GBM, com a resistência das linhagens celulares T98G e U87MG ao tratamento com TMZ. Mostramos que a linhagem T98G é a mais resistente ao tratamento com TMZ, e apresenta superexpressão de BRCA2, BRIP1, EXO1, NEIL3, RAD54L e XRCC2 e sub-expressão de ATM. Vimos também que a linhagem U87MG, mais sensível ao tratamento com TMZ, apresenta expressão reduzida dos genes ATM, BRCA2 e EXO1. Portanto, estes dados sugerem uma correlação positiva entre a expressão de genes de reparo de DNA e a resistência das células de GBM à TMZ.(AU)
Gliomas are brain tumors pathologically defined by the presence of cells with histological and immunohistochemical characteristics of glial differentiation. Among them, astrocytomas are the most common in adults. These tumors usually show diffuse infiltration into adjacent tissue, are resistant to treatment and have a natural tendency to malignant progression. The current standard treatment consists in surgical removal of the tumor followed by radiotherapy and concurrent chemotherapy. However, the prognosis remains extremely poor. The first line chemotherapy for GBM treatment is the DNA alkylating agent temozolamide (TMZ). Nevertheless, TMZ-induced damage can be reversed by the action of DNA repair machinery that prevents cell death and leads to relapse. In this study we correlated the expression of the DNA damage-signaling gene, ATM kinase, and the DNA repair genes BRCA2, BRIP1, EXO1, NEIL3, RAD54L and XRCC2, with the resistance of T98G and U87MG cell lines to TMZ. T98G cells were more resistant to TMZ treatment and showed overexpression of all DNA repair genes, while ATM kinase was down regulated. We also observed that U87MG cells, more sensitive to TMZ, have reduced expression of ATM, BRCA2 and EXO1. Therefore, these data suggest a positive correlation between the expression of DNA repair genes and the resistance of GBM cells to TMZ.(AU)
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Humains , Mâle , Adulte , Tumeurs du cerveau , Glioblastome , Agents alcoylants/usage thérapeutiqueRésumé
Objective:To detect the functional role of Fascin1 and its related molecular mechanisms in migration and invasion capacity of glioma cells,we utilized gene specific small interference RNA of Fascin1 in cell line U87 MG. Methods:Fascin1-siRNA or negative siRNA was transfected into U87 MG cells of control group or experiment group. Transwell method was employed to assess the migration and invasion capacity of glioma cells. Western blot analysis was used to detect the protein expression of Fascin1,pAKT and pSTAT3. The impact of PI3K/AKT pathway and STAT3 pathway on migration and invasion of U87 MG cells was verified,via applying LY294002 and LY294002,which was inhibitor of the two pathways respectively. Results:As compared to control groups,the migration and invasion capacity of transfected glioma cells were attenuated about 52% or 43%(P<0. 05),accompanied with the decreased phos-phorylation of AKT and STAT3. As utilizing the inhibitors of AKT and STAT3,attenuated migration and invasion capacity of U87 MG cells were observed. Conclusion:Down-regulated expression of Fascin1 could suppress the migration and invasion capacity of U87 MG cells by inhibiting the phosphorylation of PI3K/AKT pathway and STAT3 pathway.
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OBJECTIVE: With the growing interests of bacteria as a targeting vector for cancer treatment, diverse genetically engineered Salmonella has been reported to be capable of targeting primary or metastatic tumor regions after intravenous injection into mouse tumor models. The purpose of this study was to investigate the capability of the genetically engineered Salmonella typhimurium (S. typhimurium) to access the glioma xenograft, which was monitored in mouse brain tumor models using optical bioluminescence imaging technique. METHODS: U87 malignant glioma cells (U87-MG) stably transfected with firefly luciferase (Fluc) were implanted into BALB/cAnN nude mice by stereotactic injection into the striatum. After tumor formation, attenuated S. typhimurium expressing bacterial luciferase (Lux) was injected into the tail vein. Bioluminescence signals from transfected cells or bacteria were monitored using a cooled charge-coupled device camera to identify the tumor location or to trace the bacterial migration. Immunofluorescence staining was also performed in frozen sections of mouse glioma xenograft. RESULTS: The injected S. typhimurium exclusively localized in the glioma xenograft region of U87-MG-bearing mouse. Immunofluorescence staining also demonstrated the accumulation of S. typhimurium in the brain tumors. CONCLUSION: The present study demonstrated that S. typhimurium can target glioma xenograft, and may provide a potentially therapeutic probe for glioma.
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Animaux , Souris , Bactéries , Tumeurs du cerveau , Lucioles , Technique d'immunofluorescence , Coupes minces congelées , Gliome , Hétérogreffes , Injections veineuses , Luciferases , Souris nude , Salmonella , Salmonella typhimurium , VeinesRésumé
Background and purpose:Integrinαvβ3 receptor plays an important role in promoting, sustaining and regulating the angiogenesis. It is overexpressed on neovascular endothelial cells and tumor cells. RGD peptide specifically binds to integrinαvβ3, which could evaluate growth status and invasiveness of tumor. This study aimed to investigate the biodistribution in healthy KM mice and micro PET/CT imaging in U87MG tumor-bearing mice of 18F-E[c(RGDfK)2]. Methods: 18F-E[c(RGDfK)2] was produced using an automated synthesis module via a simple one-step 18F-labeling strategy of the precursor 4-NO2-3-TFMBz-E[c(RGDfK)2]. The percentage activity of injection dose per gram of tissue (%ID/g) was calculated at 0.5, 1, 2, 4 h post injection of the probe. Micro PET/CT images of U87MG tumor-bearing nude mice with or without 18F-E[c(RGDfK)2] blocking were acquired at each time point. Results: The labeling efficiency and radiochemical purity of 18F-E[c(RGDfK)2] were 10% and 98%, respectively. 18F-E[c(RGDfK)2] was excreted via renal route, with a high blood clearance. The other organs had background-level activity accumulation. At 1 h, the%ID/g of kidney, liver, intestine, muscle and blood was (1.02±0.16)%ID/g,(0.24±0.06)%ID/g, (0.35±0.03)%ID/g, (0.13±0.03)%ID/g and (0.11±0.03)%ID/g 18F-E[c(RGDfK)2] had initial high tumor uptake [(5.2±0.56)%ID/g] and good tumor-to-background contrast (5.36) at 1 h post injection. Tumor uptake for blocking group was lower than those without blocking, and T/M reduced to 1.57. Conclusion: 18F-E[c(RGDfK)2] appears a promising PET molecular imaging probe targeting integrin αvβ3, with high tumor uptake. It could be suitable for prognosis evaluation of integrin-positive tumor, selection of vascular targeting therapy and therapy effect monitoring.
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INTRODUCTION: Astrocytic gliomas are the most common intracranial central nervous system neoplasias, accounting for about 60 percent of all primary central nervous system tumors. Despite advances in the treatment of gliomas, no effective therapeutic approach is yet available; hence, the search for a more realistic model to generate more effective therapies is essential. OBJECTIVE: To develop an experimental malignant astrocytoma model with the characteristics of the human tumor. METHOD: Primary cells from subcutaneous xenograft tumors produced with malignant astrocytoma U87MG cells were inoculated intracerebrally by stereotaxis into immunosuppressed (athymic) Rowett rats. RESULTS: All four injected animals developed non-infiltrative tumors, although other glioblastoma characteristics, such as necrosis, pseudopalisading cells and intense mitotic activity, were observed. CONCLUSION: A malignant astrocytoma intracerebral xenograft model with poorly invasive behavior was achieved in athymic Rowett rats. Tumor invasiveness in an experimental animal model may depend on a combination of several factors, including the cell line used to induce tumor formation, the rat strains and the status of the animal's immune system.
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Animaux , Femelle , Humains , Rats , Astrocytome/anatomopathologie , Tumeurs du cerveau/anatomopathologie , Glioblastome/anatomopathologie , Sujet immunodéprimé , Tumeurs du cerveau/immunologie , Lignée cellulaire tumorale , Modèles animaux de maladie humaine , Glioblastome/immunologie , Transplantation tumorale , Rat nude , Transplantation hétérologueRésumé
Peroxisome proliferator-activated receptor-gamma (PPARgamma) has been implicated in the growth inhibition of a number of cancer cells. In the present study, we investigated the antitumor effect of the PPARgamma agonist rosiglitazone in U87MG human glioma cells. Rosiglitazone treatment in vitro reduced cell proliferation without induction of cell death in a dose- and time-dependent manner. Rosiglitazone decreased cell migration and mRNA level of MMP-9. Rosiglitazone treatment also induced marked changes in glioma cell morphology. Oral administration of rosiglitazone in animals with subcutaneous U87MG glioma cells reduced tumor volume. Subsequent tumor tissue analysis showed that rosiglitazone decreased the number of PCNA-positive staining cells and MMP-9 expression and induced apoptosis of tumor cells. These data suggest that rosiglitazone exerts antineoplastic effect in U87MG cells and may serve as potential therapeutic agent for malignant human gliomas.
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Animaux , Humains , Administration par voie orale , Apoptose , Mort cellulaire , Mouvement cellulaire , Prolifération cellulaire , Gliome , Péroxysomes , Récepteur PPAR gamma , ARN messager , Thiazolidinediones , Charge tumoraleRésumé
AIM: To investigate the role of transferrin/transferrin receptor system in transferrin-bound Yb 2 (Yb 2Tf) uptake by U-87 MG cells and the effect of transferrin-bound and -free Yb 2 on proliferation of U-87 MG cells. METHODS: Cell culture and ICP-MS measurement of Yb 2. RESULTS: Yb 2Tf uptake by U-87 MG cells increased with the concentrations of Yb 2Tf, and reached saturation as the concentration in the incubation medium was raised to about 2 ?mol/L. Also, Yb 2 uptake by the cells increased with increase of the mole ratio (Yb 2: apoTf), reaching a maximum at 1.5 mole ratio. Yb 2Tf in 0.4 ?mol/L significantly inhibited proliferation of U-87 MG cells, however, 10 ?mol/L Yb 3+ had no significant effect on proliferation of the cells. CONCLUSION: The uptake of Yb 2 by U-87 MG cells might be mediated by transferrin/transferrin receptor system. Transferrin-bound but not transferrin-free Yb 2 could significantly inhibit proliferation of U-87 MG cells.