Growth and characterization of normal human urothelium in vitro

Reznikoff, Catherine A.; Johnson, Mark D.; Norback, Diane H.; Bryan, George T.

doi:10.1007/bf02619511

Cited by 81 publications

(39 citation statements)

References 35 publications

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“…This pattern essentially reconstructs in two dimensions the normal 3D microarchitecture of urothelium in vivo (Reznikoff et al, 1983). Urothelial differentiation was measured in nonirradiated samples by immunostaining with antibodies against Uroplakin III.…”

Section: Resultsmentioning

confidence: 99%

A proliferation-dependent bystander effect in primary porcine and human urothelial explants in response to targeted irradiation

Belyakov

Folkard

Mothersill³

et al. 2003

Br J Cancer

104

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The aim of this study was to test whether radiation-induced bystander effects are involved in the response of multicellular systems to targeted irradiation. A primary explant technique was used that reconstructed the in vivo microarchitecture of normal urothelium with proliferating and differentiated cells present. Sections of human and porcine ureter were cultured as explants and irradiated on day 7 when the urothelial outgrowth formed a halo around the tissue fragment. The Gray Cancer Institute charge particle microbeam facility allowed the irradiation of individual cells within the explant outgrowth with a predetermined exact number of 3 He 2+ ions (which have very similar biological effectiveness to a-particles). A total of 10 individual cell nuclei were irradiated with 10 3 He 2+ ions either on the periphery, where proliferating cells are located, or at the centre of the explant outgrowth, which consisted of terminally differentiated cells. Samples were fixed 3 days after irradiation, stained and scored. The fraction of apoptotic and micronucleated cells was measured and a significant bystander-induced damage was observed. Approximately 2000 -6000 cells could be damaged by the irradiation of a few cells initially, suggesting a cascade mechanism of cell damage induction. However, the fraction of micronucleated and apoptotic cells did not exceed 1 -2% of the total number of the cells within the explant outgrowth. It is concluded that the bystander-induced damage depends on the proliferation status of the cells and can be observed in an in vitro explant model.

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Section: Resultsmentioning

confidence: 99%

A proliferation-dependent bystander effect in primary porcine and human urothelial explants in response to targeted irradiation

Belyakov

Folkard

Mothersill³

et al. 2003

Br J Cancer

104

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“…Tissue Culture-Human prostate epithelial culture (HPEC) or human urothelial cells were established on collagen-coated dishes in Ham's F-12 supplemented medium containing 1% fetal bovine serum (26,27). HPEC samples were obtained from human cystoprostatectomy specimens (ages 45-60) that did not contain prostate cancer.…”

Section: Methodsmentioning

confidence: 99%

A Loss of Insulin-like Growth Factor-2 Imprinting Is Modulated by CCCTC-binding Factor Down-regulation at Senescence in Human Epithelial Cells

Fu¹,

Schwarze²,

Kenowski³

et al. 2004

Journal of Biological Chemistry

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The imprinted insulin-like growth factor-2 (IGF2) gene is an auto/paracrine growth factor expressed only from the paternal allele in adult tissues. In tissues susceptible to aging-related cancers, including the prostate, a relaxation of IGF2 imprinting is found, suggesting a permissive role for epigenetic alterations in cancer development. To determine whether IGF2 imprinting is altered in cellular aging and senescence, human prostate epithelial and urothelial cells were passaged serially in culture to senescence. Allelic analyses using an IGF2 polymorphism demonstrated a complete conversion of the IGF2 imprint status from monoallelic to biallelic, in which the development of senescence was associated with a 10-fold increase in IGF2 expression. As a mechanism, a 2-fold decrease in the binding of the enhancer-blocking element CCCTC-binding factor (CTCF) within the intergenic IGF2-H19 region was found to underlie this switch to biallelic IGF2 expression in senescent cells. This decrease in CTCF binding was associated with reduced CTCF expression in senescent cells. No de novo increases in methylation at the IGF2 CTCF binding site were seen. The forced down-regulation of CTCF expression using small interfering RNA in imprinted prostate cell lines resulted in an increase in IGF2 expression and a relaxation of imprinting. Our data suggest a novel mechanism for IGF2 imprinting regulation, that is, the reduction of CTCF expression in the control of IGF2 imprinting. We also demonstrate that altered imprinting patterns contribute to changes in gene expression in aging cells.

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“…Some tumours were used for immediate (o24 h) adenoviral transduction. Others were used to initiate explant cultures, essentially according to the method of Reznikoff et al, 30,31 prior to transduction by adenovirus. Briefly, fresh biopsy specimens placed in Transport Medium were minced using a sterile scalpel in a glass Petri dish, collected by centrifugation, resuspended in ''TCC Growth Medium'' (Ham's F12 medium, 1% foetal bovine serum, 5 Â 10 À8 M sodium selenite, 10 À4 M ethanolamine, 2.7 mg/ml D-(+)-glucose, 2 mM Lglutamine, 10…”

Section: +mentioning

confidence: 99%

Adenovirus-mediated gene therapy for bladder cancer: efficient gene delivery to normal and malignant human urothelial cells in vitro and ex vivo

et al. 2003

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Existing local therapies for superficial transitional cell carcinoma (TCC) of the bladder have limited success in preventing progression to life-threatening, muscle-invasive disease, and novel therapies are needed. Recent studies have raised doubts concerning the feasibility of adenovirusmediated gene therapy for bladder cancer. We have therefore investigated adenoviral transduction of normal and malignant human urothelial cells, both as primary cultures and in intact epithelium. All 15 primary normal human urothelial cell lines tested were transduced in vitro by Adv-cmv-b-gal at high efficiency, and better than most human TCC cell lines. Eight primary human TCC explants were also successfully transduced. In contrast, in intact normal urothelium, transduction efficiency was lower, and occurred only in superficial epithelial layers. Expression of the hCAR adenovirus receptor, however, occurred throughout the full thickness of urothelium. Transduction of human TCC biopsy specimens was at least as efficient as intact normal urothelium. We demonstrate for the first time that adenoviral transduction of both normal and malignant human urothelial cells is feasible. A physical barrier, rather than hCAR status, may be the main determinant of transduction of intact epithelium. Clinical trials of adenovirus-mediated gene therapy for superficial bladder cancer are warranted.

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Growth and characterization of normal human urothelium in vitro

Cited by 81 publications

References 35 publications

A proliferation-dependent bystander effect in primary porcine and human urothelial explants in response to targeted irradiation

A proliferation-dependent bystander effect in primary porcine and human urothelial explants in response to targeted irradiation

A Loss of Insulin-like Growth Factor-2 Imprinting Is Modulated by CCCTC-binding Factor Down-regulation at Senescence in Human Epithelial Cells

Adenovirus-mediated gene therapy for bladder cancer: efficient gene delivery to normal and malignant human urothelial cells in vitro and ex vivo

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