Metabolic adaptations during lactogenesis. Fatty acid synthesis in rabbit mammary tissue during pregnancy and lactation

1. Mammary tissue was obtained from rabbits at various stages of pregnancy and lactation and used for tissue-slice incubations (to measure the rate of fatty acid synthesis and CO(2) production) and to determine relevant enzymic activities. A biphasic adaptation in fatty acid synthetic capacity during lactogenesis was noted. 2. The first lactogenic response occurred between day 15 and 24 of pregnancy. Over this period fatty acid synthesis (from acetate) increased 14-fold and the proportions of fatty acids synthesized changed to those characteristic of milk fat (77-86% as C(8:0)+C(10:0) acids). 3. The second lactogenic response occurred post partum as indicated by increased rates of fatty acid synthesis and CO(2) production (from acetate and glucose) and increased enzymic activities. 4. Major increases in enzymic activities between mid-pregnancy and lactation were noted for ATP citrate lyase (EC 4.1.3.8), acetyl-CoA synthetase (EC 6.2.1.1), acetyl-CoA carboxylase (EC 6.4.1.2), fatty acid synthetase, glucose 6-phosphate dehydrogenase (EC 1.1.1.49), and 6-phosphogluconate dehydrogenase (EC 1.1.1.44). Smaller increases in activity occurred with glycerol 3-phosphate dehydrogenase (EC 1.1.1.8) and NADP(+)-isocitrate dehydrogenase (EC 1.1.1.42) and the activity of NADP(+)-malate dehydrogenase (EC 1.1.1.40) was negligible at all periods tested. 5. During pregnancy and lactation there was a close temporal relationship between fatty acid synthetic capacity and the activities of ATP citrate lyase (r=0.94) and acetyl-CoA carboxylase (r=0.90).     

Metabolic adaptations during lactogenesis. Lactose synthesis in rabbit mammary tissue during pregnancy and lactation.

Lactose biosynthesis and relevant enzymatic activity in rabbit mamma ry tissue during various stages of pregnancy and lactation are investigated by using a tissue-slice incubation method in order to understand the temporal relationships. Ovulation was induced in 27 New Zealand white rabbits and they were bred by artificial insemination. Sacrifice occurred on days 15, 24, and 29 of pregnancy, and day 2, 5, 8, 15, and 22 post partum. Nucleic acids were extracted and concentratons of DNA determined spectrophotometrically at 600 nm with diphenylamine reagent and RNA determined with orcinal reagent. The tissue incubations were made with (U-14C) glucose. (14C) lactose was then separated by paper chromatography from unchanged radioactive glucose. Enzyme analysis including determining the activities of phosphoglucomutase, UDP-glucose pyrophosphorylase, and UDP-glucose 4-epimerase. Lactose synthase was determined, as well as, hexokinase. A biphasic adaptation in the rate of lactose synthesis and in the RNA concentration was noted during lactogenesis. The 1st increase in the rate of lactose biosynthes is occurred between days 15 and 24 of pregnancy. A 2nd substantial increase was noted immediately post partum. The overall rate of lactose biosynthesis increased 12-fold from day 24 of pregnancy to day 15 of lactation post partum, and then decreased from 15 to 22 days post partum. The RNA concentration/g wet weight of tissue and the ratio of RNA/DNA closely represented the biphasic ability of the mammary-tissue slice to synthesize lactose. Increases in the activities of UDP-glucose 4-epimerase and lactose synthase were most closely correlated with increases in the rate of lactose biosynthesis. UDP-glucose pyrophosphor ylase activity was unrelated with the ability to synthesize lactose, and hexokinase and phosphoglucomutase activities were variable during pregnancy and lactation. Lactose synthase activity was present by day 15 of pregnancy, but the ability to synthesize lactose was undetected until day 24 of pregnancy.     

Metabolic adaptations in goat mammary tissue during pregnancy and lactation

Metabolic adaptations of goat mammary tissue during pregnancy and lactation were monitored in serial biopsies of the tissue. Changes in the synthetic capacity of secretory cells were studied by combining measurements of enzyme activities with short-term culture of mammary explants to measure lactose, casein and total protein synthesis. By these criteria, the main phase of mammary differentiation began in late pregnancy and was essentially complete by Week 5 of lactation, coinciding with the achievement of peak milk yield. While milk yield declined after Week 5, the activities of key enzymes expressed per mg DNA and the rates of lactose and casein synthesis in mammary explants were maintained over a considerable period. The results suggest that changes in the synthetic capacity of epithelial cells may account for much of the rise in milk yield in early lactation, but are not responsible for the declining phase of milk production characteristic of lactation in ruminants.     

gamma-Glutamyl transpeptidase in rat mammary tissue. Activity during lactogenesis and regulation by prolactin.

The increased activity of gamma-glutamyl transpeptidase (EC 2.3.2.2) in rat mammary tissue at the onset of lactation was shown to involve prolactin. The timing and magnitude of these changes and its prolactin requirement suggest an important role in mammary function.     

Mammary extraction of plasma triglycerides in the cow during lactogenesis

• 1.1. In control cows, extraction of triglycerides from the circulation by the mammary gland increased abruptly and markedly, and the concentration of triglycerides in arterial plasma fell, on the day of parturition.
• 2.2. In cows that had fluid regularly removed from the mammary glands before parturition, these changes occurred at varying times from more than 4 days before parturition in an animal that secreted triglyceride copiously to 24 hr before parturition in an animal that secreted little triglyceride pre-partum
• 3.3. In all animals, the time when triglyceride extraction increased was close to the time when K⁺ concentration in the mammary secretion became maximal.

     

The onset of mammary extraction of plasma triglycerides and circulating levels of progesterone during lactogenesis in the cow and goat

In untreated cows and goats, the onset of mammary extraction of plasma triglyceride was sometimes detected several hours before parturition, prior to removal of secretion from the mammary glands by offspring and when circulating progesterone concentrations were low. In cows and goats that had secretion regularly removed from the mammary glands late in gestation, the onset of triglyceride extraction occurred up to several days before parturition, when circulating progesterone concentrations were moderately high (up to 3.9 ng/ml in the cow and 6.2 ng/ml in the goat).     

Morphology and lactose synthesis in tissue culture of mammary alveoli isolated from lactating mice

Summary Mammary epithelial cells from lactating mice synthesize and secrete lactose in culture and retain many features of their in vivo morphology if mammary glands are only partially dissociated to alveoli, rather than completely dissociated to single cells. After 5 d in culture lactose synthesis by alveoli cultured on floating collagen gels is 10 to 20 times higher than in cultures of single cells on floating collagen gels. Moreover, mammary alveoli in culture retain sensitivity to lactogenic hormones; the synthesis of lactose by alveoli depends on the continued presence of insulin and either hydrocortisone or prolactin. In addition, within alveoli the original juxtaposition of constituent epithelial cells is retained, and cells are cuboidal and have many microvilli and fat droplets. In contrast, alveoli on attached gels flatten and lose their secretory morphology. These results indicate that the shape of the cells, presence of lactogenic hormones, and maintenance of epithelial:epithelial cell contacts are required for maintenance of mammary epithelial cell differentiation in culture. This research was supported by Grants CA-16392 and AG-02909 from the National Institutes of Health and Institutional Grant IN 119 from the American Cancer Society.     

Fatty acid synthesis in chick embryonic heart and liver during development.

Fatty acid synthesis by subcellular fractions of heart and liver of chick embryos at varying stages of development has been studied. Fatty acid synthetase activity is associated with the embryonic heart at early stages of development, as suggested by substrate requirement, Schmidt decarboxylation of synthesized fatty acids and gas liquid chromatographic identification of the products as palmitic and stearic acids. The fatty acid synthetase activity decreases in heart cytosol with age of the embryo and is absent in the newly hatched chick and in older chicken. The acetyl CoA carboxylase activity is negligible in embryonic and adult chicken heart. The fatty acid synthetase activity in liver is low, but measurable during the entire embryonic development. The activity increases by about three-fold on hatching and thereafter in fed, newly hatched chicks by about 35-fold, over the basal embryonic activity. The acetyl and malonyl transacylase activities in the heart and liver cytosols during development followed closely the fatty acid synthetase activities in heart and liver, respectively. A non-coordinate induction of fatty acid synthetase and acetyl CoA carboxylase activities in liver was observed during development. The microsomal chain elongation in liver and heart followed the pattern of fatty acid synthetase activity in liver and heart, respectively. The mitochondrial chain elongation in embryonic heart is initially low and increases with age; while this activity in liver is higher in early stages of embryonic development than in the older embryos and the chicks. Measurement of lipogenesis from acetate-1-¹⁴C by liver and heart slices from chick embryos and newly hatched chicks support the conclusions reached in the studies with the subcellular fractions. The results obtained indicate that the major system of fatty acid synthesis in embryonic and adult heart is the mitochondrial chain elongation. In embryonic liver, fatty acid synthesis proceeds by chain elongation, while the de novo system is the major contributor to the lipogenic capacity of the liver after hatching.     

Fatty acid biosynthesis in rabbit mammary gland during pregnancy and early lactation

The pattern of fatty acids synthesized by mammary-gland explants from rabbits during pregnancy and early lactation has been studied. From day 12 to day 18 of pregnancy, long-chain (C(14:0)-C(18:1)) fatty acids were the major products. From day 18 to day 21 of pregnancy there was an increase of up to 12-fold in the rate of fatty acid synthesis per unit wet weight of tissue that was almost exclusively caused by the synthesis of octanoic fatty acid and decanoic fatty acid, which are characteristic of rabbit milk. These medium-chain fatty acids were mainly incorporated into triglycerides. From day 22 to day 27 of pregnancy there was little change in the rate of fatty acid synthesis and the proportions of fatty acids synthesized were essentially the same as those synthesized by the lactating gland, i.e. 80-90% octanoic acid plus decanoic acid. About 2-4 days before parturition a second lipogenic stimulus occurred, although the pattern of fatty acids synthesized did not change.     

Fatty acid synthesis from amino acids in sheep adipose tissue

The rates of incorporation of 14C from 14C labelled acetate, glucose, alanine, leucine, isoleucine and valine into fatty acids has been measured in perirenal adipose tissue from foetal lambs and 8-month-old sheep, and into both fatty acids and acylglycerol glycerol in adipose tissue from 3-year-old sheep and 220-240 g female rats. Rates of incorporation of 14C from amino acids into fatty acids were much lower in adipose tissue from sheep (at all three ages) than from rats, whereas rates of incorporation of 14C into acylglycerol glycerol were either greater in sheep adipose tissue or the same as in rat adipose tissue. The rate of incorporation of 14C from amino acids into fatty acids decreased in the order leucine greater than alanine greater than isoleucine greater than valine in adipose tissue from rats and foetal lambs, and in the order leucine greater than alanine = isoleucine greater than valine in adipose tissue from 8-month- and 3-year-old sheep. Amino acids make a very small contribution to fatty acid synthesis in adipose tissue from sheep at all stages of development examined while fatty acids are a minor product of amino acid metabolism in sheep adipose tissue. The study provides further evidence for an important role for ATP-citrate lyase in restricting the utilization of acetyl-CoA generated in the mitochondria for fatty acid synthesis.