The use of cultured cells with defects of citrulline metabolism in diagnosis and in the study of intercellular communication

 

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dc.contributor.advisor Harley, Eric H en_ZA
dc.contributor.author Davidson, James Schonland en_ZA
dc.date.accessioned 2018-02-05T12:35:48Z
dc.date.available 2018-02-05T12:35:48Z
dc.date.issued 1985 en_ZA
dc.identifier.citation Davidson, J. 1985. The use of cultured cells with defects of citrulline metabolism in diagnosis and in the study of intercellular communication. University of Cape Town. en_ZA
dc.identifier.uri http://hdl.handle.net/11427/27258
dc.description.abstract Citrullinemia and argininosuccinic aciduria are two disorders resulting from defects in two consecutive enzymes of the urea cycle, argininosuccinate synthetase and argininosuccinate lyase. Fibroblast cell lines were derived from patients with these disorders and the diagnoses, which had been made on the basis of amino acid levels in plasma and urine, were confirmed by demonstrating that the cell lines were unable to incorporate 14 c-citrulline into protein. DNA from the argininosuccinate synthetase-deficient (ASS⁻) cells was analysed by restriction enzyme digestion and hybridisation to a cDNA probe which had been cloned from human argininosuccinate synthetase mRNA. No defect in the patient's DNA could be demonstrated, indicating that no major deletions in the argininosuccinate synthetase genes were present in this patient. Co-cultures of the ASS⁻ and argininosuccinate lyase-deficient (ASL⁻) fibroblasts were able to incorporate 14 citrulline into protein at rates comparable to normal fibroblasts. This complementation did not require cell fusion, was dependent on cell contact, and was not the result of exchange of metabolites or enzymes via the culture medium. These results indicated that complementation occurred by the exchange of metabolites via intercellular junctions between the two cell types. Co-cultures of ASS⁻ and ASL⁻ cells were used as an assay system for measuring intercellular junctional communication. This allowed quantitation of the effects of pH and extracellular divalent cations on junctional communication. Tumor promoters such as phorbol esters and organochlorine pesticides have been reported to inhibit intercellular junctional communication in other systems, and this inhibitory activity may be related to the mechanism of tumor promotion. The organochlorine pesticide 1,1,1-trichloro- 2,2-bis(p-chlorophenyl)ethane (DDT) was shown to be an inhibitor of junctional communication in ASS⁻/ASL⁻ cocultures. This inhibition was reversible, of rapid onset, and independent of extracellular calcium. The tumor-promoting phorbol ester 12-0-tetradecanoyl-phorbol-13- acetate (TPA) also rapidly induced inhibition of junctional communication. However, co-cultures between Chinese hamster V79 cells, which are deficient in ASS⁻, and ASL⁻ human fibroblasts were more sensitive to inhibition by TPA than the original ASS⁻/ASL⁻ co-cultures. Refractoriness to TPA occurred following prolonged treatment with high concentrations of TPA. Retinoic acid and other retinoids also inhibited junctional communication, and the inhibitory effects of retinoic acid and TPA were additive. The significance of these results in relation to the anti-tumor-promoting activity of retinoic acid is discussed. It is concluded that co-cultures of ASS⁻ and ASL⁻ cells constitute a useful system for providing quantitative measurements of intercellular junctional communication under a wide range of experimental conditions. en_ZA
dc.language.iso eng en_ZA
dc.subject.other Cell communication en_ZA
dc.subject.other Citrulline - Metabolism en_ZA
dc.subject.other Urea - Deficiency en_ZA
dc.subject.other Arginine en_ZA
dc.title The use of cultured cells with defects of citrulline metabolism in diagnosis and in the study of intercellular communication en_ZA
dc.type Doctoral Thesis
uct.type.publication Research en_ZA
uct.type.resource Thesis en_ZA
dc.publisher.institution University of Cape Town
dc.publisher.faculty Faculty of Health Sciences en_ZA
dc.publisher.department Division of Chemical Pathology en_ZA
dc.type.qualificationlevel Doctoral
dc.type.qualificationname PhD en_ZA
uct.type.filetype Text
uct.type.filetype Image
dc.identifier.apacitation Davidson, J. S. (1985). <i>The use of cultured cells with defects of citrulline metabolism in diagnosis and in the study of intercellular communication</i>. (Thesis). University of Cape Town ,Faculty of Health Sciences ,Division of Chemical Pathology. Retrieved from http://hdl.handle.net/11427/27258 en_ZA
dc.identifier.chicagocitation Davidson, James Schonland. <i>"The use of cultured cells with defects of citrulline metabolism in diagnosis and in the study of intercellular communication."</i> Thesis., University of Cape Town ,Faculty of Health Sciences ,Division of Chemical Pathology, 1985. http://hdl.handle.net/11427/27258 en_ZA
dc.identifier.vancouvercitation Davidson JS. The use of cultured cells with defects of citrulline metabolism in diagnosis and in the study of intercellular communication. [Thesis]. University of Cape Town ,Faculty of Health Sciences ,Division of Chemical Pathology, 1985 [cited yyyy month dd]. Available from: http://hdl.handle.net/11427/27258 en_ZA
dc.identifier.ris TY - Thesis / Dissertation AU - Davidson, James Schonland AB - Citrullinemia and argininosuccinic aciduria are two disorders resulting from defects in two consecutive enzymes of the urea cycle, argininosuccinate synthetase and argininosuccinate lyase. Fibroblast cell lines were derived from patients with these disorders and the diagnoses, which had been made on the basis of amino acid levels in plasma and urine, were confirmed by demonstrating that the cell lines were unable to incorporate 14 c-citrulline into protein. DNA from the argininosuccinate synthetase-deficient (ASS⁻) cells was analysed by restriction enzyme digestion and hybridisation to a cDNA probe which had been cloned from human argininosuccinate synthetase mRNA. No defect in the patient's DNA could be demonstrated, indicating that no major deletions in the argininosuccinate synthetase genes were present in this patient. Co-cultures of the ASS⁻ and argininosuccinate lyase-deficient (ASL⁻) fibroblasts were able to incorporate 14 citrulline into protein at rates comparable to normal fibroblasts. This complementation did not require cell fusion, was dependent on cell contact, and was not the result of exchange of metabolites or enzymes via the culture medium. These results indicated that complementation occurred by the exchange of metabolites via intercellular junctions between the two cell types. Co-cultures of ASS⁻ and ASL⁻ cells were used as an assay system for measuring intercellular junctional communication. This allowed quantitation of the effects of pH and extracellular divalent cations on junctional communication. Tumor promoters such as phorbol esters and organochlorine pesticides have been reported to inhibit intercellular junctional communication in other systems, and this inhibitory activity may be related to the mechanism of tumor promotion. The organochlorine pesticide 1,1,1-trichloro- 2,2-bis(p-chlorophenyl)ethane (DDT) was shown to be an inhibitor of junctional communication in ASS⁻/ASL⁻ cocultures. This inhibition was reversible, of rapid onset, and independent of extracellular calcium. The tumor-promoting phorbol ester 12-0-tetradecanoyl-phorbol-13- acetate (TPA) also rapidly induced inhibition of junctional communication. However, co-cultures between Chinese hamster V79 cells, which are deficient in ASS⁻, and ASL⁻ human fibroblasts were more sensitive to inhibition by TPA than the original ASS⁻/ASL⁻ co-cultures. Refractoriness to TPA occurred following prolonged treatment with high concentrations of TPA. Retinoic acid and other retinoids also inhibited junctional communication, and the inhibitory effects of retinoic acid and TPA were additive. The significance of these results in relation to the anti-tumor-promoting activity of retinoic acid is discussed. It is concluded that co-cultures of ASS⁻ and ASL⁻ cells constitute a useful system for providing quantitative measurements of intercellular junctional communication under a wide range of experimental conditions. DA - 1985 DB - OpenUCT DP - University of Cape Town LK - https://open.uct.ac.za PB - University of Cape Town PY - 1985 T1 - The use of cultured cells with defects of citrulline metabolism in diagnosis and in the study of intercellular communication TI - The use of cultured cells with defects of citrulline metabolism in diagnosis and in the study of intercellular communication UR - http://hdl.handle.net/11427/27258 ER - en_ZA


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