Increased Fatty Acid Re-esterification by PEPCK Overexpression in Adipose Tissue Leads to Obesity Without Insulin Resistance

Abstract

Adipose tissue glyceroneogenesis generates glycerol 3-phosphate, which could be used for fatty acid esterification during starvation. To determine whether increased glyceroneogenesis leads to increased fat mass and to explore the role of obesity in the development of insulin resistance, we overexpressed PEPCK, a regulatory enzyme of glyceroneogenesis in adipose tissue. Transgenic mice showed a chronic increase in PEPCK activity, which led to increased glyceroneogenesis, reesterification of free fatty acids (FFAs), increased adipocyte size and fat mass, and higher body weight. In spite of increased fat mass, transgenic mice showed decreased circulating FFAs and normal leptin levels. Moreover, glucose tolerance and whole-body insulin sensitivity were preserved. Skeletal muscle basal and insulin-stimulated glucose uptake and glycogen content were not affected, suggesting that skeletal muscle insulin sensitivity is normal in transgenic obese mice. Our results indicate the key role of PEPCK in the control of FFA re-esterification in adipose tissue and, thus, the contribution of glyceroneogenesis to fat accumulation. Moreover, they suggest that higher fat mass without increased circulating FFAs does not lead to insulin resistance or type 2 diabetes in these mice. Diabetes 51: 624 -630, 2002

Chronically increased glucose uptake by adipose tissue leads to lactate production and improved insulin sensitivity rather than obesity in the mouse

“…The entire Ap2-Gck chimeric gene (8.5 kb) was microinjected into fertilised eggs as described by our group elsewhere [4]. Mice were tested for the presence of the transgene by Southern blot with a probe radiolabelled with deoxycytidine 5′-[ 32 P]triphosphate (Amersham Pharmacia Biotech, Piscataway, NJ, USA) by random oligopriming (Amersham).…”

“…RNA and protein analysis Total RNA was obtained as previously described by our group [4]. Northern blot, RT-PCR using Omniscript Reverse transcriptase kit (Qiagen, Hilden, Germany) and real-time quantitative PCR using QuantiTect SYBR Green (Qiagen) and SmartCyclerII II (Cepheid, Sunnyvale, CA, USA) was performed following the manufacturers' instructions and as detailed in the ESM.…”

“…The sequences of oligonucleotide primers are given in the ESM (ESM Table 1). Western blot analysis was performed using standard procedures with total homogenates of WAT, skeletal muscle and liver [4]. The antibodies used are described in the ESM.…”

“…Metabolic studies using radiolabelled tracers The in vivo glucose utilisation index in epididymal WAT was determined by the intravenous flash injection of deoxy-D-[ 3 H]glucose (Amersham) as previously described by Franckhauser et al [4]. Uptake of deoxy-D-[ 3 H]glucose was measured in isolated soleus muscles, in 3T3-L1 adipocytes and in isolatedadipocyte suspension incubated with or without insulin as previously described [4,28,29] and in the ESM.…”

“…The synthesis of triacylglycerol depends on the formation of glycerol 3-phosphate, which may originate either from glucose through glycolysis, or from lactate and pyruvate through glyceroneogenesis [1][2][3]. Overproduction of the key enzyme of glyceroneogenesis, phosphoenolpyruvate carboxykinase (PEPCK), in adipose tissue leads to increased glycerol 3-phosphate, NEFA re-esterification, adipocyte hypertrophy and obesity [4]. This is associated with higher susceptibility to diet-induced insulin resistance and obesity [5].…”