Influence of energy level and nicotinic acid supplementation on apoptosis of blood leukocytes of periparturient dairy cows.

The periparturient period of dairy cows is accompanied by an immunosuppression that leaves the animal more susceptible to infections and metabolic disorders. Non-esterified fatty acids (NEFA) and beta-hydroxybutyrate (BHB) which peak shortly after parturition due to lipolysis are known to impair immune cell functions. Niacin with its well-known anti-lipolytic effect may have the ability to ameliorate this situation. Additionally, niacin shows also anti-inflammatory effects that may be beneficial to the immune status of the cow. To address this 29 multiparous and 18 primiparous German Holstein cows were subjected to four different feeding groups. They were fed either a ration with a high concentrate proportion of 60% (HC), or a low concentrate proportion of 30% (LC). After parturition both concentrate levels were reduced to 30% and increased again to 50% either within 16days (LC-group) or within 24days (HC-group). Half of the animals received either 24g per day of nicotinic acid from 42days prepartum until 24days postpartum (LC-NA, HC-NA) or no supplement (LC-CON, HC-CON). Apoptosis in polymorphonuclear leukocytes (PMN) and peripheral blood mononuclear cells (PBMC) was examined with an Annexin V and propidium iodide (PI) based fluorescence flow cytometry assay and distinguished into early apoptotic (Annexin V positive and PI negative) and late apoptotic (Annexin V and PI positive) cells. Additionally, the pro-apoptotic gene BAX, the effector caspase CASP3, and the anti-apoptotic genes BCL2 and BCL-xL, as well as the NFκB subunit RELA were quantified by real-time PCR in blood leukocytes. All variables showed time dependencies that were mainly related to parturition (p<0.01). Early apoptotic PBMC were significantly affected by concentrate level showing higher numbers of apoptotic cells in the HC groups (p=0.029). PBMC were characterized by a more pronounced apoptosis than PMN and seemed to be more susceptible to the changes that occur around parturition. The genes BAX and CASP3 were positively correlated (0.631) and their peak preceded the apoptotic peak around parturition in the blood leukocytes. The LC animals showed a decrease in BCL2 expression before parturition, whereas the HC animals showed a continuous increase in BCL2 mRNA abundance (p=0.059). RELA correlated stronger with the pro-apoptotic genes (0.715 and 0.650 with BAX and CASP3 respectively) and its expression was higher in primiparous than in multiparous cows (p=0.011). Nicotinic acid supplementation did show some influence in increasing numbers of early apoptotic PMN and late apoptotic PBMC between 42 and 100 DIM.

Longitudinal changes in adipose tissue of dairy cows from late pregnancy to lactation. Part 1: The adipokines apelin and resistin and their relationship to receptors linked with lipolysis.

The transition from pregnancy to lactation is characterized by major changes in glucose and adipose tissue metabolism. Anti- and prolipolytic pathways mediated via the hydroxycarboxylic acid receptors 1 (HCAR1) and 2 (HCAR2) and tumor necrosis factor-α receptor 1 (TNFR1), as well as the adipokines apelin and resistin, are likely involved in regulating these processes. This study aimed to determine the mRNA abundance of the aforementioned receptors in both subcutaneous and visceral adipose tissue, to characterize the adipokine concentrations in serum, and to test the effects of feeding diets with either high or low portions of concentrate and a concomitant niacin supplementation from late gestation to early lactation. Twenty pluriparous German Holstein cows were all kept on the same silage-based diet until d 42 antepartum, when they were allocated to 2 feeding groups: until d 1 antepartum, 10 animals each were assigned to either a high-concentrate (60:40 concentrate-to-roughage ratio) or a low-concentrate diet (30:70). Both groups were further subdivided into a control and a niacin group, the latter receiving 24 g/d of nicotinic acid from d -42 until 24. From d 1 to 24 postpartum, the concentrate portion was increased from 30 to 50% for all cows. Biopsies of subcutaneous (SCAT) and retroperitoneal adipose tissue (RPAT) were taken at d -42, 1, 21, and 100 relative to parturition. Blood samples were drawn along with the biopsies and on d -14, 3, 7, 14, and 42. The concentrations of the adipokines apelin and resistin in serum were measured via ELISA. The mRNA of the 3 receptors in AT was quantified as well as the protein abundance of HCAR2 by Western blot. The feeding regimen did not affect the variables examined. The concentrations of apelin remained fairly constant during the observation period, whereas the resistin concentrations increased toward parturition and decreased to precalving levels within 1 wk after calving. The mRNA abundance of HCAR1, HCAR2, and TNFR1 changed in SCAT and RPAT during the considered time period. For the HCAR2 protein, time-dependent changes were restricted to SCAT. The mRNA abundance of all receptors was greater in RPAT than in SCAT. The tissue-specific correlations observed between the receptors point to a link between these factors and may indicate different regulatory roles in the respective tissues. This study provides insight into the complex metabolic adaptations during the transition period and supports a differential regulation of lipolysis among SCAT and RPAT in dairy cows.

Changes in lipid metabolism and ß-adrenergic response of adipose tissues of periparturient dairy cows affected by an energy-dense diet and nicotinic acid supplementation.

Dairy cattle will mobilize large amounts of body fat during early lactation as an effect of decreased lipogenesis and increased lipolysis. Regulation of lipid metabolism involves fatty acid synthesis from acetate and ß-adrenergic-stimulated phosphorylation of hormone-sensitive lipase (HSL) and perilipin in adipocytes. Although basic mechanisms of mobilizing fat storage in transition cows are understood, we lack a sufficiently detailed understanding to declare the exact regulatory network of these in a broad range of dairy cattle. The objective of the present study was to quantify 1) protein abundance of fatty acid synthase (FAS), 2) extent of phosphorylation of HSL and perilipin in vivo, and 3) ß-adrenergic stimulated lipolytic response of adipose tissues in vitro at different stages of the periparturient period. We fed 20 German Holstein cows an energy-dense or an energetically adequate diet prepartum and 0 or 24 g/d nicotinic acid (NA) supplementation. Biopsy samples of subcutaneous and retroperitoneal adipose tissue were obtained at d 42 prepartum (d -42) and at d 1, 21, and 100 postpartum (d +1, d +21, d +100, respectively). To assess ß-adrenergic response, tissue samples were incubated with 1 µ isoproterenol for 90 min at 37°C. The NEFA and glycerol release, as well as HSL and perilipin phosphorylation, was measured as indicators of in vitro stimulated lipolysis. In addition, protein expression of FAS and extent of HSL and perilipin phosphorylation were measured in fresh, nonincubated samples. There was no effect of dietary energy density or NA on the observed variables. The extent of HSL and perilipin phosphorylation under isoproterenol stimulation was strongly correlated with the release of NEFA and glycerol, consistent with the functional link between ß-adrenergic-stimulated protein phosphorylation and lipolysis. In the nonincubated samples, FAS protein expression was decreased at d +1 and d +21, whereas HSL and perilipin phosphorylation increased from d -42 to d +1 and remained at an increased level throughout the first 100 d of lactation. In vitro lipolytic response was significant in prepartum samples at times when in vivo lipolysis was only minimally activated by phosphorylation. These data extend our understanding of the complex nature of control of lipolysis and lipogenesis in dairy cows and could be useful to the ongoing development of systems biology models of metabolism to help improve our quantitative knowledge of the cow.