Short communication: Relationship of blood DNA methylation rate and milk performance in dairy cows.

The objective of this study was to investigate the global methylation rate in blood DNA and its relationship with lactation performance. A total of 196 mid-lactation dairy cows were fed the same diet under the same management. Milk yield was recorded and blood samples were collected from the jugular vein before morning feeding. The blood global DNA methylation rates were quantified using a methylation quantification kit. Overall, the average blood global DNA methylation rate of all cows was 12.4%. When DNA methylation rates were compared between cows with high (n = 40; 37.0 to 42.0 kg/d) and low (n = 33; 24.0 to 30.0 kg/d) milk yield, DNA methylation rates in the lower-yield cows (14.1 ± 0.7%) were significantly higher than those in the higher-yield animals (11.6 ± 0.7%). Our results indicated an association of milk and protein yields with global DNA methylation rates in lactating dairy cows. However, further research is needed to determine whether this association reflects the true influence of epigenetic mechanisms on yield or whether other factors, such as different proportions of blood cell types in high- and low-yielding cows, affect apparent global DNA methylation levels.

Assessment of rumen bacteria in dairy cows with varied milk protein yield.

The present study was conducted to assess rumen bacteria in lactating cows with different milk protein yield, aiming to understand the role of rumen bacteria in this trait. Cows with high milk protein yield (high milk yield and high milk protein content, HH; n = 20) and low milk protein yield (low milk yield and low milk protein content, LL; n = 20) were selected from 374 mid-lactation Holstein dairy cows fed a high-grain diet. Measurement of the rumen fermentation products showed that the concentrations of ruminal total volatile fatty acids, propionate, butyrate, and valerate and the proportion of isobutyrate were higher in the HH cows than in the LL cows. Amplicon sequencing analysis of the rumen bacterial community revealed that the richness (Chao 1 index) of rumen microbiota was higher in the LL cows than in the HH cows. Among the 10 predominant bacterial phyla (relative abundance being >0.10%, present in >60% of animals within each group), the relative abundance of Proteobacteria was 1.36-fold higher in the HH cows than in the LL cows. At the genus level, the relative abundance of Succinivibrio was significantly higher and that of Clostridium tended to be higher in the LL cows than in the HH cows. Sharpea was 2.28-fold enriched in the HH cows compared with the LL cows. Different relationships between the relative abundances of rumen microbial taxa and volatile fatty acid concentrations were observed in the HH and the LL animals, respectively. Succinivibrio and Prevotella were positively correlated with acetate, propionate, and valerate in the LL cows, whereas Sharpea was positively correlated with propionate and valerate concentrations in the HH cows. Collectively, our results revealed that rumen bacterial richness and the relative abundances of several bacterial taxa significantly differed between dairy cows with high and low milk protein yields, suggesting the potential roles of rumen microbiota contributing to milk protein yield in dairy cows.

Invited review: Advances and challenges in application of feedomics to improve dairy cow production and health.

Dairy cattle science has evolved greatly over the past century, contributing significantly to the improvement in milk production achieved today. However, a new approach is needed to meet the increasing demand for milk production and address the increased concerns about animal health and welfare. It is now easy to collect and access large and complex data sets consisting of molecular, physiological, and metabolic data as well as animal-level data (such as behavior). This provides new opportunities to better understand the mechanisms regulating cow performance. The recently proposed concept of feedomics could help achieve this goal by increasing our understanding of interactions between the different components or levels and their impact on animal production. Feedomics is an emerging field that integrates a range of omics technologies (e.g., genomics, epigenomics, transcriptomics, proteomics, metabolomics, metagenomics, and metatranscriptomics) to provide these insights. In this way, we can identify the best strategies to improve overall animal productivity, product quality, welfare, and health. This approach can help research communities elucidate the complex interactions among nutrition, environment, management, animal genetics, metabolism, physiology, and the symbiotic microbiota. In this review, we summarize the outcomes of the most recent research on omics in dairy cows and highlight how an integrated feedomics approach could be applied in the future to improve dairy cow production and health. Specifically, we focus on 2 topics: (1) improving milk yield and milk quality, and (2) understanding metabolic physiology in transition dairy cows, which are 2 important challenges faced by the dairy industry worldwide.

Hydrogen sulfide modulates gastric acid secretion in rats via involvement of substance P and nuclear factor-κB signaling.

Hydrogen sulfide (H2S) promotes gastric acid secretion in rats. The present study aimed to test the hypothesis that H2S regulates this response via activating TRPV1 channel and through activation of the nuclear factor-κB (NF-κB) pathway. Male Wistar rats were randomly divided into the sodium hydrosulfide (NaHS, 100 µmol/kg b.w.) group, pyrrolidine dithiocarbamate (PDTC, 100 µmol/kg b.w.) group, PDTC (100 µmol/kg b.w.) + NaHS (100 µmol /kg b.w.) group, capsazepine (0.1 mM) + NaHS (100 µmol /kg b.w.) group and L703606 (0.1 mM) + NaHS (100 µmol /kg b.w.) group. The acidity of gastric juice before injection and after injection were determined by a pH meter. The results showed that sodium hydrosulfide (NaHS), an exogenous H2S donor, significantly reduced the pH of gastric juice when injected into the enterocoelia. Further, the promotional effect of NaHS on gastric acid secretion could be attenuated by capsazepine, a transient receptor potential vanilloid 1 (TRPV1) antagonist; L703606, a neurokinin 1 (NK1) receptor antagonist; and PDTC, a NF-κB inhibitor. The data from these experiments suggest that NaHS exerts an excitatory effect on gastric acid secretion possibly mediated by TRPV1 channel activation in sensory nerve terminals with the consequent release of substance P and in a NF-κB -dependent manner.

Evaluation of Aphis glycines as an Alternative Host for Supporting Aphelinus albipodus Against Myzus persicae on Capsicum annuum cv. Ox Horn and Hejiao 13.

Bank plant systems provide effective biological control for pests infesting commercially important crops. Aphids cause physical damage to crops by feeding on the leaves, as well as transmitting damaging viral diseases. To develop a bank plant system to control aphids that damage vegetable crops, we initially reared the parasitoid Aphelinus albipodus (Hayat and Fatima) on the soybean aphid, Aphis glycines (Matsumura) reared on the soybean plant, Glycine max (L.) that was elected as the alternate host. Parasitoid adults that emerged from A. glycines were allowed to parasitize second instar nymphs of the aphid Myzus persicae (Sulzer) which were reared on sweet pepper and chili pepper leaves. The results showed that A. albipodus females feeding and parasitizing M. persicae nymphs reared on sweet pepper lived for 18.9 days, with an average fecundity of 337.3 progenies/female, while females feeding and parasitizing on M. persicae nymphs reared on chili pepper lived for 18.8 days, with an average fecundity of 356.2 progenies/female. There were no significant difference in the development time and reproduction of A. albipodus individuals parasitizing M. persicae nymphs reared on sweet pepper and chili pepper plants. The intrinsic rate of increase (r), net reproductive rate (R 0), net aphid killing rate (Z 0), and finite aphid killing rate (θ) of A. albipodus parasitizing sweet pepper and chili pepper M. persicae was 0.2258 days-1, 171.7 progeny adults, 222.6 aphids, and 0.4048 and 0.2295 days-1, 191.8 progeny adults, 243.3 aphids, and 0.4021, respectively. Our results suggested that A. glycines could serve as an effective alternative host for supporting A. albipodus against M. persicae infesting sweet pepper and chili pepper.

The Effects of Dietary Phosphorus on the Growth Performance and Phosphorus Excretion of Dairy Heifers.

The objective of this study was to investigate the effects of reducing dietary phosphorus (P) on the frame size, udder traits, blood parameters and nutrient digestibility coefficient in 8- to 10-month-old Holstein heifers. Forty-five heifers were divided into 15 blocks according to the mo of age and were randomly assigned one of three dietary treatments: 0.26% (low P [LP]), 0.36% (medium P [MP]), or 0.42% (high P [HP]) (dry matter basis). Samples were collected at the wk 1, 4, 8. The results show that low dietary P had no effect on body measurement. The blood P concentration decreased with decreasing dietary P (p<0.05), while the blood calcium content of LP was higher than that of the MP and HP groups (p<0.05), though still in the normal range. The serum contents of alkalinephosphatase, potassium, and magnesium were similar among the treatments. No differences were found in all nutrients' apparent digestibility coefficients with varied dietary P. However, with P diet decreased from HP to LP, the total fecal P and urine P concentration declined significantly, as did fecal water soluble P (p<0.05). In conclusion, reducing the dietary P from 0.42% to 0.26% did not negatively affect the heifers' growth performance but did significantly lessen manure P excretion into the environment.

Role of hydrogen sulfide within the dorsal motor nucleus of the vagus in the control of gastric function in rats.

BACKGROUND: Hydrogen sulfide (H2 S) is a gaseous messenger and serves as an important neuromodulator in the central nervous system. This study aimed to clarify the role of H2 S within the dorsal motor nucleus of the vagus (DMV) in the control of gastric function in rats. METHODS: Cystathionine ß-synthetase (CBS) is an important generator of endogenous H2 S in the brain. We investigated the distribution of CBS in the DMV using immunohistochemical method, and the effects of H2 S on gastric motility and on gastric acid secretion. KEY RESULTS: CBS-immunoreactive (IR) neurons were detected in the rostral, intermediate and caudal DMV, with the highest number of CBS-IR neurons in the caudal DMV, and the lowest in the intermediate DMV. We also found that microinjection of the exogenous H2 S donor NaHS (0.04 and 0.08 mol/L; 0.1 µL; n = 6; p < 0.05) into the DMV significantly inhibited gastric motility with a dose-dependent trend, and promoted gastric acid secretion in Wistar rats. Microinjection of the same volume of physiological saline (PS; 0.1 µL, n = 6, p > 0.05) at the same location did not noticeably change gastric motility and acid secretion. CONCLUSIONS & INFERENCES: The data from these experiments suggest that the CBS that produces H2 S is present in the DMV, and microinjection of NaHS into the DMV inhibited gastric motility and enhanced gastric acid secretion in rats.

Effects of corn and soybean meal types on rumen fermentation, nitrogen metabolism and productivity in dairy cows.

Twelve multiparous Holstein dairy cows in mid-lactation were selected for a replicated 4×4 Latin square design with a 2 ×2 factorial arrangement to investigate the effects of corn and soybean meal (SBM) types on rumen fermentation, N metabolism and lactation performance in dairy cows. Two types of corn (dry ground [DGC] and steam-flaked corn [SFC]) and two types of SBM (solvent-extracted and heat-treated SBM) with different ruminal degradation rates and extents were used to formulate four diets with the same basal ingredients. Each period lasted for 21 days, including 14 d for adaptation and 7 d for sample collection. Cows receiving SFC had a lower dry matter (DM) and total N intake than those fed DGC. However, the milk yield and milk protein yield were not influenced by the corn type, resulting in higher feed and N utilization efficiency in SFC-fed cows than those receiving DGC. Ruminal acetate concentrations was greater and total volatile fatty acids concentrations tended to be greater for cows receiving DGC relative to cows fed SFC, but milk fat content was not influenced by corn type. The SFC-fed cows had lower ruminal ammonia-N, less urea N in their blood and milk, and lower fecal N excretion than those on DGC. Compared with solvent-extracted SBM-fed cows, cows receiving heat-treated SBM had lower microbial protein yield in the rumen, but similar total tract apparent nutrient digestibility, N metabolism measurements, and productivity. Excessive supply of metabolizable protein in all diets may have caused the lack of difference in lactation performance between SBM types. Results of the present study indicated that increasing the energy degradability in the rumen could improve feed efficiency, and reduce environmental pollution.

Isolation and identification of phase I metabolites of resibufogenin in rats.

1. Resibufogenin (1), a major bufadienolide of Chinese medicine Chan Su, had a wide range of pharmacological activities. In present work, the metabolism of 1 in male Sprague-Dawley rats was investigated by identifying the metabolites of resibufogenin excreted in rat bile. 2. Following an oral dose of 60 mg/kg resibufagenin, nine metabolites were isolated from bile of rats, and their structures were identified as 3-keto- resibufogenin (2), 3-epi-resibufogenin (3), 5ß-hydroxy-3-epi-resibufogenin (4), 1α, 5ß-dihydroxy-3-epi-resibufogenin (5), 3α, 5ß, 14α, 15ß-tetrahydroxyl-bufa- 20, 22-dienolide (6), 3α, 14α, 15ß-trihydroxy-bufa-20, 22-dienolide (7), 3-epi- 5ß-hydroxy-bufalin (8), 12α, 16ß-dihydroxy-3-epi-resibufogenin (9), and 5ß, 16ß-dihydroxy-3-epi-resibufogenin (10), respectively, on the basis of widely spectroscopic methods including 2D-NMR technology. It is first time to describe the metabolites of 1 in vivo, and metabolites 5-7 and 9-10 are novel. 3. On the basis of these identified metabolites, a possible metabolism pathway for 1 in rats has been proposed. This is the first systematic study on the phase I metabolites of resibufogenin.

Role of nitric oxide in the control of the gastric motility within the nucleus ambiguus of rats.

This study aims to investigate whether exogenous nitric oxide (NO) plays a role in controlling gastric motility within the nucleus ambiguus (NA). Experiments were performed on male Wistar rats anaesthetized with chloral hydrate. A latex balloon, connected to a pressure transducer, was inserted into the pylorus through the fundus for continuous recording of the change of gastric smooth muscle contractile curves. Microinjection of the NO-donor sodium nitroprusside (SNP; 5 nmol) or L-arginine (L-Arg; 5 nmol) into the NA significantly inhibited gastric motility, whereas the treatment of NO-synthase inhibitor N-nitro-L-arginine methylester (L-NAME) increased gastric motility remarkably. The negative effect of SNP or L-Arg on gastric motility was abolished by bilateral subdiaphragmatic vagotomy as well as by intravenous injection of ganglionic blocker, hexamethonium bromide (Hb). These results demonstrated that NO inhibited gastric motility by activating the cholinergic preganglionic neurons in the NA and through the mediation of vagus nerves.

Online coupling of reverse-phase and hydrophilic interaction liquid chromatography for protein and glycoprotein characterization.

We have developed a novel system for coupling reverse-phase (RP) and hydrophilic interaction liquid chromatography (HILIC) online in a micro-flow scheme. In this approach, the inherent solvent incompatibility between RP and HILIC is overcome through the use of constant-pressure online solvent mixing, which allows our system to perform efficient separations of both hydrophilic and hydrophobic compounds for mass spectrometry-based proteomics applications. When analyzing the tryptic digests of bovine serum albumin, ribonuclease B, and horseradish peroxidase, we observed near-identical coverage of peptides and glycopeptides when using online RP-HILIC--with only a single sample injection event--as we did from two separate RP and HILIC analyses. The coupled system was also capable of concurrently characterizing the peptide and glycan portions of deglycosylated glycoproteins from one injection event, as confirmed, for example, through our detection of 23 novel glycans from turkey ovalbumin. Finally, we validated the applicability of using RP-HILIC for the analysis of highly complex biological samples (mouse chondrocyte lysate, deglycosylated human serum). The enhanced coverage and efficiency of online RP-HILIC makes it a viable technique for the comprehensive separation of components displaying dramatically different hydrophobicities, such as peptides, glycopeptides, and glycans.

Helicases as antiviral drug targets.

1. We have demonstrated for the first time that the helicase of a ribonucleic acid virus, the SARS coronavirus (SARS-CoV), is a valid target for drug development. 2. Using high throughput screen and chemical synthesis, several lead compounds targeting the SARS-CoV helicase have been identified. We have shown that these compounds can inhibit SARS-CoV helicase activity and viral growth in cell culture systems. These compounds can potentially be used to target other viruses.

Inhibition of cultured rat prostatic epithelial cell growth by epristeride in vitro.

AIM: To study the molecular mechanism of rat prostate atrophy induced by epristeride. METHODS: MTT test was used to determine the effect of epristeride on the growth of prostatic epithelial cell induced by exogenous epithelial growth factor (EGF) or insulin-like growth factor-I (IGF-I). RT-PCR and flow cytometry were then used to quantitatively detect the mRNA and protein expressions of EGFR and IGF-I R of the epithelial cells treated or untreated with epristeride. RESULTS: Epristeride attenuated growth of epithelial cells induced by exogenous EGF, IGF-I. Epristeride 360 nmol/L inhibited EGFR and IGF-I R expression at mRNA level, while epristeride 180 nmol/L had no marked effect on EGFR and IGF-I R mRNA expression. Both epristeride 180 nmol/L and 360 nmol/L could down regulate EGFR and IGF-I R protein levels. CONCLUSION: The molecular mechanisms of prostatic epithelial cell atrophy induced by epristeride might be associated with alteration in the expression of growth factor receptors such as EGF and IGF-I.

Effect of epristeride on expression of TGF-beta receptor in rat prostatic epithelial cells in vitro.

AIM: To assess the effect of epristeride on the expression of transforming growth factor beta type II receptor (TbetaR II) in rat prostatic epithelial cells in vitro. METHODS: RT-PCR and Western blot were used to quantitatively detect the mRNA and protein expressions of TbetaR II in rat prostatic epithelial cells treated or untreated with epristeride. Immunocytochemical staining method was used to qualitatively analyze the expression of TbetaR II protein. RESULTS: After treatment with epristeride 180 or 360 nmol/L, TbetaR II mRNA expression levels were 0.56 +/- 0.08 and 0.59 +/- 0.07, respectively, which were significantly up-regulated compared with control cells (0.38 +/- 0.04, P < 0.05); expression level of TbetaR II protein were 3163 +/- 920 and 6769 +/- 1941, respectively, which were also markedly up-regulated compared with control cells (536 +/- 240, P < 0.05). Immunostaining showed weak positive reaction in control cells, while strong staining of TbetaR II was found in epristeride-treated cells. CONCLUSION: Epristeride may up-regulate the expression of TbetaR II to induce apoptosis of prostatic epithelial cells.

Knockdown of IGF-IR by Antisense Oligodeoxynucleotide auguments the sensitivity of bladder cancer cells to mitomycin.

AIM: To investigate whether insulin-like growth factor I receptor (IGF-IR) was involved in drug resistance of bladder cancer cells. METHODS: RT-PCR was used to detect the mRNA expression of IGF-I, IGF-II, and IGF-IR in T24 cells and normal urothelial cells. Flow cytometry and MTT tests were used to assess the effect of antisense oligodeoxynucleotide (ODN) on drug sensitivities and apoptosis of T24 cells to mitomycin (MMC). Western blot was used to analyze the effect of ODN on expression of IGF-I R protein. RESULTS: mRNA of IGF-I, IGF-II, and IGF-IR were strongly expressed in serum-free cultured T24 cell line, whereas normal urothelial cells did not express these factors/receptors or only in trace levels; knockdown of IGF-IR by antisense ODN greatly inhibited the growth of bladder cancer cells and enhanced sensitivity and apoptosis of T24 cells to MMC. CONCLUSION: These results suggested that blockage of IGF-IR signaling might potentially contribute to the treatment of bladder cancer cells which were insensitive to chemotherapy.

Effect of epristeride on the expression of IGF-1 and TGF-beta receptors in androgen-induced castrated rat prostate.

The development of benign prostatic hyperplasia (BPH) is an androgen-dependent process that may be mediated by a number of locally produced growth factors. Among them, insulin-like growth factor 1 (IGF-1) and transforming growth factor beta (TGF beta) are thought important in regulating prostate growth and homeostasis, and their expression undergoes changes in proliferative prostatic disease. Epristeride, a 5 alpha-reductase inhibitor, is an effective drug in the treatment of BPH, inducing regressive changes in the prostate. This study was designed to assess the effects of epristeride on expression of these two factors at mRNA and protein levels in castrated rats maintained with exogenous testosterone. Epristeride treatment caused significant reduction in ventral prostate weight in a dose-dependent manner. There was a positive correlation between IGF-1 mRNA expression and ventral prostate weight and an inverse correlation between TGF-beta 1 mRNA expression and ventral prostate weight. Immunohistochemistry showed strong IGF-1 receptor immunoreactivity in the prostatic epithelial cells of untreated animals. In situ hybridization demonstrated high levels of IGF-1 mRNA expression both in the prostatic stromal and epithelial cells of untreated rats. In treated rats, both IGF-1 receptor protein and IGF-1 mRNA levels decreased significantly, and IGF-1 mRNA was mainly expressed in prostatic stromal cells. Weak expression of TGF beta receptors at the protein level and TGF beta at the mRNA level were found in the prostatic hyperplastic epithelial cells of untreated rats. In treated animals, intense T beta RII immunoreactivity was observed in epithelial cells, and a higher level of TGF beta mRNA was observed in both epithelial cells and stromal cells compared with control animals. In our opinion, the effect of epristeride on rat prostatic atrophy might be mediated via local growth factor(s).

Blockage of IGF-1R signaling sensitizes urinary bladder cancer cells to mitomycin-mediated cytotoxicity.

A major problem which is poorly understood in the management of bladder cancer is low sensitivity to chemotherapy and high recurrence after transurethral resection. Insulin-like growth factor 1 receptor (IGF-1R) signaling plays a very important role in progression, invasion and metastasis of bladder cancer cells. In this study, we investigated whether IGF-1R was involved in the growth stimulating activity and drug resistance of bladder cancer cells. The results showed: The mRNAs of IGF-1, IGF-2 and IGF-1R were strongly expressed in serum-free cultured T24 cell line, whereas normal urothelial cells did not express these factors/receptors or only in trace levels; T24 cell responded far better to growth stimulation by IGF-1 than did normal urothelial cells; blockage of IGF1R by antisense oligodeoxynucleotide (ODN) significantly inhibited the growth of T24 cell and enhanced sensitivity and apoptosis of T24 cells to mitomycin (MMC). These results suggested that blockage of IGF-IR signaling might potentially contribute to the treatment of bladder cancer cells which are insensitive to chemotherapy.