Gut microbiota contributes to bisphenol A-induced maternal intestinal and placental apoptosis, oxidative stress, and fetal growth restriction in pregnant ewe model by regulating gut-placental axis.

BACKGROUND: Bisphenol A (BPA) is an environmental contaminant with endocrine-disrupting properties that induce fetal growth restriction (FGR). Previous studies on pregnant ewes revealed that BPA exposure causes placental apoptosis and oxidative stress (OS) and decreases placental efficiency, consequently leading to FGR. Nonetheless, the response of gut microbiota to BPA exposure and its role in aggravating BPA-mediated apoptosis, autophagy, mitochondrial dysfunction, endoplasmic reticulum stress (ERS), and OS of the maternal placenta and intestine are unclear in an ovine model of gestation. RESULTS: Two pregnant ewe groups (n = 8/group) were given either a subcutaneous (sc) injection of corn oil (CON group) or BPA (5 mg/kg/day) dissolved in corn oil (BPA group) once daily, from day 40 to day 110 of gestation. The maternal colonic digesta and the ileum and placental tissue samples were collected to measure the biomarkers of autophagy, apoptosis, mitochondrial dysfunction, ERS, and OS. To investigate the link between gut microbiota and the BPA-induced FGR in pregnant ewes, gut microbiota transplantation (GMT) was conducted in two pregnant mice groups (n = 10/group) from day 0 to day 18 of gestation after removing their intestinal microbiota by antibiotics. The results indicated that BPA aggravates apoptosis, ERS and autophagy, mitochondrial function injury of the placenta and ileum, and gut microbiota dysbiosis in pregnant ewes. GMT indicated that BPA-induced ERS, autophagy, and apoptosis in the ileum and placenta are attributed to gut microbiota dysbiosis resulting from BPA exposure. CONCLUSIONS: Our findings indicate the underlying role of gut microbiota dysbiosis and gut-placental axis behind the BPA-mediated maternal intestinal and placental apoptosis, OS, and FGR. The findings further provide novel insights into modulating the balance of gut microbiota through medication or probiotics, functioning via the gut-placental axis, to alleviate gut-derived placental impairment or FGR. Video Abstract.

Dietary rumen-protected L-arginine or N-carbamylglutamate enhances placental amino acid transport and suppresses angiogenesis and steroid anabolism in underfed pregnant ewes.

This study aimed to investigate the effects of dietary supplementation of underfed Hu ewes from d 35 to 110 of gestation with either rumen-protected L-arginine (RP-Arg) or N-carbamylglutamate (NCG) on placental amino acid (AA) transport, angiogenic gene expression, and steroid anabolism. On d 35 of gestation, 32 Hu ewes carrying twin fetuses were randomly divided into four treatment groups, each consisting of eight ewes, and were fed the following diets: A diet providing 100% of NRC's nutrient requirements for pregnant ewes (CON); A diet providing 50% of NRC's nutrient requirements for pregnant ewes (RES); RES diet plus 5 g/d NCG (RES + NCG); or RES diet plus 20 g/d RP-Arg (RES + ARG). On the d 110 of pregnancy, blood samples were taken from the mother, and samples were collected from type A cotyledons (COT; the fetal portions of the placenta). The levels of 17ß-estradiol and progesterone in the maternal serum and both the capillary area density (CAD) and capillary surface density (CSD) in type A COT were decreased in response to Arg or NCG supplementation when compared to the RES group. The concentrations of arginine, leucine, putrescine and spermidine in type A COT were higher (P < 0.05) in the RES + ARG or RES + NCG group than in the RES group. The mRNA expression levels of inducible nitric oxide synthase (iNOS) and solute carrier family 15, member 1 (SLC15A1) were increased (P < 0.05) while those of progesterone receptor (PGR) and fibroblast growth factor 2 (FGF2) were decreased in type A COT by supplementation with either NCG or RP-Arg compared to the RES group. The results suggest that providing underfed pregnant ewes from d 35 to 110 of gestation with a diet supplemented with NCG or RP-Arg improves placental AA transport, and reduces the expression of angiogenic growth factor genes and steroid anabolism, leading to better fetal development.

Dietary rumen-protected choline supplementation regulates blood biochemical profiles and urinary metabolome and improves growth performance of growing lambs.

This study aimed to assess the growth performance and blood metabolites, as well as metabolic profiles in the urine of lambs fed on dietary rumen-protected choline (RPC). Thirty-six Dorper × Hu lambs weighing approximately 20 kg were equally assigned to three groups, and fed on three diets supplemented with different RPC concentrations (0, 0.25% and 0.75%) for 45 days. Supplementation of RPC significantly increased average daily gain (ADG) and decreased feed-to-gain ratio (F/G) of lambs (p < 0.05). Dietary RPC was significantly associated with elevated plasma high-density lipoprotein (HDL) and suppressed low-density lipoprotein (LDL) levels when compared to the control group (p < 0.05). Moreover, concentrations of very-low-density lipoprotein (VLDL) exhibited an increasing trend (p = 0.065), whereas ß-hydroxybutyrate (BHBA) levels decreased (p = 0.086) in plasma. Analysis of urine metabolome revealed that RPC supplementation significantly suppressed urinary concentrations of pyruvate (p < 0.05), while increased urinary concentrations of trimethylamine oxide, p-cresol, phenylacetylglycine and hippurate (p < 0.05). These findings suggest that RPC supplementation can promote weight gain, alter plasma lipid metabolism and modify urinary metabolome which is correlated with energy metabolism, lipid metabolism and intestinal microbial metabolism in lambs. In conclusion, based on our findings, we recommend 0.25% RPC as a supplement for growing lambs.

Dietary N-carbamylglutamate and L-arginine supplementation improves redox status and suppresses apoptosis in the colon of intrauterine growth-retarded suckling lambs.

Previous studies have revealed that dietary N-carbamylglutamate (NCG) or L-arginine (Arg) improves small intestinal integrity and immune function in suckling Hu lambs that have experienced intrauterine growth retardation (IUGR). Whether these nutrients alter redox status and apoptosis in the colon of IUGR lambs is still unknown. This study, therefore, aimed at investigating whether dietary supplementation of Arg or NCG alters colonic redox status, apoptosis and endoplasmic reticulum (ER) stress and the underlying mechanism of these alterations in IUGR suckling Hu lambs. Forty-eight 7-d old Hu lambs, including 12 with normal birth weight (4.25 ± 0.14 kg) and 36 with IUGR (3.01 ± 0.12 kg), were assigned to 4 treatment groups (n = 12 each; 6 males and 6 females) for 3 weeks. The treatment groups were control (CON), IUGR, IUGR + Arg and IUGR + NCG. Relative to IUGR lambs, superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) content, as well as proliferation index, were higher (P < 0.05) whereas reactive oxygen species (ROS), malondialdehyde (MDA) levels and apoptotic cell numbers were lower (P < 0.05) in colonic tissue for both IUGR + Arg and NCG lambs. Both mRNA and protein levels of C/EBP homologous protein 10 (CHOP10), B-cell lymphoma/leukaemia 2 (Bcl-2) -associated X protein (Bax), apoptosis antigen 1 (Fas), activating transcription factor 6 (ATF6), caspase 3, and glucose-regulated protein 78 (GRP78) were lower (P < 0.05) while glutathione peroxidase 1 (GPx1), Bcl-2 and catalase (CAT) levels were higher (P < 0.05) in colonic tissue for IUGR + Arg and IUGR + NCG lambs compared with IUGR lambs. Based on our results, dietary NCG or Arg supplementation can improve colonic redox status and suppress apoptosis via death receptor-dependent, mitochondrial and ER stress pathways in IUGR suckling lambs.

Dietary L-Arginine or N-Carbamylglutamate Alleviates Colonic Barrier Injury, Oxidative Stress, and Inflammation by Modulation of Intestinal Microbiota in Intrauterine Growth-Retarded Suckling Lambs.

Our previous studies have revealed that dietary N-carbamylglutamate (NCG) and L-arginine (Arg) supplementation improves redox status and suppresses apoptosis in the colon of suckling Hu lambs with intrauterine growth retardation (IUGR). However, no studies have reported the function of Arg or NCG in the colonic microbial communities, barrier function, and inflammation in IUGR-suckling lambs. This work aimed to further investigate how dietary Arg or NCG influences the microbiota, barrier function, and inflammation in the colon of IUGR lambs. Forty-eight newborn Hu lambs of 7 d old were assigned to four treatment groups (n = 12 per group; six male, six female) as follows: CON (normal birth weight, 4.25 ± 0.14 kg), IUGR (3.01 ± 0.12 kg), IUGR + Arg (2.99 ± 0.13 kg), and IUGR + NCG (3.03 ± 0.11 kg). A total of 1% Arg or 0.1% NCG was supplemented in a basal diet of milk replacer, respectively. Lambs were fed the milk replacer for 21 d until 28 d after birth. Compared to the non-supplemented IUGR lambs, the transepithelial electrical resistance (TER) was higher, while fluorescein isothiocyanate dextran 4 kDa (FD4) was lower in the colon of the NCG- or Arg-supplemented IUGR lambs (p < 0.05). The IUGR lambs exhibited higher (p < 0.05) colonic interleukin (IL)-6, IL-1ß, tumor necrosis factor (TNF)-α, reactive oxygen species (ROS), and malondialdehyde (MDA) levels than the CON lambs; the detrimental effects of IUGR on colonic proinflammatory cytokine concentrations and redox status were counteracted by dietary Arg or NCG supplementation. Both IUGR + Arg and IUGR + NCG lambs exhibited an elevated protein and mRNA expression of Occludin, Claudin-1, and zonula occludens-1 (ZO-1) compared to the IUGR lambs (p < 0.05). Additionally, the lipopolysaccharide (LPS) concentration was decreased while the levels of acetate, butyrate, and propionate were increased in IUGR + Arg and IUGR + NCG lambs compared to the IUGR lambs (p < 0.05). The relative abundance of Clostridium, Lactobacillus, and Streptococcus was lower in the colonic mucosa of the IUGR lambs than in the CON lambs (p < 0.05) but was restored upon the dietary supplementation of Arg or NCG to the IUGR lambs (p < 0.05). Both Arg and NCG can alleviate colonic barrier injury, oxidative stress (OS), and inflammation by the modulation of colonic microbiota in IUGR-suckling lambs. This work contributes to improving knowledge about the crosstalk among gut microbiota, immunity, OS, and barrier function and emphasizes the potential of Arg or NCG in health enhancement as feed additives in the early life nutrition of ruminants.

Thiamine Supplementation Alleviates Lipopolysaccharide-Triggered Adaptive Inflammatory Response and Modulates Energy State via Suppression of NFκB/p38 MAPK/AMPK Signaling in Rumen Epithelial Cells of Goats.

Studies have shown that exogenous thiamine (THI) supplementation can alleviate inflammation and promote rumen epithelial development in goats and cows. This research aimed to evaluate the effect of THI supplementation on LPS-induced inflammation and energy metabolic dysregulation in RECs of goats. Cells were stimulated with either 5 µg/mL THI for 18 h (THI group) or with 5 µg/mL LPS for 6 h (LPS group). The CON group was stimulated with DMEM/F-12 medium without THI for 18 h. The LPTH group was pretreated with THI for 18 h, followed by LPS stimulation for 6 h. THI supplementation decreased the ROS content (p < 0.05), as well as the ratios of phosphorylated (p)-p65 to p65 (p < 0.05) and p-AMPKα to AMPKα (p < 0.05). Interestingly, when the p38 gene was overexpressed in the LPTH group, the ratio of p-p65 to p65 and p-AMPKα to AMPKα proteins significantly increased, and ATP content decreased (p < 0.05). Our results suggest that THI possesses anti-inflammatory and metabolic-modulatory effects in RECs. The mechanism is largely related to the suppression of the NF-κB/p38 MAPK/AMPK signaling pathway. Additionally, we also revealed that THI supplementation can inhibit LPS-induced oxidative damage and apoptosis to protect mitochondrial function in RECs.

Effects of the Gut Microbiota and Barrier Function on Melatonin Efficacy in Alleviating Liver Injury.

Environmental cadmium (Cd) exposure has been associated with severe liver injury. In contrast, melatonin (Mel) is a candidate drug therapy for Cd-induced liver injury due to its diverse hepatoprotective activities. However, the precise molecular mechanism by which Mel alleviates the Cd-induced liver injury, as well as the Mel-gut microbiota interaction in liver health, remains unknown. In this study, mice were given oral gavage CdCl2 and Mel for 10 weeks before the collection of liver tissues and colonic contents. The role of the gut microbiota in Mel's efficacy in alleviating the Cd-induced liver injury was evaluated by the gut microbiota depletion technique in the presence of antibiotic treatment and gut microbiota transplantation (GMT). Our results revealed that the oral administration of Mel supplementation mitigated liver inflammation, endoplasmic reticulum (ER) stress and mitophagy, improved the oxidation of fatty acids, and counteracted intestinal microbial dysbiosis in mice suffering from liver injury. It was interesting to find that neither Mel nor Cd administration induced any changes in the liver of antibiotic-treated mice. By adopting the GMT approach where gut microbiota collected from mice in the control (CON), Cd, or Mel + Cd treatment groups was colonized in mice, it was found that gut microbiota was involved in Cd-induced liver injury. Therefore, the gut microbiota is involved in the Mel-mediated mitigation of ER stress, liver inflammation and mitophagy, and the improved oxidation of fatty acids in mice suffering from Cd-induced liver injury.

Extra arginine supplementation during the suckling period alleviates weaning stress through the regulation of dendritic cells and Notch2 signaling in piglets.

This study aims to study the effects of extra arginine (Arg) supplementation during the suckling period on the weaning stress and intestinal barrier function of breastfed piglets. Forty 7-day-old breastfed piglets divided into the control group (CON) and Arg group (Arg) were fed with extra saline or Arg (250 mg per kg per d body weight), respectively. All piglets were weaned when they were 21 days old. Eight piglets from each group were sacrificed before weaning and on the 3rd-day after weaning, respectively. The results showed that Arg improved the average daily weight gain of piglets before weaning (P < 0.01) and decreased the average daily weight loss after weaning (P < 0.05). Weaning decreased the ratio of the villus length versus crypt depth (V/C) in the SI (P < 0.001), while Arg increased the V/C of the jejunum (P < 0.05). Arg increased the levels of immunoglobulins in the serum and SI (P < 0.05), decreased pro-inflammatory cytokines and increased anti-inflammatory cytokines in the SI (P < 0.05). In addition, Arg supplementation increased the numbers of SWC3a+CD40+ (P < 0.01) and SWC3a+SLAII+ DCs (P < 0.05), down-regulated Notch2 expression and up-regulated Jagged1 expression in the ilea of weaning piglets (P < 0.05). In conclusion, Arg supplementation during the suckling period decreased the LDH leakage in the SI, improved the intestinal morphology, down-regulated the contents of pro-inflammatory cytokines, accelerated the accumulation of DC precursors before weaning and increased the number of mature DCs after weaning, and thus improved the growth performance and reduced the weaning stress of piglets, and this might be associated with the regulation of Notch2 signaling.

Subacute ruminal acidosis in dairy herds: Microbiological and nutritional causes, consequences, and prevention strategies.

Dairy cattle are frequently fed high-concentrate (HC) diets in modern intensive feeding systems, especially in the transition period. During this period, cows face many alterations that include hormonal changes and shifting to a lactating state. Switching to a HC diet that may disrupt the ruminal microbiota balance can lead to subacute ruminal acidosis (SARA). Moreover, the main factor shaping the rumen microbiota is dietary composition, especially the ratio of starch to fibrous carbohydrates. Feeding highly fermentable carbohydrate diets after adaptation to forage diets leads to a rumen fermentation rate that exceeds rumen absorption and buffering rates, resulting in a reduction in ruminal pH. As a result of Gram-negative bacterial cell lysis, an increase in harmful ruminal bacterial metabolites, including lipopolysaccharide, lactic acid, and histamine, is observed. The interactions between the host immune system and the ruminal microbiota play an essential role in many physiological processes and the development of the disorder. Progress in DNA sequencing and bioinformatics platforms provides new opportunities to investigate the composition of ruminal microbes and yields unique advances in understanding ecology of the rumen. Subacute ruminal acidosis is linked with a change in the ruminal microbiota structure and richness and with other metabolic disorders; such as rumenitis, milk fat depression, laminitis, and liver abscesses. Therefore, this review aims to explore a better understanding of the crosstalk between diet and microbiota in the prevalence of rumen acidosis and its consequences, which is crucial for control strategies such as feeding management, and supplementation with thiamine, prebiotics, and probiotics.

Dietary supplementation of thiamine enhances colonic integrity and modulates mucosal inflammation injury in goats challenged by lipopolysaccharide and low pH.

The current study aimed to investigate the protective effects of dietary thiamine supplementation on the regulation of colonic integrity and mucosal inflammation in goats fed a high-concentrate (HC) diet. Twenty-four Boer goats (live weight of 35·62 (sem 2·4) kg) were allocated to three groups (CON: concentrate/forage = 30:70; HC; concentrate/forage = 70:30 and HCT: concentrate/forage = 70:30 with 200 mg thiamine/kg DMI) for 12 weeks. Results showed that compared with the HC treatment, the HCT group had a significantly higher ruminal pH value from 0 to 12 h after the feeding. The haematoxylin-eosin staining showed that desquamation and severe cellular damage were observed in the colon epithelium of the HC group, whereas the HCT group exhibited more structural integrity of the epithelial cell morphology. Compared with the HC treatment, the HCT group showed a markedly increase in pyruvate dehydrogenase and α-ketoglutarate dehydrogenase enzymes activity. The mRNA expressions in the colonic epithelium of SLC19A2, SLC19A3, SLC25A19, Bcl-2, occludin, claudin-1, claudin-4 and ZO-1 in the HCT group were significantly increased in comparison with the HC diet treatment. Compared with the HC treatment, the HCT diet significantly increased the protein expression of claudin-1 and significantly decreased the protein expression of NF-κB-related proteins p65. The results show that dietary thiamine supplementation could improve the colon epithelial barrier function and alleviate mucosal inflammation injury in goats after lipopolysaccharide and low pH challenge.

Dietary N-carbamylglutamate or L-arginine improves fetal intestinal amino acid profiles during intrauterine growth restriction in undernourished ewes.

Our previous studies demonstrated that prenatal in utero growth restriction impairs postnatal intestinal function. Thus, improving postpartal intestinal absorption capacity and growth by manipulating the maternal diet prepartum is of importance. This work was conducted to determine whether supplementation of N-carbamylglutamate (NCG) or rumen-protected L-arginine (RP-Arg) increased fetal intestinal amino acid (AA) profiles in intrauterine growth retardation (IUGR) fetuses. On d 35 of gestation, Hu ewes (n = 32) carrying twin fetuses were randomized into 4 groups (8 ewes and 16 fetuses in each group), where diets were as follows: 100% of nutrient requirements recommended by National Research Council (NRC, 2007) (CON); 50% of nutrient requirements recommended by NRC (2007) (RES); RES + RP-Arg (20 g/d), (RES + ARG); and RES + NCG (5 g/d), (RES + NCG). On d 110 of gestation, both fetal and maternal tissues were collected and weighed. Compared with RES, solute carrier family 1, member 5 (SLC1A5) was upregulated (P < 0.05) within fetal jejunum, duodenum and ileum when supplementing NCG and RP-Arg. Relative to RES, RP-Arg or NCG supplementation to RES resulted in upregulation (P < 0.05) of peptide transporter 1 protein abundance within the fetal ileum. NCG or RP-Arg supplementation to RES also upregulated phosphorylated mechanistic target of rapamycin (pmTOR)-to-mTOR ratio in the fetal ileum induced by IUGR (P < 0.05). As a result, during IUGR, supplementation of Arg or NCG affected intestinal AA profiles in the fetus in part through controlling mTOR signal transduction as well as AA and peptide transport. Future studies should be conducted to understand the role (if any) of the placenta on the improvement of growth and AA profiles independent of the fetal intestine. This would help demonstrate the relative contribution of intestinal uptake in fetal life.

Dietary rumen-protected L-arginine or N-carbamylglutamate attenuated fetal hepatic inflammation in undernourished ewes suffering from intrauterine growth restriction.

This study aimed to explore whether dietary rumen-protected L-arginine (RP-Arg) or N-carbamylglutamate (NCG) supplementation to feed-restricted pregnant ewes counteracts fetal hepatic inflammation and innate immune dysfunction associated with intrauterine growth retardation (IUGR) in ovine fetuses. On d 35 of pregnancy, twin-bearing Hu ewes (n = 32) were randomly assigned to 4 treatment groups (8 ewes and 16 fetuses per group) and fed diets containing 100% of the NRC requirements (CON), 50% of the NRC requirements (RES), RES + RP-Arg (20 g/d) (RESA), or RES + NCG (5 g/d) (RESN). At 08:00 on d 110 of gestation, fetal blood and liver tissue samples were collected. The levels of triglyceride, free fatty acid, cholesterol and ß-hydroxybutyrate in the fetal blood of RESA and RESN groups were lower (P < 0.05) than those of the RES group, but were higher (P < 0.05) than those of the CON group. The interleukin (IL)-6 and IL-1 levels in fetal blood and liver tissue as well as the myeloid differentiation primary response 88 (MyD88), transforming growth factor ß (TGFß), and nuclear factor kappa B (NF-κB) mRNA levels in the fetal liver were decreased (P < 0.05) by the NCG or RP-Arg supplementation compared to the RES treatment. Similarly, the toll-like receptor (TLR)-4, MyD88, TGFß, and p-c-Jun N-terminal kinase (JNK) protein levels in the fetal liver were reduced (P < 0.05) in the NCG and RP-Arg -supplemented groups compared to the RES group. These results showed that dietary supplementation of RP-Arg or NCG to underfed pregnant ewes could protect against IUGR fetal hepatic inflammation via improving lipid metabolism, down-regulating the TLR-4 and the inflammatory JNK and NF-κB signaling pathways, and decreasing cytokine production in ovine fetal blood and liver tissue.

Dietary N-carbamylglutamate or l-arginine supplementation improves hepatic energy status and mitochondrial function and inhibits the AMP-activated protein kinase-peroxisome proliferator-activated receptor γ coactivator-1α-transcription factor A pathway in intrauterine-growth-retarded suckling lambs.

The objective of this study was to investigate the effects of dietary administration of l-arginine (Arg) or N-carbamylglutamate (NCG) on hepatic energy status and mitochondrial functions in suckling Hu lambs with intrauterine growth retardation (IUGR). Forty-eight newborn Hu lambs of 7 d old were allocated into 4 treatment groups of 12 lambs each, in triplicate with 4 lambs per replicate (2 males and 2 females) as follows: CON (lambs of normal birth weight, 4.25 ± 0.14 kg), IUGR (3.01 ± 0.12 kg), IUGR + 1% Arg (2.99 ± 0.13 kg), or IUGR + 0.1% NCG (3.03 ± 0.11 kg). The experiment lasted for 21 d, until d 28 after birth, and all lambs were fed milk replacer as a basal diet. Compared with IUGR lambs, NCG or Arg administration increased (P < 0.05) the adenosine triphosphate (ATP) level and the activities of complexes I/III/IV, isocitrate dehydrogenase and citrate synthase in the liver. Compared with CON lambs, the relative mRNA levels of adenosine monophosphate-activated protein kinase α1 (AMPKα1), peroxisome proliferator-activated receptor γ coactivator-1α (PGC1α) and transcription factor A (TFAM) were increased (P < 0.05) in the liver of IUGR lambs, but were decreased (P < 0.05) in the liver of NCG- or Arg-treated lambs compared with those in the IUGR lambs. Compared with IUGR lambs, NCG or Arg administration decreased (P < 0.05) the total AMPKα (tAMPKα)-to-phosphorylated AMPKα (pAMPKα) ratio and the protein expression of PGC1α and TFAM. The results suggested that dietary Arg or NCG supplements improved hepatic energy status and mitochondrial function and inhibited the AMPK-PGC1α-TFAM pathway in IUGR suckling lambs.

Effects of irrigation and nitrogen on chlorophyll content, dry matter and nitrogen accumulation in sugar beet (Beta vulgaris L.).

A 2-year field experiment was conducted to analyze the growth conditions, physical features, yield, and nitrogen use efficiency (NUE) of sugar-beet under limited irrigation conditions in northeast of China. A cultivar H003 was used as plant materials; six treatments (C1-C6) were included: C1, no nitrogen applied, rain-fed; C2, nitrogen (120.00 kg ha-1), rain-fed; C3, no nitrogen applied, hole irrigation for seeding; C4, nitrogen (120.00 kg ha-1), hole irrigation for seeding; C5, no nitrogen applied, hole irrigation for seeding; and C6, nitrogen (120.00 kg ha-1), hole irrigation for seeding, and irrigation at foliage rapid growth stage. The irrigation supply was only 500 mL/plant once. Results showed C6 showed the highest chlorophyll content, dry matter accumulation, yield, etc. and had the best NUE among all the treatments. In conclusion, under the routine fertilization conditions of northeast of China, the cultivation measure of hole irrigation 500 mL/plant for seeding combined with irrigation 500 mL/plant at foliage rapid growth stage greatly improved sugar-beet yield and NUE.

Thiamine Alleviates High-Concentrate-Diet-Induced Oxidative Stress, Apoptosis, and Protects the Rumen Epithelial Barrier Function in Goats.

High-concentrate diets are continually used in ruminants to meet the needs of milk yield, which can lead to the occurrence of subacute rumen acidosis in ruminants. This study investigated the protective effects of dietary thiamine supplementation on the damage of the ruminal epithelium barrier function in goats fed a high-concentrate diet. Twenty-four healthy Boer goats (live weight of 35.62 ± 2.4 kg; age, 1 year) were randomly assigned into three treatments, with eight goats in each treatment, consuming one of three diets: a low-concentrate diet (CON; concentrate/forage, 30:70), a high-concentrate diet (HC; concentrate/forage, 70:30), or a high-concentrate diet with 200 mg of thiamine/kg of dry matter intake (HCT; concentrate/forage, 70:30) for 12 weeks. The additional dose of thiamine was based on our previous study wherein thiamine ameliorates inflammation. Compared with HC treatment, the HCT treatment had markedly higher concentrations of glutathione, superoxide dismutase, and glutathione peroxidase and total antioxidant capacity (P < 0.05) in plasma and rumen epithelium. The results showed that the apoptosis index was lower (P < 0.05) in the HCT treatment than in that of the HC treatment. Compared with the HC treatment, permeability and the electrophysiology parameter short circuit current for ruminal epithelial tissue were significantly decreased (P < 0.05) in the HCT treatment. The immunohistochemical results showed that the expression distribution of tight junctions including claudin-1, claudin-4, occludin, and zonula occludin-1 (ZO-1) was greater (P < 0.05) in the HCT treatments than in the HC treatment. The mRNA expression in the rumen epithelium of ZO-1, occludin, claudin-1, B-cell lymphoma/leukemia 2, nuclear factor erythroid-2 related factor 2 (Nrf2), superoxide dismutase 2 (SOD2), glutathione peroxidase 1, and the phase II metabolizing enzymes quinone oxidoreductase and heme oxygenase in the HCT group was significantly increased in comparison with the HC diet treatment (P < 0.05), whereas the mRNA expression of caspase 3, caspase 8, caspase 9, bcl-2 associated X protein, lipopolysaccharide binding protein, toll-like receptor 4, nuclear factor kappa-B (NFκB), tumor necrosis factor alpha, interleukin-1ß, interleukin, and tumor necrosis factor receptor-associated factor 6 decreased significantly in the HCT treatment (P < 0.05). Compared with the HC treatment, the HCT diet significantly increased the protein expression of ZO-1, occludin, claudin-1, NQO1, HO-1, SOD2, serine/threonine kinase, p-Akt, Nrf2, and p-Nrf2; conversely, the expression of NFκB-related proteins p65 and pp65 was significantly decreased (P < 0.05). In addition, thiamine relieved the damage on the ruminal epithelium caused by the HC diet. The results show that dietary thiamine supplementation improves the rumen epithelial barrier function by regulating Nrf2-NFκB signaling pathways during high-concentrate-diet feeding.

Metagenomic Insight: Dietary Thiamine Supplementation Promoted the Growth of Carbohydrate-Associated Microorganisms and Enzymes in the Rumen of Saanen Goats Fed High-Concentrate Diets.

Subacute ruminal acidosis (SARA) is often caused by feeding a high-concentrate diet in intensive ruminant production. Although previous studies have shown that dietary thiamine supplementation can effectively increase rumen pH and modify rumen fermentation, the effect of thiamine supplementation on rumen carbohydrate-related microorganisms and enzymes in goats under SARA conditions remain unclear. Therefore, the objective of the present study was to investigate the effects of dietary thiamine supplementation on carbohydrate-associated microorganisms and enzymes in the rumen of Saanen goats fed high-concentrate diets. Nine healthy mid-lactating Saanen goats in parity 1 or 2 were randomly assigned into three treatments: A control diet (CON; concentrate:forage (30:70)), a high-concentrate diet (HC; concentrate:forage (70:30)), and a high-concentrate diet with 200 mg of thiamine/kg of DMI (HCT; concentrate:forage (70:30)). Compared with the HC group, dietary thiamine supplementation improved ruminal microbes associated with fiber, including Prevotella, Fibrobacter, Neocallimastix, and Piromyces (p < 0.05). In addition, an increase in the relative abundance of enzymes involved in both fiber degradation and starch degradation, such as CBM16, GH3, and GH97, was observed in the HCT treatment. (p < 0.05). Thus, thiamine supplementation can improve carbohydrate metabolism by increasing the abundance of the microorganisms and enzymes involved in carbohydrate degradation. In conclusion, this study revealed the relationship between ruminal microbiota and enzymes, and these findings contributed to solving the problems arising from the high-concentrate feeding in ruminant production and to providing a new perspective on ruminant health.

Illumina Sequencing and Metabolomics Analysis Reveal Thiamine Modulation of Ruminal Microbiota and Metabolome Characteristics in Goats Fed a High-Concentrate Diet.

Long-term supplementation of a high-concentrate diet enhances the accumulation of lactate and decrease in pH in goat rumen, thereby disrupting the composition of microbial community. Studies have shown that incorporation of thiamine in high-concentrate diet increases ruminal pH and decreases rumen lactate concentration. To explore the effects of thiamine supplementation with a high-concentrate diet on alteration of the whole ruminal microbiota and their metabolites, 18 mid-lactating Saanen goats were randomly fed with one of three diets: (1) control diet (CON; n = 6; concentrate:forage 30:70), (2) high-concentrate diet (HG; n = 6; concentrate:forage 70:30), and (3) high-concentrate diet with 200 mg of thiamine/kg of DMI (HGT; n = 6; concentrate:forage 70:30). The goats received experimental diets for 8 weeks. Ruminal samples were collected on the last day of the 8 weeks for 16S rRNA gene sequencing and the liquid chromatograph-mass spectrometer (LC-MS) analysis. The results revealed significant alterations of the ruminal bacterial community structure and diversity in HGT groups compared to HG groups, with an overall dominance of Bacteroidetes at the phylum level and Oribacterium (P < 0.05), Anaerobiospirillum (P < 0.01), and Fibrobacter (P < 0.01) at genus level in the HGT group. The LC-MS analysis revealed that thiamine supplementation resulted in lower levels of propionate (P < 0.05), pyruvate (P < 0.01), lactate (P < 0.05), putrescine (P < 0.05), tyramine (P < 0.05), and histamine (P < 0.01) and higher levels of acetate (P < 0.05), succinates (P < 0.01), oxaloacetic acid (P < 0.01), leucine (P < 0.01), valine (P < 0.05), linoleic acid (P < 0.05), docosahexaenoic acid (P < 0.05), and 4-phenylbutyric acid (P < 0.05) in the HGT group than in the HG group. The decrease in these compounds enhanced homeostasis in the rumen environment and suppressed epithelial inflammation. Correlation analysis revealed the potential relationships between ruminal metabolites and microbial community. These findings demonstrate that thiamine supplementation can alleviate subacute ruminal acidosis (SARA) by stabilizing the microbial community and reducing toxic unnatural compounds.

l-Arginine Alleviates Hydrogen Peroxide-Induced Oxidative Damage in Ovine Intestinal Epithelial Cells by Regulating Apoptosis, Mitochondrial Function, and Autophagy.

BACKGROUND: Previous studies demonstrated that dietary l-arginine (Arg) alters the equilibrium between reactive oxygen species (ROS) generation and biological defenses to resist oxidant-induced toxicity. Whether supplying Arg can protect ovine intestinal epithelial cells (OIECs) from hydrogen peroxide (H2O2)-induced oxidative damage is unclear. OBJECTIVES: The current study aimed to examine the effect of Arg on mitophagy, mitochondrial dysfunction, and apoptosis induced by H2O2 in OIECs. METHODS: The OIECs were incubated in Arg-free DMEM supplemented with 100 µM Arg (CON) or 350 µM Arg (ARG) alone or with 150 µM H2O2 (CON + H2O2, ARG + H2O2) for 24 h. Cellular apoptosis, mitochondrial function, autophagy, and the related categories of genes and proteins were determined. All data were analyzed by ANOVA using the general linear model procedures of SAS (SAS Institute) for a 2 × 2 factorial design. RESULTS: Relative to the CON and ARG groups, H2O2 administration resulted in 44.9% and 26.5% lower (P < 0.05) cell viability but 34.7% and 61.8% greater (P < 0.05) ROS concentration in OIECs, respectively. Compared with the CON and CON + H2O2 groups, Arg supplementation led to 40.7% and 28.8% lower (P < 0.05) ROS concentration but 14.9%-49.0% and 29.3%-64.1% greater (P < 0.05) mitochondrial membrane potential, relative mitochondrial DNA content, and complex (I-IV) activity in OIECs, respectively. Compared with the CON and CON + H2O2 groups, Arg supplementation led to 33.9%-53.1% and 22.4%-49.1% lower (P < 0.05) mRNA abundance of proapoptotic genes, respectively. Relative to the CON and CON + H2O2 groups, Arg supplementation resulted in 33.0%-59.2% and 14.6%-37.7% lower (P < 0.05) abundance of proapoptotic, mitophagy, and cytoplasmic cytochrome c protein, respectively. CONCLUSIONS: Supply of Arg protects OIECs against H2O2-induced damage partly by improving mitochondrial function and alleviating cellular apoptosis and autophagy.

Melatonin ameliorates ochratoxin A induced liver inflammation, oxidative stress and mitophagy in mice involving in intestinal microbiota and restoring the intestinal barrier function.

The mycotoxin ochratoxin A (OTA) is a widespread contaminant in human and animal food products. Previous studies in rats revealed that melatonin (Mel) exhibits a preventive effect against OTA-induced oxidative stress in liver. However, it remains unknown whether gut microbiota respond to Mel and, if so, whether it can prevent OTA-induced inflammation and mitophagy in the liver. In the present study, mice received an oral gavage of Mel and OTA for 3 weeks before harvesting colonic digesta and liver tissue for analyses. In another study, the role of intestinal microbiota on the effects of Mel on OTA-induced liver inflammation and mitophagy was assessed through clearance of intestinal microbiota with antibiotics followed by gut microbiota transplantation (GMT). Oral Mel supplementation ameliorated mitophagy in the liver and reversed gut microbiota dysbiosis. Intriguingly, in antibiotic-treated mice, Mel and OTA failed to induce mitophagy in the liver. Using the GMT approach in which mice were colonised with intestinal microbiota from control-, OTA-, or Mel + OTA-treated mice led us to elucidated the involvement of intestinal microbiota in liver inflammation and mitophagy induced by OTA. The findings suggested that intestinal microbiota play some role in the Mel-induced amelioration of liver inflammation and mitophagy induced by OTA.

Dietary supplementation of l-arginine and N-carbamylglutamate enhances duodenal barrier and mitochondrial functions and suppresses duodenal inflammation and mitophagy in suckling lambs suffering from intrauterine-growth-restriction.

The current work aimed at investigating the effects of the dietary supplementation of N-carbamylglutamate (NCG) or l-arginine (Arg) on the duodenal mitophagy, mitochondrial function, inflammation, and barrier function in suckling lambs suffering from intrauterine-growth-retardation (IUGR). Forty-eight neonate Hu lambs were used in this study: 12 lambs with normal birth weight (NBW: 4.25 ± 0.14 kg) and 36 lambs with IUGR (3.01 ± 0.13 kg). Seven day old lambs were assigned to 4 treatment groups (12 lambs in each group) as follows: control group (CON), IUGR group, IUGR + Arg, and IUGR + NCG. Lambs were fed the experimental diets for 21 days from 7 days to 28 days of age. Compared with IUGR lambs, the Arg or NCG-treated IUGR lambs had a markedly higher duodenal transepithelial electrical resistance (TER) and lower fluorescein isothiocyanate dextran (FD4) (P < 0.05), respectively. The duodenal mitochondrial membrane potential change (ΔΨm), relative mitochondrial DNA (mtDNA) content, adenosine triphosphate (ATP) level, together with the activities of the respiratory complexes I, III, and IV were markedly higher in Arg or NCG-treated IUGR lambs than those in non-supplemented IUGR lambs (P < 0.05). The expressions of the integrity-related proteins (occludin and zonula occludens-1 (ZO-1)), antioxidant- and apoptosis-related proteins (B-cell lymphoma/leukaemia 2 (Bcl2), superoxide dismutase 2 (SOD2), catalase (CAT), and glutathione peroxidase 1 (GPx1)), and the nitric oxide-dependent pathway-related proteins (epithelial NO synthase (eNOS) and inducible NO synthase (iNOS)) were higher in NCG or Arg-supplemented IUGR lambs than those in nontreated IUGR lambs (P < 0.05). The duodenal expressions of the mitophagy-related proteins (microtubule-associated protein light chain 3 (LC3) I, LC3 II, Belin1, PTEN induced putative kinase 1 (PINK1), and Parkin) and the immune function-related proteins (myeloid differentiation factor 88 (MyD88), IL-6, nuclear factor kappa B (p65), toll-like receptor (TLR4) and TNF-α) were reduced (P < 0.05) in NCG or Arg-supplemented IUGR lambs compared with non-supplemented IUGR lambs. These results demonstrated that the dietary supplementation of Arg or NCG enhanced the duodenal barrier function and mitochondrial function, mitigated duodenal inflammation, and suppressed mitophagy in suckling lambs suffering from IUGR.