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AMPK-ChREBP axis mediates de novo milk fatty acid synthesis promoted by glucose in the mammary gland of lactating goats.

ABSTRACT: To investigate the role of glucose in regulating milk fatty acid synthesis, 6 lactating Guanzhong dairy goats were infused with 0, 60, or 100 g/d glucose via the external pubic artery in a 3 × 3 repeated Latin square experiment. A concomitant in vitro experiment was conducted to investigate possible mechanisms whereby glucose regulates milk fatty acid synthesis. RNA sequencing was used for cellular transcriptome analysis. Drugs, MK-2206, rapamycin, and dorsomorphin were used to block cellular mammalian AMP-activated protein kinase (AMPK), AKT serine/threonine kinase 1, and mechanistic target of rapamycin kinase signaling pathways, respectively. Carbohydrate response element binding protein (ChREBP) was knockdown and overexpressed to investigate its role in regulating milk fatty acid synthesis in mammary epithelial cells. Glucose infusion linearly elevated the concentration of C8:0 (P = 0.039) and C10:0 (P = 0.041) in milk fat while it linearly decreased (P = 0.049) that of C16:0. This result was in agreement with the upregulation of genes related to de novo synthesis of fatty acids and lipid droplet formation, including adipose differentiation-related protein, butyrophilin subfamily 1 member A1, fatty acid synthase (FASN) and ChREBP. Their expression increased (P < 0.05) linearly in the lactating goat mammary gland. In vitro, glucose linearly stimulated the expression of genes related to de novo synthesis of fatty acids and cellular triacylglycerol in cultured mammary epithelial cells. RNA sequencing and inhibition studies revealed that glucose induced transcriptomic changes increasing lipogenic pathways, with AMPK responding to glucose by controlling ChREBP and FASN. Knockdown and overexpression of ChREBP highlighted its essential role in lipogenesis. The knockdown and overexpression of ChREBP protein also revealed an essential role in regulating the de novo synthesis of fatty acids. Collectively, our data highlight that glucose supplementation promotes de novo fatty acid synthesis via the AMPK-ChREBP axis, hence increasing milk fat yield in the goat mammary gland. Results from the current study provide possible strategies to manipulate the fatty acid composition as well as improve ruminant milk quality.

SUBMITTER: Shi H

PROVIDER: S-EPMC9213698 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:The physiology of the sow mammary gland is qualitatively well described and understood. However, the quantitative effect of various biological mechanisms contributing to the synthesis of colostrum and milk is lacking and more complicated to obtain. The objective of this study was to integrate physiological and empirical knowledge of the production of colostrum and milk in a dynamic model of a single sow mammary gland to understand and quantify parameters controlling mammary gland output. In 1983, Heather Neal and John Thornley published a model of the mammary gland in cattle, which was used as a starting point for the development of this model. The original cattle model was reparameterized, modified, and extended to describe the production of milk by the sow mammary gland during lactation and the prepartum production of colostrum as the combined output of immunoglobulins (Ig) and milk. Initially, the model was reparameterized to simulate milk synthesis potential of a single gland by considering biological characteristics and empirical estimations of sows and piglets. Secondly, the model was modified to simulate more accurately the responses to changes in milk removal rates. This was done by linking the ejectable milk storage capacity to the number of secretory cells rather than being constant throughout lactation. Finally, the model was extended to include the prepartum synthesis of milk and the kinetics of Ig into and out of the mammary gland. A progressive capacity of secretory cells to synthesize milk was used to differentiate the time between the onset of milk synthesis and Ig transfer. Changes in maximum milk removal rate, duration of milk ejection, and nursing interval exerted a great impact on the modeled milk output. Changes by ±60% in one of these parameters were capable of increasing milk output by 28% to 39% during the first 4 wk in lactation compared with the reference parameterization. This suggests that the ability of the piglet to remove milk from the gland exerts a key control on milk synthesis during lactation. Modeling colostrum as the combined output of Ig and milk allowed to represent the rapid decline in Ig concentration observed during the first hours after farrowing. In conclusion, biological and empirical knowledge was integrated into a model of the sow mammary gland and constitutes a simple approach to explore in which conditions and to what extent individual parameters influence Ig kinetics and milk production.

Project description:BackgroundIt is well known that feeding a high concentrate (HC) diet to lactating ruminants likely induces subacute ruminal acidosis (SARA) and leads to a decrease in milk fat production. However, the effects of feeding a HC diet for long periods on milk fatty acids composition and the mechanism behind the decline of milk fat still remains poorly understood. The aim of this study was to investigate the impact of feeding a HC diet to lactating dairy goats on milk fat yield and fatty acids composition with an emphasis on the mechanisms underlying the milk fat depression. Seventeen mid-lactating dairy goats were randomly allocated to three groups. The control treatment was fed a low-concentrate diet (35% concentrate, n = 5, LC) and there were two high-concentrate treatments (65% concentrate, HC), one fed a high concentrate diet for a long period (19 wks, n = 7, HL); one fed a high concentrate diet for a short period of time (4 wk, n = 5, HS). Milk fat production and fatty acids profiles were measured. In order to investigate the mechanisms underlying the changes in milk fat production and composition, the gene expression involved in lipid metabolism and DNA methylation in the mammary gland were also analyzed.ResultsMilk production was increased by feeding the HC diet in the HS and HL groups compared with the LC diet (P < 0.01), while the percentage of milk fat was lower in the HL (P < 0.05) but not in the HS group. The total amount of saturated fatty acids (SFA) in the milk was not changed by feeding the HC diet, whereas the levels of unsaturated fatty acids (UFA) and monounsaturated fatty acids (MUFA) were markedly decreased in the HL group compared with the LC group (P < 0.05). Among these fatty acids, the concentrations of C15:0 (P < 0.01), C17:0 (P < 0.01), C17:1 (P < 0.01), C18:1n-9c (P < 0.05), C18:3n-3r (P < 0.01) and C20:0 (P < 0.01) were markedly lower in the HL group, and the concentrations of C20:0 (P < 0.05) and C18:3n-3r (P < 0.01) were lower in the HS group compared with the LC group. However, the concentrations of C18:2n-6c (P < 0.05) and C20:4n-6 (P < 0.05) in the milk fat were higher in the HS group. Real-time PCR results showed that the mRNA expression of the genes involved in milk fat production in the mammary gland was generally decreased in the HL and HS groups compared with the LC group. Among these genes, ACSL1, ACSS1 & 2, ACACA, FAS, SCD, FADS2, and SREBP1 were down-regulated in the mammary gland of the HL group (P < 0.05), and the expressions of ACSS2, ACACA, and FADS2 mRNA were markedly decreased in the HS goats compared with the LC group (P < 0.05). In contrast to the gene expression, the level of DNA methylation in the promoter regions of the ACACA and SCD genes was increased in the HL group compared with the LC group (P < 0.05). The levels of ACSL1 protein expression and FAS enzyme activity were also decreased in the mammary gland of the HL compared with the LC group (P < 0.05).ConclusionsLong-term feeding of a HC diet to lactating goats induced milk fat depression and FAs profile shift with lower MUFAs but higher SFAs. A general down-regulation of the gene expression involved in the milk fat production and a higher DNA methylation in the mammary gland may contribute to the decrease in milk fat production in goats fed a HC diet for long time periods.

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AMPK-ChREBP axis mediates de novo milk fatty acid synthesis promoted by glucose in the mammary gland of lactating goats.

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