Deletion of prolyl carboxypeptidase attenuates the metabolic effects of diet-induced obesity

Abstract

α-Melanocyte-stimulating hormone (α-MSH) is a critical regulator of energy metabolism. Prolyl carboxypeptidase (PRCP) is an enzyme responsible for its degradation and inactivation. PRCP-null mice (PRCP(gt/gt)) showed elevated levels of brain α-MSH, reduced food intake, and a leaner phenotype compared with wild-type controls. In addition, they were protected against diet-induced obesity. Here, we show that PRCP(gt/gt) animals have improved metabolic parameters compared with wild-type controls under a standard chow diet (SD) as well as on a high-fat diet (HFD). Similarly to when they are exposed to SD, PRCP(gt/gt) mice exposed to HFD for 13 wk showed a leaner phenotype due to decreased fat mass, increased energy expenditure, and locomotor activity. They also showed improved insulin sensitivity and glucose tolerance compared with WT controls and a significant reduction in fasting glucose levels. These improvements occured before changes in body weight and composition were evident, suggesting that the beneficial effect of PRCP ablation is independent of the adiposity levels. In support of a reduced gluconeogenesis, liver PEPCK and G-6-Pase mRNA levels were reduced significantly in PRCP(gt/gt) compared with WT mice. A significant decrease in liver weight and hepatic triglycerides were also observed in PRCP(gt/gt) compared with WT mice. Altogether, our data suggest that PRCP is an important regulator of energy and glucose homeostasis since its deletion significantly improves metabolic parameters in mice exposed to both SD and HFD.

Fig. 1.

Effect of high-fat diet (HFD) on wild-type (WT) and PRCP^gt/gt mice. A: graphs showing body weight changes in WT and PRCP^gt/gt mice during HFD. B and C: graphs showing the %fat (B) and %lean mass (C) in 13-wk HFD mice. *P < 0.05 compared with WT mice; **P < 0.01. PRCP, prolylcarboxypeptide.

Fig. 2.

Effect of PRCP deletion of energy metabolism during HFD. A: results of the energy expenditure in WT and PRCP^gt/gt mice exposed to HFD for 13 wk. Gray area represents dark phases. B: results of the energy expenditure as total in the 24-h cycle and in the dark and light phases of the cycle. C–E: results of the O₂ consumed (V̇o₂), CO₂ produced (V̇co₂), and their ratio. F: results of the locomotor activity of the WT and PRCP^gt/gt mice in 48 h and in the dark and light phases of 2 cycles (48 h). *P < 0.05; **P < 0.01. RQ, respiratory quotient.

Fig. 3.

Effect of PRCP ablation on brown adipose tissue (BAT). Graph showing the results of uncoupling protein-1 (UCP1) mRNA levels measured by real-time PCR in the BAT of WT and PRCP^gt/gt mice on standard chow diet (SD) or HFD for 13 wk. *P < 0.05.

Fig. 4.

Circulating leptin (A), adiponectin (B), active ghrelin (C), and total ghrelin (D) levels in WT and PRCP^gt/gt mice on either SD or HFD for 13 wk. A: corresponding to the results on the reduced fat mass, PRCP^gt/gt mice showed lower serum leptin levels than the WT controls when exposed to either SD or HFD. B: adiponectin levels were found reduced in PRCP^gt/gt mice compared with their WT controls when exposed to SD; no difference was observed in adiponectin levels in mice exposed to HFD. C: HFD reduced active ghrelin levels in WT and PRCP^gt/gt mice significantly. However, whereas HFD reduced total ghrelin levels in WT mice significantly, no decrease in total ghrelin levels was observed in PRCP^gt/gt mice on HFD compared with PRCP^gt/gt mice on SD. *P < 0.05; **P < 0.01; ***P < 0.001.

Fig. 5.

Effect of PRCP deletion on glucose homeostasis. Glucose (left) and insulin tolerance tests (right) in WT and PRCP^gt/gt mice exposed to either SD (A and B) or 13-wk HFD (C and D). A and C: both PRCP^gt/gt experimental groups showed significantly lower fasting glucose levels compared with their WT control groups. B and D: increased insulin sensitivity was found in PRCP^gt/gt mice when exposed to SD and 13-wk HFD. E–G: no difference in body weight (E), %fat (F), or lean mass (G) between WT and PRCP^gt/gt mice exposed to HFD for 7–8 wk. When glucose and insulin tolerance tests were performed in these mice, a significant difference in glucose tolerance (H) and insulin sensitivity (I) was noticed between WT and PRCP^gt/gt mice, suggesting that the differences in glucose levels observed in SD and 13-wk HFD were not due to changes in adiposity levels.

Fig. 6.

A: results of circulating insulin levels measured in fasted WT and PRCP^gt/gt mice on SD. Insulin levels were significantly lower in PRCP^gt/gt mice compared with WT controls. B: results of insulin levels in fed WT and PRCP^gt/gt mice on SD or 13-wk HFD. Although increased, insulin levels in PRCP^gt/gt mice on 13-wk HFD were not statistically significant compared with WT and PRCP^gt/gt mice on SD. Circulating nonesterified fatty acid (NEFA; C) and triglyceride levels (D) were measured in mice on 13-wk HFD. Note that NEFA levels were reduced significantly in PRCP^gt/gt mice on HFD compared with all other experimental groups. D: HFD induced a significant increase in triglyceride levels in both WT and PRCP^gt/gt mice compared with mice on SD. **P < 0.01; ***P < 0.001.

Fig. 7.

Effect of PRCP deletion on liver. A: liver weight in WT and PRCP^gt/gt mice on SD and HFD for either 7–8 or 13 wk. Note that whereas HFD induced increase in liver weight in WT mice, no difference was observed in PRCP^gt/gt mice. B: hepatic triglyceride (TGL) levels. PRCP^gt/gt mice showed statistically lower TGL levels on both diets (SD and 13 wk-HFD) compared with their WT controls. Thirteen weeks of HFD induced a significant increase in hepatic TGL in both WT and PRCP^gt/gt mice. However, the levels of TGL in 13-wk HFD PRCP^gt/gt mice were significantly lower than those of WT mice exposed to the same diet. C–F: representative pictures of liver histology from PRCP^gt/gt mice (C and E) and WT mice (D and F) on HFD for 7–8 (C and D) or 13 wk (E and F). Note the increase in lipid accumulation in the 13-wk HFD WT mouse (F) compared with the 13-wk HFD PRCP^gt/gt mouse (E). G and H: results from the real-time PCR for liver phosphoenolpyruvate carboxykinase (PEPCK; G) and glucose-6-phosphatase (G-6-Pase; H) in WT and PRCP^gt/gt mice exposed to SD and HFD for 7 and 13 wk. *P < 0.05; **P < 0.01; ***P < 0.001. Bar scale in F (for C–F) represents 50 μm.