Wild Type p53 Gene Sensitizes Rat C6 Glioma Cells to HSV-TK/ACV Treatment In Vitro and In Vivo

2010 ◽  
Vol 16 (4) ◽  
pp. 509-514 ◽  
Author(s):  
Qiang Huang ◽  
Zhibo Xia ◽  
Yongping You ◽  
Peiyu Pu
Keyword(s):  
Glioma Cells ◽  
P53 Gene ◽  
C6 Glioma ◽  
C6 Glioma Cells ◽  
Wild Type ◽  
Wild Type P53 ◽  
Rat C6 Glioma

Effects of Gamma Knife surgery on C6 glioma in combination with adenoviral p53 in vitro and in vivo

2006 ◽  
Vol 105 (Supplement) ◽  
pp. 208-213 ◽  
Author(s):  
Desheng Xu ◽  
Qiang Jia ◽  
Yanhe Li ◽  
Chunsheng Kang ◽  
Peiyu Pu
Keyword(s):  
Glioma Cells ◽  
P53 Gene ◽  
Treated Group ◽  
C6 Glioma ◽  
C6 Glioma Cells ◽  
Survival Duration ◽  
Wild Type ◽  
Wild Type P53

ObjectThe authors sought to study the combined potential of wild-type p53 gene transfer and Gamma Knife surgery (GKS) for the treatment of glioblastomas multiforme. Modification of the radiation response in C6 glioma cells in vitro and in vivo by the wild-type p53 gene was investigated.MethodsStable expression of wild-type p53 in C6 cells was achieved by transduction of the cells with adenoviral p53. Two days later, some cells were treated with GKS. Forty-eight hours after irradiation, the comparative survival rate was assessed by monotetrazolium (MTT) assays. Treated and control C6 glioma cells (4 × 103 per well) were plated into a 96-well plate in octuplicate and tested every 24 hours. Meanwhile, immunohistopathological examination of proliferating cell nuclear antigen (PCNA) and terminal deoxynucleotidyl transferase—mediated deoxyuridine triphosphate (TUNEL) assays were performed. The MTT assays indicated the p53, GKS, and combined treated cells proliferated at a significantly lower rate than those of the control group (p < 0.01, Days 2–6) and the positive fraction of PCNA in p53-treated group and GKS-treated group was 70.18 ± 3.61 and 50.71 ± 2.61, respectively, whereas the percentage in the combined group was 30.68 ± 1.49 (p < 0.01).Fifty-six male Sprague–Dawley rats were anesthetized and inoculated with 106 cultured C6 glioma cells into the cerebrum. Forty-eight hours after transduction with adenoviral p53, some rats underwent GKS. A margin dose of 15 Gy was delivered to the 50% isodose line. Two days later, six rats in each group were killed. Their brains were removed and paraffin-embedded section were prepared for immunohistopathological examination and TUNEL assays. The remaining rats were observed for the duration of the survival period. The survival curve indicated that a modest but significant enhancement of survival duration was seen in the p53-treated or GKS alone groups, whereas a more marked and highly significant enhancement of survival duration was achieved when these two treatment modalities were combined. When PCNA expression was downregulated, apoptotic cells become obvious after TUNEL staining.Conclusions The findings of this study suggest that p53-based gene therapy in combination with GKS may be superior to single-modality treatment of C6 glioma.

scholarly journals Gene Therapy with HSV1-sr39TK/GCV Exhibits a Stronger Therapeutic Efficacy Than HSV1-TK/GCV in Rat C6 Glioma Cells

2013 ◽  
Vol 2013 ◽  
pp. 1-10 ◽  
Author(s):  
Lei-qing Li ◽  
Fang Shen ◽  
Xiao-yan Xu ◽  
Hong Zhang ◽  
Xiao-feng Yang ◽  
...  
Keyword(s):  
Gene Therapy ◽  
Therapeutic Efficacy ◽  
Gene Transfection ◽  
Glioma Cells ◽  
C6 Glioma ◽  
C6 Glioma Cells ◽  
Transfection Efficacy ◽  
Rat C6 Glioma

Although the combination of herpes simplex virus type 1 (HSV-1) thymidine kinase (TK) with ganciclovir (GCV) has been shown as a promising suicide gene treatment strategy for glioma, the almost immunodepressive dose of GCV required for its adequatein vivoefficacy has hampered its further clinical application. Therefore, In order to reduce the GCV dose required, we aim to compare the therapeutic efficacy of HSV1-sr39TK, an HSV1-TK mutant with increased GCV prodrug catalytic activity, with wildtype TK in C6 glioma cells. Accordingly, rat C6 glioma cells were first transfected with pCDNA-TK and pCDNA-sr39TK, respectively, and the gene transfection efficacy was verified by immunocytochemistry and western blot analysis. Then thein vivosensitivity of these transfected C6-TK and C6-sr39TK cells to GCV was determined by 3-(4,5)-dimethylthiahiazo-(-z-y1)-3,5-di-phenytetrazoliumromide (MTT) colorimetric assay and Hoechst-propidium iodide (PI) staining. Finally, a subcutaneously C6 xenograft tumor model was established in the nude mice to test thein vitroefficacy of TK/GCV gene therapy. Our results showed that, as compared with wildtype TK, HSV1-sr39TK/GCV demonstrated a stronger therapeutic efficacy against C6 glioma bothin vitroandin vivo, which, by reducing the required GCV dose, might warrant its future use in the treatment of glioma under clinical setting.

Radiosensitivity of glioma to Gamma Knife treatment enhanced in vitro and in vivo by RNA interfering Ku70 that is mediated by a recombinant adenovirus

2010 ◽  
Vol 113 (Special_Supplement) ◽  
pp. 228-235 ◽  
Author(s):  
Qiang Jia ◽  
Yanhe Li ◽  
Desheng Xu ◽  
Zhenjiang Li ◽  
Zhiyuan Zhang ◽  
...  
Keyword(s):  
Western Blot ◽  
Gamma Knife ◽  
Blot Analysis ◽  
Western Blot Analysis ◽  
Glioma Cells ◽  
C6 Glioma ◽  
C6 Glioma Cells ◽  
C6 Cells

Object The authors sought to evaluate modification of the radiation response of C6 glioma cells in vitro and in vivo by inhibiting the expression of Ku70. To do so they investigated the effect of gene transfer involving a recombinant replication-defective adenovirus containing Ku70 short hairpin RNA (Ad-Ku70shRNA) combined with Gamma Knife treatment (GKT). Methods First, Ad-Ku70shRNA was transfected into C6 glioma cells and the expression of Ku70 was measured using Western blot analysis. In vitro, phenotypical changes in C6 cells, including proliferation, cell cycle modification, invasion ability, and apoptosis were evaluated using the MTT (3′(4,5-dimethylthiazol-2-yl)2,5-diphenyltetrazolium bromide) assay, Western blot analysis, and cell flow cytometry. In vivo, parental C6 cells transfected with Ad-Ku70shRNA were implanted stereotactically into the right caudate nucleus in Sprague-Dawley rats. After GKS, apoptosis was analyzed using the TUNEL (terminal deoxynucleotidyl transferase–mediated deoxyuridine triphosphate nick-end labeling) method. The inhibitory effects on growth and invasion that were induced by expression of proliferating cell nuclear antigen and matrix metalloproteinase–9 were determined using immunohistochemical analyses. Results The expression of Ku70 was clearly inhibited in C6 cells after transfection with Ad-Ku70shRNA. In vitro following transfection, the C6 cells showed improved responses to GKT, including suppression of proliferation and invasion as well as an increased apoptosis index. In vivo following transfection of Ad-Ku70shRNA, the therapeutic efficacy of GKT in rats with C6 gliomas was greatly enhanced and survival times in these animals were prolonged. Conclusions Our data support the potential for downregulation of Ku70 expression in enhancing the radiosensitivity of gliomas. The findings of our study indicate that targeted gene therapy–mediated inactivation of Ku70 may represent a promising strategy in improving the radioresponsiveness of gliomas to GKT.

scholarly journals Adenovirus mediated p53 tumour suppressor gene therapy for human gastric cancer cells in vitro and in vivo

Gut ◽  
1999 ◽  
Vol 44 (3) ◽  
pp. 366-371 ◽  
Author(s):  
M Ohashi ◽  
F Kanai ◽  
H Ueno ◽  
T Tanaka ◽  
K Tateishi ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
Gene Therapy ◽  
P53 Gene ◽  
Human Gastric Cancer ◽  
Wild Type ◽  
Gastric Cancer Cells ◽  
Gastric Cancer Cell Lines ◽  
Wild Type P53

BACKGROUND/AIMSGastric cancer is one of the most prevalent forms of cancer in East Asia. Point mutation of the p53 gene has been reported in more than 60% of cases of gastric cancer and can lead to genetic instability and uncontrolled cell proliferation. The purpose of this investigation was to evaluate the potential of p53 gene therapy for gastric cancer.METHODSThe responses of human gastric cancer cell lines, MKN1, MKN7, MKN28, MKN45, and TMK-1, to recombinant adenoviruses encoding wild type p53 (AdCAp53) were analysed in vitro. The efficacy of the AdCAp53 treatment for MKN1 and MKN45 subcutaneous tumours in nude mice was assessed in vivo.RESULTSp53-specific growth inhibition was observed in vitro in two of four gastric cancer cell lines with mutated p53, but not in the wild type p53 cell line. The mechanism of the killing of gastric cancer cells by AdCAp53 was found, by flow cytometric analysis and detection of DNA fragmentation, to be apoptosis. In vivo studies showed that the growth of subcutaneous tumours of p53 mutant MKN1 cells was significantly inhibited by direct injection of AdCAp53, but no significant growth inhibition was detected in the growth of p53 wild type MKN45 tumours.CONCLUSIONSAdenovirus mediated reintroduction of wild type p53 is a potential clinical utility in gene therapy for gastric cancers.

scholarly journals Culture of C6 glioma cells and establishment of model

2018 ◽  
Vol 1 (1) ◽  
Author(s):  
Shumin Liang ◽  
Yijun Chen ◽  
Kaifei Wang
Keyword(s):  
Formation Rate ◽  
Glioma Cells ◽  
C6 Glioma ◽  
Tumor Formation ◽  
C6 Glioma Cells ◽  
Rat Glioma ◽  
Survival Status ◽  
The Right ◽  
Rat C6 Glioma

Objectives: To establish a simple, reliable, stable rat C6 glioma model, and to provide an operating platform for studying the pathogenesis and prevention and treatment of glioma. Methods: C6 rat glioma cells were cultured in vitro and cultured in the right caudate putamen of SD rats. After implantation, the survival status of rats was observed and the pathological sections were made. HE is dyeing under light microscope. Results: After 6 days of inoculation of C6 glioma cells, the survival status was good at about 7 days, and the intracranial hypertension was more obvious at about 14 days. Most of them were in the endangered state. In the 18 specimens of the rats, except for the accidental death, the other tumor formation rate was 75%. The tumor structure was obviously displaced with the prolongation of the age of the tumor. HE staining was used to observe the formation of glioma in rat brain. Conclusion: The animal model of C6 glioma is reliable and stable. Its tumor growth characteristics and pathological features are similar to those of human glioma, which can be used as an ideal model for clinical study of glioma.

scholarly journals The isolation of an RNA aptamer targeting to p53 protein with single amino acid mutation

2015 ◽  
Vol 112 (32) ◽  
pp. 10002-10007 ◽  
Author(s):  
Liang Chen ◽  
Farooq Rashid ◽  
Abdullah Shah ◽  
Hassaan M. Awan ◽  
Mingming Wu ◽  
...  
Keyword(s):  
P53 Protein ◽  
P53 Gene ◽  
P53 Mutation ◽  
Single Amino Acid ◽  
Human Lung Cancer ◽  
Rna Aptamer ◽  
Wild Type ◽  
Wild Type P53

p53, known as a tumor suppressor, is a DNA binding protein that regulates cell cycle, activates DNA repair proteins, and triggers apoptosis in multicellular animals. More than 50% of human cancers contain a mutation or deletion of the p53 gene, and p53R175 is one of the hot spots of p53 mutation. Nucleic acid aptamers are short single-stranded oligonucleotides that are able to bind various targets, and they are typically isolated from an experimental procedure called systematic evolution of ligand exponential enrichment (SELEX). Using a previously unidentified strategy of contrast screening with SELEX, we have isolated an RNA aptamer targeting p53R175H. This RNA aptamer (p53R175H-APT) has a significantly stronger affinity to p53R175H than to the wild-type p53 in both in vitro and in vivo assays. p53R175H-APT decreased the growth rate, weakened the migration capability, and triggered apoptosis in human lung cancer cells harboring p53R175H. Further analysis actually indicated that p53R175H-APT might partially rescue or correct the p53R175H to function more like the wild-type p53. In situ injections of p53R175H-APT to the tumor xenografts confirmed the effects of this RNA aptamer on p53R175H mutation in mice.

Retinoids Induce t-PA Synthesis by C6 Glioma Cells -Role in Tumoral Haemorrhagic Necrosis

1996 ◽  
Vol 75 (02) ◽  
pp. 332-338 ◽  
Author(s):  
Gilles Pernod ◽  
Guy Amalfltano ◽  
Brigitte Le Magueresse ◽  
Francois Berger ◽  
Benolt Polack ◽  
...  
Keyword(s):  
Cell Line ◽  
Fibrinolytic Activity ◽  
Glioma Cells ◽  
C6 Glioma ◽  
C6 Glioma Cells ◽  
C6 Cell Line ◽  
Haemorrhagic Necrosis ◽  
Retinoid Treatment ◽  
Rat C6 Glioma

SummaryTreatment of rat C6 glioma with high doses of 13 cis-retinoic acid (cRA) was responsible for death related to haemorrhagic necrosis localized to the tumor. Our aim was to explore this adverse effect of retinoid treatment. We show that cRA-treated C6 glioma at 25 mg/kg/day for 18 days exhibits in vivo an increased t-PA activity, which is responsible for a localized tumor fibrinolytic activity. Production of t-PA is supported by specific enhancement of gene expression, as was shown by the increase in t-PA mRNA (X 2.3). This production is a direct effect of cRA when treating the tumor, since tumor cells themself do not produce enough t-PA and treatment of control rats does not increase the t-PA level. t-PA production by rat C6 glioma is in vivo related to the specific synthesis of t-PA by the C6 cell-line. The stimulation of C6 cell-line by cRA in vitro is dose-dependent and reached a maximum for 3 and 30 |iM at the 72nd h. So cRA-treated C6 glioma cells produce t-PA which appears to be the major species associated with the fibrinolytic activity-induced intra-tumoral haemorrhage after exposure to retinoid treatment.

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