Cd36 and molecular mechanisms of insulin resistance in the stroke-prone spontaneously hypertensive rat.

Abstract

Insulin resistance is of pathogenic importance in several common human disorders including type 2 diabetes, hypertension, obesity and hyperlipidemia, but the underlying mechanisms are unknown. The spontaneously hypertensive rat (SHR) is a model of these human insulin resistance syndromes. Quantitative trait loci (QTLs) for SHR defects in glucose and fatty acid metabolism, hypertriglyceridemia, and hypertension map to a single region on rat chromosome 4. Genetic analysis of an SHR derived from a National Institutes of Health colony led to the identification of a causative mutation in the SHR Cd36. We have investigated glucose and fatty acid metabolism in the stroke-prone SHR (SHRSP). We demonstrate defects in insulin action on 2-deoxy-D-glucose transport (SHRSP 3.3 ± 1.5 vs. 21.0 ± 7.4 pmol · min -1 · [20 µl packed cells] -1 , SHRSP vs. WKY, respectively, P = 0.01) and inhibition of catecholaminestimulated lipolysis (P < 0.05 at all concentrations of insulin) in adipocytes isolated from SHRSP. In contrast, basal levels of catecholamine-stimulated nonesterified free fatty acid (NEFA) release and plasma levels of NEFA are similar in SHRSP and WKY. These results are in agreement with the data on the SHR.4 congenic strain, which suggested that the QTL containing Cd36 mutations accounted for the entire defect in basal catecholamine action but only for ~40% of the SHR defect in insulin action. In the SHR, both abnormalities appear consequent of defective Cd36 expression. Because Cd36 sequence and expression are apparently normal in SHRSP, it is likely that the molecular mechanism for defective insulin action in this strain is caused by a gene(s) different than Cd36. Diabetes

Reduced insulin-stimulated GLUT4 bioavailability in stroke-prone spontaneously hypertensive rats

“…We have recently extended these observations in a close relative of the SHR, the stroke-prone spontaneously hypertensive rat (SHRSP). We demonstrated that adipocytes from these animals similarly exhibit attenuated insulin-stimulated glucose uptake and a reduced ability to suppress isoproterenol-stimulated lipolysis [5]. Furthermore, we demonstrated that insulin-stimulated glucose uptake is also attenuated in isolated skeletal muscles of SHRSP rats [6].…”

“…Mouse anticaveolin antibodies were from BD Biosciences (Oxford, UK). All other reagents were from sources described previously [5,6].…”

“…Animals Rat strains used in this study were inbred SHRSP and WKY rats obtained from the NIH and maintained in the Division of Cardiovascular and Medical Sciences, BHF Glasgow Cardiovascular Research Centre, University of Glasgow, as described previously [5,6,9]. Male rats were housed under controlled conditions of temperature (21°C) and light (12 h light-dark cycle), were fed on normal rat diet (rat and mouse no.…”

“…Isolation of adipocytes Epididymal adipocytes were prepared from WKY and SHRSP rats using the collagenase method as described previously [5]. Buffer was removed and cells were resuspended in 2 vol lysis buffer (50 mmol/l Tris-HCl pH 7.4 at 4°C, 50 mmol/l NaF, 5 mmol/l Na pyrophosphate, 1 mmol/l Na orthovanadate, 1 mmol/l EDTA, 1 mmol/l EGTA, 250 mmol/l mannitol, 1% [v/v] Triton X-100, 1 mmol/l dithiothreitol, 0.1 mmol/l PMSF, 0.1 mmol/l benzamidine, 5 μg/ml soybean trypsin inhibitor).…”

“…Electrophoresis and immunoblotting Proteins were electrophoresed on SDS polyacrylamide gels and transblotted onto nitrocellulose membranes as described previously [5,6]. Immunolabelled proteins were visualised using horseradish peroxidase-conjugated secondary antibody and the enhanced chemiluminescence system (Amersham Biosciences).…”