Chicken LEAP2 Level Substantially Changes with Feed Intake and May Be Regulated by CDX4 in Small Intestine

Abstract

Ghrelin O-acyltransferase (GOAT), ghrelin, and GHSR have been reported to play important roles that influence feed intake in mammals. LEAP2, an endogenous antagonist of GHSR, plays an important role in the regulation of feed intake. However, chicken ghrelin has also been reported to have an inhibitory effect on feed intake. The role of the GOAT–Ghrelin–GHSR–LEAP2 axis in chicken-feed intake remains unclear. Therefore, it is necessary to systematically evaluate the changes in the tissue expression levels of these genes under different energy states. In this study, broiler chicks in different energy states were subjected to starvation and feeding, and relevant gene expression levels were measured using quantitative real-time PCR. Different energy states significantly modulated the expression levels of LEAP2 and GHSR but did not significantly affect the expression levels of GOAT and ghrelin. A high expression level of LEAP2 was detected in the liver and the whole small intestine. Compared to the fed group, the fasted chicks showed significantly reduced LEAP2 expression levels in the liver and the small intestine; 2 h after being refed, the LEAP2 expression of the fasted chicks returned to the level of the fed group. Transcription factor prediction and results of a dual luciferase assay indicated that the transcription factor CDX4 binds to the LEAP2 promoter region and positively regulates its expression. High expression levels of GHSR were detected in the hypothalamus and pituitary. Moreover, we detected GHSR highly expressed in the jejunum—this finding has not been previously reported. Thus, GHSR may regulate intestinal motility, and this aspect needs further investigation. In conclusion, this study revealed the function of chicken LEAP2 as a potential feed-intake regulator and identified the potential mechanism governing its intestine-specific expression. Our study lays the foundations for future studies on avian feed-intake regulation.

The dietary regulation of LEAP2 depends on meal composition in mice

“…The fact that LEAP2 may show an acute regulation according to glycogen or lipid levels in the liver and intestine, respectively, supports the notion that LEAP2 is a peptide that is upregulated by a positive energy balance (i.e., an increase in plasma LEAP2 or Leap2 expression levels reported to be associated with food intake/refeeding and obesity) 7,10,15,20 . Since LEAP2 deletion has been reported to both increase food intake, body weight, and exaggerate hepatic fat accumulation in female mice, 16 the increase in LEAP2 that occurs with a meal challenge may act to counteract further expansion of energy stores.…”

“…7 The fact that LEAP2 may show an acute regulation according to glycogen or lipid levels in the liver and intestine, respectively, supports the notion that LEAP2 is a peptide that is upregulated by a positive energy balance (i.e., an increase in plasma LEAP2 or Leap2 expression levels reported to be associated with food intake/refeeding and obesity). 7,10,15,20 Since LEAP2 deletion has been reported to both increase food intake, body weight, and exaggerate hepatic fat accumulation in female mice, 16 the increase in LEAP2 that occurs with a meal challenge may act to counteract further expansion of energy stores. It should however be mentioned that although the correlation between energy substrates and Leap2 expression suggests that LEAP2 is regulated according to local energy availability, a causal link as well as the mechanism behind this regulation (e.g., the role of insulin-regulated glucose disposal), remains to be determined.…”

“…14,26,27 However, the expression of ghrelin receptors in the liver is not consistently reported. 28 A definitive role of SREBP1 in LEAP2 regulation in addition to the potential regulation by other transcription factors (e.g., CDX4 which also binds to the LEAP2 promotor region and has been suggested to regulate LEAP2 in chicken 20 ) remains to be investigated.…”

“…Plasma LEAP2 levels have for example been shown to decrease with fasting (and exposure to ketone bodies), whereas concentrations increase with glucose administration or subsequent refeeding in rodents 7,10,15,19 . Decreased Leap2 expression levels in the liver and small intestine have also been reported in fasted versus fed broiler chicks, with expression levels returning to the levels observed in the fed chicks upon refeeding, although it should be mentioned that ghrelin exerts opposite functions on food intake in avian species 20 . In humans, increased circulating levels of LEAP2 are found in obese individuals, and postprandial increases in LEAP2 have been reported to correlate with BMI 7,19 .…”

“…7,10,15,19 Decreased Leap2 expression levels in the liver and small intestine have also been reported in fasted versus fed broiler chicks, with expression levels returning to the levels observed in the fed chicks upon refeeding, although it should be mentioned that ghrelin exerts opposite functions on food intake in avian species. 20 In humans, increased circulating levels of LEAP2 are found in obese individuals, and postprandial increases in LEAP2 have been reported to correlate with BMI. 7,19 Plasma LEAP2 levels have also been reported to be increased in ob/ob mice and in mice with diet-induced obesity-an increase that in the latter could be reversed by weight loss.…”