Phytogenic feed additives alleviate pathogenic Escherichia coli-induced intestinal damage through improving barrier integrity and inhibiting inflammation in weaned pigs

Background:This study was conducted to investigate the effects of each phytogenic feed additive(PFA;PFA1,bitter citrus extract;PFA2,a microencapsulated blend of thymol and carvacrol;PFA3,a mixture of bitter citrus extract,thymol,and carvacrol;PFA4,a premixture of grape seed,grape marc extract,green tea,and hops;PFA5,fenugreek seed powder)on the growth performance,nutrient digestibility,intestinal morphology,and immune response in weaned pigs infected with Escherichia coli(E.coli).Results:A total of 634-week-old weaned pigs were placed in individual metabolic cages and assigned to seven treatment groups.The seven treatments were as follows:1)NC;basal diet without E.coli challenge,2)PC;basal diet with E.coli challenge,3)T1;PC+0.04%PFA1,4)T2;PC+0.01%PFA2,5)T3;PC+0.10%PFA3,6)T4;PC+0.04%PFA4,7)T5;PC+0.10%PFA5.The experiments lasted in 21 d,including 7 d before and 14 d after the first E.coli challenge.In the E.coli challenge treatments,all pigs were orally inoculated by dividing a total of 10 mL of E.coli F18 for 3 consecutive days.The PFA-added groups significantly increased(P<0.05)average daily gain and feed efficiency and decreased(P<0.05)the fecal score at d 0 to 14 post-inoculation(PI).Tumor necrosis factorαwas significantly lower(P<0.05)in the PFA-added groups except for T1 in d 14 PI compared to the PC treatment.The T3 had a higher(P<0.05)immunoglobulin G and immunoglobulin A concentration compared to the PC treatment at d 7 PI.Also,T3 showed significantly higher(P<0.05)villus height:crypt depth and claudin 1 expression in ileal mucosa,and significantly downregulated(P<0.05)the expression of calprotectin compared to the PC treatment.Conclusions:Supplementation of PFA in weaned pigs challenged with E.coli alleviated the negative effects of E.coli and improved growth performance.Among them,the mixed additive of bitter citrus extract,thymol,and carvacrol showed the most effective results,improving immune response,intestinal morphology,and expression of tight junctions.

Effects of ursolic acid and oleanolic acid on human colon carcinoma cell line HCT15

AIM: Ursolic acid (UA) and oleanolic acid (OA) are triperpene acids having a similar chemical structure and are distributed wildly in plants all over the world. In recent years, it was found that they had marked anti-tumor effects. There is little literature currently available regarding their effects on colon carcinoma cells. The present study was designed to investigate their inhibitory effects on human colon carcinoma cell line HCT15. METHODS: HCT15 cells were cultured with different drugs. The treated cells were stained with hematoxylin-eosin and their morphologic changes observed under a light microscope. The cytotoxicity of these drugs was evaluated by tetrazolium dye assay. Cell cycle analysis was performed by flow cytometry (FCM). Data were expressed as means +/-SEM and Analysis of variance and Student' t-test for individual comparisons. RESULTS: Twenty-four to 72 h after UA or OA 60 micromol/L treatment, the numbers of dead cells and cell fragments were increased and most cells were dead at the 72nd hour. The cytotoxicity of UA was stronger than that of OA. Seventy-eight hours after 30 micromol/L of UA or OA treatment, a number of cells were degenerated, but cell fragments were rarely seen. The IC(50) values for UA and OA were 30 and 60 micromol/L, respectively. Proliferation assay showed that proliferation of UA and OA-treated cells was slightly increased at 24h and significantly decreased at 48 h and 60 h, whereas untreated control cells maintained an exponential growth curve. Cell cycle analysis by FCM showed HCT15 cells treated with UA 30 and OA 60 for 36 h and 72 h gradually accumulated in G(0)/G(1) phase (both drugs P【0.05 for 72 h), with a concomitant decrease of cell populations in S phase (both drugs P【0.01 for 72 h) and no detectable apoptotic fraction. CONCLUSION: UA and OA have significant anti-tumor activity. The effect of UA is stronger than that of OA. The possible mechanism of action is that both drugs have an inhibitory effect on tumor cell proliferation through cell-cycle arrest.

Paclitaxel induces apoptosis in human gastric carcinoma cells

AIM;To investigate the apoptosis in gastric cancer cells induced by paclitaxel,and the relation between this apoptosis and expression of Bcl-2 and Bax. METHODS:In in vitro experiments,MTT assay was used to determine the cell growth inhibitory rate.Transmission electron microscope and TUNEL staining method were used to quantitatively and qualitively detect the apoptosis status of gastric cancer cell line SGC-7901 before and after the paclitaxel treatment.Immunohistochemical staining was used to detect the expression of apoptosis-regulated gene Bcl-2 and Bax. RESULTS:Paclitaxel inhibited the growth of gastric cancer cell line SGC-7901 in a dose-and time-dependent manner. Paclitaxel induced SGC-7901 cells to undergo apoptosis with typically apoptotic characteristics,including morphological changes of chromatin condensation,chromatin crescent formation,nucleus fragmentation and apoptotic body formation.Paclitaxel could reduce the expression of apoptosis-regulated gene Bcl-2,and improve the expression of apoptosis-regulated gene Bax. CONCLUSION:Paclitaxel is able to induce the apoptosis in gastric cancer.This apoptosis may be mediated by down- expression of apoptosis-regulated gene Bcl-2 and up- expression of apoptosis-regulated gene Bax.

Mechanical properties of hepatocellular carcinoma cells

AIM: To study the viscoelastic properties of human hepatocytes and hepatocellular carcinoma (HCC) cells under cytoskeletal perturbation, and to further to study the viscoelastic properties and the adhesive properties of mouse hepatoma cells (HTC) in different cell cycle. METHODS: Micropipette aspiration technique was adopted to measure viscoelastic coefficients and adhesion force to collagen coated surface of the cells. Three kinds of cytoskeleton perturbing agents, colchicines (Col), cytochalasin D (CD) and vinblastine (VBL), were used to treat HCC cells and hepatocytes and the effects of these treatment on cell viscoelastic coefficients were investigated. The experimental results were analyzed with a three-element standard linear solid. Further, the viscoelastic properties of HTC cells and the adhesion force of different cycle HTC cells were also investigated. The synchronous G(1) and S phase cells were achieved through thymine-2-desoryriboside and colchicines sequential blockage method and thymine-2-desoryriboside blockage method respectively. RESULTS: The elastic coefficients, but not viscous coefficient of HCC cells (K(1)=103.6+/-12.6N.m(-2), K(2)=42.5 +/ 10.4N.m(-2), mu=4.5 +/- 1.9Pa.s), were significantly higher than the corresponding value for hepatocytes (K(1)=87.5 +/- 12.1N.m(-2), K(2)=33.3+/-10.3N.m(-2), mu=5.9+/-3.0Pa.s, P【0.01). Upon treatment with CD, the viscoelastic coefficients of both hepatocytes and HCC cells decreased consistently, with magnitudes for the decrease in elastic coefficients of HCC cells (K(1): 68.7 N.m(-2) to 81.7N.m(-2), 66.3% to 78.9%; K(2): 34.5N.m(-2) to 37.1N.m(-2), 81.2% to 87.3%, P【0.001) larger than those for normal hepatocytes (K(1): 42.6N.m(-2) to 49.8N.m(-2), 48.7% to 56.9%; K(2): 17.2N.m(-2) to 20.4N.m(-2), 51.7% to 61.3%, P【0.001). There was a little decrease in the viscous coefficient of HCC cells (2.0 to 3.4Pa.s, 44.4 to 75.6%, P【0.001) than that for hepatocytes (3.0 to 3.9Pa.s, 50.8 to 66.1% P【0.001). Upon treatment with Col and VBL, the elastic coefficients of hepatocytes generally increased or tended to increase while those of HCC cells decreased. HTC cells with 72.1% of G(1) phase and 98.9% of S phase were achieved and high K(1), K(2) value and low mu value were the general characteristics of HTC cells. G(1) phase cells had higher K(1) value and lower mu value than S phase cells had, and G(1) phase HTC cells had stronger adhesive forces ((275.9 +/- 232.8) x 10(-10)N) than S phase cells ((161.2 +/- 120.4) x 10(-10)N, P【0.001). CONCLUSION: The difference in both the pattern and the magnitude of the effect of cytoskeletal perturbing agent on the viscoelastic properties between HCC cells and hepatocytes may reflect differences in the state of the cytoskeleton structure and function and in the sensitivity to perturbing agent treatment between these two types of cells. Change in the viscoelastic properties of cancer cells may affect significantly tumor cell invasion and metastasis as well as interactions between tumor cells and their micro-mechanical environments.

Taxotere resistance in SUIT Taxotere resistance in pancreatic carcinoma cell line SUIT 2 and its sublines

AIM: To investigate the specific mechanisms of intrinsic and acquired resistance to taxotere (TXT) in pancreatic adenocarcinoma (PAC). METHODS: MTT assay was used to detect the sensitivity of PAC cell line SUIT-2 and its sublines (S-007, S-013, S-020, S-028 and TXT selected SUIT-2 cell line, S2/TXT) to TXT. Mdr1 (P-gp), multidrug resistance associated protein (MRP), lung resistance protein (LRP) and beta-tubulin isotype gene expressions were detected by RT-PCR. The functionality of P-gp and MRP was tested using their specific blocker verapamil (Ver) and indomethacin (IMC), respectively. The transporter activity of P-gp was also confirmed by Rhodamine 123 accumulation assay. RESULTS: S-020 and S2/TXT were found to be significantly resistant to TXT(19 and 9.5-fold to their parental cell line SUIT-2, respectively). RT-PCR demonstrated strong expression of Mdr1 in these two cell lines, but weaker expression or no expression in other cells lines. MRP and LRP expressions were found in most of these cell lines. The TXT-resistance in S2-020 and S2/TXT could be reversed almost completely by Ver, but not by IMC. Flow cytometry showed that Ver increased the accumulation of Rhodamine-123 in these two cell lines. Compared with S-020 and SUIT-2, the levels of beta-tubulin isotype II, III expressions in S-2/TXT were increased remarkably. CONCLUSION: The both intrinsic and acquired TXT-related drug resistance in these PAC cell lines is mainly mediated by P-gp, but had no relationship to MRP and LRP expressions. The increases of beta-tubulin isotype II, III might be collateral changes that occur when the SUIT-2 cells are treated with TXT.

Effects of Taxotere on invasive potential and multidrug resistance phenotype in pancreatic carcinoma cell line SUIT-2

INTRODUCTIONDevelopment of drug-resistance to chemotherapyand subsequent metastasis of tumor are primarilyresponsible for treatment failure and the death fromcancer. There have been many previous studies onthe relationship between expression of multidrugresistance (MDR) phenotype P-glycoprotein (P-gp)and the malignant properties of tumors, but theresults are often conflicting[1-8]. The difference intumor types or MDR phenotype induced by specificagents might account for this discrepancy. Taxotere(TXT), a member of the family of taxanes, hasantitumor activity through its effect of promotingthe polymerization of tubulin[9,10].

Removal of lead and cadmium ions by single and binary systems using phytogenic magnetic nanoparticles functionalized by 3-marcaptopropanic acid

The present research study is focused on green fabrication of superparamagnetic Phytogenic Magnetic Nanoparticles(PMNPs), and then its surface functionalization with 3-Mercaptopropionic acid(3-MPA). The resulting material(i.e. 3-MPA@PMNPs) characterized by FTIR, powder XRD, SEM, TEM, EDX, VSM, BET and TGA techniques and then further employed for the investigation of the adsorptive removal of lead(Pb^2+) and cadmium(Cd^2+) ions from aqueous solutions in single and binary systems. The material showed fastest adsorptive rate(98.23%) for Pb^2+ and(96.5%) Cd2+within the contact time of 60 min at pH 6.5 in the single system. The experimental data were fitted well to Langmuir isotherm, indicated monolayer adsorption of both metal ions onto 3-MPA@PMNPs and an estimated comparable adsorptive capacity of 68.41 mg·g^-1(Pb2+) and 79.8 mg·g^-1(Cd2+) at p H 6.5. However, kinetic data agreed well with pseudo-second-order model, and indicated that the removal mainly supported chemisorption and/or ion-exchange mechanism. Thermodynamic parameters such asΔGo,ΔHo, and ΔSo, were-3259.20, 119.35 and 20.73 for Pb^2+, and-1491.10, 45.441 and 7.87 for Cd^2+ at temperature 298.15 K, confirmed that adsorption was endothermic, spontaneous and favorable. The material demonstrated higher selectivity of Pb2+ and its removal efficiency was(98.20 ± 0.3)% in binary system experiments. The material persisted performance up-to seven(07) consecutive treatment cycles without losing their stability and offered comparable fastest magnetic separation(35 s) from aqueous solutions. Therefore, it is recommended that the prepared material can be employed to remove toxic heavy metal ions from water/wastewaters and this "green" method can easily be implemented at large scale in low economy countries.

Phytogenic Feed Additive for Sows： Effects on Sow Feed Intake, Serum Metabolite Concentrations, IgG Level, Lysozyme Activity and Milk Quality

The objective of this study was to investigate the influence ofa phytogenic feed additive （PFA） to late-gestation （d 90） and lactation sows on their reproduction performance. Sixty Large White × Landrace primiparous sows were divided into four groups （fifteen sows per group）. The control group was fed with basal diet, the others were fed with basal diet supplemented with 0.02%, 0.04% and 0.06% PFA, respectively. Compared with the control, the sows in 0.04% PFA group had higher feed intake during lactation and higher litter weaning weight （P 〈 0.05）. At farrowing, glucose level in 0.04% PFA sows was higher than the control and sows in 0.02% PFA had the highest IgG content among the treatments （P 〈 0.05）. On day 7 of lactation, serum urea nitrogen contents were lower in response to PFA supplementation compared to the control （P 〈 0.05）. At the same time, sows fed the 0.06% PFA diet increased the lysozyme activity （P 〈 0.05）. The levels of milk lactose and IgG were increased in 0.02% and 0.04% PFA groups （P 〈 0.05）. In conclusion, feeding PFA improved sows and litter performance, serum metabolite concentrations, lgG level and lysozyme activity at postpartum and milk quality.

Adeno-associated virus mediated endostatin gene therapy in combination with topoisomerase inhibitor effectively controls liver tumor in mouse model

AIM:rAAV mediated endostatin gene therapy has been examined as a new method for treating cancer.However, a sustained and high protein delivery is required to achieve the desired therapeutic effects.We evaluated the impact of topoisomerase inhibitors in rAAV delivered endostatin gene therapy in a liver tumor model. METHODS:rAAV containing endostatin expression cassettes were transduced into hepatoma cell lines.To test whether the topoisomerase inhibitor pretreatment increased the expression of endostatin,Western blotting and ELISA were performed.The biologic activity of endostatin was confirmed by endothelial cell proliferation and tube formation assays. The anti-tumor effects of the rAAV-endostatin vector combined with a topoisomerase inhibitor,etoposide,were evaluated in a mouse liver tumor model. RESULTS:Topoisomerase inhibitors,including camptothecin and etoposide,were found to increase the endostatin exPression level in vitro.The over-expressed endostatin, as a result of pretreatment with a topoisomerase inhibitor, was also biologically active.In animal experiments,the combined therapy of topoisomerase inhibitor,etoposide with the rAAV-endostatin vector had the best tumor- suppressive effect and tumor foci were barely observed in livers of the treated mice.Pretreatment with an etoposide increased the level of endostatin in the liver and serum of rAAV-endostatin treated mice.Finally,the mice treated With rAAV-endostatin in combination with etoposide showed the longest survival among the experimental models. CONCLUSION:rAAV delivered endostatin gene therapy in combination with a topoisomerase inhibitor pretreatment is an effective modality for anticancer gene therapy.

Altered expression of nuclear matrix proteins in etoposide induced apoptosis in HL-60 cells

The events of cell death and the expression of nuclear matrix protein (NMP) have been investigated in a promyelocytic leukemic cell line HL-60 induced with etoposide. By means of TUNEL assay, the nuclei displayed a characteristic morphology change, and the amount of apoptotic cells increased early and reached maximun about 39% after treatment with etoposide for 2 h. Nucleosomal DNA fragmentation was observed after treatment for 4 h. The morphological change of HL-60 cells, thus, occurred earlier than the appearance of DNA ladder. Total nuclear matrix proteins were analyzed by 2-dimensional gel electrophoresis. Differential expression of 59 nuclear matrix proteins was found in 4 h etoposide treated cells. Western blotting was then performed on three nuclear matrix acssociated proteins, PML, HSC70 and NuMA. The expression of the suppressor PML protein and heat shock protein HSC70 were significantly upregulated after etoposide treatment, while NuMA, a nuclear mitotic apparatus protein, was down regulated. These results demonstrate that significant biochemical alterations in nuclear matrix proteins take place during the apoptotic process.

Effects of broccoli extract and various essential oils on intestinal and faecal microflora and on xenobiotic enzymes and the antioxidant system of piglets

Objective: Since the ban of antibiotics as growth promoting feed additives in the EU in 2006 research in alternatives has gained importance. Phytogenic feed additives represent a heterogenous class of different plant derived substances that are discussed to improve the health of farm animals by direct and indirect antioxidant effects and by influencing microbial eubiosis in the gastrointestinal tract. Consequently our study aimed to investigate the influence of broccoli extract and the essential oils of tur- meric, oregano, thyme and rosemary, as selected individual additives, on intestinal and faecal microflora, on xenobiotic enzymes, and on the antioxidant system of piglets. Methods: 48 four weeks old male weaned piglets were assigned to 6 groups of 8. The piglets were housed individually in stainless steel pens with slatted floor. The control group (Con) was fed a diet without an additive for 4 weeks. The diet of group BE contained 0.15 g/kg sulforaphane in form of a broccoli extract. 535, 282, 373 and 476 mg/kg of the essential oils of turmeric (Cuo), oregano (Oo), thyme (To) and rosemary (Ro) were added to the diets of the remaining 4 groups to stan-dardise supplementation to 150 mg/kg of the oils’ key terpene compounds ar-turmerone, carvacrol, thymol and 1,8-cineole. The composition of bacterial microflora was examined by cultivating samples of jejeunal and colonic mucosa and of faeces under specific conditions. The mRNA expression of xenobiotic and antioxidant enzymes was determined by reversing transcrip- tase real time detection PCR (RT-PCR). Total antioxidant status was assayed using the Trolox Equivalent Antioxidant Capacity (TEAC), and lipid peroxidation was determined by measuring thiobarbioturic acid reactive substances (TBA- RS). Results: Compared to Con piglets all additives positively influenced weight gain and feed conversion in week 1. Over the whole trial period no significant differences in performance parameters existed between the experimental groups. Compared to group Con performance of Ro piglets was, however, slightly impaired. Com- pared to Con piglets Cuo, Oo and To increased the ratio of Lactobacilli:E. coli attached to the jejunal mucosa, whereas BE and Ro impaired this ratio slightly. In contrast in colonic mucosa Ro improved Lactobacilli:E. coli ratio. In faecal samples an improvement of Lactobacilli:E. coli ratio could be analysed for To and Ro. Ro was the only additive that reduced the incidence rate of piglets tested positive for enterotoxic E. coli (ETEC). All additives significantly increased jejunal TEAC and reduced TBA-RS. In the liver BE, Cuo, Oo and To increased TEAC in tendency and Ro significantly. Liver TBA-RS were slightly reduced by all additives compared to Con piglets. Whereas the influence of BE, To and Ro on jejunal TEAC mainly was derived from the induction of xenobiotic and antioxidant enzymes (indirect antioxidant effects), Cuo and Oo influenced TEAC by direct antioxidant effects. Discussion and Conclusions: Our results have shown: That within the labiatae oils Oo and To have the potential to improve performance slightly. That phytogenic substances have a small but not sig- nificant influence on intestinal microflora. That phytogenic feed additives up-regulate the anti- oxidant system of piglets either by direct or by indirect antioxidant effects and that they may thereby improve health status. That within the labiatae oils Oo has a high direct antioxidant potential whereas Ro potently induces xenobiotic and antioxidant enzymes. That broccoli extract is an attractive new phytogenic additive, improving antioxidant status by indirect antioxidant effects. That defined combinations of selected phytogenic substances may produce additive effects. That health promoting effects of phytogenic additives in the future should be studied systematically under the challenge with pathogenic microorganisms or food derived to-xins.

Advances in the Researches on the Blocking Effect of Chinese Drugs on Tumors

Malignant tumors are caused by multiple carcinogenic factors undergoing several stages. The occurrence and development of tumors may be prevented and blocked if some effective interference factors are brought into play.1 At present, there are two main subjects for the researches, that is, blocking the precancerous lesions and blocking the develop-ment of tumors. The former focuses on the removing of carcinogenic factors and on the chemoprophylaxis of cancer, while the latter on the inhibition of cancer cell infiltration and cancerometastasis. These are summarized as follows.

Turmeric on Poultry Production: A Review

In the commercial poultry chain, a great diversity of production systems is found to meet the needs of the market, since the consumer has become increasingly aware of the importance of the relationship between diet and health, which has encouraged researchers and the food industry to develop products enriched with nutrients capable of producing beneficial effects on health. There has been increasing progress in research aimed at including viable alternative products, such as plant extracts, in poultry diets, which have been shown to have an antimicrobial and antioxidant action and to promote improvements in animal performance. Turmeric (Curcuma longa L.) is a popular medicinal herb, which shows a wide range of pharmacological properties, such as antioxidant, antiangiogenic, antitumor, antivenom, antinflammatory, antimicrobial, antiproliferative, antiprotozoal, and antiaging. The use of turmeric in poultry production has been reported by many researchers. The aim of this review is to describe and report the most recent research on the use of turmeric as feed supplement for birds and so its effect on animal health and welfare.

Turmeric and Vitamin E on the Performance and Quality of Eggs from White Layers

A 112-day experiment was conducted to evaluate turmeric and vitamin E levels on the productive performance and quality of hen eggs. A total of 432 20-week-old white layers were used, distributed in an entirely randomized design, in a 4 × 3 factor scheme (turmeric extract levels and vitamin E levels), with 6 repetitions of 6 birds per cage. The treatments consisted of four levels of turmeric extract inclusion (0;0.1;0.2 and 0.3) and three levels of vitamin E inclusion (0;50 and 100 IU/kg). Egg quality was evaluated at each 28-day cycle through the variables specific gravity, yolk percentage, albumen percentage, shell percentage and thickness, Haugh unit, yolk index and staining, and production performance: weight, egg production and mass, feed intake, feed conversion (kg/kg;kg/dz), and mortality. The addition of vitamin E alone worsened feed intake, egg weight and feed conversion of layers. Turmeric added to 100 IU of Vit E improved egg mass indices, Haugh Unit and yolk index and intensified yolk staining. The amount of turmeric used in this experiment was not sufficient to improve the performance or egg quality of the birds.

Laying Egg Production Supplemented with Antioxidants: An Economical View

A 112-day experiment was conducted to economically evaluate the production of eggs from commercial layers submitted to diets supplemented with levels of plant extracts (dried turmeric powder) added or not to vitamin E. A total of 432 20-week-old white layers were used, distributed in an entirely randomized design, in a 4 × 3 factor scheme (turmeric extract levels and vitamin E levels), with 6 repetitions of 6 birds per cage. The treatments consisted of four levels of turmeric extract inclusion (0;0.1%;0.2% and 0.3%) and three levels of vitamin E inclusion (0;50 and 100 IU/kg). The results calculated at work were favorable to the activity, and the items that most influenced the expenses in the effective production cost were: poultry, feed and labor, respectively. The best economic response in egg production was obtained by layers that consumed 100 IU/kg vitamin E and 0.2% turmeric;because the leveling point represents that the activity equals the revenues with the costs. The best profitability index was found in treatment that contained 50 IU/kg vitamin E and 0.1% turmeric com (16.78%).

Pin Oak (Quercus palustris) Leaf Extract Mediated Synthesis of Triangular, Polyhedral and Spherical Gold Nanoparticles

Fabrication of phytogenic metal nanoparticles is a developing area of nanobiotechnology which has economic and potentially eco-friendly advantages over conventional chemical and physical synthesis methods. We report the synthesis of anisotropic gold nanostructures using a phytoextract from the leaves of the pin oak tree, Quercus palustris. We used 2.5%, 5%, and 10% m/v extract concentrations to determine the importance of pin oak phytoextract concentration on the properties of the nanomaterials synthesized. Characterization of the resulting nanomaterials revealed that the concentration of the phytoextract is a key factor controlling the size and relative frequency of shapes the gold nanostructures formed. TEM micrographs demonstrate that triangular, spherical, and polyhedral gold nanostructures were formed in all treatments, although UV-Vis spectra indicated that the formation of fewer nanostructures in the 2.5% m/v phytoextract treatment We found that anisotropic nanostructures are formed at the highest concentrations in the preparation using 10% m/v phytoextract. UV-Vis, TEM, and DLS data indicate that the 5% m/v phytoextract results in the synthesis of the smallest sized nanoparticles.

Broiler gut microbiota and expressions of gut barrier genes affected by cereal type and phytogenic inclusion

The present study assessed the effects of cereal type and the inclusion level of a phytogenic feed additive(PFA) on broiler ileal and cecal gut microbiota composition, volatile fatty acids(VFA) and gene expression of toll like receptors(TLR), tight junction proteins, mucin 2(MUC2) and secretory immunoglobulin A(sIgA). Depending on cereal type(i.e. maize or wheat) and PFA inclusion level(i.e. 0, 100 and 150 mg/kg diet), 450 one-day-old male broilers were allocated in 6 treatments according to a 2 × 3 factorial arrangement with 5 replicates of 15 broilers each, for 42 d. Significant interactions(P 0.05) between cereal type and PFA were shown for cecal digesta Bacteroides and Clostridium cluster XIVa, ileal digesta propionic and branched VFA, ileal s IgA gene expression, as well as cecal digesta branched and other VFA molar ratios. Cereal type affected the cecal microbiota composition. In particular, wheat-fed broilers had higher levels of mucosa-associated Lactobacillus(P_(CT)= 0.007) and digesta Bifidobacterium(P_(CT)< 0.001),as well as lower levels of total bacteria(P_(CT)= 0.004) and Clostridia clusters I, IV and XIVa(P_(CT) 0.05),compared with maize-fed ones. In addition, cereal type gave differences in fermentation intensity(P_(CT)= 0.021) and in certain individual VFA molar ratios. Wheat-fed broilers had higher(P 0.05) ileal zonula occluden 2(ZO-2) and lower ileal and cecal TLR2 and sIgA levels, compared with maize-fed broilers. On the other hand, PFA inclusion at 150 mg/kg had a stimulating effect on microbial fermentation at ileum and a retarding effect in ceca with additional variable VFA molar patterns. In addition, PFA inclusion at 100 mg/kg increased the ileal mucosa expression of claudin 5(CLDN5)(PPFA= 0.023) and MUC2(PPFA= 0.001) genes, and at 150 mg/kg decreased cecal TLR2(PPFA= 0.022) gene expression compared with the un-supplemented controls. In conclusion, cereal type and PFA affected in combination and independently broiler gut microbiota composition and metabolic activity as well as the expression of critical gut barrier genes including TLR2. Further exploitation of these properties in cases of stressor challenges is warranted.

Ophiopogonin D inhibits cell proliferation,causes cell cycle arrest at G_2/M,and induces apoptosis in human breast carcinoma MCF-7 cells

OBJECTIVE： To investigate the effects of ophiopogonin D on human breast cancer MCF-7 cells METHODS： Cell viability was assessed by 3-（4,5-dimethylthiazol-2-yl）-2,5-diphenyltetrazolium bromide and colony formation experiments. Cell cycle was measured with cell cycle flow cytometry and a living cell assay. Apoptosis and terminal deoxynucleoitidyl transferase-mediated dUTP nick end labeling assays were performed to detect the apoptosis of MCF-7 cells induced by ophiopogonin D. Finally, Western blotting was used to explore the mechanism. RESULTS： Exposure of cells to ophiopogonin D resulted in marked decreases in viable cells and colony formation with a dose-dependent manner. Treatment of these cells with ophiopogonin D also resulted in cell cycle arrest at the G2/M phase, and increased apoptosis. Mechanistically, ophiopogonin D-induced G2/M cell cycle arrest was associated with down-regulation of cyclin BI. Furthermore, activation of caspase-8 and caspase-9 was involved in ophiopogonin D-induced apoptosis. CONCLUSION： The data suggested that ophiopogonin D inhibits MCF-7 cell growth via the induction of cell cycle arrest at the G2/M phase.

Growth performance, nutrient digestibility, antioxidant capacity,blood biochemical biomarkers and cytokines expression in broiler chickens fed different phytogenic levels

The effects of inclusion levels of a phytogenic feed additive(PFA), characterized by menthol anethol and eugenol, on broiler growth performance, nutrient digestibility, biochemical biomarkers and total antioxidant capacity(TAC) of plasma and meat, as well as on the relative expression of selected cytokines,were studied in a 42-d experiment. A total of 225 one-day-old male Cobb broiler chickens were assigned into 3 treatments, with 5 replicates of 15 chickens each. Chickens were fed maize-soybean meal basal diets following a 3 phase(i.e., starter, grower and finisher) feeding program. Depending on PFA inclusion level, treatments were: no PFA(PFA-0), PFA at 100 mg/kg(PFA-100) and PFA at 150 mg/kg(PFA-150).Feed and water were available ad libitum. Feed conversion ratio(FCR) during finisher phase was improved quadratically(P < 0.05) with increasing PFA level. Overall, increasing PFA level increased body weight gain(BWG) in a linear(P < 0.05) and quadratic(P < 0.05) manner with treatments PFA-100 and PFA-150 being greater(P < 0.05) compared with PFA-0. Total tract apparent digestibility of dry matter increased linearly(P < 0.05) and quadratically(P < 0.05) with increasing PFA level. The apparent metabolizable energy corrected for nitrogen(AMEn) also increased linearly(P < 0.05). Increasing PFA level resulted in a linear(P < 0.05) increase in blood plasma TAC. Expression of pro-inflammatory cytokine interleukin-18(IL-18) was reduced linearly(P < 0.05) in spleen with increasing PFA level. In conclusion, PFA inclusion at 100 mg/kg diet positively influenced performance, whereas PFA inclusion at150 mg/kg resulted in a stronger improvement in AMEn and plasma TAC. Finally, PFA inclusion resulted in a pattern of reduced pro-inflammatory biomarker IL-18 at spleen. Overall, this study provides evidence for the beneficial role of PFA as a natural growth and health promoter in broiler chickens that needs to be further confirmed in field studies.

Effect of supplementing layer hen diet with phytogenic feed additives on laying performance, egg quality, egg lipid peroxidation and blood biochemical constituents

This study was conducted to evaluate the effects of supplementing laying hen diet with phytogenic additives on laying performance, egg quality, blood constituents and egg lipid peroxidation. Two hundred Lohmann Brown Lite laying hens were randomly allotted to 4 dietary treatments: control(without phytogenic additive), fennel seeds(5 g/kg), black cumin seeds(5 g/kg) and hot red pepper(5 g/kg).Each of the 4 diets was fed to 5 replicates of 10 hens for 8 weeks. No significant differences were observed in body weight or feed intake between the groups. Dietary inclusion of fennel followed by red pepper improved(P < 0.05) egg weight, egg production, egg mass and feed conversion ratio compared with control. Higher yolk shape index, shell and albumen weight percentages and Haugh unit(P < 0.05)were recorded in the fennel supplemented group compared with control. The egg yolk color score increased by the addition of fennel or hot red pepper in laying hen diets compared with control. The inclusion of black cumin or hot red pepper decreased serum and egg yolk cholesterol and malondialdehyde concentrations(P < 0.05) compared with control. Serum aspartate aminotransferase concentration was lower in black cumin group(P < 0.05) than in other treatments. In conclusion, the best laying performance and egg quality were obtained by dietary inclusion of fennel, followed by hot red pepper and black cumin. Dietary supplementation of black cumin or red pepper may lead to the development of low-cholesterol concentration and better antioxidant capacity of eggs.