316 Maternal citrulline supplementation increases fetal growth and programs postnatal pancreatic development in the lamb.

Abstract Arginine, a conditionally essential amino acid, has been shown to reduce the incidence of fetal growth restriction following maternal malnutrition, while stimulating development of the pancreas. Despite these beneficial outcomes, supplementing pregnant ruminants with arginine is challenging and cost-prohibitive due to its extensive catabolism by ruminal microbes. Interestingly citrulline, the precursor for arginine synthesis, bypasses ruminal degradation, making it a potentially valuable alternative to increase availability of arginine in gestating ewes. While maternal arginine supplementation has been shown to have beneficial effects on the developing sheep fetus, postnatal development of these lambs has not been evaluated. This study aimed to determine if maternal citrulline supplementation from days 28 to 140 of gestation would increase fetal growth rates and program postnatal growth and performance of the offspring. Superovulated Rambouillet donor ewes were synchronized and bred to a single Hampshire sire by laparoscopic insemination. Embryos were collected by uterine flushing on day 6 and a single grade one blastocyst was transferred into Hampshire recipient ewes. Beginning on day 21 of gestation ewes were individually housed on concrete to control dietary intake. From gestational day 28-140, pregnant ewes received 50% of their NRC requirements in addition to one of two dietary supplements, alanine (boxnutra.com) at 0.61% of their diet (isonitrogenous control; n=17) or citrulline (boxnutra.com) at 0.40% of their diet (CIT, n=17). On day 140 of gestation (term=~147), amino acid supplementation ceased, and ewes returned to a 100% NRC dietary intake level. At lambing, birth weight and sternal circumference were assessed, and a blood sample was collected from each lamb prior to suckling. Lambs were reared on their mother until necropsy at 60 days of age (PND60). Results indicate that lambs born to CIT-treated ewes were heavier (P<0.05) at birth, and this increase in weight remained through weaning on PND60 (P<0.05). At necropsy, the absolute weight of the pancreas, brain, liver, and small intestine were greater (P<0.05) in CIT lambs. Further, when expressed as a proportion of total body mass, pancreatic mass was greater per gram of body weight (P<0.05) than controls. Examination of pancreatic morphometry showed no difference in percentage of endocrine (P=0.29) or exocrine pancreas (P=0.33), but when adjusted for overall pancreas weight, both endocrine (P=0.03) and exocrine (P<0.01) pancreas areas showed an increase in weight. As a likely consequence of increased endocrine pancreas mass, circulating insulin levels were greater (P=0.05) in CIT lambs at birth compared to controls. In contrast, circulating glucose concentrations were increased (P<0.05) in CIT lambs on PND60. These results suggest that maternal citrulline supplementation can serve as an effective alternative to arginine to improve fetal growth during periods of maternal malnutrition, with benefits persisting through weaning. (Supported by a USDA/NIFA grant #2022-67015-37272)