Expression of genes related to energy metabolism and the unfolded protein response in dairy cow mammary cells is affected differently during dietary supplementation with energy from protein and fat.

Secretory capacity of bovine mammary glands is enabled by a high number of secretory cells and their ability to use a range of metabolites to produce milk components. We isolated RNA from milk fat to measure expression of genes involved in energy-yielding pathways and the unfolded protein response in mammary glands of lactating cows given supplemental energy from protein (PT) and fat (FT) tested in a 2 × 2 factorial arrangement. We hypothesized that PT and FT would affect expression of genes in the branched-chain AA catabolic pathway and tricarboxylic acid (TCA) cycle based on the different energy types (aminogenic versus lipogenic) used to synthesize milk components. We also hypothesized that the response of genes related to endoplasmic reticulum (ER) homeostasis via the unfolded protein response would reflect the increase in milk production stimulated by PT and FT. Fifty-six multiparous Holstein-Friesian dairy cows were fed a basal total mixed ration (34% grass silage, 33% corn silage, 5% grass hay, and 28% concentrate on a dry matter basis) for a 28-d control period. Experimental rations were then fed for 28 d, consisting of (1) low protein, low fat (LP/LF); (2) high protein, low fat (HP/LF); (3) low protein, high fat (LP/HF); or (4) high protein and high fat (HP/HF). To obtain the high-protein (HP) and high-fat (HF) diets, intake of the basal ration was restricted and supplemented isoenergetically (net energy basis) with 2.0 kg/d rumen-protected protein (soybean + rapeseed, 50:50 mixture on dry matter basis) and 0.68 kg/d hydrogenated palm fatty acids on a dry matter basis. RNA from milk fat samples collected on d 27 of each period underwent real-time quantitative PCR. Energy from protein increased expression of BCAT1 (branched-chain amino acid transferase 1) mRNA, but only at the LF level, and tended to decrease expression of mRNA encoding the main subunit of the branched-chain keto-acid dehydrogenase complex. mRNA expression of malic enzyme, a proposed channeling route for AA though the TCA cycle, was decreased by PT, but only at the LF level. Expression of genes associated with de novo fatty acid synthesis was not affected by PT or FT. Energy from fat had no independent effect on genes related to ER homeostasis. At the LF level, PT activated XBP1 (X-box binding protein 1) mRNA. At the HF level, PT increased mRNA expression of the gene encoding GADD34 (growth arrest and DNA damage-inducible 34). These findings support our hypothesis that mammary cells use aminogenic and lipogenic precursors differently for milk component production when dietary intervention alters AA and fatty acid supply. They also suggest that mammary cells respond to increased AA supply through mechanisms of ER homeostasis, dependent on the presence of FT.

Mammary gland utilization of amino acids and energy metabolites differs when dairy cow rations are isoenergetically supplemented with protein and fat.

Mammary gland utilization of AA and other metabolites in response to supplemental energy from protein (PT) and supplemental energy from fat (FT) was tested in a 2 × 2 factorial arrangement using a randomized complete block design. Fifty-six Holstein-Friesian dairy cows were adapted during a 28-d control period to a basal total mixed ration consisting of 34% grass silage, 33% corn silage, 5% grass hay, and 28% concentrate on a dry matter (DM) basis. Experimental rations were fed for 28 d immediately following the control period and consisted of (1) low protein, low fat (LP/LF), (2) high protein, low fat (HP/LF), (3) low protein, high fat (LP/HF), and (4) high protein, high fat (HP/HF). To obtain the high-protein (HP) and high-fat (HF) diets, intake of the basal ration was restricted and supplemented isoenergetically [net energy (MJ/d) basis] with 2.0 kg/d rumen-protected protein (soybean + rapeseed, 50:50 mixture on a DM basis) and 0.68 kg/d hydrogenated palm fatty acids on a DM basis. Arterial and venous blood samples were collected on d 28 of both periods. Isoenergetic supplements (MJ/d) of protein and fat independently and additively increased milk yield, PT increased protein yield, and FT increased fat yield. A PT × FT interaction affected arterial concentration of all essential AA (EAA) groups, where they increased in response to PT by a greater magnitude at the LF level (on average 35%) compared with the HF level (on average 14%). Mammary gland plasma flow was unaffected by PT or FT. Supplementation with PT tended to decrease mammary clearance of total EAA and decreased group 1 AA clearance by 19%. In response to PT, mammary uptake of total EAA and group 2 AA increased 12 and 14%, respectively, with significantly higher uptake of Arg, Ile, and Leu. Energy from fat had no effect on mammary clearance or uptake of any AA group. The mammary gland uptake:milk protein output ratio was not affected by FT, whereas PT increased this ratio for EAA and group 2 AA. Arterial plasma insulin concentration decreased in response to FT, in particular on the HP/HF diet, as indicated by a PT × FT interaction. Arterial concentrations of nonesterified fatty acids, triacylglycerol, and long-chain fatty acids increased in response to FT, and concentrations of ß-hydroxybutyrate and acetate decreased in response to FT only at the HP level. Mammary clearance and uptake of triacylglycerol and long-chain fatty acids increased in response to FT. Energy from PT and FT increased lactose yield despite no change in arterial glucose concentration or mammary glucose uptake. Mammary-sequestered glucose with PT or FT was used in the same amount for lactose synthesis, and a positive net mammary glucose balance was found across all treatments. Results presented here illustrate metabolic flexibility of the mammary gland in its use of aminogenic versus lipogenic substrates for milk synthesis.

Feed and nitrogen efficiency are affected differently but milk lactose production is stimulated equally when isoenergetic protein and fat is supplemented in lactating dairy cow diets.

Fifty-six Holstein-Friesian cows were used in a randomized complete block design to test the effects of supplemental energy from protein (PT) and fat (FT) on lactation performance and nutrient digestibility in a 2 × 2 factorial arrangement. During the control period, cows were adapted for 28 d to a basal total mixed ration consisting of 34% grass silage, 33% corn silage, 5% grass hay, and 28% concentrate on a dry matter (DM) basis. Experimental rations were fed for 28 d immediately following the control period and consisted of (1) low protein, low fat (LP/LF), (2) high protein, low fat (HP/LF), (3) low protein, high fat (LP/HF), or (4) high protein and high fat (HP/HF). To obtain the HP and HF diets, intake of the basal ration was restricted and supplemented isoenergetically (net energy basis) with 2.0 kg/d of rumen-protected protein (soybean + rapeseed, 50:50 mixture on DM basis) and 0.68 kg/d of hydrogenated palm fatty acids (FA) on a DM basis. Milk production and composition, nutrient intake, and apparent digestibility were measured during the final 7 d of the control and experimental periods. No interaction was found between PT and FT on milk production and composition. Yields of milk, fat- and protein-corrected milk, and lactose increased in response to PT and FT and lactose concentration was unaffected by treatment. Milk protein concentration and yield increased in response to PT, and protein yield tended to increase in response to FT. Milk fat concentration and yield increased in response to FT and were unaffected by PT. Milk urea concentration increased and nitrogen efficiency decreased in response to PT. Feed and nitrogen efficiency were highest on the LP/HF diet and both parameters increased in response to FT, whereas milk urea concentration was not affected by FT. Energy from fat increased the concentration and yield of ≥16-carbon FA in milk and decreased the concentration of FA synthesized de novo, but had no effect on their yield. Concentration and yield of de novo-synthesized FA increased in response to PT. Concentration and yield of polyunsaturated FA increased and decreased in response to PT and FT, respectively. Apparent total-tract digestibility of crude fat decreased in response to PT, and FT increased crude protein digestibility. Energy supplementation through rumen-inert hydrogenated palm FA appears to be an efficient feeding strategy to stimulate milk production with regard to feed and nitrogen efficiency compared with supplementing an isoenergetic level of rumen-protected protein.