Effects of concentrate feeding sequence on VFA production, and cecal microbiota of Dezhou donkeys by metagenomic technology.

Microorganisms residing in the cecum of donkeys are crucial for physiological processes, nutrient metabolism, and immune function. Feeding methods can affect the dynamic balance of animal gut microbiota, thereby affecting indicators such as volatile fatty acids. This study explores suitable feeding methods to promote actual production by changing the feeding order of concentrate. Fifteen Dezhou donkeys with similar age and weight profiles were randomly divided into three groups with the concentrate feeding sequence: fiber-to-concentrate (FC), concentrate-to-fiber (CF), and total mixed ration (TMR). The experiment spanned a duration of 82 days. The analyses conducted were primarily aimed at determining the effects of feeding on gut microbes, primarily using metagenomic sequencing techniques. The experimental findings revealed that the levels of valeric acid were notably higher in the CF and TMR groups compared to the FC group (p < 0.05). These results suggest that the feeding sequence exerts a certain impact on the microbial composition within the cecum of Dezhou donkeys. At the phylum level, the predominant microbiota consisted of Firmicutes and Bacteroidetes, with the CF group displaying a higher relative abundance of Firmicutes compared to both the FC and TMR groups. At the genus level, Prevotella, Bacteroides, and Fibrobacter were the dominant bacterial genera identified in cecum. The functional gene annotation analysis indicated a significantly lower abundance of lacZ (K01190), Por/nifJ (K03737), and ppdK (K01006) genes in CF group relative to the FC and TMR groups (p < 0.05), highlighting their roles in galactose metabolism and glycolysis, respectively. Moreover, the CF group exhibited a higher concentration of antibiotic resistance genes (tetO and tet44) in the gut microbiota compared to the TMR and FC groups (p < 0.05), underscoring the presence of numerous antibiotic resistance genes within the phyla Bacteroidetes, Firmicutes, and Proteobacteria. In conclusion, different precision feed sequences significantly impact the levels of volatile fatty acids in Dezhou fattening donkeys, modify the composition and functional genes of the cecal microbiota, and elucidate the microbial mechanisms influenced by the feeding sequence on the growth and metabolism. These insights are anticipated to provide a foundation for the rational design of precision feed sequences in practical agricultural settings.

Effects of Concentrate Feeding Sequence on Growth Performance, Nutrient Digestibility, VFA Production, and Fecal Microbiota of Weaned Donkeys.

In this study, effects on the growth performance, nutrient digestibility, volatile fatty acids (VFA) production, and fecal microbiota of weaned donkeys were observed using different concentrate feeding sequences. Fifteen healthy 6-month-old weaned male donkeys with a body weight of 117.13 ± 10.60 kg were randomly divided into three treatment groups, including group C1 (roughage-then-concentrate), group C2 (concentrate-then-roughage), and group C3 (total mixed ration, TMR). The experiment lasted 35 d. We measured nutrient digestion by the acid-insoluble ash method and analyzed the fecal microbiota of the weaned donkeys by high-throughput sequencing of 16s rRNA genes in the V3-V4 region. The results show that group C3 obtained the best growth performance, and the digestibility of crude protein (CP) and crude extract (EE) was significantly higher than that of group C1 (p < 0.05). Acetic acid, isobutyric acid, valeric acid, isovaleric acid, and caproic acid were notably different among all groups (p < 0.05). In addition, we observed that Firmicutes and Bacteroidetes were dominant in the fecal microbes of each group, and Firmicutes was significantly higher in group C3 (p < 0.05). At the genus level, the different genera were Treponema, Rikenellaceae-RC9-gut-group, Unidentified-F082, and Bacteroidales-RF16-group (p < 0.05). The prediction of fecal microbiota function by PICRUSt indicated that different feeding sequences had minimal impact on the function of the fecal microbiota, particularly on the high-abundance pathway. In summary, the concentrate feeding sequence changed the composition of the fecal microbe of weaned donkeys.

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.

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.

Ambient pH regulates lactate catabolism pathway of the ruminal Megasphaera elsdenii BE2-2083 and Selenomonas ruminantium HD4.

AIMS: To explore the impact of ambient pH on lactate catabolism by Megasphaera elsdenii BE2-2083 and Selenomonas ruminantium HD4 in both pure culture and in binary mixed culture. METHODS AND RESULTS: The growth rate, substrate consumption, product formation, enzymatic activity and gene expression of M. elsdenii and S. ruminantium at various pHs were examined. Furthermore, the metabolism of lactate catabolism pathways for M. elsdenii and S. ruminantium in the co-culture system was investigated by chasing the conversion of sodium L-[3-13 C]-lactate in nuclear magnetic resonance. In the pure culture systems, ambient pH had significant effects on the growth of M. elsdenii, whereas S. ruminantium was less sensitive to pH changes. In addition, lactate metabolic genes and activities of key enzymes were affected by ambient pH in M. elsdenii and S. ruminantium. In the co-culture system, low ambient pH reduced the contribution lactate catabolism by M. elsdenii. CONCLUSION: M. elsdenii BE2-2083 and S. ruminantium HD4 lactate degradation affected by ambient pH. SIGNIFICANCE AND IMPACT OF THE STUDY: This study demonstrates the regulatory mechanisms of lactate decomposing bacteria in lactate catabolism under the condition of subacute ruminal acidosis.

Transcriptome Analysis Reveals Catabolite Control Protein A Regulatory Mechanisms Underlying Glucose-Excess or -Limited Conditions in a Ruminal Bacterium, Streptococcus bovis.

Ruminants may suffer from rumen acidosis when fed with high-concentrate diets due to the higher proliferation and overproduction of lactate by Streptococcus bovis. The catabolite control protein A (CcpA) regulates the transcription of lactate dehydrogenase (ldh) and pyruvate formate-lyase (pfl) in S. bovis, but its role in response to different carbon concentrations remains unclear. To characterize the regulatory mechanisms of CcpA in S. bovis S1 at different levels of carbon, herein, we analyzed the transcriptomic and physiological characteristics of S. bovis S1 and its ccpA mutant strain grown in glucose-excess and glucose-limited conditions. A reduced growth rate and a shift in fermentation pattern from homofermentation to heterofermentation were observed under glucose-limited condition as compared to glucose-excess condition, in S. bovis S1. Additionally, the inactivation of ccpA significantly affected the growth and end metabolites in both conditions. For the glycolytic intermediate, fructose 1,6-bisphosphate (FBP), the concentration significantly reduced at lower glucose conditions; its concentration decreased significantly in the ccpA mutant strain. Transcriptomic results showed that about 46% of the total genes were differentially transcribed between the wild-type strain and ccpA mutant strain grown in glucose-excess conditions; while only 12% genes were differentially transcribed in glucose-limited conditions. Different glucose concentrations led to the differential expression of 38% genes in the wild-type strain, while only half of these were differentially expressed in the ccpA-knockout strain. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses showed that the substrate glucose concentration significantly affected the gene expression in histidine metabolism, nitrogen metabolism, and some carbohydrate metabolism pathways. The deletion of ccpA affected several genes involved in carbohydrate metabolism, such as glycolysis, pyruvate metabolism, fructose and mannose metabolism, as well as in fatty acid biosynthesis pathways in bacteria grown in glucose-excess conditions; this effect was attenuated under glucose-limited conditions. Overall, these findings provide new information on gene transcription and metabolic mechanisms associated with substrate glucose concentration and validate the important role of CcpA in the regulation of carbon metabolism in S. bovis S1 at differential glucose availability.

Regulation of CcpA on the growth and organic acid production characteristics of ruminal Streptococcus bovis at different pH.

BACKGROUND: Catabolite control protein A (CcpA) regulates the transcription of lactate dehydrogenase and pyruvate formate-lyase in Streptococcus bovis, but knowledge of its role in response to different pH is still limited. In this study, a ccpA-knockout strain of S. bovis S1 was constructed and then used to examine the effects of ccpA gene deletion on the growth and fermentation characteristics of S. bovis S1 at pH 5.5 or 6.5. RESULTS: There was a significant interaction between strain and pH for the maximum specific growth rate (µmax) and growth lag period (λ), which caused a lowest µmax and a longest λ in ccpA-knockout strain at pH 5.5. Deletion of ccpA decreased the concentration and molar percentage of lactic acid, while increased those of formic acid. Strains at pH 5.5 had decreased concentrations of lactic acid and formic acid compared to pH 6.5. The significant interaction between strain and pH caused the highest production of total organic acids and acetic acid in ccpA-knockout strain at pH 6.5. The activities of α-amylase and lactate dehydrogenase decreased in ccpA-knockout strain compared to the wild-type strain, and increased at pH 5.5 compared to pH 6.5. There was a significant interaction between strain and pH for the activity of acetate kinase, which was the highest in the ccpA-knockout strain at pH 6.5. The expression of pyruvate formate-lyase and acetate kinase was higher in the ccpA-knockout strain compared to wild-type strain. The lower pH improved the relative expression of pyruvate formate-lyase, while had no effect on the relative expression of acetate kinase. The strain × pH interaction was significant for the relative expression of lactate dehydrogenase and α-amylase, both of which were highest in the wild-type strain at pH 5.5 and lowest in the ccpA-knockout strain at pH 6.5. CONCLUSIONS: Overall, low pH inhibited the growth of S. bovis S1, but did not affect the fermentation pattern. CcpA regulated S. bovis S1 growth and organic acid fermentation pattern. Moreover, there seemed to be an interaction effect between pH and ccpA deletion on regulating the growth and organic acids production of S. bovis S1.

Ursolic acid ameliorates adipose tissue insulin resistance in aged rats via activating the Akt-glucose transporter 4 signaling pathway and inhibiting inflammation.

Ageing often results in insulin resistance (IR) and chronic inflammation, and adipose is one of the tissues in which inflammation and IR occur earliest during this process. The present study investigated the effect and underlying mechanisms of ursolic acid (UA) on adipose IR and inflammation in ageing rats. Specific pathogen-free male Sprague-Dawley rats were randomly divided into 4 groups: i) Young normal (young); ii) untreated ageing (aged); and groups supplemented with UA either iii) low-UA 10 mg/kg (UA-L) or iv) high-50 mg/kg (UA-H). Animals in the UA-treated groups received 10 or 50 mg/kg UA (suspended in 5% Gum Arabic solution). The rats in the corresponding aged group and young groups received vehicle (5% Gum Arabic) alone. All rats were intragastrically treated once daily by oral gavage for 7 weeks. The day before the experiment terminated, overnight fasting blood (~700 µl) was collected and plasma was prepared to measure biochemical indicators; western blotting was performed to analyze the expression of insulin signaling proteins [(insulin receptor substrate 1 (IRS-1), phosphorylated (p)-IRS-1, PI3K, glucose transporter 4 (GLUT4), Akt and p-Akt)] and inflammatory factors (NF-κB, IL-6 and IL-1ß) in the epididymis white adipose tissue (eWAT). The results revealed that treatment with UA-H decreased eWAT weight, the ratio of eWAT weight/body weight, fasted insulin and triglyceride levels, the homeostasis model assessment of insulin resistance and adipose tissue insulin resistance index in ageing rats, indicating the amelioration of systemic and adipose tissue IR, compared with the aged group. Mechanistically, UA-H administration upregulated p-protein kinase B, the ratio of p-Akt to protein kinase B and total and cellular membrane GLUT4 protein levels in eWAT of ageing rats. Conversely, UA inhibited the increase in NF-κB expression and proinflammatory cytokines IL-6 and IL-1ß. However, these alterations were not observed in the rats of the aged group. Taken together, the findings of the present study indicated that UA may ameliorate adipose IR, which is associated with activation of the Akt-GLUT4 signaling pathway and inhibition of inflammation in ageing rats. These data provide a basis for the development of effective and safe drugs or functional substances, such as UA, for the prevention and treatment of metabolic diseases.

The Effect of Replacing Wildrye Hay with Mulberry Leaves on the Growth Performance, Blood Metabolites, and Carcass Characteristics of Sheep.

The objective of this study was to evaluate the effects of partially substituting for conventional forage, Chinese wildrye (CW), with mulberry leaves (ML) on the growth, digestion, ruminal fermentation, blood metabolites, and meat quality of sheep in a 65-day feedlot study. Thirty-two four-month-old male small-tailed Han sheep (25.15 ± 1.03 kg) were randomly assigned to one of four treatments. The dietary treatments consisted of four proportions of ML (0, 8, 24, and 32%) as a substitute for CW (designated as ML0, ML8, ML24, and ML32, respectively). Rumen digesta and blood samples were collected at day 63 of the trial. Carcass traits were assessed after slaughter at the end of performance period. The results from this study revealed no differences in average daily bodyweight gain (ADG), feed conversion ratio (FCR), and final body weight (FBW) among treatments. The apparent digestibility of dry matter (DM), organic matter (OM), and acid detergent fiber (ADF) was higher in the sheep fed with ML than in those fed CW. The ML24 treatment had a higher digestibility of crude protein (CP) and ether extract (EE). There were no differences (p = 0.13) in ruminal pH values among the treatments. However, there was more microbial protein (p < 0.01) in ML24 and ML32 treatments than the ML0 treatment. Ruminal concentrations of acetate and butyrate were significantly different among treatments, although no difference in concentrations of total volatile fatty acid were found. Additionally, no differences were detected for serum parameters except blood urea nitrogen (BUN). No differences were observed for carcass weight (p = 0.62), dressing percentage (p = 0.31) or longissimus dorsi muscle (LM) area (p = 0.94) among treatments. However, intramuscular fat was higher in the ML24 treatment than in the ML0 treatment. (p < 0.01). There were higher pH values of the 24-h longissimus dorsi in the ML24 treatment than in the ML0 treatment. In addition, the saturated fatty acid (SFA) content was lower (p < 0.01) and the monounsaturated fatty acid (MUFA) content higher (p < 0.01) in the ML24 treatment than in the ML0 treatment. In conclusion, the partially substitution of mulberry leaves for Chinese wildrye in the diet of sheep had a beneficial influence on the growth performance, blood metabolites and carcass characteristics. The inclusion of 24% (air dry basis) mulberry leaf hay in the ration of sheep is recommended based on these findings.

Ginsenoside Rg3 Inhibits the Growth of Osteosarcoma and Attenuates Metastasis through the Wnt/ß-Catenin and EMT Signaling Pathway.

Osteosarcoma (OS) is the most common primary malignant bone cancer. An increasing number of studies have demonstrated that ginsenoside Rg3 (Rg3), which is extracted from the roots of the traditional Chinese herb Panax ginseng, plays a tumor suppression role in various malignant tumors. In the present study, we aimed at investigating the role of Rg3 in the proliferation, migration, and invasion of OS and at exploring the underlying mechanisms. Cell viability and proliferation were observed by MTT assay and crystal violet staining. The migration and invasion of cells were measured by wound-healing assay and Transwell method. Signaling pathway screening was investigated using luciferase reporter gene assay. qRT-PCR and western blot were performed to measure the expression of molecules involved in cell epithelial-mesenchymal transition (EMT), and Wnt/ß-catenin pathway. Results suggested that Rg3 could not only inhibit proliferation but also hamper the migration and invasion of OS. qRT-PCR and western blot demonstrated that a reduced level of MMP2/MMP7/MMP9 was induced after Rg3 treatment. In addition, the expression levels of proteins related to EMT and the Wnt/ß-catenin pathway were downregulated. In summary, our data revealed that Rg3 could inhibit the proliferation, migration, and invasion of OS cells. This effect of Rg3 might be mediated by downregulating MMP2, MMP7, and MMP9 expression and suppressing EMT as well as the Wnt/ß-catenin pathway. Thus, Rg3 might be a potential agent for the treatment of OS.

Effects of diet supplementation with rumen-protected betaine on carcass characteristics and fat deposition in growing lambs.

This study evaluated the effects of dietary rumen-protected betaine (RPB) supplementation on the fat deposition of lambs. Sixty Hu sheep were randomly divided into 5 groups. The lambs were fed a control diet (CON) or diets supplemented with 1.1 g/d unprotected betaine (UPB), 1.1 g/d RPB, 2.2 g/d RPB or 3.3 g/d RPB for 70 days. Compared with UPB, the abdominal fat in 2.2 g/d RPB supplemented group was decreased (P < .05). Compared with CON and UPB, the fat contents in longissimus dorsi (LD) of RPB treatments were increased (P < .01). With increasing of RPB levels, the fat content in the LD was quadratically increased (P < .05). Compared with CON, genes expression of PI3K, mTOR and S6K1 in the LD of RPB treatments were up-regulated (P < .05). In conclusion, RPB supplementation decreased the abdominal fat in lambs but increased the fat content in lamb meat, and this effect might be regulated by mTOR signaling.

Rhodiola crenulata root extract ameliorates fructose-induced hepatic steatosis in rats: Association with activating autophagy.

BACKGROUND: Increasing evidence has shown the beneficial effects of Rhodiola species on metabolic disorders, but their mechanisms are not clear. Hepatic steatosis is closely related to metabolic disorders, we aim to investigate the therapeutic effects of Rhodiola crenulata root (RCR) on fructose-induced hepatic steatosis and explore the underlying mechanisms. PURPOSE: To observe the effect of Rhodiola crenulata root extract (RCR) on fructose-induced hepatic steatosis in Sprague-Dawley (SD) rats and explore its possible mechanism. METHODS: Male Sprague-Dawley rats were treated with liquid fructose in their drinking water over 18 weeks. The extract of RCR was co-administered (once daily by oral gavage) during the last 5 weeks. Liver lipid deposition and morphological changes were observed by Oil red O staining. Real-time fluorescence quantitative PCR, Western blot and immunoprecipitation were used to detect gene and protein expression in liver. RESULTS: RCR (50 mg/kg) reversed liquid fructose-induced increase in hepatic triglyceride content in rats. Attenuation of the increased vacuolization and Oil Red O staining area was evident on histological examination of liver in RCR-treated rats. However, RCR treatment did not affect chow intake and body weight of rats. Although some genes of the pathways involved in DNL (ChREBP, SREBP-1c, FAS, ACC1, SCD1, DGAT1, DGAT2 and MGAT2), fatty acid ß-oxidation (PPARα, CPT1a, ACO and FGF21), VLDL-export (MTTP) and decomposition (HSL, ATGL) in the liver of fructose-fed rats were not changed significantly after RCR administration, the decrease in PPARα and PGC-1α proteins was reversed by RCR. Notably, SIRT1 mRNA and protein expression increased significantly with RCR administration. Furthermore, RCR increased expression of ATG4B, Beclin1 and decreased expression of Bcl2-Beclin1 complex dramatically. Meanwhile, RCR decreased the acetylation of beclin1. Moreover, RCR increased expression of autophagosome markers including LC3B and ATG5-ATG12-ATG16L1, and decreased expression of autophagolysosome marker p62 in the livers of fructose-fed rats. CONCLUSIONS: RCR has a certain improvement effect on fructose-induced hepatic steatosis, which is related to the activation of autophagy.

ß-Sitosterol Attenuates High Grain Diet-Induced Inflammatory Stress and Modifies Rumen Fermentation and Microbiota in Sheep.

ß-sitosterol (BSS) is a plant-derived natural bioactive compound, its cellular mechanism of anti-inflammatory activity has been proven recently. Little information is available regarding the application of BSS on ruminants under high grain diet. The objective of this study was to evaluate the effects of dietary BSS supplementation on inflammatory response, ruminal fermentation characteristics and the composition of the ruminal bacterial community under high grain diet. Eight rumen-cannulated Hu sheep (59.7 ± 4.8 kg of initial body weight) were randomly assigned into a replicated 4 × 4 Latin square design trial. Sheep were fed a high grain diet (non-fiber carbohydrate: neutral detergent fiber = 2.03) supplemented either with 0.25 (LBS), 0.5 (MBS), 1.0 (HBS) or without (CON) g BSS /kg dry matter diet. On day 21 of each period, rumen content samples were obtained at 6 h postfeeding, and blood samples were obtained before morning feeding. The data showed that compared with control group, Dietary BSS supplementation decreased serum concentrations of tumor necrosis factor, interleukin (IL)-6, and IL-1ß. The ruminal pH and acetate concentration for BSS treatment were improved, while concentration of propionate, butyrate and lactate was decreased. The result of Illumina MiSeq sequencing of 16S rRNA gene revealed that BSS addition can increase the proportion of Prevotella_1, Rikenellaceae_RC9_gut_group, Prevotella_7, and Selenomonas_1, and decrease the proportion of Lachnospiraceae_NK3A20_group. These results indicated that BSS attenuates high grain diet-induced inflammatory response and modifies ruminal fermentation. In addition, the BSS dietary supplementation at the level of 0.5 g/kg is recommended in sheep.

The microRNA­708­5p/ZEB1/EMT axis mediates the metastatic potential of osteosarcoma.

MicroRNA­708­5p (miR­708­5p) and epithelial­â€‹to­mesenchymal transition (EMT) have been widely identified to contribute to the pathogenesis and progression of multiple cancers. However, the connection between miR­708­5p and EMT has not been sufficiently clarified. Therefore, our research aimed to investigate the impact of miR­708­5p on EMT and the metastasis of osteosarcoma (OS). We first analyzed the differentially expressed microRNAs (DEmiRNAs) from the GSE70367 dataset. We found that the expression of miR­708­5p was lower in OS cells. Overexpression of miR­708­5p was able to impair the migration and invasion of OS cells. Moreover, miR­708­5p inhibited EMT of OS cells MG63 and SaOS­2, wherein E­cadherin was increased, and N­cadherin, vimentin, and Snail were decreased. Semaphorin 4C (SEMA4C), mitogen­activated protein kinase kinase kinase 3 (MAP3K3), and zinc finger E­box­binding homeobox 1 (ZEB1) were predicted as target genes of miR­708­5p by bioinformatics method. Only ZEB1, one of the EMT­inducing transcription factors, was validated as the direct target gene of miR­708­5p in OS cells through dual­luciferase reporter assay and Western blot analysis. Knockdown of ZEB1 was found to inhibit the metastasis of MG63 and SaOS­2 cells, whereas ZEB1 over-expression promoted their metastasis. In summary, miR­708­5p impaired the metastasis and EMT of OS, which was found to be mediated by inhibition of ZEB1.

N-Carbamylglutamate and l-arginine supplementation improve hepatic antioxidant status in intrauterine growth-retarded suckling lambs.

The influence of dietary supplementation of l-arginine (Arg) or N-carbamylglutamate (NCG) on the hepatic antioxidant status in intrauterine-growth-retarded (IUGR) suckling lambs remains unclear. The current work aimed to investigate the regulatory mechanisms whereby dietary Arg or NCG alter hepatic antioxidant status in suckling lambs suffering from IUGR. Forty-eight newborn Hu lambs of normal birth weight (CON) and IUGR were allocated randomly into four groups of 12 animals each: CON (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). All lambs were raised for a period of 21 days from 7 to 28 days after birth. Compared with the IUGR suckling animals, glutathione peroxidase (GSH-Px), superoxide dismutase (SOD), and reduced glutathione (GSH) content were greater (P < 0.05), and protein carbonyl and malondialdehyde (MDA) levels were reduced (P < 0.05) in the livers of both IUGR + 1% Arg and 0.1% NCG suckling animals. Relative to IUGR suckling lambs, supplementing with Arg or NCG markedly reduced (P < 0.05) reactive oxygen species (ROS) levels, apoptosis, and necrosis in liver. Relative to IUGR suckling lambs, protein and mRNA expression of GSH-Px1, SOD2, catalase (CAT), heme oxygenase-1 (HO-1), inducible nitric oxide (NO) synthase (iNOS), and epithelial NO synthase (eNOS) increased in IUGR animals receiving Arg or NCG (P < 0.05). Both Arg and NCG can protect neonates from IUGR-induced hepatic oxidative damage through promoting the expression of antioxidative enzymes (including SOD, CAT, and GSH-Px), phase II metabolizing enzymes, and activation of the NO pathway.

Biological analysis of cancer specific microRNAs on function modeling in osteosarcoma.

Osteosarcoma (OS) is the most common bone tumor characterized with a high risk of amputation and malignant morbidity among teenagers and adolescents. However, relevant pathogenic/biological mechanisms underlying OS-genesis remains to be ambiguous. The aim of this study was to elucidate functional relationship about microRNAs-mRNAs networks and to identify potential molecular markers via a computational method. Gene expression profile (GSE70415) was recruited from Gene Expression Omnibus. 3856 differentially expressed genes and 250 significantly expressed microRNAs were identified by using GCBI. The results of GO and KEGG pathways associated proteomics analysis indicated that extracellular matrix organization, small molecule metabolic process, cell adhesion (GO IDs: 0030198, 0044281, 0007155) and pathways in cancer, PI3K-Akt signaling pathway, metabolic pathways (pathway IDs: 5200, 4151, 1100) were significantly enriched. In addition, CKMT2, miR-93b-5p, miR-29b-3p were found to be positively/negatively correlated with TP53, EGFR, and MMP members mediated OS development, including angiogenesis, migration and invasion. Further visualization of collective effect of 1181 microRNAs-mRNAs pairs and protein-protein interactions was realized by applying with cytosacpe. In summary, our work provided a better understanding of non-coding regulatory mechanisms of transcriptomics and unraveled essential molecular biomarkers in osteosarcoma.

The effect of dietary grape pomace supplementation on epididymal sperm quality and testicular antioxidant ability in ram lambs.

Wine grape pomace (WGP) contains a rich source of polyphenols that can act as powerful antioxidants. The objective of this study was to investigate the effect of dietary WGP supplementation on antioxidative activity and epididymal sperm quality in rams. The rams were raised either under free-range or pen conditions, and the pen-raised rams were fed a WGP-containing diet (0, 5% and 10% of dry matter basis) for 74 days. An increase in the concentrations of reactive oxygen species (ROS, P < 0.05) and malondialdehyde (MDA, P < 0.05) were observed in the testes of rams subjected to restraint stress, and dietary WGP supplementation effectively decreased their contents (P < 0.05). Restraint stress reduced both weight and volume of testes, and impaired sperm quality. Dietary WGP supplementation increased testes weight, sperm concentration, motility and acrosomal integrity, and decreased sperm deformity in pen-raised animals (P < 0.05). The total antioxidative capacity (T-AOC) and catalase activity were decreased in the testes of pen-raised lambs (P < 0.05), and T-AOC, catalase, glutathione peroxidase 4 (GPx4) and superoxide dismutase (SOD) activity were increased when rams were fed the WGP-containing diet (P < 0.05). With the exception of SOD and GPx4, the mRNA contents of catalase and nuclear factor-like-2 factor (Nrf2) did not vary among the groups, and greater protein levels of SOD, catalase and GPx4 were observed in WGP-treated lambs (P < 0.05). Taken together, these results suggest that WGP can be used as a feed ingredient in rams to alleviate restraint induced oxidative stress and improve epididymal sperm quality.