Expression and localisation of calbindin D28k in all intestinal segments of the laying hen

1. The aim of the present study was to investigate expression and localisation of a 28-kDa calcium-binding protein (CaBP-D28k) related to active calcium (Ca) absorption, in the entire intestine of egg-laying hens. 2. Western blotting analysis showed that the entire intestine expressed CaBP-D28k to the following degree: duodenum > jejunum > caecum > ileum > colon. Immunohistochemistry showed strong CaBP-D28k localisation in enterocytes along the villus tip-crypt axis in the duodenum and in villus tips in the caecum and colon. The jejunum and ileum had moderate localisation with respect to the number of immunoreactive cells and staining intensity. 3. These results suggest that laying hens actively absorb Ca in both the large and small intestines.     

不同日龄乳鸽肠道CaBP-D28k基因表达的变化规律研究

为了解乳鸽肠道钙吸收规律,以满足乳鸽生产中钙的营养需要,试验将48对健康种鸽随机分为8个重复,每个重复6对,产蛋后由种鸽自然孵化和哺育乳鸽,在乳鸽7、14、21、28日龄时称量体重,平均体重分别为(120.27±31.37)g、(264.35±88.77)g、(399.22±75.04)g和(454.16±38.39)g,分别从每个重复选取1只体重适中的乳鸽,取其嗉囊内容物(鸽乳),并对其十二指肠和空肠肠段进行采样,测定鸽乳钙含量和肠道CaBP-D28k的相对表达量。结果表明:7日龄鸽乳钙含量显著低于21、28日龄鸽乳钙含量(P<0.01);7日龄时十二指肠和空肠中CaBP-D28k的表达量显著高于14、21、28日龄的表达量(P<0.01);且CaBP-D28k在7～28日龄乳鸽十二指肠的表达量显著高于在空肠的表达量(P<0.05)。由此可见:鸽乳的钙含量随着乳鸽日龄的增大而增加,乳鸽肠道CaBP-D28k基因表达量随着日龄增大而减少,十二指肠是鸽进行钙载体吸收的主要部位。     

Genetic variation in the time of oviposition in the laying hen

1. The distribution of oviposition times in modern layers is relatively narrow. Consequently, in management systems where hens have to compete for nestboxes, some eggs will be laid on the floor. 2. The purpose of this study was to investigate the possibility of using genetic methods to change oviposition time while maintaining a high production rate. 3. The mean oviposition times of 464 hens from three selection lines were calculated using time-of-lay records for every egg laid until the hens reached 51 weeks of age. 4. Heritability estimates for mean oviposition time were high for each of the three lines: 0.38, 0.68 and 0.78. 5. The three lines differed in terms of the genetic correlations between mean oviposition time and certain important production traits. It should, however, be possible to shift the mean oviposition time while maintaining a high production rate. 6. It is suggested that the spread in oviposition times be increased by performing divergent selection for mean oviposition time and by mixing the different genotypes in the same commercial flock, kept on the floor or in other non-cage systems.     

Influence of dietary magnesium content on laying performance and egg shell magnesium content in the domestic hen

1. The effects of feeding diets containing various amounts of magnesium on laying performance in the domestic hen were investigated.

2. A reduction in egg number, egg mass, egg shell thickness and egg shell magnesium content occurred in birds receiving diets containing either 207 or 132 mg Mg²⁺/kg even though the duration of egg formation was unaffected.

3. The progressive increase in magnesium content normally observed towards the outer surface of the hen's egg shell was not present in birds receiving a diet containing 207 mg Mg²⁺/kg.     

蛋鸡TRPV6基因的原核表达、纯化及抗体制备

为制备蛋鸡TRPV6蛋白多克隆抗体,根据其基因序列,设计1对特异性引物,以卵巢组织中提取的总RNA为模板,扩增蛋鸡TRPV6基因1 801～2 176nt的375bp序列,构建原核表达质粒pET-32a（＋）-TRPV6;将重组质粒转化BL21（DE3）,经IPTG诱导表达TRPV6融合蛋白,通过镍离子螯合柱纯化后免疫新西兰大白兔,获得兔抗鸡TRPV6多克隆抗体,分别通过酶联免疫吸附试验（ELISA）法和Western blot检测抗体的效价和抗体特异性。结果表明,试验成功构建原核表达载体pET-32a（＋）-TRPV6,SDS-PAGE蛋白电泳检测发现目的蛋白大小约35 000;Western blot分析显示,表达的TRPV6融合蛋白具有良好的免疫原性,其抗体可与大肠杆菌表达的产物特异性结合;ELISA显示其抗体效价达1∶100 000。获得纯化的融合蛋白和多克隆抗体对研究TRPV6钙离子通道在蛋鸡髓质骨形成的作用机理具有重要作用。     

Calcitonin receptor binding in the hen anterior pituitary during an oviposition cycle

The equilibrium dissociation constant (K_d) and the maximum binding capacity (B_max) of calcitonin (CT) receptor in the plasma membrane of the anterior pituitary in hens were examined by Scatchard analysis of specific binding of ¹²⁵I‐labeled chicken CT. Values of K_d and B_max of CT receptor were smaller in laying hens than in non‐laying hens. A decrease in the K_d and B_max value of CT receptor was observed in the anterior pituitary after the injection of estradiol‐17β and progesterone into nonlaying hens, but not changed after the injection of 5α‐dihydrotestosterone. During an oviposition cycle, the K_d and the B_max value decreased 3 h before oviposition. In non‐laying hens, neither the K_d nor the B_max value changed during a full day period. The present study suggests that the CT action on the anterior pituitary may increase 3 h before oviposition by the effect of estradiol‐17β and progesterone in laying hens.     

Effect of metyrapone on the timing of oviposition and ovarian steroidogenesis in the laying hen

1. The purpose of this study was to observe the effects of metyrapone on the time of oviposition and LH-stimulated steroidogenesis by granulosa cells and small yellow follicles. 2. In experiment 1, White Leghorn hens were injected for 11 d with 240 mg metyrapone 5 h before 'lights off'. Control hens were injected with 1 ml of vehicle (PEG-400). Metyrapone treatment resulted in a 28% decrease in the rate of lay and the modal frequency of the time of oviposition was phase-shifted by 15 h. 3. In experiment 2, hens were injected with 240 mg metyrapone 5 h before 'lights off' or at 'lights on'. While metyrapone treatment reduced the rate of lay, a clear phase-shift in the distribution of oviposition was not observed. Basal and LH-stimulated progesterone synthesis by the granulosa cells of the largest follicle and oestradiol synthesis by small yellow follicles was significantly reduced. 4. Metyrapone treatment significantly reduced basal, but not LH-stimulated output of androstenedione by whole small yellow follicles compared to that observed in control hens. 5. The addition of metyrapone in vitro to isolated granulosa cells from the three largest preovulatory follicles inhibited LH-stimulated progesterone production in a dose-specific manner. 6. The results of this study suggest that the ability of metyrapone to perturb the open-period is a pharmacological effect mediated through inhibition of ovarian and adrenal steroidogenesis.     

Modelling the changes in the proportions of the egg components during a laying cycle

1. As hens age, egg weight increases but the eggs contain proportionally more yolk and less albumen and shell. However, at a given age, larger eggs contain proportionally more albumen. When modelling the nutrient requirements of the hen over a production cycle, based on the daily outputs of each nutrient, egg weight needs to be predicted as the sum of the three components, since each has a unique chemical composition, and these proportional changes will therefore influence the nutrient requirements of the hen. 2. Yolk weight is related to hen age and may be calculated using a logistic or Gompertz function. Allometric functions are used to predict albumen weight from yolk weight and shell weight from the weight of the egg contents. 3. A mechanistic, stochastic population model for layers may be used to verify that these functions correctly reflect the proportional changes in the egg components with advancing hen age and at a given age, over a range of egg weights. 4. The various parameters used in the equations need to be defined for the available genotypes.     

Role of magnesium in egg shell formation in the domestic hen

1. The effects of feeding diets containing various amounts of magnesium on plasma concentration of calcium and magnesium in the domestic hen were investigated. 2. Plasma concentrations of calcium and magnesium decreased during shell formation in all birds. 3. Plasma magnesium content and egg shell thickness were severely reduced in birds given diets containing either 207 or 132 mg Mg++/kg. 4. Using electron microscopy, a precise correlation was observed between the normal distributions of magnesium and organic material across the egg shell of the domestic hen.     

Activities of amylase, trypsin and lipase in the pancreas and small intestine of the laying hen during egg formation

1. During a cycle of 14 h artificial light and 10 h darkness, dry matter and enzyme concentrations were measured in the pancreas and gut of egg-forming hens. 2. Dry or fresh pancreas weight did not show significant variation but pancreatic enzyme activities increased gradually from 2 to 22 h after oviposition. 3. Total wet and dry matter contents of the small intestine gradually increased from 2 to 18 h after oviposition and then decreased significantly. Intestinal tryptic activity followed the same pattern whereas intestinal amylase gradually decreased from 2 h after oviposition; intestinal lipase activity tended to decrease during the light period (2 to 10 h after oviposition) and to increase during the dark period (14 to 22 h after oviposition). 4. By contrast, patterns of enzyme activity in the pancreas were similar. It is proposed that modification of the chyme and transit rates during the day may affect pancreatic enzyme fate and distribution in the small intestine. 5. In non-egg-forming birds intestinal contents and enzyme activities were less than in egg-forming birds. The reverse was observed in the case of the pancreas where contents and enzyme activities were increased. A relationship with food intake pattern is discussed.     

Staining properties of the luminal epithelium of the isthmus and shell gland of the oviduct of the hen Gallus domesticus during the passage of an egg

The effect of a developing egg on the staining properties of the luminal epithelium of the isthmus and shell gland of the oviduct of the hen Gallus domesticus is described, with special reference to the red region. Using histochemical methods for the demonstration of complex mucopolysaccharides, the staining reactions in the surface epithelium adjacent to an egg were found to be similar to those seen in an empty oviduct. With the empirical methods for intracellular calcium, a staining reaction was seen in cells which were adjacent to an egg. There was no reaction in an empty oviduct. If an egg lay between the red region and the shell gland, methods for the demonstration of acid mucopolysaccharides or intracellular calcium showed a more pronounced reaction in an area of surface cells at the neck of the shell gland. These results are based on recently defined anatomical regions.     

The distribution and carbohydrate composition of the organic matrix in hen egg shell

After the removal of membranes and cuticle, successive arbitrary layers were dissolved with dilute HC1 from shells laid by a single Thornber “ 404 “ hen. The rate of solution was inversely related to the nitrogen content at a given level but, for a given nitrogen content, was faster from the inside of the shell than from the outside. Matrix is distributed unevenly through the mineralised shell, the concentration of nitrogen and carbohydrates increasing to a maximum about two‐thirds of the way through the shell from the inside. With the exception of hexose, these concentrations then decrease to values as low, or lower than those found on the inside of the shell. The content of chondroitin sulphates in the matrix is fairly constant while that of sialic acid decreases from the inside to the outside. Non‐chondroitin sulphate polysaccharides are more highly concentrated in matrix in the inner and outer strata of the shell than in the region of maximum matrix concentration. Aspects of shell composition and calcification are discussed in relation to these results.     

A study of the progressive deposition of shell in the shell gland of the domestic hen

It is known that the organic matrix of the shell is not distributed evenly throughout the calcified portion and an experiment has been described in which incompletely shelled eggs were removed from a group of domestic hens after being in the shell gland for various known periods.

Rate of shell deposition and the timing of the deposition of the various layers in the shell has thus been determined, and it was found that shell deposition was initially slow for the first 3 h in the gland and thereafter more rapid, at a constant rate of 322 mg h^?1, until termination after 17 or 18 h. The narrow layer low in matrix at the top of the mammillary layer of the shell was laid down during the 7th to 9th hour in the gland and did not appear to be associated with any change in shell deposition rate. It was also found that the characteristic pole to pole thickness patterns of shells were present at the earliest measurable stage, that is after about 8 h in the gland.     

蛋鸡产蛋性能和蛋壳品质耐受低水平磷的应用研究

本研究选择健康49周龄褐壳蛋鸡1296只,随机分为6组,每组2个重复,每个重复108只。A为对照组,含0.20%有效磷(AP)和0.9‰植酸酶;B、C、D、E、F组分别含AP0.16%、0.12%、0.20%、0.16%和0.12%,其中B、C组添加0.9‰植酸酶。试验期8周。结果显示,试验4、6、8周和试验全期,F组比对照组产蛋率分别降低6.67%、7.78%、7.87%和6.67%,差异均显著(P0.05);各组平均蛋重和采食量无显著差异;试验6、8周和试验全期,F组料蛋比较对照组分别高10.19%、11.30%和8.00%,差异均显著(P0.05)。8周试验全期,C、D、E、F组软破蛋率和各组斑蛋率均高于对照组(P0.05);F组出现9只瘫鸡,其他各组均没有出现。提示日粮AP降至0.12%时,对蛋鸡产蛋性能、蛋壳品质自第3～4周开始产生影响。     

Studies of membrane,mammilary cores and cuticle of the hen egg shell

Mammillary cores and outer shell membrane have been studied by comparing the weight and chemical composition of outer membrane with cores still attached and of outer membrane free from cores, from the same shells. Inner membrane and cuticle have also been investigated.

The weights of inner and outer membrane are significantly correlated (P < 0.05). Both membranes contain galactosamine, glucosamine, sialic acid, glucose, mannose and fucose but in different proportions. Uronic acid is probably absent.

An average of 3.3 mg of organic mammillary core material per shell (52 μg/cm²) was present in the shells investigated, but there was a trend suggesting that eggs laid by more mature hens may contain rather more. Cores are rich in hexo‐samine, sialic acid and hexose, but the probable absence of uronic acid suggests that chondroitin sulphates play no part in shell initiation. There were no major differences in the amino acid spectrum of membrane samples with and without cores.

Galactosamine, glucosamine, sialic acid, galactose, glucose, mannose, fucose and an unidentified pentose are present in the cuticle. In some respects these sugars are combined differently from their combination in the adjacent outer levels of the matrix.     

Expression and regulation of Enpp2 in rat uterus during the estrous cycle

Ectonucleotide pyrophosphatase/phosphodiestrase 2 (Enpp2) isolated from the supernatant of human melanoma cells is a lysophospholipase D that transforms lysophosphatidylcholine into lysophospatidic acid. Although multiple analyses have investigated the function of Enpp2 in the hypothalamus, its role in the uterus during the estrous cycle is not well understood. In the present study, rat uterine Enpp2 was analyzed by RT-PCR, Western blotting, and immunohistochemistry. Quantitative PCR analysis demonstrated that uterine Enpp2 mRNA was decreased during estrus compared to proestrus and diestrus. To determine whether uterine Enpp2 expression is affected by sex steroid hormones, immature rats were treated with 17β-estradiol (E2), progesterone, or both on postnatal days 14 to 16. Interestingly, the expression of Enpp2 mRNA and protein were down-regulated by E2 in the uterus during estrus but not during proestrus or diestrus, suggesting that Enpp2 may play a role in uterine function during estrus. Enpp2 is primarily localized in the stromal cells of the endometrium during proestrus and estrus. During diestrus, Enpp2 was highly expressed in the epithelial cells of the endometrium. Taken together, these results suggest that uterine Enpp2 may be regulated by E2 and plays a role in reproductive functions during female rat development.     

鲎素抗菌肽对蛋鸡产蛋后期蛋品质及蛋壳超微结构的影响

旨在探索鲎素抗菌肽对海兰褐蛋鸡产蛋后期蛋品质及子宫CaBP-D28k mRNA表达量的影响及其机理探讨。随机选取600只385日龄海兰褐蛋鸡,分为对照组、试验组,每组各3个重复,每个重复100只鸡。对照组饲喂基础日粮,试验组在基础日粮中添加鲎素抗菌肽,鲎素抗菌肽发酵液最适添加剂量为0.01 L/kg,按饲料所需量配比添加,预饲期7 d,试验期35 d。结果显示,与对照组相比,试验组产蛋率提高8.86%(P0.05),料蛋比、破蛋率分别降低2.68%和12.76%(P0.05);试验组蛋壳强度比对照组提高了18.97%(P0.05),哈氏单位、蛋壳相对质量和蛋壳厚度均有上升的趋势;试验组蛋壳表面皴裂纹数较少,裂隙变小,纵切面结构中垂直晶体层完整,栅栏层结构整齐,上下层面平滑,栅栏层气孔数量少,乳头层乳头数目少且宽;试验组蛋鸡子宫内CaBP-D28k mRNA表达水平显著提升(P0.01)。可见,鲎素抗菌肽可提高CaBP-D28k参与产蛋过程中钙的代谢过程,调控蛋鸡体内钙代谢发挥功能作用。