Ghrelin O-acyltransferase (GOAT) is essential for growth hormone-mediated survival of calorie-restricted mice

Abstract

Ghrelin O-acyltransferase (GOAT) attaches octanoate to proghrelin, which is processed to ghrelin, an octanoylated peptide hormone that stimulates release of growth hormone (GH) from pituitary cells. Elimination of the gene encoding ghrelin or its receptor produces only mild phenotypes in mice. Thus, the essential function of ghrelin is obscure. Here, we eliminate the Goat gene in mice, thereby eliminating all octanoylated ghrelin from blood. On normal or high fat diets, Goat(-/-) mice grew and maintained the same weights as wild-type (WT) littermates. When subjected to 60% calorie restriction, WT and Goat(-/-) mice both lost 30% of body weight and 75% of body fat within 4 days. In both lines, fasting blood glucose initially declined equally. After 4 days, glucose stabilized in WT mice at 58-76 mg/dL. In Goat(-/-) mice, glucose continued to decline, reaching 12-36 mg/dL on day 7. At this point, WT mice showed normal physical activity, whereas Goat(-/-) mice were moribund. GH rose progressively in calorie-restricted WT mice and less in Goat(-/-) mice. Infusion of either ghrelin or GH normalized blood glucose in Goat(-/-) mice and prevented death. Thus, an essential function of ghrelin in mice is elevation of GH levels during severe calorie restriction, thereby preserving blood glucose and preventing death.

Fig. 1.

Body composition of WT and Goat^−/− mice fed a chow or high fat diet. Male littermates (4-wk-old) were fed ad libitum either the chow diet or the high fat diet for the indicated time. (A) Mice were weighed at weekly intervals. (B and C) Fourteen weeks after diet treatments, the total fat (B) and lean (C) body masses were measured by NMR. Each value represents mean ± SEM of data from six mice.

Fig. 2.

Comparison of WT and Goat^−/− mice fed the chow diet ad libitum or subjected to 60% calorie restriction. Male littermates (8-wk-old) were housed in individual cages and fed ad libitum with the chow diet or subjected to 60% calorie restriction as described in Materials and Methods. Body weight (A) was measured daily at 5:30 p.m., and total fat mass (B) was measured by NMR every 2 or 3 days at 5 p.m. (C) Blood glucose was measured daily with a Bayer Glucometer at 5:30 p.m. (30 min before feeding). (D–F) the mice were euthanized at 5:30 p.m. (before feeding) on the seventh day of calorie restriction, and plasma levels of ghrelin (D), des-acyl ghrelin (E), and GH (F) were determined. Each value represents mean ± SEM of data from six mice. Asterisks (*) denote level of statistical significance (Student's t test) between the WT and Goat^−/− mice under calorie-restricted conditions. *, P < 0.05; **, P < 0.01; ***, P < 0.001.

Fig. 3.

Differential changes in blood glucose and GH levels in WT and Goat^−/− mice subjected to calorie restriction. Male WT and Goat^−/− littermates (8-wk-old) were subjected to a 60% calorie restriction as described in Materials and Methods . The concentrations of blood glucose (A), plasma ghrelin (B), plasma des-acyl ghrelin (C), and plasma GH (D) were measured at 5:30 p.m. (30 min before feeding) on the indicated day. Each value represents mean ± SEM of data from five mice. Asterisks (*) denote level of statistical significance (Student's t test) between WT and Goat^−/− mice. *, P < 0.05; **, P < 0.01; ***, P < 0.001.

Fig. 4.

Comparison of calorie-restricted WT and Goat^−/− mice undergoing continuous s.c. infusion of saline or ghrelin. Alzet osmotic pumps (delivery rate of 0.25 μL/h) filled with saline or saline (●) containing 5 mg/mL ghrelin (○) were inserted s.c. in the interscapular regions of 8-wk-old male littermate WT and Goat^−/− mice. Three days after initiation of the infusion, both groups of mice were placed under 60% calorie restriction that was continued for 7 days. (A and B) Blood glucose levels were measured daily at 5:30 p.m. (30 min before feeding). (C–E) Mice were euthanized at 5:30 p.m. on the seventh day of calorie restriction. Plasma levels of ghrelin (C), des-acyl ghrelin (D), and GH (E) were determined. Each value represents mean ± SEM of data from 6 mice. Asterisks (*) denote level of statistical significance (Student's t test) between WT and Goat^−/− mice infused with saline or ghrelin. *, P < 0.05; **, P < 0.01; ***, P < 0.001.

Fig. 5.

Comparison of calorie-restricted WT and Goat^−/− mice undergoing continuous s.c. infusion of vehicle or GH. Alzet osmotic pumps (delivery rate of 0.25 μL/h) filled with vehicle (●) or vehicle containing 2.5 mg/mL GH (○) were inserted s.c. in the interscapular regions of 8-wk-old male littermate WT and Goat^−/− mice. Three days after initiation of the infusion, both groups of mice were placed under 60% calorie restriction that was continued for 8 days. (A and B) Blood glucose levels were measured daily at 5:30 p.m. (30 min before feeding). (C–E) Mice were euthanized at 5:30 p.m. on the eighth day of calorie restriction. Plasma levels of ghrelin (C), des-acyl ghrelin (D), and GH (E) were determined. Each value represents mean ± SEM of data from five mice. Asterisks (*) denote level of statistical significance (Student's t test) between WT and Goat^−/− mice infused with vehicle or GH. *, P < 0.05; **, P < 0.01; ***, P < 0.001.