Untersuchungen zur ernährungsphysiologischen Bedeutung von NPN-Verbindungen

The aims of the experiment were to investigate the variation in ruminal fermentation, milk performance and milk fatty acid profile triggered by induced subacute ruminal acidosis (SARA); and to evaluate the ability of beet pulp (BP) as a replacement for ground maize in order to alleviate SARA. Eight Holstein-Friesian cows were fed four diets (total mixed rations) during four successive periods (each of 17 d): (1) without wheat (W0); (2) with 10% finely ground wheat (FGW) (W10); (3) with 20% FGW (W20); (4) with 20% FGW and 10% pelleted BP (BP10). Inducing SARA by diet W20 decreased the daily mean ruminal pH (6.37 vs. 5.94) and the minimum ruminal pH (5.99 vs. 5.41) from baseline to challenge period. Ruminal concentrations of total volatile fatty acid, propionate, butyrate, valerate and isovalerate increased with the W20 compared with the W0 and W10 treatments. The substitution of BP for maize increased the minimum ruminal pH and molar percentage of acetate and decreased the molar percentage of butyrate. The diets had no effect on dry matter intake (DMI) and milk yield, but the milk fat percentage and yield as well as the amount of fat-corrected milk was reduced in the W20 and BP10 treatments. The cows fed the W20 diet had greater milk concentrations of C11:0, C13:0, C15:0, C14:1, C16:1, C17:1, C18:2n6c, C20:3n6, total polyunsaturated fatty acids (FA) and total odd-chain FA, and lower concentrations of C18:0 and total saturated FA compared with the cows fed the W0 diet. Therefore, it can be concluded that changes in ruminal fermentation, milk fat concentration and fatty acid profile are highly related to SARA induced by feeding high FGW diets, and that the substitution of BP for maize could reduce the risk of SARA in dairy cows.     

Relative reticulo-rumen pH indicators for subacute ruminal acidosis detection in dairy cows

Subacute ruminal acidosis (SARA) is usually characterized by abnormal and intermittent drops in rumen pH. Nevertheless, high individual animal variability in rumen pH and the difference in measurement methods for pH data acquisition decrease the sensitivity and accuracy of pH indicators for detecting SARA in ruminants. The aim of this study was to refine rumen pH indicators in long-term SARA based on individual dairy cow reticulo-rumen pH kinetics. Animal performances and rumen parameters were studied weekly in order to validate SARA syndrome and rumen pH was continuously measured using reticulo-rumen sensors. In total, 11 primiparous dairy cows were consecutively fed two different diets for 12 successive weeks: a control diet as low-starch diet (LSD; 13% starch for 4 weeks in period 1), an acidotic diet as high-starch diet (HSD; 32% starch for 4 weeks in period 2), and again the LSD diet (3 weeks in period 3). There was a 1-week dietary transition between LSD and HSD. Commonly used absolute SARA pH indicators such as daily average, area under the curve (AUC) and time spent below pH<5.8 and pH<6 were processed from absolute (raw) daily kinetics. Then signal processing was applied to raw pH values in order to calculate relative pH indicators by filtering and normalizing data to remove inter-individual variability, sensor drift and sensor noise. Normalized AUC, times spent below NpH<−0.3 and NpH<−0.5, NpH range and NpH standard deviation were calculated. Those relative pH indicators were compared with commonly used pH indicators to assess their ability to detect SARA. This syndrome induced by HSD was confirmed by consistent expected changes in milk quality, dry matter intake and acetate : propionate ratio in the rumen, whereas the ruminal concentration of lipopolysaccharide was increased. Commonly used pH SARA indicators were not able to discriminate SARA syndrome due to high animal variability and sensor drift and noise, whereas relative pH indicators developed in this study appeared more relevant for SARA detection as assessed by receiver operating characteristic tests. This work shows that absolute pH kinetics should be corrected for drift, noise and animal variability to produce relative pH indicators that are more robust for SARA detection. These relative pH indicators could be more relevant for identifying affected animals in a herd and also for comparing SARA risk among studies.     

Diet-induced bacterial immunogens in the gastrointestinal tract of dairy cows: Impacts on immunity and metabolism

Dairy cows are often fed high grain diets to meet the energy demand for high milk production or simply due to a lack of forages at times. As a result, ruminal acidosis, especially subacute ruminal acidosis (SARA), occurs frequently in practical dairy production. When SARA occurs, bacterial endotoxin (or lipopolysaccharide, LPS) is released in the rumen and the large intestine in a large amount. Many other bacterial immunogens may also be released in the digestive tract following feeding dairy cows diets containing high proportions of grain. LPS can be translocated into the bloodstream across the epithelium of the digestive tract, especially the lower tract, due to possible alterations of permeability and injuries of the epithelial tissue. As a result, the concentration of blood LPS increases. Immune responses are subsequently caused by circulating LPS, and the systemic effects include increases in concentrations of neutrophils and the acute phase proteins such as serum amyloid-A (SAA), haptoglobin (Hp), LPS binding protein (LBP), and C-reactive protein (CRP) in blood. Entry of LPS into blood can also result in metabolic alterations. Blood glucose and nonesterified fatty acid concentrations are enhanced accompanying an increase of blood LPS after increasing the amount of grain in the diet, which adversely affects feed intake of dairy cows. As the proportions of grain in the diet increase, patterns of plasma β-hydoxybutyric acid, cholesterol, and minerals (Ca, Fe, and Zn) are also perturbed. The bacterial immunogens can also lead to reduced supply of nutrients for synthesis of milk components and depressed functions of the epithelial cells in the mammary gland. The immune responses and metabolic alterations caused by circulating bacterial immunogens will exert an effect on milk production. It has been demonstrated that increases in concentrations of ruminal LPS and plasma acute phase proteins (CRP, SAA, and LBP) are associated with declines in milk fat content, milk fat yield, 3.5% fat-corrected milk yield, as well as milk energy efficiency.     

GC–MS analysis of the ruminal metabolome response to thiamine supplementation during high grain feeding in dairy cows

Introduction

Thiamine is known to attenuate high-concentrate diet induced subacute ruminal acidosis (SARA) in dairy cows, however, the underlying mechanisms remain unclear.

Objectives

The major objective of this study was to investigate the metabolic mechanisms of thiamine supplementation on high-concentrate diet induced SARA.

Methods

Six multiparous, rumen-fistulated Holstein cows were used in a replicated 3?×?3 Latin square design. The treatments included a control diet (CON; 20% starch, dry matter basis), a SARA-inducing diet (SAID; 33.2% starch, dry matter basis) and SARA-inducing diet supplemented with 180 mg of thiamine/kg of dry matter intake (SAID?+?T). On d21 of each period, ruminal fluid samples were collected at 3 h post feeding, and GC/MS was used to analyze rumen fluid samples.

Results

PCA and OPLS-DA analysis demonstrated that the ruminal metabolite profile were different in three treatments. Compared with CON treatment, SAID feeding significantly decreased rumen pH, acetate, succinic acid, increased propionate, pyruvate, lactate, glycine and biogenic amines including spermidine and putrescine. Thiamine supplementation significantly decreased rumen content of propionate, pyruvate, lactate, glycine and spermidine; increase rumen pH, acetate and some medium-chain fatty acids. The enrichment analysis of different metabolites indicated that thiamine supplementation mainly affected carbohydrates, amino acids, pyruvate and thiamine metabolism compared with SAID treatment.

Conclusions

These findings revealed that thiamine supplementation could attenuate high-concentrate diet induced SARA by increasing pyruvate formate-lyase activity to promote pyruvate to generate acetyl-CoA and inhibit lactate generation. Besides, thiamine reduced biogenic amines to alleviate ruminal epithelial inflammatory response.

     

Expression of MCT1, MCT2 and MCT4 in the rumen, small intestine and liver of reindeer (Rangifer tarandus tarandus L.)

The expression of monocarboxylate transporters MCT1, MCT2 and MCT4 in the rumen, small intestine and liver was examined in free-ranging and captive reindeer. In addition, expression of chaperone protein CD147, which is needed for the activity of MCT1 and MCT4, was studied in the rumen of suckling calves. Immunoblotting of cell membrane proteins showed the expression of MCT1 and MCT4, but not that of MCT2 in the rumen of reindeer. In free-ranging reindeer the amount of MCT1 was higher than in the captive ones (P<0.01). Developing rumen of suckling calves expressed MCT1 and MCT4 and positive correlation was found between MCT1 and CD147. Both MCT1 and CD147 correlated also with age in calves less than 10 days. In the small intestine all the isoforms studied were expressed, but the amounts were lower than in the rumen (P<0.05). In the liver MCT1 and MCT2 were found while MCT4 was nearly undetectable. The expression of MCT isoforms in the rumen and small intestine reflects the site of absorption and concentrations of short chain fatty acids (SCFA). In the liver the expression of high affinity transporters, MCT1 and MCT2, is in accordance with almost complete uptake of propionate from portal blood.     

Effect of short-chain fatty acids on the expression of genes involved in short-chain fatty acid transporters and inflammatory response in goat jejunum epithelial cells

Short-chain fatty acids (SCFAs) produced by microbial fermentation of dietary fibers are utilized by intestinal epithelial cells to provide an energy source for the ruminant. Although the regulation of mRNA expression and inflammatory response involved in SCFAs is established in other animals and tissues, the underlying mechanisms of the inflammatory response by SCFAs in goat jejunum epithelial cells (GJECs) have not been understood. Therefore, the objective of the study is to investigate the underlying mechanisms of the effects of SCFAs on SCFA transporters and inflammatory response in GJECs. These results showed that the acetate, butyrate, and SCFA concentration were markedly reduced in GJECs (p?<?0.01). In addition, the propionate concentration was significantly decreased in GJECs (p?<?0.05). The mRNA abundance of monocarboxylate transporter 1 (MCT1), MCT4, NHE1, and putative anion transporter 1 (PAT1) was elevated (p?<?0.05) by 20 mM SCFAs at pH 7.4 compared with exposure to the pH group. The anion exchanger 2 (AE2) was increased (p?<?0.05) by 20 mM SCFAs at pH 6.2. The mRNA abundance of vH⁺ ATPase B subunit (vH⁺ ATPase) was attenuated by SCFAs. For inflammatory responses, IL-1β and TNF-α were increased with SCFAs (p?<?0.05). In addition, IκBα involved in NF-κB signaling pathways was disrupted by SCFAs. Consistently, p-p65 signaling molecule was enhanced by adding SCFAs. However, IL-6 was attenuated by adding SCFAs (p?<?0.05). Furthermore, p-ErK1/2 mitogen-activated protein kinase (MAPK) signaling pathway was downregulated by adding SCFAs. In conclusion, these novel findings demonstrated that mRNA abundance involved in SCFA absorption is probably associated to SCFAs and pH value, and mechanism of the inflammatory response by SCFAs may be involved in NF-κB and p-ErK1/2 MAPK signaling pathways in GJECs. These pathways may mediate protective inflammation response in GJECs.     

Effect of oat hay provision method on growth performance,rumen fermentation and blood metabolites of dairy calves during preweaning and postweaning periods

The comparison of the effects of all forage offering methods would be particularly useful information in modeling growth performance and rumen fermentation of dairy calves. Therefore, this study attempted to evaluate the effects of methods of oat hay provision on growth performance, rumen fermentation and biochemical blood indices of dairy calves during preweaning and postweaning periods. At birth, 40 female Polish Holstein-Friesian calves (3 days of age; 39.6 ± 0.39 kg BW) were randomly assigned to four treatment groups differing in the access to chopped oat hay: CON (control, starter without oat hay), OH (starter feed containing 10% DM basis oat hay), OH-FC (starter feed containing 10% DM basis oat hay and oat hay fed as free-choice provision in different buckets) and FC (starter feed and oat hay fed as free-choice provision in different buckets). The calves were weaned on day 56, and then the study continued until day 84. Intakes of starter feed and oat hay were recorded daily, whereas BW and hip height (HH) on day 3 and then every 14 days. Samples of blood were collected on the initiation of experiment and then every 14 days, and rumen contents on day 28, 56 and 84. No treatment effects were found for starter, starch, CP, total DM intake, average daily gain, feeding efficiency, change in HH, ruminal fluid pH, concentrations of ruminal propionate and NH₃-N, concentrations of urea nitrogen and non-esterified fatty acids in the blood. There were differences between treatments in terms of ruminal total volatile fatty acids and molar concentrations of acetate, butyrate and acetate to propionate ratio; highest in OH and OH-FC groups, especially during the postweaning period. On the other hand, lower concentrations of iso-valerate were found in OH and OH-FC groups on day 56 and 84. The concentrations of IGF-I throughout the experiment and β-hydroxybutyrate during the postweaning period in the blood were influenced by treatment, with the greatest values observed in OH and OH-FC calves. Results of this study indicate that starter feed containing chopped oat hay improves rumen fermentation parameters, which might allow successful transition from preruminant to mature ruminant state. Also, providing chopped oat hay with pelleted starter feed seems to be a better method than free-choice supplementation.     

Effects of In-Feed Inclusion of Clinoptilolite on Blood Serum Concentrations of Aluminium and Inorganic Phosphorus and on Ruminal Ph and Volatile Fatty Acid Concentrations in Dairy Cows

The experiment investigated the effects of the dietary inclusion of 200 g of the natural zeolite, clinoptilolite on the blood serum concentrations of aluminium (Al) and inorganic phosphorus (P) as well as on the ruminal pH and the ruminal concentrations of Al and P and of certain volatile fatty acids. Sixteen Holstein cows with a rumen fistula were randomly assigned to one of two groups. Group A cows (n = 8) were fed the basal ration supplemented with 200 g of clinoptilolite per day, and group B cows (n = 8) were fed the basal ration and served as controls. Blood and rumen fluid samples were collected at the same day of each week and at the same time (5 h after morning feeding) for 12 weeks. Clinoptilolite supplementation had no significant effect on ruminal and blood serum concentrations of Al and P. However, clinoptilolite significantly increased ruminal pH and acetate, and decreased ruminal propionate and valerate.     

Impact of subacute ruminal acidosis on the diversity of liquid and solid-associated bacteria in the rumen of goats

This study was aimed to investigate the impact of subacute ruminal acidosis (SARA) on the diversity of liquid (LAB) and solid-associated bacteria (SAB) following high-grain feeding. Six ruminally cannulated goats were divided into two groups: one group was fed a hay diet (COD), and the other group was fed a high grain diet (SAID). Rumen liquids and rumen solids were sampled after 2 weeks adaption. SARA was diagnosed with a pH below 5.8 for 8 h. SAID decreased ruminal pH (P < 0.001) and increased the acetate (P = 0.017), propionate (P = 0.001), butyrate (P < 0.001) and total volatile fatty acid (P < 0.001) concentration in rumen compared with the COD. Denaturing gradient gel electrophoresis fingerprints analysis revealed a clear separation between both the diet and the fraction of rumen digesta in bacterial communities. Pyrosequencing analysis showed that the proportion of phylum Bacteroidetes in the SAID-LAB and SAID-SAB communities was less than in the COD group, whereas the SAID group had a greater percentage of Firmicutes in both the LAB and SAB libraries. UniFrac analyses and a Venn diagram revealed a large difference between the two diets in the diversity of rumen bacterial communities. Overall, our findings revealed that SARA feeding did alter the community structure of rumen liquids and rumen solids. Thus, manipulation of dietary factors, such as ratio of forage to concentrate may have the potential to alter the microbial composition of rumen liquid and rumen solid.     

Effects of the acid–base treatment of corn on rumen fermentation and microbiota,inflammatory response and growth performance in beef cattle fed high-concentrate diet

Beef cattle are often fed high-concentrate diet (HCD) to achieve high growth rate. However, HCD feeding is strongly associated with metabolic disorders. Mild acid treatment of grains in HCD with 1% hydrochloric acid (HA) followed by neutralization with sodium bicarbonate (SB) might modify rumen fermentation patterns and microbiota, thereby decreasing the negative effects of HCD. This study was thus aimed to investigate the effects of treatment of corn with 1% HA and subsequent neutralization with SB on rumen fermentation and microbiota, inflammatory response and growth performance in beef cattle fed HCD. Eighteen beef cattle were randomly allocated to three groups and each group was fed different diets: low-concentrate diet (LCD) (concentrate : forage = 40 : 60), HCD (concentrate : forage = 60 : 40) or HCD based on treated corn (HCDT) with the same concentrate to forage ratio as the HCD. The corn in the HCDT was steeped in 1% HA (wt/wt) for 48 h and neutralized with SB after HA treatment. The animal trial lasted for 42 days with an adaptation period of 7 days. At the end of the trial, rumen fluid samples were collected for measuring ruminal pH values, short-chain fatty acids, endotoxin (or lipopolysaccharide, LPS) and bacterial microbiota. Plasma samples were collected at the end of the trial to determine the concentrations of plasma LPS, proinflammatory cytokines and acute phase proteins (APPs). The results showed that compared with the LCD, feeding the HCD had better growth performance due to a shift in the ruminal fermentation pattern from acetate towards propionate, butyrate and valerate. However, the HCD decreased ruminal pH and increased ruminal LPS release and the concentrations of plasma proinflammatory cytokines and APPs. Furthermore, feeding the HCD reduced bacterial richness and diversity in the rumen. Treatment of corn increased resistant starch (RS) content. Compared with the HCD, feeding the HCDT reduced ruminal LPS and improved ruminal bacterial microbiota, resulting in decreased inflammation and improved growth performance. In conclusion, although the HCD had better growth performance than the LCD, feeding the HCD promoted the pH reduction and the LPS release in the rumen, disturbed the ruminal bacterial stability and increased inflammatory response. Treatment of corn with HA in combination with subsequent SB neutralization increased the RS content and helped counter the negative effects of feeding HCD to beef steers.     

Bovine rumen epithelium undergoes rapid structural adaptations during grain-induced subacute ruminal acidosis

Alterations in rumen epithelial structure and function during grain-induced subacute ruminal acidosis (SARA) are largely undescribed. In this study, four mature nonlactating dairy cattle were transitioned from a high-forage diet (HF; 0% grain) to a high-grain diet (HG; 65% grain). After feeding the HG diet for 3 wk, the cattle were transitioned back to the original HF diet, which was fed for an additional 3 wk. Continuous ruminal pH was measured on a weekly basis, and rumen papillae were biopsied during the baseline and at the first and final week of each diet. The mean, minimum, and maximum daily ruminal pH were depressed (P < 0.01) in the HG period compared with the HF period. During the HG period, SARA was diagnosed only during week 1, indicating ruminal adaptation to the HG diet. Microscopic examination of the papillae revealed a reduction (P < 0.01) in the stratum basale, spinosum, and granulosum layers, as well as total depth of the epithelium during the HG period. The highest (P < 0.05) papillae lesion scores were noted during week 1 when SARA occurred. Biopsied papillae exhibited a decline in cellular junctions, extensive sloughing of the stratum corneum, and the appearance of undifferentiated cells near the stratum corneum. Differential mRNA expression of candidate genes, including desmoglein 1 and IGF binding proteins 3, 5, and 6, was detected between diets using qRT-PCR. These results suggest that the structural integrity of the rumen epithelium is compromised during grain feeding and is associated with the differential expression of genes involved in epithelial growth and structure.     

Urea and short-chain fatty acids metabolism in Holstein cows fed a low-nitrogen grass-based diet

Three ruminally cannulated and multicatheterised lactating dairy cows were used to investigate the effect of different supplement strategies to fresh clover grass on urea and short-chain fatty acid (SCFA) metabolism in a zero-grazing experiment with 24-h blood and ruminal samplings. Fresh clover grass was cut every morning and offered from 0800 to 1500 h. Maize silage was fed at 1530 h. The three treatments, arranged in a Latin square, differed by timing of feeding rolled barley and soya-bean hulls relative to fresh clover grass. All diets had the same overall composition. Treatments were soya-bean hulls fed at 0700 h and barley fed at 1530 h (SAM), barley fed at 0700 h and soya-bean hulls fed at 1530 h (BAM), and both soya-bean hulls and barley fed at 1530 h (SBPM). The grass had an unexpectedly low content of crude protein (12.7%) and the cows were severely undersupplied with rumen degradable protein. The treatment effects were numerically small; greater arterial ammonia concentration, net portal flux of ammonia and net hepatic flux of urea during part of the day were observed when no supplementary carbohydrate was fed before grass feeding. A marked diurnal variation in ruminal fermentation was observed and grass feeding increased ruminal concentrations of propionate and butyrate. The net portal fluxes of propionate, butyrate, isovalerate and valerate as well as the net hepatic uptake of propionate, butyrate, valerate and caproate increased after feeding at 0700 h. The hepatic extraction of butyrate showed a relatively large depression with grass feeding with nadir at 1200 to 1330 h. The increased net portal absorption and the decreased hepatic extraction resulted in an approximately six-fold increase in the arterial blood concentration of butyrate. The gut entry rate of urea accounted for 70 ± 10% of the net hepatic production of urea. Saliva contributed to 14% of the total amount of urea recycled to the gut. Urea recycling to the gut was equivalent to 58% of the dietary nitrogen intake. Despite the severe undersupply of rumen degradable protein, the portal-drained viscera did not extract more than 4.3% of the urea supplied with arterial blood. This value is in line with the literature values for cows fed diets only moderately deficient in rumen degradable protein and indicates that cows maximise urea transfer across gut epithelia even when the diet is moderately deficient in rumen degradable protein.     

Calcium propionate supplementation improves development of rumen epithelium in calves via stimulating G protein-coupled receptors

In the present study, calcium propionate (CaP) was used as feed additive in the diet of calves to investigate their effects on rumen fermentation and the development of rumen epithelium in calves. To elucidate the mechanism in which CaP improves development of calf rumen epithelium via stimulating the messenger RNA (mRNA) expression of G protein-coupled receptors, a total of 54 male Jersey calves (age=7±1 days, BW=23.1±1.2 kg) were randomly divided into three treatment groups: control without CaP supplementation (Con), 5% CaP supplementation (5% CaP) and 10% CaP supplementation (10% CaP). The experiment lasted 160 days and was divided into three feeding stages: Stage 1 (days 0 to 30), Stage 2 (days 31 to 90) and Stage 3 (days 91 to 160). Calcium propionate supplementation percentages were calculated on a dry matter basis. In total, six calves from each group were randomly selected and slaughtered on days 30, 90 and 160 at the conclusion of each experimental feeding stage. Rumen fermentation was improved with increasing concentration of CaP supplementation in calves through the first 30 days (Stage 1). No effects of CaP supplementation were observed on rumen fermentation in calves during Stage 2 (days 31 to 90). Supplementation with 5% CaP increased propionate concentration, but not acetate and butyrate in calves during Stage 3 (days 91 to 160). The rumen papillae length of calves in the 5% CaP supplementation group was greater than that of Con groups in calves after 160 days feeding. The mRNA expression of G protein-coupled receptor 41 (GPR41) and GPR43 supplemented with 5% CaP were greater than the control group and 10% CaP group in feeding 160 days calves. 5% CaP supplementation increased the mRNA expression of cyclin D1, whereas did not increase the mRNA expression of cyclin-dependent kinase 4 compared with the control group in feeding 160-day calves. These results indicate that propionate may act as a signaling molecule to improve rumen epithelium development through stimulating mRNA expression of GPR41 and GPR43.     

Impact of subacute ruminal acidosis (SARA) adaptation and recovery on the density and diversity of bacteria in the rumen of dairy cows

Subacute ruminal acidosis (SARA) is characterized by ruminal pH depression and microbial perturbation. The impact of SARA adaptation and recovery on rumen bacterial density and diversity was investigated following high-grain feeding. Four ruminally cannulated dairy cows were fed a hay diet, transitioned to a 65% grain diet for 3 weeks, and returned to the hay diet for 3 weeks. Rumen fluid, rumen solids, and feces were sampled during weeks 0 (hay), 1 and 3 (high grain), and 4 and 6 (hay). SARA was diagnosed during week 1, with a pH below 5.6 for 4.6±1.4 h. Bacterial density was significantly lower in the rumen solids with high grain (P=0.047). Rumen fluid clone libraries from weeks 0, 3, and 6 were assessed at the 98% level and 154 operational taxonomic units were resolved. Week 3 diversity significantly differed from week 0, and community structure differed from weeks 0 and 6 (P<0.0001). Clones belonging to the phylum Firmicutes predominated. Compared with the hay diet, the high-grain diet contained clones from Selenomonas ruminantium and Succiniclasticum ruminis, but lacked Eubacterium spp. SARA adaptation was found to significantly alter bacterial density, diversity, and community structure, warranting further investigation into the role bacteria play in SARA adaptation.     

Short-chain fatty acids administration is protective in colitis-associated colorectal cancer development

Reduced short-chain fatty acids (SCFAs) have been reported in patients with ulcerative colitis, and increased intake of dietary fiber has shown to be clinically beneficial for colitis. Whether SCFAs suppress tumorigenesis in colitis-associated colorectal cancer remains unknown. The chemopreventive effect of SCFAs in colitis-associated colorectal cancer was evaluated in this study. Model of colitis-associated colorectal cancer in male BALB/c mice was induced by azoxymethane (AOM) and dextran sodium sulfate (DSS). SCFAs mix (67.5 mM acetate, 40 mM butyrate, 25.9 mM propionate) was administered in drink water during the study period. Macroscopic and histological studies were performed to examine the colorectal inflammation and tumorigenesis in AOM/DSS-induced mice treated with or without SCFA mix. The effects of SCFAs mix on colonic epithelial cellular proliferation were also assessed using Ki67 immunohistochemistry and TUNEL staining. The administration of SCFAs mix significantly reduced the tumor incidence and size in mice with AOM/DSS-induced colitis associated colorectal cancer. SCFAs mix protected from AOM/DSS-induced colorectal cancer by improving colon inflammation and disease activity index score as well as suppressing the expression of proinflammatory cytokines including IL-6, TNF-α and IL-17. A decrease in cell proliferation markers and an increase in TUNEL-positive tumor epithelial cells were also demonstrated in AOM/DSS mice treated with SCFAs mix. SCFAs mix administration prevented development of tumor and attenuated the colonic inflammation in a mouse model of colitis-associated colorectal cancer. SCFAs mix may be a potential agent in the prevention and treatment of colitis-associated colorectal cancer.     

Effect of disodium fumarate on ruminal metabolism and rumen bacterial communities as revealed by denaturing gradient gel electrophoresis analysis of 16S ribosomal DNA

The aim of the present study was to examine the effects of feeding diets with addition of disodium fumarate (DF) to goats on ruminal metabolism and changes of rumen bacterial communities. Four cannulated goats were used in a 4 × 4 Latin square design. The results showed that ruminal pH increased linearly (P<0.01)as the amount of DF added increased, while lactate production decreased linearly (P<0.01). DF addition did not affect the production of acetate, propionate, butyrate, TVFA and NH₃-N. The effect of DF on the changes in rumen bacterial-community structure of goats was analyzed using 16S rDNA-based approaches. Amplicons of the V6-V8 variable regions of bacterial 16S rDNA were analyzed by denaturing gradient gel electrophoresis (DGGE), cloning and sequencing. Differences in rumen bacterial community structure were determined based on the Shannon index of diversity for pairwise comparison of the DGGE fingerprints and revealed significant changes in rumen microbiota after DF addition. As compared with those fed with the control diet, goats fed on the diets with DF addition showed a higher bacterial diversity. The sequences of seven amplicons in total 11 clones showed less than 97% similarity with those of previously identified or unidentified bacteria, suggesting that most bacteria in the gastrointestinal tract have not been cultured or identified. Amplicons related to Succinivibrio dextrinosolvens species were found in most DGGE fingerprints derived from goats on the diet containing DF, but not in goats on the control diet. These results demonstrated the ability of DF to improve the metabolism of rumen lactate fermentation and to influence the bacterial composition of the rumen in goats.     

In Vitro Lactate Metabolism by Ruminal Ingesta

Ruminal ingesta (300 ml) obtained from a fistulated cow fed alfalfa hay (H), 3.6 kg of grain mixture with corn silage fed ad libitum (S), 2.5:1 grain-alfalfa hay mixture (G), or a 2.5:1 grain-alfalfa hay mixture providing 545 g of sodium and calcium lactate daily (L) were incubated for 8 hr with nonpolymerized sodium lactate or 17% polymerized lactic acid neutralized to pH 6.7. Polymerization had no effect on the rate of lactate utilization. The initial rates of lactate metabolism for the H, G, S, and L ingesta were 0.72, 0.95, 1.8, and 3.4 meq per 100 ml of rumen fluid per hr, respectively. Lactate-2-(14)C was incubated for 4 hr with each type of ruminal ingesta. Of the label recovered in the volatile fatty acids (VFA), 74.1, 61.2, 49.3, and 38.9% was recovered in acetate, and 9.4, 19.8, 23.3, and 51.9% was recovered in propionate with H, G, S, and L ingesta, respectively. The balance of label was distributed between butyrate and valerate. The titratable VFA did not follow this pattern of production. With the hay ingesta, lactate metabolism resulted in a net loss of acetate and a large increase in butyrate. Little propionate was produced. The G, S, and L ingesta metabolized lactate to yield progressively more propionate and less butyrate. Evidence was gathered to suggest that acetate was the primary end product of lactate metabolism but that oxidation of lactate to pyruvate dictated the synthesis of butyrate from acetate to maintain an oxidation-reduction balance. It was noted that acetate and butyrate production from lactate was pH-dependent, with acetate production maximal at pH 7.4 and butyrate at 6.2. Propionate production was largely unaffected within this pH range.     

Ruminal transport and metabolism of short-chain fatty acids (SCFA) in vitro: effect of SCFA chain length and pH

The unidirectional transport and metabolism of 14C-labeled acetate, propionate and butyrate across the isolated bovine rumen epithelium was measured in vitro by the Ussing chamber technique. There was a significant, but relatively small, net secretion of acetate and propionate, and a large and significant net absorption of butyrate. The results demonstrate that the mucosal-serosal (MS) pathway for short-chain fatty acids (SCFA) is different from the serosal-mucosal (SM) pathway, and that butyrate is treated differently from acetate and propionate by the epithelium. The results support that the main route for epithelial SCFA transport is transcellular. The correlation between SCFA lipophility and the flux rate was positive but weak at both pH 7.3 and 6.0. Decreasing pH increased all SCFA fluxes significantly, but not proportionally to the increase of protonized SCFA in the bathing solution. There was a significant and apparently non-competitive interaction between the transport of acetate, propionate and butyrate. It seems that mediated transport mechanisms must be involved in epithelial SCFA transport in the bovine rumen, but the data do not exclude that passive diffusion could account for a significant part of the flux. The metabolism of SCFA in the Ussing chamber system was considerable, and there was a clear preference for excretion of CO2 from this metabolism to the mucosal side, while side preference for non-CO2 metabolite excretion was not studied. Of the propionate and butyrate transported in the MS direction, 78 and 95% was metabolised, while only 37 and 38% was metabolised in the SM direction (acetate metabolism could not be measured). There was, however, no simple relation between the degree of metabolism and the transport rate or the transport asymmetry of the SCFA.