The objective of this study was to explore the metabolic profiles of pregnancy malnutrition induced by feed restriction (FR) and the counteracting effects of glycerol and rumen-protected choline chloride supplementation. Two feeding trials were conducted. In the first experiment, twenty pregnant Hu sheep carrying multiple fetuses with a gestation period of 108 d were randomly divided into two groups. The ewes in the control (CON) group were offered 100 % of their nutritional requirements as recommended by the National Research Council (NRC), while the FR group was offered 30 % of feed intake of CON for 15 d. In the second experiment, eighteen pregnant Hu sheep were offered a feed intake comprising 30 % of the NRC-recommended nutritional requirements twice daily. The sheep were randomly divided into three groups: the FR group in the second experiment (FR2), with no supplementation, the glycerol (GLY) group, which received 40 ml of glycerol per d, and the rumen-protected choline chloride (RPC) group, which received 10 g of rumen-protected choline chloride per d for 9 d. In the first experiment, the urine metabolome of sixteen ewes showed significant difference between the CON group and FR group. Compared with the CON group, FR decreased the level of d-glucose, lactic acid, levoglucosan, α-ketoglutarate, phosphohydroxypyruvic acid, glucose 6-phosphate and the methyl donors, while increasing the level of pyruvate, fumaric acid and carnitines in urine. Both the GLY and RPC treatments counteracted some of these changes and modulated the urine metabolome in advanced pregnant ewes suffering from malnutrition.

Pathogenesis of pregnancy toxemia (PT) is believed to be associated with the disruption of lipid metabolism. The present study aimed to explore the underlying mechanisms of lipid metabolism disorder in the livers of ewes with PT. In total, 10 pregnant ewes were fed normally (control group) whereas another 10 were subjected to 70% level feed restriction for 15 days to establish a pathological model of PT. Results showed that, as compared with the controls, the levels of blood β-hydroxybutyrate (BHBA), non-esterified fatty acids (NEFAs) and cholesterol were greater (P<0.05) and blood glucose level was lower (P<0.05) in PT ewes. The contents of NEFAs, BHBA, cholesterol and triglyceride were higher (P<0.05) and glycerol content was lower (P<0.05) in hepatic tissues of PT ewes than those of the controls. For ewes with PT, excessive fat vacuoles were observed in liver sections stained with hematoxylin–eosin; furthermore, inner structures of hepatocytes including nuclei, mitochondria and endoplasmic reticulum were damaged seriously according to the results of transmission electron microscope. Real-time PCR data showed that compared with the controls, the expression of hepatic genes involved in fatty acid oxidation (FAO) and triglyceride synthesis (TGS) was enhanced (P<0.05) whereas that related to acetyl-CoA metabolism (ACM) was repressed (P<0.05) in PT ewes. Generally, our results showed that negative energy balance altered the expression of genes involved in FAO, ACM and TGS, further caused lipid metabolism disorder in livers, resulting in PT of ewes. Our findings may provide the molecular basis for novel therapeutic strategies against this systemic metabolic disease in sheep.