Relationships Between Gastrointestinal Permeability, Heat Stress, And Milk Production in Lactating Dairy Cows.

Heat stress (HS) is a global issue that decreases farm profits and compromises animal welfare. To distinguish between the direct and indirect effects of HS, 16 multiparous Holstein cows approximately 100 d in milk were assigned to one of 2 treatments: pair fed to match HS cow intake, housed in thermoneutral conditions (PFTN, n = 8) or cyclical HS (n = 8). All cows were subjected to 2 experimental periods. P1 consisted of a 4 d thermoneutral period with ad libitum intake. During P2, the HS cows were housed in cyclical HS conditions with a temperature humidity index (THI) ranging from 76 to 80 and the PFTN cows were exposed to a constant THI of 64 for 4 d. DMI of the PFTN cow was intake matched to the HS cows. Milk yield, milk composition, rectal temperature, and respiration rate were recorded twice daily, blood was collected daily via a jugular catheter, and cows were fed twice daily. On d 3 of each period, Cr-EDTA and sucralose were orally administered and recovered via 24 h total urine collection to assess gastrointestinal permeability (GIP). All data were analyzed using the GLIMMIX procedure in SAS. The daily data collected in P1 was averaged and used as a covariate if deemed significant in the model. HS decreased voluntary intake by 35% and increased rectal temperature and respiration rate (38.4 vs 39.4°C and 40 vs 71 respirations/min, respectively). HS reduced dry matter intake (DMI) by 35% which accounted for 66% of the decrease in milk yield. The yield, and not concentration, of milk protein, fat, and other solids were lower in the HS cows on d 4 of P2. Milk urea nitrogen (MUN) was higher and plasma urea nitrogen (PUN) tended to be higher on d 3 and d 4 of HS. Glucose was 7% lower in the HS cows and insulin was 71% higher in the HS cows than the PFTN cows on d 4 of P2. No difference in lipopolysaccharide binding protein (LBP) was observed. HS cows produced 7 L/d more urine than PFTN cows. No differences were detected in the urine concentration or percentage of the oral dose recovered for Cr-EDTA or sucralose. In conclusion, HS was responsible for 34% of the reduction of milk yield. The elevated MUN and the tendency for elevated PUN indicate a whole-body shift in nitrogen metabolism. No differences in GIP or LBP were observed. These results indicate that, under conditions of this experiment, activation of the immune system by gut derived lipopolysaccharide was not responsible for the decreased milk yield observed during HS.

Milk production and anatomical udder capacity changes of udder halves subjected to increased milking frequency at two stages of lactation.

This study aimed to characterize the effects of increased milking frequency (IMF) at early and mid-lactation on milk yield and its association with changes in cistern and alveolar capacity. Fourteen multiparous Holstein cows were subjected to IMF using the unilateral frequent milking method from 3 to 24 d in milk (DIM). At mid-lactation, cows were randomly assigned to 1 of 2 treatments: control or repeated. From 150 to 170 DIM, IMF treatment was reimposed in the repeated group. During IMF, left udder halves were milked 2× and right udder halves were milked 4× daily. To separate individual milk yields of udder halves, separate buckets were used to collect samples from each udder half. Milk samples and milk yield from right and left udder halves were collected on d 150, 170, 200, 230, 260, and 290 of lactation. Alveolar and cistern capacity were measured 26 h after the last milking at 140 and 172 DIM using an oxytocin inhibitor. Cistern and alveolar capacity were measured by evaluating the milk harvested after oxytocin inhibitor and oxytocin administration, respectively. Udder half difference yields were calculated by subtracting left half yield from right half yield. At 170 DIM, the udder half difference in repeated was 2.27 kg greater than the udder half difference in control. Udder halves milked 4× produced more milk and protein than 2× udder halves in the repeated group at 170, 200, 230, and 260 DIM. Cumulative (150 to 290 DIM) and carry over (200 to 290 DIM) udder half differences in milk yield were similar between the control and repeated treatments. Alveolar volume was similar between udder halves milked 2× or 4× at 140 DIM, while cistern volume was larger for udder halves milked 4× than 2× in early lactation. There was no difference between alveolar or cistern volume proportion in udder halves milked 2× or 4× before mid-lactation IMF. After 20 d IMF for the repeated group, alveolar volume was similar between control and repeated independent of udder half milking frequency. However, repeated held 4.9 kg more cistern milk than control. Control treatment udder halves had a greater alveolar proportion than repeated treatment udder halves. As expected, the cistern proportion was smaller in control and larger in repeated after mid-lactation IMF. IMF at early and mid-lactation enhances milk and protein yield largely during differential milking frequency regimens. The lack of enhancement in milk yield after IMF might be associated with a different response to IMF in the mammary gland at early versus mid-lactation. Based on our results, we conclude that udder halves subjected to early and mid-lactation IMF had increased cistern volume capacity.

Uneven milking intervals are adequate to achieve the benefits of increased milking frequency in early lactation.

Increasing milking frequency (MF) increases milk yield (MY) and farm profit, and optimal milking intervals (MI) prevent milk production decline. The objective of this experiment was to compare the MY effect of even and uneven 4 times daily (4×) MI in early lactation under increased MF. Fourteen multiparous and 6 primiparous cows were milked using unilateral frequent milking, with right udder halves milked 4× and left udder halves milked 2 times daily (2×) for 20 d in early lactation starting on d 5 postpartum. Ten (7 multiparous and 3 primiparous) cows were allocated evenly based on parity and assigned to either the even or the uneven MI groups distinguished by intervals of 9:3:9:3 h or 6:6:6:6 h. The left and right udder halves were milked at 0100 and 1300 h. The right udder glands were additionally milked at 0400 and 1600 h for the uneven MI group and at 0700 and 1900 h for the even MI group. Milk from each udder half was weighed and sampled for components on the final day of treatment and at 60, 120, 180, 240, and 300 d in milk. The overall effect of 4× milking on the right udder halves was a 5.96 ± 0.70 kg/d increase in MY on d 21 of unilateral frequent milking compared with the 2× udder halves. This elevated MY continued through 300 d in milk and averaged 1.56 ± 0.70 kg/d. Increased MF in early lactation increased the udder half difference in total yield throughout a 300-d lactation by 508 kg for milk, 25 kg for milk fat, and 15 kg for milk protein. Increased MF in early lactation increased milk component yields, but there were no differences between MI groups. The lack of treatment difference may be beneficial to farmers. The ability to achieve the same increased MY effect with uneven MI may optimize labor efficiency because early-lactation cows could be milked at the beginning and end of milking sessions. Farmers would not have to add additional milking sessions to achieve the enhanced MY response regardless of normal milking session length.

A flow cytometric method for measuring and isolating mammary epithelial cells from bovine milk.

Sampling frequent time points of mammary signaling pathways is not possible with tissue biopsies. We have validated a flow cytometry and cell sorting procedure for isolating live bovine mammary epithelial cells from somatic cell populations in milk using butyrophilin 1A1 as a marker for mammary epithelial cells and CD45 as a marker for hematopoietic cells. Hoechst 33342 staining and propidium iodide exclusion were used to select for nucleated live cells. Positive selection of butyrophilin (BTN)-expressing cells was performed by fluorescence-activated cell sorting. Quantitative real-time PCR performed on mRNA isolated from these cells showed a 226-fold increase in κ-casein (CSN3) mRNA expression in BTN single-positive cells compared with unsorted cells, whereas CD45 single-positive cells showed a significant decrease. A negative selection strategy for cells not expressing the hematopoietic cell marker CD45 also resulted in a cell population with a 196-fold increase in CSN3 mRNA expression compared with unsorted cells. We found no enrichment of CSN3 mRNA expression after sorting cells using cytokeratin antibodies. The noninvasive assays described here allow for daily or more frequent sampling time points for measurement of mammary epithelial cells during the course of lactation.

Characterization of raft microdomains in bovine mammary tissue during lactation: How they are modulated by fatty acid treatments.

The objective of the current study was first to characterize lipid raft microdomains isolated as detergent-resistant membranes (DRM) from mammary gland tissue, and second to determine how dietary fatty acids (FA) such as conjugated linoleic acid (CLA), 19:1 cyclo, and long-chain n-3 polyunsaturated FA affect lipid raft markers of mammary cells, and to finally establish relationships between these markers and lactation performance in dairy cows. Eight Holstein cows were used in a replicated 4 × 4 Latin square design with periods of 28 d. For the first 14 d, cows received daily an abomasal infusion of (1) 406 g of a saturated FA supplement (112 g of 16:0 + 230 g of 18:0) used as a control; (2) 36 g of a CLA supplement (13.9 g of trans-10,cis-12 18:2) + 370 g of saturated FA; (3) 7 g of Sterculia fetida oil (3.1 g of 19:1 cyclo, STO) + 399 g of saturated FA; or (4) 406 g of fish oil (55.2 g of cis-5,cis-8,cis-11,cis-14,cis-17 20:5 + 59.3 g of cis-4,cis-7,cis-10,cis-13,cis-16,cis-19 22:6, FO). Mammary biopsies were harvested on d 14 of each infusion period and were followed by a 14-d washout interval. Cholera toxin subunit B, which specifically binds to ganglioside M-1 (GM-1), a lipid raft marker, was used to assess its distribution in DRM. Infusions of CLA, STO, and FO were individually compared with the control, and significance was declared at P ≤ 0.05. Milk fat yield was decreased with CLA and FO, but was not affected by STO. Milk lactose yield was decreased with CLA and STO, but was not affected by FO. Mammary tissue shows a strong GM-1-signal enrichment in isolated DRM from mammary gland tissue. Caveolin (CAV) and flotillin (FLOT) are 2 proteins considered as lipid raft markers and they are present in DRM from mammary gland tissue. Distributions of GM-1, CAV-1, and FLOT-1 showed an effect of treatments determined by their subcellular distributions in sucrose gradient fractions. Regardless of treatments, data showed positive relationships between the yield of milk fat, protein, and lactose, and the abundance GM-1 in DRM fraction. Milk protein yield was positively correlated with relative proportion of FLOT-1 in the soluble fraction, whereas lactose yield was positively correlated with relative proportion of CAV-1 in the DRM fractions. Infusion of CLA decreased mRNA abundance of CAV-1, FLOT-1, and FLOT-2. Regardless of treatments, a positive relationship was observed between fat yield and mRNA abundance of FLOT-2. In conclusion, although limited to a few markers, results of the current experiment raised potential links between variation in specific biologically active component of raft microdomains in bovine mammary gland and lactation performances in dairy cows.

Abomasally infused SFA with varying chain length differently affect milk production and composition and alter hepatic and mammary gene expression in lactating cows.

The aim of the present study was to compare the effects of post-ruminally infused fat supplements, varying in fatty acid (FA) chain length, on animal performance, metabolism and milk FA. Eleven multiparous Holstein dairy cows were used in a replicated incomplete 3 × 3 Latin square design with 7-d periods, separated by 7-d washouts. Treatments were administered as abomasal infusions of enrichments providing 280 g/d of FA: (1) palmitic acid (98·4 % 16 : 0; PA), (2) caprylic and capric acids (56·2 % 8 : 0, 43·8 % 10 : 0; medium-chain TAG (MCT)) and (3) stearic acid (99·0 % 18 : 0; SA). Relative to PA, SA decreased the efficiency of fat-corrected milk production, which was associated with a tendency for higher DM intake and lower FA absorption with SA, whereas MCT was not different from PA for these variables. Milk fat concentration and yield were increased by PA relative to SA, but only fat yield tended to be greater relative to MCT. Relative to PA, MCT increased milk fat concentration of FA < 16 C, whereas SA increased FA > 16 C. Expression of mammary stearoyl-coA desaturase 1 was lower with SA than with PA. Relative to PA, liver expression of adenosine monophosphate-activated protein kinase-1 and pyruvate kinase was increased with MCT, whereas expression of these genes tended to be increased by SA. The mechanism of increased fat secretion with PA does not seem to be related to a modulation of the expression of lipogenesis-related genes, but rather to increased substrate availability as reflected by milk FA profile.

Production performance, nutrient digestibility, and milk fatty acid profile of lactating dairy cows fed corn silage- or sorghum silage-based diets with and without xylanase supplementation.

The objective of this study was to evaluate the effects of supplementing xylanase on production performance, nutrient digestibility, and milk fatty acid profile in high-producing dairy cows consuming corn silage- or sorghum silage-based diets. Conventional corn (80,000 seeds/ha) and brown midrib forage sorghum (250,000 seeds/ha) were planted, harvested [34 and 32% of dry matter (DM), respectively], and ensiled for more than 10 mo. Four primiparous and 20 multiparous Holstein cows were randomly assigned to 1 of 4 diets in a replicated 4 × 4 Latin square design with a 2 × 2 factorial arrangement of treatments and 19-d periods. Treatment diets consisted of (1) corn silage-based diet without xylanase, (2) corn silage-based diet with xylanase, (3) sorghum silage-based diet without xylanase, and (4) sorghum silage-based diet with xylanase. The xylanase product was supplemented at a rate of 1.5 g of product/kg of total DM. Corn silage had higher concentrations of starch (31.2 vs. 29.2%), slightly higher concentrations of crude protein (7.1 vs. 6.8%) and fat (3.7 vs. 3.2%), and lower concentrations of neutral detergent fiber (36.4 vs. 49.0%) and lignin (2.1 vs. 5.7%) than sorghum silage. Xylanase supplementation did not affect DM intake, milk yield, milk fat percentage and yield, milk protein percentage and yield, lactose percentage and yield, and 3.5% fat-corrected milk yield. Cows consuming corn silage-based diets consumed 13% more DM (28.8 vs. 25.5 kg/d) and produced 5% more milk (51.6 vs. 48.9 kg/d) than cows consuming sorghum silage-based diets. Milk from cows consuming sorghum silage-based diets had 16% greater fat concentrations (3.84 and 3.30%) than milk from cows consuming corn silage-based diets. This resulted in 8% greater fat yields (1.81 vs. 1.68 kg/d). Silage type did not affect milk protein and lactose concentrations. Xylanase supplementation did not affect nutrient digestibility. Cows consuming corn silage-based diets showed greater DM (77.3 vs. 73.5%), crude protein (78.0 vs. 72.4), and starch (99.2 vs. 96.5%) digestibilities than cows consuming sorghum silage-based diets. In conclusion, xylanase supplementation did not improve production performance when high-producing dairy cows were fed corn silage- or sorghum silage-based diets. In addition, production performance can be sustained by feeding sorghum silage in replacement of corn silage.

Effects of feeding hulled and hull-less barley with low- and high-forage diets on lactation performance, nutrient digestibility, and milk fatty acid composition of lactating dairy cows.

The objective of this study was to evaluate lactation performance, nutrient digestibility, and milk fatty acid composition of high-producing dairy cows consuming diets containing hulled or hull-less barley as the grain source when feeding low-forage (LF) or high-forage (HF) diets. Eight primiparous (610 ± 40 kg of body weight and 72 ± 14 d in milk) and 16 multiparous (650 ± 58 kg of body weight and 58 ± 16 d in milk) Holstein cows were randomly assigned to 1 of 4 diets in a replicated 4 × 4 Latin square design with a 2 × 2 factorial arrangement of treatments and 21-d periods. Cows were assigned to squares based on parity (1, 2, and ≥3) and days in milk. Diets were formulated to contain on a dry matter basis (1) 45% forage and hulled barley as the sole grain source, (2) 65% forage and hulled barley as the sole grain source, (3) 45% forage and hull-less barley as the sole grain source, and (4) 65% forage and hull-less barley as the sole grain source. Dry matter intake tended to be lower for the diet with 65% forage and hulled barley than for the rest of the diets (24.4 vs. 26.6 kg/d). Neither the type of barley nor the forage-to-concentrate ratio affected milk yield (41.7 kg/d). Barley type did not affect milk fat or protein concentrations. Feeding LF diets decreased milk fat concentration from 3.91% to 3.50%. This decrease was less than anticipated and resulted in a 7% decrease in milk fat yield relative to cows consuming HF diets (1.60 and 1.49 kg/d for HF and LF diets, respectively). Feeding LF diets increased the concentration of C18:1 trans-10 in milk fat, suggesting that feeding LF diets may have marginally altered rumen function. In conclusion, LF diets containing barley grains can marginally decrease milk fat concentration. Overall, and based on the conditions of this study, there is limited evidence to anticipate a dramatic or acute milk fat depression when feeding hull-less barley as the grain source in diets for high-producing dairy cows.

Effects of feeding hull-less barley on production performance, milk fatty acid composition, and nutrient digestibility of lactating dairy cows.

The objectives of this study were to evaluate production performance, milk fatty acid composition, and nutrient digestibility in high-producing dairy cows consuming diets containing corn and hull-less barley (cultivar Amaze 10) in different proportions as the grain source. Eight primiparous and 16 multiparous Holstein cows were assigned to 1 of 4 diets in a replicated 4 × 4 Latin square design with 21-d periods. Cows were fed once daily (1200 h) by means of a Calan gate system (American Calan Inc., Northwood, NH). All diets contained ∼20% grain (dry matter basis). Treatments consisted of 100% corn (0B), 67% corn and 33% hull-less barley (33B), 33% corn and 67% hull-less barley (67B), and 100% hull-less barley (100B) as the grain sources. Total-tract nutrient digestibility was estimated using lanthanum chloride (LaCl3) as an external marker. Dry matter intake differed quadratically among treatments, being lowest for 67B and highest for 0B and 100B. Feeding hull-less barley did not affect milk yield, and milk fat concentration differed cubically among treatments. The cubic response was attributed to the higher milk fat concentration observed for the diet containing 67B. Neither the concentrations in milk of protein and lactose nor the yields of protein and lactose differed among treatments. The proportion of de novo synthesized fatty acids in milk did not differ among treatments. The apparent total-tract digestibility of dry matter, crude protein, and neutral detergent fiber did not differ among treatments. Although a quadratic effect was observed, starch digestibility was minimally affected by treatments. In conclusion, this study indicates that hull-less barley grain is as good as corn grain as an energy source when formulating diets for high-producing dairy cows.

Bovine brain region-specific stearoyl-CoA desaturase expression and fatty acid composition.

In this study, we sought to determine the relationship between stearoyl-CoA desaturase (SCD) gene isoform expression in the bovine brain and the accumulation of 18:1n-9. Two SCD gene isoforms are found in cows-SCD1 and SCD5. Samples of six brain regions (cerebellum, frontal cortex, hippocampus, hypothalamus, midbrain, and pons) were collected from animals at four different ages (neonates, weanlings, yearlings, and adults) for mRNA isolation and fatty acid analysis. Expression of SCD1 and SCD5 mRNA was compared across age groups to determine its developmental regulation. Fatty acid composition and SCD isoform mRNA expression were compared to examine the correlation of SCD1 and SCD5 with 18:1n-9 content in different brain regions. We found statistically significant correlations between SCD1 and SCD5 mRNA expression and the ratio of 18:1n-9 to 18:0 across age groups, with stronger correlations observed for SCD5. Similarly, there was a significant correlation between the ratio of 18:1n-9 to 18:0 and SCD5 mRNA expression across brain regions. SCD1 mRNA and the 18:1n-9 to 18:0 ratio were negatively correlated in the hippocampus. There was no trend of increasing 18:1n-9 content or SCD expression with age. Correlations indicated a stronger relationship between SCD5 mRNA expression and the 18:1n-9 to 18:0 ratio, potentially indicating a strong contribution of the SCD5 isoform to brain 18:1n-9 content. This is the first study examining a potential role for SCD5 in providing 18:1n-9 for brain lipids.

Supplementing antioxidants to pigs fed diets high in oxidants: II. Effects on carcass characteristics, meat quality, and fatty acid profile.

The study was conducted to determine effects of dietary supplementation with a blend of antioxidants (ethoxyquin and propyl gallate) on carcass characteristics, meat quality, and fatty acid profile in finishing pigs fed a diet high in oxidants. A total of 100 crossbred barrows (10.9±1.4 kg BW, 36±2 d of age) were randomly allotted to 5 diet treatments (5 replicate pens per treatment, 4 pigs per pen). Treatments included: 1) HO: high oxidant diet containing 5% oxidized soy oil and 10% PUFA source which contributed 5.56% crude fat and 2.05% docosahexanoic acid (DHA) to the diet; 2) VE: the HO diet with 11 IU/kg of added vitamin E; 3) AOX: the HO diet with antioxidant blend (135 mg/kg); 4) VE+AOX: the HO diet with both vitamin E and antioxidant blend; and 5) SC: a standard corn-soy control diet with nonoxidized oil and no PUFA source. The trial lasted for 118 d; on d 83, the HO diet pigs were switched to the SC diet due to very poor health. From that point, the VE pigs displayed the poorest performance. On d 118, 2 pigs from each pen were harvested for sampling. Compared to pigs fed SC diet, the HO and VE pigs (P<0.05) showed lighter carcass weight, less back fat, less lean body mass, and smaller loin eye area. In addition, the VE pigs had decreased dressing percentage than the AOX and VE+AOX pigs (65.7 vs. 75.3 and 74.2%). Compared to the SC pigs, greater moisture percentage (74.7 vs. 77.4%) and less extractable lipid content (2.43 vs. 0.95%) were found in VE fed pigs (P<0.05). Drip loss of loin muscle in VE pigs was less than SC pigs (0.46 vs. 3.98%, P=0.02), which was associated with a trend for a greater 24-h muscle pH (5.74 vs. 5.54, P=0.07). The antioxidant blend addition in the high oxidant diet attenuated all of these effects to levels similar to SC (P>0.05), except a* value (redness) and belly firmness. Visible yellow coloration of backfat and lipofuscin in HO and VE pigs was observed at harvest at d 118. The high oxidant diet resulted in greater concentration of DHA in backfat (P<0.001); switching the diet on d 83 resulted in HO pigs having a similar fatty acid profile to SC at d 118 pigs. Vitamin E concentration in plasma and muscle was greater in HO and SC than VE, AOX, and VE+AOX on d 118. Feeding the high oxidant diet caused a series of changes in carcass characteristics and meat quality. Addition of antioxidant blend attenuated many of these, whereas the protective effects of supplemental vitamin E at 11 IU/kg were minimal during the finisher phase of the study.

Short communication: Characteristics of student success in an undergraduate physiology and anatomy course.

Several factors affect the success of students in college classes. The objective of this research was to determine what factors affect success of undergraduate students in an anatomy and physiology class. Data were collected from 602 students enrolled in the Agriculture and Life Sciences (ALS) 2304 Animal Physiology and Anatomy course from 2005 through 2012. The data set included 476 females (79.1%) and 126 males (20.9%). Time to complete exams was recorded for each student. For statistical analyses, students' majors were animal and poultry sciences (APSC), agricultural sciences, biochemistry, biological sciences, dairy science, and "other," which combined all other majors. All analyses were completed using the GLIMMIX procedure of SAS (SAS Institute Inc., Cary, NC). Gender, major, matriculation year, major by year interaction, gender by year interaction, and time to complete the exam affected final course grade. The significant gender effect was manifested in the final grade percentage of 75.9 ± 0.4 for female students compared with 72.3 ± 0.6 for male students. Junior males had final course grades comparable with those of females, but sophomore and senior males had lower final course grades than other combinations. Biology majors had a final grade of 82.4 ± 0.6 and this grade was greater than all other majors. Students classified as "other" had a final score of 74.4 ± 0.8, which was greater than agricultural science majors (69.5 ± 0.9). The APSC grade (72.6 ± 0.5) was higher than the agricultural science majors. Junior students had significantly greater final grades (76.1 ± 0.5) than sophomores (73.3 ± 0.6) and seniors (72.9 ± 0.9). All biology students had greater final grades than all other majors, but biochemistry juniors had greater final course grades than APSC, agricultural science, and dairy science juniors. "Other" seniors had greater final course grades than agricultural science seniors. The regression for time to complete the exam was curvilinear and suggests that highest exam scores were at about 90-min completion time. It may be that some male students need better preparation for anatomy and physiology and their educational preparation should mimic that of female students more in terms of advance-placement biology in high school. These results suggest that biology majors might be better prepared for animal anatomy and physiology than other students.

Effects of abomasal infusion of conjugated linoleic acids, Sterculia foetida oil, and fish oil on production performance and the extent of fatty acid Δ9-desaturation in dairy cows.

The purpose of this study was to determine the effects of conjugated linoleic acid (CLA), Sterculia foetida oil (STO), and fish oil (FO) on milk yield and composition, milk FA profile, Δ(9)-desaturation activity, and mammary expression of 2 isoforms of stearoyl-coenzyme A desaturase (SCD-1 and SCD-5) in lactating dairy cows. Eight multiparous Holstein cows (69 ± 13 d postpartum) were used in a double 4 × 4 Latin square design with 28-d periods. For the first 14 d of each period, cows received an abomasal infusion of (1) 406 g of a saturated fatty acid (SFA) supplement (112 g of 16:0 + 230 g of 18:0) used as a control (CTL), (2) 36 g of a CLA supplement (13.9 g of trans-10,cis-12 18:2) + 370 g of SFA, (3) 7 g of STO (3.1g of 19:1 cyclo) + 399 g of SFA, or (4) 406 g of FO (55.2 g of cis-5,-8,-11,-14,-17 20:5 + 59.3 g of cis-4,-7,-10,-13,-16,-19 22:6). Infusions were followed by a 14-d washout interval. Compared with CTL, STO decreased milk yield from 38.0 to 33.0 kg/d, and increased milk fat concentration from 3.79 to 4.45%. Milk fat concentration was also decreased by CLA (2.23%) and FO (3.34%). Milk fat yield was not affected by STO (1,475 g/d) compared with CTL (1,431 g/d), but was decreased by CLA (774 g/d) and FO (1,186 g/d). Desaturase indices for 10:0, 12:0, and 20:0 were decreased, whereas the extent of desaturation of 14:0, 16:0, 17:0, and 18:0 was not affected by CLA treatment compared with CTL. Infusion of STO significantly decreased all calculated desaturase indices compared with CTL; the 14:0 index was reduced by 80.7%. Infusion of FO decreased the desaturase indices for 10:0, 14:0, 20:0, trans-11 18:1, and 18:0. The effect of FO on the 14:0 index indicates a decrease in apparent Δ(9)-desaturase activity of 30.2%. Compared with CTL, mammary mRNA abundance of SCD-1 was increased by STO (+30%) and decreased by CLA (-24%), whereas FO had no effect. No effect was observed on mRNA abundance of SCD-5. In conclusion, abomasal infusion of CLA, STO, and FO were shown to exhibit varying and distinct effects on desaturase indices, an indicator of apparent SCD activity, and mammary mRNA abundance of SCD-1.

Short communication: effect of trans-10,cis-12 conjugated linoleic acid on activation of lipogenic transcription factors in bovine mammary epithelial cells.

The objective of this study was to examine the effect of trans-10,cis-12 conjugated linoleic acid (t10c12CLA) on the activation of transcription factors that potentially regulate lipid synthesis in a bovine mammary epithelial cell line (MAC-T). Cells were transfected with luciferase reporter constructs containing sterol response element (SRE and SRE complex) for sterol regulatory element binding protein-1, peroxisome proliferator response element for peroxisome proliferator-activated receptor γ, or liver X receptor response element for liver X receptor. Different concentrations of t10c12CLA (0, 25, 50, 75, or 100µM) were applied to cells to determine the activation of transcription factors. The influence of t10c12CLA bond structure on transcription factor activation was also investigated by treating cells with different 18:1 fatty acid isomers (trans-10 18:1 or cis-12 18:1) at 100µM. Cells were harvested for luciferase assay after 24h of treatment. Compared with linoleic acid and cis-9,trans-11 CLA controls, the SRE reporters had significantly lower activity in t10c12CLA-treated cells at 50 and 75µM for SRE complex and SRE, respectively. Lower SRE and SRE complex activation was observed in t10c12CLA treatment at 25, 50, and 75µM compared with 0µM. The peroxisome proliferator response element and liver X receptor response element reporters did not respond differently between the t10c12CLA treatment and controls. Compared with t10c12CLA, both trans-10 18:1 and cis-12 18:1 increased the activities of SRE and SRE complex reporters by 1.3- to 4.2-fold. In conclusion, t10c12CLA has an inhibitory role in lipogenic transcription factor activation of SRE, and this negative effect is due to the conjugation of trans-10 and cis-12 double bonds in the fatty acid. Furthermore, we found no support for a regulatory role of response elements for peroxisome proliferator-activated receptor γ or liver X receptor in the t10c12CLA inhibition of mammary lipid synthesis.

Effects of a dietary antioxidant blend and vitamin E on fatty acid profile, liver function, and inflammatory response in broiler chickens fed a diet high in oxidants.

The aim of the current study was to determine the effects of a dietary antioxidant blend and vitamin E on fatty acid profile, inflammatory response, and liver function. Cobb 500 male broilers (n = 1,200, d 0) were randomly distributed into 6 treatments with 10 replicate floor pens. Treatments included (1) a high-oxidant diet, with vitamin E at 10 IU/kg, 3% oxidized oil, 3% polyunsaturated fatty acids (PUFA) source (HO); (2) the HO diet with vitamin E at 200 IU/kg (VE); (3) the HO diet with an antioxidant blend at 135 mg/kg (AOX); (4) the HO diet with both vitamin E at 200 IU/kg and an antioxidant blend at 135 mg/kg (VE+AOX); (5) standard control (SC); and (6) a positive control, which was the SC diet with an antioxidant blend at 135 mg/kg. The concentrations of 20:4, 20:5, 22:5, 22:6, and all the n-3 fatty acids were greater in the abdominal fat of HO, VE, AOX, and VE+AOX birds than SC and positive control birds on d 21 and 42 (P < 0.001). Compared with HO treatment, AOX and VE+AOX preserved the deposition of PUFA better (P < 0.001). The HO birds had greater concentrations of aspartate aminotransferase on d 21 and 42, and γ-glutamyl transferase on d 21, whereas AOX and VE+AOX chickens had restored γ-glutamyl transferase concentration (P < 0.01). The inflammation scores of abdominal fat of AOX and VE+AOX birds were lower than the HO on d 21 (P < 0.001). Compared with SC, the VE and VE+AOX birds exhibited greater vacuole scores on d 21 and 42 (P < 0.01). The lower vacuoles score in SC was associated with a greater expression of peroxisome proliferator activated receptor -γ and -α (P < 0.05). The expression of inflammatory genes in the liver did not differ among treatments. In conclusion, the AOX and AOX+VE diets were effective in preserving PUFA in the abdominal fat, moderately improved liver function, and reduced inflammation in fat.

Effect of growth hormone on the differentiation of bovine preadipocytes into adipocytes and the role of the signal transducer and activator of transcription 5b.

We evaluated the effect of GH on the differentiation of primary bovine preadipocytes into adipocytes. Bovine preadipocytes, derived from adipose tissue explants, were induced to differentiate into adipocytes in the presence or absence of recombinant bovine GH. The differentiation status of adipocytes was assessed by Oil Red O staining and by measuring the activity of glycerol-3-phosphate dehydrogenase (G3PDH) and the rate of acetate incorporation. Fewer preadipocytes became adipocytes in the presence of GH than in the absence of GH; adipocytes formed in the presence of GH had lower G3PDH activity and lower rate of acetate incorporation than those formed without GH treatment (P < 0.05). These data suggest an inhibitory effect of GH on the differentiation of bovine preadipocytes into adipocytes. Growth hormone decreased the expression of C/EBPα and PPARγ mRNA in bovine adipocytes (P < 0.05). Because C/EBPα and PPARγ are the master regulators of adipocyte differentiation, this data suggests that GH might inhibit the differentiation of bovine preadipocytes into adipocytes by inhibiting the expression of C/EBPα and/or PPARγ. Because the signal transducer and activator of transcription 5 (STAT5) is a major component of signaling from the GH receptor, we next determined the potential role of STAT5 in GH inhibition of bovine adipocyte differentiation. Overexpression of a constitutively active form of STAT5b (STAT5bCA) in bovine preadipocytes through adenoviral transduction mimicked the effects of GH on the formation of lipid-containing adipocytes, G3PDH activity, and acetate incorporation rate. Overexpression of STAT5bCA was associated with decreased expression of C/EBPα mRNA (P < 0.05) but not that of PPARγ mRNA in bovine adipocytes. These results support a role of STAT5b in mediating GH inhibition of C/EBPα expression but not that of PPARγ expression in bovine preadipocytes. Overall, the present study suggests that GH may inhibit adipose growth in cattle in part by inhibiting adipogenesis and that GH inhibits the differentiation of bovine preadipocytes to adipocytes through STAT5b-dependent inhibition of C/EBPα expression and STAT5b-independent inhibition of PPARγ expression.

Mammary uptake of fatty acids supplied by intravenous triacylglycerol infusion to lactating dairy cows.

Supplementing dairy cows with n-3 fatty acid-rich feeds does not easily increase quantities in milk fat. Previous results demonstrated very long-chain n-3 fatty acids are primarily transported in the PL fraction of blood, making them largely unavailable to the mammary gland for enrichment of milk fat. Our objective was to compare mammary uptake of fatty acids of increasing chain length and unsaturation delivered intravenously as TAG emulsions. Late lactation dairy cows were assigned to a completely randomized block design. Treatments were intravenous TAG emulsions enriched with oleic acid (OLA), linoleic acid (LNA), alpha-linolenic acid (ALA), or docosahexaenoic acid (DHA) and were delivered continuously at 16 mL/h for 72 h. Each treatment supplied 30 g/day of the target fatty acid. Treatment did not affect feed intake, milk yield, or milk composition, but all treatments reduced intake and yield. The proportion of DHA increased in plasma FFA, TAG, and PL with infusion. Increases of n-3 fatty acids, ALA, EPA, and DHA, were evident in the plasma PL fraction, suggesting re-esterification in the liver. Transfer efficiencies were 37.8 ± 4.1, 27.6 ± 5.4, and 10.9 ± 4.1 %, and day 3 total milk fatty acyl yields were 37.0 ± 3.4, 10.8 ± 0.4, and 3.3 ± 0.3 g for LNA, ALA, and DHA. Variation in oleic acyl yield prevented calculation of OLA transfer efficiency. Mammary uptake of fatty acids was reduced with increased chain length and unsaturation. Both liver and mammary mechanisms may regulate transfer of long-chain polyunsaturates.

An explant based-method for differentiating adipocytes from equine adipose tissue.

Culturing adipocytes enables fine control of experimental conditions and helps minimise animal use. This report describes an explant-based method for isolating stromal-vascular cells from equine adipose tissue that enables use of small amounts of tissue. Subcutaneous and mesenteric adipose tissues were harvested post mortem and stromal-vascular cells grown from explants, prior to testing the capacity of several differentiation media to induce lipid droplet formation and increase transcript abundance of adipocyte markers. Inclusion of rosiglitazone at 1 and 5 µmol/l concentrations, along with other media components, induced differentiation of cultured equine stromal-vascular cells derived from subcutaneous and mesenteric adipose tissues.

Regulation of lipid synthesis by liver X receptor α and sterol regulatory element-binding protein 1 in mammary epithelial cells.

The objectives of this experiment were to characterize the roles of the transcription factors liver X receptor α (LXRα) and sterol regulatory element-binding protein 1 (SREBP1) in the transcriptional regulation of lipid synthesis in a bovine mammary epithelial cell line. Whereas many lipid synthesis genes contain a response element in their promoters for SREBP1, a few also contain a response element for LXR, suggesting that both transcription factors could directly regulate transcription of these genes. However, the promoter of SREBP1 contains a response element for LXR, indicating the additional potential for indirect transcriptional regulation by LXR, through SREBP1, on lipogenic genes. To characterize these effects, small interfering RNA (siRNA) directed against LXRα and SREBP1 were used to knockdown gene expression, and then, in the presence of SREBP1 siRNA, T 4506585 (T09) was used to specifically activate LXRα. Reducing LXRα mRNA abundance in mammary alveolar T cells did not alter mRNA abundance of genes involved in de novo lipogenesis or the rate of de novo lipogenesis, suggesting that LXRα is not required for basal transcription of genes required for fatty acid synthesis. Knockdown of SREBP1 reduced the mRNA abundance of acetyl-coenzyme A (CoA) carboxylase, fatty acid synthase, diacylglycerol acyltransferase, and stearoyl-CoA desaturase-1, indicating that these genes are regulated in part by SREBP1. When SREBP1 was reduced, T09 increased the mRNA abundance of acetyl-CoA carboxylase, fatty acid synthase, and diacylglycerol acyltransferase, potentially indicating that these genes are directly regulated by LXR. The results of the present study provide insight into the transcriptional regulatory mechanisms involved in lipid synthesis by mammary epithelial cells, and suggest that several transcription factors may be required for full lipogenic activation.

Relationships between body condition score and plasma inflammatory cytokines, insulin, and lipids in a mixed population of light-breed horses.

BACKGROUND: Obesity and hyperinsulinemia increase the risk of laminitis in horses and ponies. In mares, obesity also has been associated with increased circulating concentrations of the proinflammatory cytokine, tumor necrosis factor (TNF)-α. The association of other proinflammatory cytokines with body condition score (BCS) and insulin requires further determination. HYPOTHESIS: Plasma concentrations of TNF, interleukin (IL)-1ß, IL-6, and serum amyloid A (SAA) will positively correlate with BCS or insulin or both in horses. Furthermore, inflammatory protein concentrations will correlate with age and variables associated with BCS, including plasma insulin, triglycerides, nonesterified fatty acids, and leptin concentrations. ANIMALS: One hundred and ten mixed light-breed horses, including mares, geldings, and stallions, aged 4-20 years. METHODS: Samples were selected from a larger population of plasma samples previously collected during June-July of 2006. Samples were analyzed for TNF, IL-1ß, IL-6, and SAA using commercially available ELISAs and simple correlations were used to determine relationships with BCS, insulin, age, and sex. RESULTS: Plasma TNF (P = .047) and IL-6 (P = .021) concentrations were higher in females than males, whereas IL-6 concentrations correlated (P = .001) with age. Plasma SAA concentrations correlated with both insulin (P < .001) and BCS (P = .007). CONCLUSIONS AND CLINICAL IMPORTANCE: This study provides evidence for factors, including age and sex, that may be associated with plasma concentrations of inflammatory proteins. Concentrations of SAA correlated with BCS and insulin, independent of age or sex. Because BCS and insulin correlate with increased SAA, it is possible that SAA is a component of laminitis pathophysiology.