Changes in the expression of gallinacins, antimicrobial peptides, in ovarian follicles during follicular growth and in response to lipopolysaccharide in laying hens (Gallus domesticus)

The aim of this study was to identify the types of gallinacin genes (GALs) expressed in ovarian follicles and to determine the changes in their expression during follicular growth and in response to lipopolysaccharide (LPS). Follicles at different stages of growth were collected from laying hens (n = 5) and LPS-injected hens (n = 3). The expression of GALs in the theca and granulosa layers was examined by semi-quantitative RT-PCR. The expression of GAL-1, -2, -7, -8, -10, and -12 in the theca layer and GAL-1, - 8, -10, and -12 in the granulosa layer was identified in white and yellow follicles. The expression of these genes was not changed in the theca and granulosa layers during follicular growth except for a decrease in that of GAL-1 in theca. The expression of GAL-1, -7, and -12 in the theca layer of the third largest follicles was increased in response to LPS at a dose of 1 mg/kg body weight and this increase was induced within 3 h and maintained until 12h postinjection. Granulosa layers did not respond to LPS until 12h injection. These results show that six and four types of GALs are expressed in the theca and granulosa layers of healthy follicles respectively, and their levels do not change with follicular growth except for GAL-1 in theca. Elevated levels of GAL-1, -7, and -12 expression in theca in response to LPS suggest that the theca cells expressing these GALs function to eliminate LPS-containing bacteria.     

The thiocyanate ion (SCN−) content of eggs from hens (Gallus domesticus) fed on a diet containing rapeseed meal

The antimicrobial effect of the lactoperoxidase-thiocyanate-hydrogen peroxide system has been recognised in milk and other biological fluids. In this study the main object was to determine whether the inclusion of 10% rapeseed meal (RSM) in layers' diet could enhance the levels of the thiocyanate ion (SCN^?) in eggs to a level required by the LP system to promote bactericidal activity in egg products. Twelve ?Abor Acre’? pullets were divided into two lots, separately housed and maintained under similar conditions. One lot was fed on a basal diet and the other on an experimental diet containing 10% RSM. Both diets were of known composition, isoenergetic and isonitrogenous. Throughout the feeding trials (15 weeks) a record was kept of egg productivity, individual egg weight and layers weight. Subsequently, the weekly-lots of eggs were analysed for thiocyanate ions (SCN^?) by gas chromatography. The colour of each yolk was measured using the ?Roche Colour Fan’? and organoleptic assessments were made for the detection of trimethylamine taint since the feeding of RSM has been reported to influence these factors. No significant difference was recorded in the average weight of eggs obtained from both groups. However, egg production was 15% higher with RSM fed hens which lost a little weight. No taint was detected in either set of eggs and no significant variation in yolk colour was noted. After 4 weeks the levels of SCN^? reached 1.6 mg kg^?1 in the experimental eggs and then remained almost constant. The control set had an average of 0.5 mg kg^?1 SCN^?. Hence an enhancement of the SCN^? levels can be achieved in shell eggs by incorporating 10% RSM in the layers' diet. This enhancement is of the order claimed necessary for the operation of this system.     

Stimulatory effects of fasting on vascular endothelial growth factor (VEGF) production by growing pig ovarian follicles

The aim of this study was to investigate the effect of fasting on both vascular endothelial growth factor (VEGF) production and VEGF mRNA expression in growing ovarian follicles (>5 mm in diameter) from gilts at 48 h after equine chorionic gonadotrophin (eCG) treatment. The concentrations of VEGF and albumin were measured in the follicular fluid of single follicles, and VEGF mRNA was determined in the follicle wall. Fasting resulted in a significant increase in VEGF concentrations in follicular fluid (20.64+/-0.72 versus 10.79+/-0.86 ng ml(-1), P<0.001), but it did not affect the total amount of VEGF mRNA in the follicle wall compared with that of fed animals. However, VEGF mRNA in the theca and granulosa compartments increased and decreased, respectively, compared with that of fed animals. The concentrations of albumin measured in follicular fluid as an index of vessel permeability were higher in fasted than in animals fed normally, most likely as a result of the increased VEGF production. Follicular steroidogenesis was impaired in fasted animals. Progesterone was the most abundant steroid in the follicular fluid and oestradiol was present in lower concentrations, thus indicating an alteration in the steroidogenic enzymatic cascade. In conclusion, fasting induces an increase in both VEGF production and vessel permeability. Such a reaction is unable under severe food deprivation to preserve follicle function, but may represent a mechanism that regulates blood vessel extension and distribution in relation to tissue requirements and availability of systemic nutrient.     

Organochlorine pesticide residues in domestic fowl (Gallus domesticus) eggs from central Kenya

In 367 domestic fowl (Gallus domesticus) eggs collected from 61 farms, residues of 10 pesticides were detected in various combinations and in the following order of frequency: p,p′-DDE (in 100% of the eggs), p,p′-DDT (98%), dieldrin (95%), Indiane (66%), p,p′-DDD (46%), o,p′-DDT (17%), β-HCH (9%), γ-HCH (5%), endrin (4%) and aldrin (0–5%). No residues of heptachlor, heptachlor epoxide, HCB or PCBs were found. The mean concentration (0–70 mg kg^?1 eggs; range <0–01–10–25) of total DDT exceeded the extraneous residue limit (ERL) of 0–50 mg kg^?1. The mean dieldrin residue level (0–35 mg kg^?1; range 0–01–14–90) was 3–5 times higher than the ERL (0–10 mg kg^?1). Only 3% of the eggs exceeded the ERL for Indane. The 156 eggs from free-range hens had significantly (P<0–05) higher residue concentrations of total DDT, dieldrin and Iindane than eggs collected from hens kept in enclosures. The mean ratio [p,p′-DDT]/[p,p′-DDE] in eggs from enclosed hens (0–97) was significantly higher (P<0–01) than in eggs from free-range hens (0–53), indicating that the former had a more direct exposure to p,p′-DDT, whereas the latter obtained more of it after environmental conversion to p,p′-DDE. Eggs from a rice-growing area had the highest concentrations of all pesticide residues detected. Accumulation ratios indicated that the levels of DDT and Iindane in the feed of enclosed hens could account for the levels in the corresponding eggs. The much higher accumulation ratios calculated for the free-range hens demonstrated that the feed ingested by these chickens obviously contained ingredients additional to those sampled, and revealed probable extensive environmental contamination by these persistent pesticides. The present results indicate that there is a need to identify sources of dieldrin in the eggs of domestic fowls and, where necessary to investigate local wildlife samples. The amounts of total DDT and dieldrin in eggs in this study seem to be higher than reported from any other country. Toxicological evaluation of the results indicates that, at lest in parts of KEnya there is a need for improved practices in the use of some organochlorine pesticides.     

Determination of carnosine in Black-Bone Silky Fowl (Gallus gallus domesticus Brisson) and common chicken by HPLC

Black-Bone Silky Fowl (Gallus gallus domesticus Brisson) is both consumed as a healthy food and used particularly as a kind of traditional Chinese medicine. Carnosine in the muscles of Black-Bone Silky Fowl was identified by HPLC–MS/PAD. The carnosine content in the meat of Black-Bone Silky Fowl was determined by HPLC and compared with that contained in the meat of White Plymouth Rock, which were bred under the same condition. The results showed that the contents of carnosine in the mixed meat, breast meat and thigh meat of Black-Bone Silky Fowl were all remarkably higher than that in the White Plymouth Rock. These findings indicate that Black-Bone Silky Fowl would be a better chicken breed for carnosine supplement.     

Direct in vivo effects of leptin on ovarian steroidogenesis in sheep

Leptin, the metabolic fat hormone, has been shown to have effects on reproduction in mice and to modulate steroid production by cultured ovarian somatic cells in a number of species. However, a direct role of leptin on normal ovarian function in vivo has not been shown. In this paper the effect of passive immunisation against leptin (experiment 1; 20 ml antiserum or non-immune plasma i.v.; n = 6/treatment) and direct ovarian infusion of leptin (experiment 2; 0, 2 or 20 mug recombinant ovine leptin; n = 4/treatment) during the early follicular phase was investigated in sheep with ovarian autotransplants, which allow recovery of ovarian venous blood and regular non-invasive scanning of the ovary. Passive immunisation against leptin resulted in an acute increase (P < 0.05) in ovarian oestradiol secretion but had no effect on gonadotrophin concentrations, ovulation or subsequent luteal function. Conversely, direct ovarian arterial infusion of the low dose of leptin resulted in an acute decline (P < 0.05) in ovarian oestradiol secretion whereas the high dose, which resulted in supra-physiological leptin concentrations, had no effect on oestradiol production compared with the controls. Neither dose of leptin had any effect on gonadotrophin concentrations or ovulation but both doses resulted in an increase (P < 0.05) in progesterone concentrations over the subsequent luteal phase. In conclusion, together these data provide strong in vivo evidence that leptin can modulate ovarian steroidogenesis directly and acutely in ruminants and suggest that leptin is an alternate regulatory system whereby nutritional status can regulate reproductive activity.     

乌鸡肽的营养成分及其功能特性研究

探讨乌鸡肽的营养成分及功能,为其深度开发打下基础。利用木瓜蛋白酶和风味蛋白酶水解乌骨鸡肌肉、活性炭脱色法制备乌鸡肽,高效液相色谱(high performance liquid chromatography,HPLC)法检测乌鸡肽的总氨基酸及游离氨基酸含量及相对分子质量分布区间、纳升级液相色谱串联质谱(Nano liquid chromatography tandem mass spectrometry,Nano LC-MS/MS)技术鉴定其组成成分。结果表明,游离氨基酸占总氨基酸的比例为1.74%,总氨基酸含量较高者为Leu、Ala、His、Pro、Met和Phe,游离氨基酸含量较高者为Glu、Pro、Asp、Arg、Gly和Ala;相对分子质量分布范围为0至6 918 Da,其中1 084 Da以下的比例达到88.91%;Nano LC-MS/MS鉴定出源于107种蛋白质的2 287种肽段序列,其中种类最为丰富的蛋白为参与骨骼肌修复的蛋白及酶类。该研究较为全面的分析乌鸡肽的营养成分,并表征了其功能,为乌鸡肽的潜在应用价值研究打下基础。     

Purification and identification of novel antioxidative peptide released from Black-bone silky fowl (Gallus gallus domesticus Brisson)

Papain-treated Black-bone silky fowl (BSF) muscle hydrolysate was subjected to 6 kDa cutoff membrane ultrafiltration, and the resulting BSF peptides (<6 kDa) were purified by two-step reverse-phase high-performance liquid chromatography. The molecular weight (MW) distribution and amino acid composition were investigated for characterization of the BSF peptides. The results showed that the major amino acids of BSF peptides were Glu, Tyr, Lys, Asp, Leu, Ala, Thr and Pro, and the MW was from 281 to 7,982 Da. BSF peptides exhibited a strong antioxidant capacity. At 10 mg/mL, they displayed more powerful $ {\text{O}}_{2}^{ \cdot - } $ , DPPH^· and ABTS^·+ scavenging activity and reducing power than carnosine. The peptide fraction 8 with more hydrophilicity revealed stronger $ {\text{O}}_{2}^{ \cdot - } $ and ABTS^·+ scavenging activity and reducing power than BSF peptides and carnosine. Besides, a peptide, separated from fraction 8 and showed the strongest antioxidant capacity, was purified and identified by LC-ESI-MS/MS to be Glu-Pro-Asp-Arg-Tyr (678 Da).     

Neonatal programming by immunological challenge: effects on ovarian function in the adult rat

Neonatal exposure to an immunological challenge (lipopolysaccharide, LPS) increases the activity of hypothalamo-pituitary-adrenal axis and sensitises the GNRH pulse generator to the inhibitory influence of stress in adult rats. We investigated the effects of neonatal exposure to LPS on various reproductive parameters during puberty and into adulthood in female rats. LPS (50?μg/kg, i.p.) or saline was administered on postnatal days 3 and 5. Vaginal opening was recorded, and oestrous cyclicity was monitored immediately post puberty and again at 8-9 weeks of age. At 10 weeks of age, the ovaries were removed and the number of follicles was counted, together with the thickness of the theca interna of the largest antral follicles. Ovarian sympathetic nerve activity was assessed immunohistochemically by measurement of the levels of ovarian low-affinity receptor of nerve growth factor (p75NGFR). In rats exposed to LPS in early life, there was a significant delay in puberty and disruption of oestrous cyclicity immediately post puberty, which persisted into adulthood. The follicle reserve was decreased, the thickness of the theca interna increased and the expression profile of ovarian p75NGFR increased in the neonatal LPS-treated animals. These data suggest that exposure to LPS during early neonatal life can have long-term dysfunctional effects on the female reproductive system, which might involve, at least in part, increased ovarian sympathetic nerve activity.     

泰和乌骨鸡活性肽的分离及其体外抗氧化作用

利用制备型HPLC 对泰和乌骨鸡活性肽进行分离,流动相为0.01mol/L 的磷酸盐溶液,采用强酸型离子交换树脂和强碱型离子交换树脂两步脱盐法对泰和乌骨鸡活性肽各分离组分进行脱盐处理。以肌肽和抗坏血酸为对照,通过清除羟自由基作用、抑制脂质过氧化作用以及对Fe2+ 和Cu2+ 螯合能力4 个体系测定泰和乌骨鸡活性肽及其分离组分的体外抗氧化能力。结果表明：泰和乌骨鸡活性肽分离所得13 个组分均具有一定的抗氧化能力,且与质量浓度呈量效关系,绝大部分分离活性肽组分比活性总肽有更强的羟自由基清除能力、脂质过氧化抑制能力以及Fe2+ 螯合能力。     

泰和乌骨鸡活性肽对小鼠骨髓有核细胞的增殖作用

采用MTT法测定活性肽对正常小鼠骨髓有核细胞的增殖作用,利用高效液相色谱-2,4-二硝基氟苯柱前衍生法测定4个活性肽组分的氨基酸组成。结果表明:小鼠骨髓细胞培养时间为24 h,泰和乌骨鸡活性肽质量浓度为0.1、1、10、100μg/mL,组分7、8、9、10可极显著(P<0.01)促进小鼠骨髓有核细胞增殖,增殖率达130%以上。在低质量浓度(0.1μg/mL)条件下,组分7能显著促进小鼠骨髓有核细胞增殖(P<0.01)且重现性较好,组分8可刺激小鼠骨髓有核细胞增殖298%。氨基酸组成分析表明,组分7、8、9、10均含有丰富的酪氨酸、甘氨酸、谷氨酸、天冬氨酸和丙氨酸,可以推测该5种氨基酸组成了刺激小鼠骨髓有核细胞增殖活性成分的物质基础。     

Chorioallantoic membranes indicate avian exposure and biomarker responses to environmental contaminants: a laboratory study with White Leghorn chickens (Gallus domesticus)

PCB and endosulfan concentrations in the chorioallantoic membrane (CAM) of white leghorn chickens (Gallus domesticus) were evaluated as indicators of hepatic cytochrome P450 isozyme activity in hens and chicks as well as toxicant concentrations in eggs and hens. Sixteen hens were randomly divided into four groups of four and dosed with a mixture of PCB105 (2,3,3',4,4'-pentachlorobiphenyl), PCB156(2,3,3',4,4',5-hexachlorobiphenyl), PCB189 (2,3,3',4,4',5,5'-heptachlorobiphenyl), and technical grade endosulfan (3:1 ratio of alpha and beta isomers) at three different dose groups. The first 10 fertile eggs laid by each hen were collected, the even-numbered eggs incubated until hatched, and the odd numbered eggs were analyzed for test chemicals. Strong (r2), significantly positive (p value) relationships were found between total PCB mass (ng) in CAMs and both total PCB concentrations (ng/g wet wt) in adults (r2 = 0.91, p = 0.0001) and eggs (r2 = 0.87, p = 0.0001). The relationship between total PCB mass in CAMs and hepatic cytochrome p450 isozyme activity in chicks (r2 = 0.49, p = 0.0001) and hens (r2 = 0.45, p = 0.014) was also significant but not as strong. This study shows that CAMs can be used to estimate avian exposure to PCBs and resultant biological response.     

玛咖粉对小鼠免疫功能的影响研究

目的明确玛咖粉对小鼠免疫功能的调节作用。方法 40只小鼠进行玛咖粉抗体生成细胞测定、对小鼠HC50的影响、碳廓清能力以及鸡红细胞吞噬率及吞噬指数的影响实验。实验设4组,灌胃的玛咔粉分别为0、0.3、0.6、0.9 g/(kg·bw),连续灌胃30 d后测定各项指标。结果 0.3 g/(kg·bw)及0.9 g/(kg·bw)组玛咔粉可以显著促进小鼠B细胞分泌抗体的能力(P0.05),0.9 g/(kg·bw)组玛咔粉具有显著增加小鼠半数溶血值(HC50)(P0.01),0.6 g/(kg·bw)及0.9 g/(kg·bw)组可以显著提高小鼠单核-巨噬细胞碳廓清能力(P0.05),0.9 g/(kg·bw)组可以显著提高小鼠腹腔巨噬细胞鸡红细胞吞噬率及吞噬指数(P0.05)。结论玛咔粉可以增强小鼠体液免疫功能和单核-巨噬细胞功能的作用。     

Regulation of ovarian function by the TGF-beta superfamily and follistatin

The role of follistatin as an activin-binding protein has dominated the study of this molecule for the last 10 years. However, there is emerging evidence that follistatin has a role in modulating the biology of other members of the transforming growth factor beta (TGF-beta) superfamily. This review summarizes the current concepts encompassing follistatin biochemistry as well as molecules with which it is functionally associated. Moreover, the importance of the two follistatin isoforms (follistatin-288 and follistatin-315) is discussed with particular emphasis on the regulation of the ovary. In addition to activin, this review discusses the functions of other members of the TGF-beta superfamily, for example growth differentiation factor 9 (GDF-9), bone morphogenetic protein 15 (BMP-15), BMP-6, BMP-4 and BMP-7, in the ovary, and the potential interactions between follistatin and these growth factors. The complex network of TGF-beta superfamily growth factor members involved in the modulation of ovarian function and the interactions of follistatin with these proteins is highlighted.     

Endocrine-disrupting chemicals in ovarian function: effects on steroidogenesis, metabolism and nuclear receptor signaling

Endocrine-disrupting chemicals (EDCs) are exogenous agents with the ability to interfere with processes regulated by endogenous hormones. One such process is female reproductive function. The major reproductive organ in the female is the ovary. Disruptions in ovarian processes by EDCs can lead to adverse outcomes such as anovulation, infertility, estrogen deficiency, and premature ovarian failure among others. This review summarizes the effects of EDCs on ovarian function by describing how they interfere with hormone signaling via two mechanisms: altering the availability of ovarian hormones, and altering binding and activity of the hormone at the receptor level. Among the chemicals covered are pesticides (e.g. dichlorodiphenyltrichloroethane and methoxychlor), plasticizers (e.g. bisphenol A and phthalates), dioxins, polychlorinated biphenyls, and polycyclic aromatic hydrocarbons (e.g. benzo[a]pyrene).     

Determination of Fermentability (Final Attenuation) of Wort

Two methods for the determination of the apparent fermentability (attenuation limit) of wort, the regular fermentation as a reference method as well as a more rapid method have been collaboratively tested by members of the Analysis Committee of the European Brewery Convention to obtain repeatability (r95)and reproducibility (R95) values and to form an idea of the equivalence of these two procedures based on the comparison of results obtained from identical samples. In the collaborative trials the 14th EBC Standard Malt and two industrial worts were analysed. Repeatability (r95) and reproducibility (R95) values of 0.5% and 2.9% respectively at a mean level of 83.9% were obtained for laboratory wort from malt using the reference fermentation method. Using the rapid fermentation method the values were 0.9% and 2.4% respectively at the mean level of 82.8%. The precision values for industrial wort using the reference fermentation method were r95 = 0.4% and R95 = 2.4% for mean levels of 81.3 and 82.4%, using the rapid method 0.3% and 3.5% were obtained respectively for mean levels of 80.5 and 81.7%. The reference fermentation is more precise in terms of reproducibility and yields higher results. The rapid method is recommended for guideline purposes only.     

Effects of leptin administration and feed restriction on thecal leucocytes in the preovulatory rat ovary and the effects of leptin on meiotic maturation,granulosa cell proliferation,steroid hormone and PGE2 release in cultured rat ovarian follicles

Leptin is expressed by adipocytes and is thought to play a role in regulating food intake and in reproduction. It has been demonstrated that acute leptin administration to immature gonadotrophin-primed rats in vivo inhibits ovulation and causes a decline in food intake. However, feed restriction alone does not inhibit ovulation. Two experiments were designed to investigate the mechanism of leptin-induced inhibition of ovulation. In the first experiment, which was prompted by the importance of ovarian leucocytes in ovulation, the role of leucocytes in leptin-induced inhibition of ovulation was investigated. The second experiment investigated whether high leptin concentrations could inhibit other factors important to ovulation, such as meiotic competence of oocytes, granulosa cell proliferation, steroid or PGE(2) release, and interleukin 1beta production, in vitro. In the first experiment, the populations of neutrophils and monocytes-macrophages in the preovulatory follicles of gonadotrophin-primed, leptin-treated and -untreated rats were examined. A decrease in food intake, as a result of either leptin treatment or feed restriction, specifically reduced the numbers of neutrophils and monocytes-macrophages infiltrating the theca interna of preovulatory follicles without affecting the numbers found in the stroma. The findings show that reduced infiltration of thecal neutrophils and macrophages into preovulatory follicles is a response to reduced food intake. Furthermore, this reduction is not the direct cause of the leptin-induced inhibition of ovulation. In the second experiment, ovarian follicles were cultured for 4 or 12 h in the presence or absence of the following hormones: FSH (500 miu), insulin-like growth factor I (IGF-I) (50 ng ml(-1)), LH (100 ng ml(-1)) and leptin (300 ng ml(-1)). The results demonstrated that high concentrations of leptin in follicle culture do not affect meiotic maturation or steroid release, but tend to inhibit release of PGE 2 (although this result was not significant). DNA synthesis in granulosa cells was not inhibited by leptin in FSH- and IGF-I-supplemented culture media. These results are in agreement with previous studies that have shown that leptin inhibits the stimulatory effects of IGF-I on FSH-stimulated oestradiol production in rat granulosa cells without affecting progesterone production. In summary, leptin does not appear to have an adverse effect on the components of ovulation tested in this study, and therefore must impact on the ovulatory cascade in a way that remains to be defined.     

Bacteriological characteristics of dressed young pigeon (squabs) Columba livia domesticus

A total of 50 frozen squabs carcasses were collected from different retail markets in Cairo and Giza governorates. The collected samples were examined bacteriologically. The aerobic plate count, the enterobacteriaceae and the Staphylococcus count were 6.6 x 10(5), 6.3 x 10(2) and 1.4 x 10(3) cfu./gram, respectively. The results revealed no positive samples for Salmonella Yersinia entercolitica, Listeria monocytogenes and Campylobacter jejuni. Staphylococcus aureus was isolated from 2% of samples. The sources of contamination of processed squabs carcasses and measures to minimize the bacterial load and safe guard the consumer are mentioned.