Decreased pancreatic islet response to L-leucine in the spontaneously diabetic GK rat: enzymatic, metabolic and secretory data
M.-H. Giroix
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C. Saulnier
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B. Portha
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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
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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)