Highly fermentable diets are rapidly converted to organic acids i.e., short-chain fatty acids (SCFA) and lactic acid within the rumen. The resulting release of protons can constitute a challenge to ...the ruminal ecosystem and animal health. Health disturbances, resulting from acidogenic diets, are classified as subacute and acute acidosis based on the degree of ruminal pH depression. Although increased acid production is a nutritionally desired effect of increased concentrate feeding, the accumulation of protons in the rumen is not. Consequently, mechanisms of proton removal and their quantitative importance are of major interest. Saliva buffers (i.e., bicarbonate, phosphate) have long been identified as important mechanisms for ruminal proton removal. An even larger proportion of protons appears to be removed from the rumen by SCFA absorption across the ruminal epithelium, making efficiency of SCFA absorption a key determinant for the individual susceptibility to subacute ruminal acidosis. Proceeding initially from a model of exclusively diffusional absorption of fermentation acids, several protein-dependent mechanisms have been discovered over the last 2 decades. Although the molecular identity of these proteins is mostly uncertain, apical acetate absorption is mediated, to a major degree, via acetate-bicarbonate exchange in addition to another nitrate-sensitive, bicarbonate-independent transport mechanism and lipophilic diffusion. Propionate and butyrate also show partially bicarbonate-dependent transport modes. Basolateral efflux of SCFA and their metabolites has to be mediated primarily by proteins and probably involves the monocarboxylate transporter (MCT1) and anion channels. Although the ruminal epithelium removes a large fraction of protons from the rumen, it also recycles protons to the rumen via apical sodium-proton exchanger, NHE. The latter is stimulated by ruminal SCFA absorption and salivary Na⁺ secretion and protects epithelial integrity. Finally, SCFA absorption also accelerates urea transport into the rumen, which via ammonium recycling, may remove protons from rumen to the blood. Ammonium absorption into the blood is also stimulated by luminal SCFA. It is suggested that the interacting transport processes for SCFA, urea, and ammonia represent evolutionary adaptations of ruminants to actively coordinate energy fermentation, protein assimilation, and pH regulation in the rumen.
To investigate, using culture-independent methods, whether the ruminal bacterial structure, population and fermentation parameters differed between sampling locations and time. The detectable ...bacteria and fermentation parameters in the digesta from five locations in the rumen of three cows at three time points were analysed. The PCR-denaturing gradient gel electrophoresis (PCR-DGGE) profiles were similar among digesta samples from five locations (95·4%) and three time points (93·4%) within cows; however, a lower similarity was observed for samples collected from different host animals (85·5%). Rumen pH and concentration of volatile fatty acids (VFA) were affected by time points of sampling relative to feeding. The detectable bacterial structure in the rumen is highly conserved among different locations and over time, while the quantity of individual bacterial species may change diurnally in response to the feeding. This study supplies the fundamental understanding of the microbial ecology in the rumen, which is essential for manipulation of ruminal microflora and subsequent improvement in animal production.
The current study was undertaken to investigate the effect of feeding diets varying in sugar concentration to postpartum transition cows on productivity, ruminal fermentation, and nutrient ...digestibility. We hypothesized that the high-sugar diet would increase dry matter intake and lactation performance. The secondary objective was to characterize changes in ruminal fermentation and nutrient digestibility over the first 4 wk of lactation. Fifty-two Holstein cows, including 28 primiparous and 24 multiparous cows, 10 of which were previously fitted with a ruminal cannula, were assigned to the experimental diets containing either high sugar (HS=8.4%) or low sugar (LS=4.7%) immediately after calving, based on their expected calving date. Data and samples were collected on d 5.2 ± 0.3, 12.2 ± 0.3, 19.2 ± 0.3, and 26.1 ± 0.3 relative to parturition for wk 1, 2, 3, and 4 respectively. Cows fed HS had increased dry matter intake compared with those fed LS (18.3. vs. 17.2 kg/d). Further, cows fed HS sorted for particles retained on the pan of the Penn State Particle Size Separator to a greater extent than cows fed LS. Feeding HS tended to increase nadir (5.62 vs. 5.42), mean (6.21 vs. 6.06), and maximum pH (6.83 vs. 6.65). The duration (h/d) and area (pH × min/d) that ruminal pH was below pH 5.8 were not affected by treatment. Ruminal volatile fatty acid concentration and molar proportions of individual volatile fatty acids were not affected by treatment. The digestibility of dry matter, organic matter, neutral detergent fiber, and starch were not affected by treatment, averaging 63.3, 65.2, 43.2, and 93.5%, respectively. Feeding HS decreased plasma glucose concentration compared with feeding LS (51.3 vs. 54.0 mg/dL), but concentration of plasma insulin was not affected by treatment, averaging 4.17μIU/mL. Cows fed HS had higher concentrations of plasma β-hydroxybutrate (17.5 vs. 10.5 mg/dL) and nonesterified fatty acids (344 vs. 280μEq/L). Milk yield and milk composition were not affected by treatment, but a tendency for increased milk fat yield was observed for cows fed HS compared with LS (1.44 vs. 1.35 kg/d). The results of the current study imply that replacing cracked corn grain with sucrose may improve dry matter intake, ruminal pH status, and lactation performance.
Inflammation of ruminal epithelium may occur during ruminal acidosis as a result of translocation and interaction of ruminal epithelial cells (REC) with molecules such as lipopolysaccharide (LPS). ...Such inflammation has been reported to alter cellular processes such as nutrient absorption, metabolic regulation, and energy substrate utilization in other cell types but has not been investigated for REC. The objectives of this study were to investigate the effects of LPS on metabolism of short-chain fatty acids by primary REC, as well as investigating the effects of media containing short-chain fatty acids on the proinflammatory response. Ruminal papillae from 9 yearling Speckle Park beef heifers were used to isolate and culture primary REC. Cells were grown in minimum essential medium (MEM) for 12 d before use and then reseeded in 24-well culture plates. The study was conducted as a 2 × 2 factorial, where cells were grown in unaltered MEM (REG) or medium containing 2 mM butyrate and 5 mM propionate (SCFA) with (50,000 EU/mL; +LPS) or without LPS (−LPS) for 24 h. Supernatant samples were collected for analysis of glucose and SCFA consumption. Cells were collected to determine the expression of mRNA for genes associated with inflammation (TNF, IL1B, CXCL2, CXCL8, PTGS2, and TLR4), purinergic signaling (P2RX7, ADORAB2, and CD73), nutrient transport SLC16A1 (MCT1), SLC16A3 (MCT4), SLC5A8, and MCU, and cell metabolism ACAT1, SLC2A1 (GLUT1), IGFBP3, and IGFBP5. Protein expression of TLR4 and ketogenic enzymes (BDH1 and HMGCS1) were also analyzed using flow cytometry. Statistical analysis was conducted with the MIXED model of SAS version 9.4 (SAS Institute Inc., Cary, NC) with medium, LPS exposure, and medium × LPS interaction as fixed effects and animal within plate as a random effect. Cells tended to consume more glucose when exposed to LPS as opposed to no LPS exposure (31.8 vs. 28.7 ± 2.7), but consumption of propionate and butyrate was not influenced by LPS. Expression of TNF and IL1B was upregulated when exposed to LPS, and expression of CXCL2 and CXCL8 increased following LPS exposure with SCFA (medium × LPS). For cells exposed to LPS, we found a downregulation of ACAT1 and IGFBP5 and an upregulation of SLC2A1, SLC16A3, MCU, and IGFBP3. Medium with SCFA led to greater expression of MCU. SLC16A1 was upregulated in cells incubated with SCFA and without LPS compared with the other groups. Protein expression of ketogenic enzymes was not affected; however, BDH1 mean fluorescence intensity (MFI) expression tended to be less in cells exposed to LPS. These data are interpreted to indicate that when REC are exposed to LPS, they may increase glucose metabolism. Moreover, transport of solutes was affected by SCFA in the medium and by exposure to LPS. Overall, the results suggest that metabolic function of REC in vitro is altered by a proinflammatory response, which may lead to a greater glucose requirement.
The objective of this study was to investigate whether cultured ruminal epithelial cells (REC) responded to lipopolysaccharide (LPS) stimulation and determine whether LPS induced a proinflammatory ...response. Primary bovine REC were isolated and grown in culture for 2 studies. In study 1, REC were isolated from Holstein bull calves (n = 8) and grown in culture for 10 to 12 d. Cells were then exposed to 0, 10,000, 50,000, or 200,000 endotoxin (E)U/mL of LPS (Escherichia coli O55:B5) for either 6 or 24 h. The effect of LPS exposure on cell viability was analyzed by flow cytometry using a propidium iodide stain. In study 2, cells were isolated from Holstein bull calves (n = 5) and yearling beef heifers (n = 4). Cells were exposed to either 1,000 or 50,000 EU/mL of LPS using the following conditions: (1) medium alone time-matched controls, (2) 12-h LPS exposure, (3) 24 h of LPS exposure, (4) 36 h of LPS exposure, (5) 12 h of LPS exposure followed by LPS removal for 24 h before restimulating with LPS for an additional 12 h (RPT), and (6) 12 h of LPS exposure followed by LPS removal for 36 (RVY). For both experiments, total RNA was extracted from REC and real-time quantitative PCR was performed to determine relative expression of genes for toll-like receptors (TLR2 and TLR4), proinflammatory cytokines (TNF and IL1B), chemokines (CXCL2 and CXCL8), a lipid mediator (PTGS2), and growth factor-like cytokines (CSF2 and IL7). In study 1, LPS exposure did not negatively affect cell viability. Treatment of cells with LPS resulted in increased transcript abundance for all genes analyzed. The TLR2, IL7, and TLR4 had a greater magnitude of change at 6 h compared with 24 h. Quadratic expression patterns were detected for TNF, IL1B, CXCL2, CXCL8, and CSF2. These results suggested that REC increase expression of proinflammatory genes following exposure to LPS. In study 2, all genes analyzed were upregulated in a quadratic manner following exposure to LPS for different time intervals. The TLR4, TNF, CXCL2, CXCL8, CSF2, and IL7 gene expression was significantly greater after a single 12 h of LPS exposure than after RPT exposure, suggesting repeated exposure of REC to LPS may induce a tolerogenic effect. When LPS was removed from the medium (RVY), transcript abundance for all genes analyzed decreased and expression of TLR2, TLR4, and IL7 returned to baseline levels, suggesting REC recovered following exposure to LPS. Overall, the data suggest cultured REC respond to LPS stimulation by increasing transcription of proinflammatory genes and this transcriptional response was influenced by the dose, duration, and frequency of LPS exposure.
The objectives of this study were: 1) to determine the effect of providing additional prepartum concentrate on the occurrence and severity of ruminal acidosis (RA) and lactational performance during ...the periparturient period in primiparous cows; and 2) to characterize the occurrence and severity of RA during the periparturient period. We hypothesized that providing additional concentrate prepartum would reduce postpartum RA. Fourteen ruminally cannulated Holstein heifers were paired by expected calving date and body condition score. The heifers were assigned to 1 of 2 prepartum feeding regimens: 1) a control treatment consisting of a far-off diet (forage:concentrate, F:C=80:20) fed from d −60 to d −25 and a close-up diet (F:C=54:46) fed from d −24 until parturition; or 2) a high-concentrate (HC) feeding program consisting of 4 prepartum diets, HC-1 (F:C=68:32) fed from d −60 to d −43, HC-2 (F:C=60:40) fed from d −42 to d −25, HC-3 (F:C=52:48) fed from d −24 to d −13, and HC-4 (F:C=46:54) fed from d −12 until parturition. All cows received the same lactation diet postpartum. Ruminal pH was measured continuously from d −5 to d +5, and for 3 consecutive days starting on d +17±1.2, d +37±1.4, and d +58±1.5 relative to parturition using an indwelling ruminal pH system. Ruminal acidosis was considered to occur when ruminal pH was <5.8 (total RA). Ruminal acidosis was further partitioned into: 1) mild RA (5.8>ruminal pH>5.5), 2) moderate RA (5.5>ruminal pH>5.2), and 3) acute RA (ruminal pH<5.2). Feeding additional concentrate prepartum did not reduce postpartum RA. In fact, cows fed the HC treatment had more daily episodes of acute RA than cows fed the control treatment. Day relative to parturition affected the occurrence and severity of RA; RA increased following parturition and was sustained thereafter. The DM intake during the last 5 d of gestation was lower for cows fed the HC treatment compared with cows fed the control treatment, but lactational performance was not affected. We conclude that, under the conditions imposed, feeding additional concentrate prepartum does not reduce postpartum RA. Furthermore, the incidence and severity of RA increases immediately postpartum, emphasizing the need to develop and implement feeding strategies that reduce this risk.
Feeding highly fermentable diets to ruminants is one strategy to increase energy intake. The increase in short-chain fatty acid (SCFA) production and reduced ruminal pH associated with highly ...fermentable diets imposes a challenge to the metabolism and the regulation of intracellular pH homeostasis of ruminal epithelia. The ruminal epithelia respond to these challenges in a coordinated manner. Whereas the enlargement of absorptive surface area is well documented, emerging evidence at the mRNA and transporter and enzyme activity levels indicate that changes in epithelial cell function may be the initial response. It is not surprising that gene expression analysis has identified pathways involved in fatty acid metabolism, ion transport, and intracellular homeostasis to be the pathways dominantly affected during adaptation and after adaptation to a highly fermentable diet. These findings are important because the intraepithelial metabolism of SCFA, particularly butyrate, helps to maintain the concentration gradient between the cytosol and lumen, thereby facilitating absorption. Butyrate metabolism also controls the intracellular availability of butyrate, which is widely regarded as a signaling molecule. Current data indicate that for butyrate metabolism, 3-hydroxy-3-methylglutaryl-CoA synthase and acetyl-CoA acetyltransferase are potential regulatory points with transient up- and downregulation during diet adaptation. In addition to nutrient transport and utilization, genes involved in the maintenance of cellular tight junction integrity and induction of inflammation have been identified as differentially expressed genes during adaptation to highly fermentable diets. This may have important implications on ruminal epithelial barrier function and the inflammatory response often associated with subacute ruminal acidosis. The objective of this review is to summarize ruminal epithelial adaptation to highly fermentable diets focusing on the changes at the enzyme and transporter activity levels, as well as the underlying molecular changes at the mRNA and protein expression levels.
Rumen health is of vital importance in ensuring healthy and efficient dairy cattle production. Current feeding programs for cattle recommend concentrate-rich diets to meet the high nutritional needs ...of cows during lactation and enhance cost-efficiency. These diets, however, can impair rumen health. The term “subacute ruminal acidosis” (SARA) is often used as a synonym for poor rumen health. In this review, we first describe the physiological demands of cattle for dietary physically effective fiber. We also provide background information on the importance of enhancing salivary secretions and short-chain fatty acid absorption across the stratified squamous epithelium of the rumen; thus, preventing the disruption of the ruminal acid–base balance, a process that paves the way for acidification of the rumen. On-farm evaluation of dietary fiber adequacy is challenging for both nutritionists and veterinarians; therefore, this review provides practical recommendations on how to evaluate the physical effectiveness of the diet based on differences in particle size distribution, fiber content, and the type of concentrate fed, both when the latter is part of total mixed ration and when it is supplemented in partial mixed rations. Besides considering the absolute amount of physically effective fiber and starch types in the diet, we highlight the role of several feeding management factors that affect rumen health and should be considered to control and mitigate SARA. Most importantly, transitional feeding to ensure gradual adaptation of the ruminal epithelium and microbiota; monitoring and careful management of particle size distribution; controlling feed sorting, meal size, and meal frequency; and paying special attention to primiparous cows are some of the feeding management tools that can help in sustaining rumen health in high-producing dairy herds. Supplementation of feed additives including yeast products, phytogenic compounds, and buffers may help attenuate SARA, especially during stress periods when the risk of a deficiency of physically effective fiber in the diet is high, such as during early lactation. However, the usage of feed additives cannot fully compensate for suboptimal feeding management.
The objective of the study was to investigate the fractional rate of volatile fatty acid (VFA) absorption and the expression of genes encoding for transporters and enzymes involved in the absorption ...and metabolism of VFA in ruminal tissue when cattle were fed high or low concentrate diets. Twelve ruminally cannulated Holstein cows were used in a randomized complete block design. The low concentrate (LC) and high concentrate (HC) diets contained 8 and 64% dietary concentrate (dry matter basis), respectively. Cows were fed their respective diet for at least 28 d, following which data and samples were collected over 6 d. Ruminal pH was measured continuously for 72h, and the in vivo VFA absorption and passage rates were measured using Co-EDTA and n-valeric acid as markers. Ruminal tissue was collected postslaughter from the ventral sac of the rumen, and gene expression was evaluated using quantitative real-time PCR. Dry matter intake was not affected by treatment, averaging 14.9 kg/d, but cows fed HC had lower mean ruminal pH (6.03 vs. 6.48), and a greater duration (376 vs. 10 min/d) that ruminal pH was <5.8. Ruminal VFA concentration was 24mM higher for cows fed HC compared with LC; however, the fractional rate of VFA absorption and passage from the rumen was not affected by dietary treatment, averaging 23.4 and 9.6%/h, respectively. The expression of genes encoding for enzymes involved in VFA activation and ketogenesis were not affected by treatment. Cows fed HC tended to have a relative abundance of pyruvate dehydrogenase lipoamide α 1 mRNA transcripts that was 1.4 times lower than that of cows fed LC, but other enzymes involved in pyruvate metabolism or regulation of the citric acid cycle were not affected. Collectively, these results suggest that the dietary forage to concentrate ratio does not affect the fractional rate of VFA absorption in vivo, but potentially alters energy metabolism in ruminal tissue.
PDF
