TY - JOUR
T1 - Respiratory chain activity and mitochondrial DNA content of nonpurified and purified pancreatic islet cells
AU - Conget, Ignacio
AU - Barrientos, Antoni
AU - Manzanares, Josep M.
AU - Casademont, Jordi
AU - Viñas, Odette
AU - Barceló, J.
AU - Nunes, Virginia
AU - Gomis, Ramon
AU - Cardellach, Francesc
N1 - Funding Information:
From the Servei d'Endocrinologia i Diabetis, Grup d'Investigaci6 Muscular. Departament de Medicina. and Servei d'Immunologia, Hospital Clfnic. Universitat de Barcelona, Barcelona: and Departa-ment de Genktica Molecular, lnstitut de Recerca Oncol6gica, Hosl~italet de Llobregat, Barcelona, Spain. Submitted December 26, 1996; accepted March 26. 1997. Supported by Direccion General de Investigacion Ciencia y Tecnogia Grants No. PB93-0019 from the Ministerio de Educaci6n y Ciencia (MEC), SAF913-93 from Comision InterdepartamentaI de Ciencia y Tecnogia, and 95/0267 from Fondo lnvestigaciones de l.a Seguridad Social, and ir~ part by Grant No. PF92-37289410 from the MEC 'A.B.) and a postresidency fellowship from the Hospital Clinic de Barcelona (J.M.M.). Address reprint requests to lgnacio Conger, MD. Servei d'Endocrinologia i Diabetis. Hospital Clfnic, Universitat de Barcelona. Villarroel 170. Barcelona 08036. Spain. Copyright © 1997 by W.B. Saunders Company 0026-0495/97/4609-0002503.00/0
PY - 1997
Y1 - 1997
N2 - Considerable interest has recently focused on the possible role of alterations in mitochondrial activity and mutations in the mitochondrial genome for the development of non-insulin-dependent diabetes. Our study aimed at investigating the normal mitochondrial respiratorY chain activity of nonpurified and purified islet cells to further explore whether some diabetic states are associated with alterations of mitochondrial oxidative processes. For this purpose, pancreatic islets were isolated from Wistar rats. Unpurified islet cells were obtained in the presence of trypsin and DNAse, and purified β and non-β cells were prepared by autofluorescence-activated sorting using a flowcytometer. Intact cell respiration and substrate oxidation in digitonin-permeabilized cells were measured polarographically with a Clark oxygen electrode in a micro-water-jacketed cell. Specific activity of the individual complexes of the respiratory chain was determined spectrophotometrically in unpurified islet cells. The relative amount of mitochondrial (mtDNA) and nuclear (nDNA) DNA in all three cell populations and in rat brain and skeletal muscle was estimated by dot blotting. The intact cell respiration of unpurified islet cells corresponds to the mean of values obtained for β and non-β islet cells. Oxidation rates of different substrates by permeabilized β cells were lower than those for unpurified and non-β cells. The amount of mtDNA relative to nDNA was similar in all three groups of cells, and was also similar to that obtained from brain and skeletal muscle. In summary, we have described mitochondrial respiratory chain activity in unpurified, β, and non-β islet cells. Our results represent an initial step in investigating the potential pathogenic role that alterations in oxidative phosphorylation could play in some diabetic states.
AB - Considerable interest has recently focused on the possible role of alterations in mitochondrial activity and mutations in the mitochondrial genome for the development of non-insulin-dependent diabetes. Our study aimed at investigating the normal mitochondrial respiratorY chain activity of nonpurified and purified islet cells to further explore whether some diabetic states are associated with alterations of mitochondrial oxidative processes. For this purpose, pancreatic islets were isolated from Wistar rats. Unpurified islet cells were obtained in the presence of trypsin and DNAse, and purified β and non-β cells were prepared by autofluorescence-activated sorting using a flowcytometer. Intact cell respiration and substrate oxidation in digitonin-permeabilized cells were measured polarographically with a Clark oxygen electrode in a micro-water-jacketed cell. Specific activity of the individual complexes of the respiratory chain was determined spectrophotometrically in unpurified islet cells. The relative amount of mitochondrial (mtDNA) and nuclear (nDNA) DNA in all three cell populations and in rat brain and skeletal muscle was estimated by dot blotting. The intact cell respiration of unpurified islet cells corresponds to the mean of values obtained for β and non-β islet cells. Oxidation rates of different substrates by permeabilized β cells were lower than those for unpurified and non-β cells. The amount of mtDNA relative to nDNA was similar in all three groups of cells, and was also similar to that obtained from brain and skeletal muscle. In summary, we have described mitochondrial respiratory chain activity in unpurified, β, and non-β islet cells. Our results represent an initial step in investigating the potential pathogenic role that alterations in oxidative phosphorylation could play in some diabetic states.
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U2 - 10.1016/S0026-0495(97)90265-1
DO - 10.1016/S0026-0495(97)90265-1
M3 - Article
C2 - 9284883
AN - SCOPUS:0030800845
VL - 46
SP - 984
EP - 987
JO - Metabolism: Clinical and Experimental
JF - Metabolism: Clinical and Experimental
SN - 0026-0495
IS - 9
ER -