Decreased pancreatic islet response to L-leucine in the spontaneously diabetic GK rat: enzymatic, metabolic and secretory data

Diabetologia, Jul 1999

Aims/hypothesis. Pancreatic islets from hereditarily non-insulin-dependent diabetic Goto-Kakizaki (GK) rats have a deficient insulin response not only to d-glucose but also to l-leucine. Our aim was to explain the cellular mechanism(s) underlying the beta-cell unresponsiveness to this amino acid. Methods. Freshly collagenase isolated islets from GK rats and healthy Wistar control rats matched with them for sex and age were compared. Leucine uptake, metabolic fluxes and insulin secretory capacity were investigated on batch incubated-islets. Enzymatic activities were measured on sonicated islets. Results. In GK rat islets, neither leucine transport nor leucine transaminase activity was disturbed. By contrast, 14CO2 production from either l-[U-14C]leucine or l-[1-14C]leucine was decreased. The l-[U-14C]leucine oxidation : l -[1- 14C]leucine decarboxylation ratio was unaffected, indicating that the acetyl-CoA generated from leucine undergoes normal oxidation in the Krebs cycle. The leucine non-metabolizable analogue 2-amino-bicyclo[2,2,1]heptane-2-carboxylic acid induced insulin release and enhanced the secretory response to leucine as in controls, whereas leucine failed to amplify the response to the leucine analogue. Moreover, the potentiating action of l-glutamine on leucine-mediated insulin release was preserved. This coincided with normal glutamate dehydrogenase activity and l-[U-14C]glutamine oxidation. Finally, the secretory response to the leucine deamination product 2-ketoisocaproate was decreased, as was the 2-keto[1-14C]isocaproate oxidation. Conclusion/interpretation. In islet beta cells from GK rats, the defective secretory response to leucine cannot be ascribed to a deteriorated leucine-stimulated glutamate metabolism but rather to an impaired leucine catabolism. A reduced generation of acetyl-CoA from 2-ketoisocaproate, due to the defective oxidative decarboxylation of this keto-acid by the mitochondrial branched-chain 2-ketoacid dehydrogenase, is incriminated. [Diabetologia (1999) 42: 965–977]

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Decreased pancreatic islet response to L-leucine in the spontaneously diabetic GK rat: enzymatic, metabolic and secretory data

M.-H. Giroix 0 C. Saulnier 0 B. Portha 0 0 Laboratory of Nutrition Physiopathology, University of Paris 7 (Denis Diderot) , Paris, France Aims/hypothesis. Pancreatic islets from hereditarily non-insulin-dependent diabetic Goto-Kakizaki (GK) rats have a deficient insulin response not only to d-glucose but also to l-leucine. Our aim was to explain the cellular mechanism(s) underlying the betacell unresponsiveness to this amino acid. Methods. Freshly collagenase isolated islets from GK rats and healthy Wistar control rats matched with them for sex and age were compared. Leucine uptake, metabolic fluxes and insulin secretory capacity were investigated on batch incubated-islets. Enzymatic activities were measured on sonicated islets. Results. In GK rat islets, neither leucine transport nor leucine transaminase activity was disturbed. By contrast, 14CO2 production from either l-[U-14C]leucine or l-[1-14C]leucine was decreased. The l-[U-14C]leucine oxidation : l-[1-14C]leucine decarboxylation ratio was unaffected, indicating that the acetyl-CoA generated from leucine undergoes normal oxidation in the Krebs cycle. The leucine non-metabolizable analogue 2-amino-bicyclo[2,2,1]heptane-2-carboxylic - Corresponding author: Dr. M.-H. Giroix, Laboratoire de Physiopathologie de la Nutrition, ESA CNRS 7059, Universit Paris 7 (Denis Diderot), tour 23/33, 1 er tage, 2 place Jussieu, F-75251 Paris cedex 05, France Abbreviations: BCH, 2-Amino-bicyclo[2,2,1]heptane-2-carboxylic acid; BCKDH, branched-chain 2-ketoacid dehydrogenase; DAB, 3,3-diaminobenzidine-tetrahydrochloride; FAD, flavine adenine dinucleotide; GLUT 2, glucose transporter isoform 2; GK rat, Goto-Kakizaki rat; KIC, 2-ketoisocaproate; PDH, pyruvate dehydrogenase. acid induced insulin release and enhanced the secretory response to leucine as in controls, whereas leucine failed to amplify the response to the leucine analogue. Moreover, the potentiating action of l-glutamine on leucine-mediated insulin release was preserved. This coincided with normal glutamate dehydrogenase activity and l-[U-14C]glutamine oxidation. Finally, the secretory response to the leucine deamination product 2-ketoisocaproate was decreased, as was the 2-keto[1-14C]isocaproate oxidation. Conclusion/interpretation. In islet beta cells from GK rats, the defective secretory response to leucine cannot be ascribed to a deteriorated leucine-stimulated glutamate metabolism but rather to an impaired leucine catabolism. A reduced generation of acetylCoA from 2-ketoisocaproate, due to the defective oxidative decarboxylation of this keto-acid by the mitochondrial branched-chain 2-ketoacid dehydrogenase, is incriminated. [Diabetologia (1999) 42: 965 977] One of the major characteristic features of Type II (non-insulin-dependent) diabetes mellitus is the decreased ability of pancreatic beta cells to release insulin in response to stimulation by the carbohydrate d-glucose, the most potent physiological insulin secretagogue. Such a disturbance in islet function occurs in the GK rat [17], a genetic non-overweight model of Type II diabetes which was obtained through repetitive selective inbreeding of normal Wistar rats with a plasma glucose concentration in the upper normal range, as shown by an oral glucose tolerance test [8, 9]. Even though the reasons for the Fig. 1. Simplified representation for the mechanisms of action of l-leucine in pancreatic islet beta cells. The process by which l-leucine stimulate insulin release is thought to rely on an increase in catabolic fluxes in the islet beta cells via two main routes, first, the own catabolism of l-leucine [1720] and second, the leucine-stimulated glutamate catabolism [2025]. l-leucine entered the islet cells using mainly a Na+-independent transport known as system L [26, 27]. In the first pathway, the amino-acid is metabolized in the beta-cell mitochondria by first undergoing deamination to yield 2-ketoisocaproate (KIC). This reaction (transamination), catalysed by a branched-chain amino-acid aminotransferase (mentioned in the text as leucine transaminase), is coupled with the conversion of a suitable 2-keto-acid to the corresponding amino-acid (e. g. the conversion of 2-ketoglutarate to l-glutamate). Then, an oxidative decarboxylation, catalysed by the branched-chain 2-ketoacid dehydrogenase (BCKDH) multienzyme complex, convertes the deamination product of leucine, KIC, into isovaleryl-coenzyme A. This latter product is further degraded and, after several conversions, forms free acetoacetate and acetylcoenzyme A which enter the citric acid cycle (Krebs cycle) for complete oxidation to CO2 and H2O. In the second pathway, l-leucine acts as an allosteric activator of glutamate dehydrogenase in beta cells. By doing so, it increases the oxidative deamination of endogenous glutamate and, hence, the formation of 2-ketoglutarate which is further oxidized in the Krebs cycle. The accelerated flux through each of these two pathways leads to an increase in necessa (...truncated)


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M.-H. Giroix, C. Saulnier, B. Portha. Decreased pancreatic islet response to L-leucine in the spontaneously diabetic GK rat: enzymatic, metabolic and secretory data, Diabetologia, 1999, pp. 965-977, Volume 42, Issue 8, DOI: 10.1007/s001250051255