温度、排蛋方向及翻蛋对鸡胚的影响

从卵子在输卵管前端受精时起到种蛋的贮存及整个孵化期间,各种环境因素都影响着鸡胚的发育。在卵产出后,畜牧业人员至少能部分地控制象温度、湿度、通风、排蛋方向及翻蛋此类的环境因素。本文对其中的温度、排蛋方向及翻蛋三个因素讨论如下: 1.种蛋孵化前的贮存合格种蛋在良好的条件下可贮存7天左右而对孵化率影响很小。

立式孵化箱不同编组排蛋方式对孵化成绩的影响

近几年来,由于立式电孵箱安全、便于操作,因而逐步代替了旧式滚桶式电孵箱。为了了解立式孵化箱不同编组排蛋方式对孵化成绩的影响,以便指导孵化生产,取得最佳经济效益。

Classification and Nomenclature of Plant Metallothionein-like Proteins Based on Their Cysteine Arrangement Patterns

随着植物基因组研究的进展 ,在基因文库和蛋白文库登录的植物类金属硫蛋白基因已超过 5 0个 ,接近金属硫蛋白总数的 1/ 3,而且有不断上升的趋势。鉴于目前植物类金属硫蛋白命名与分类随意性太大 ,很有必要建立一个统一合理的命名与分类法。对植物类金属硫蛋白一级结构进行详细分析后 ,发现该蛋白两端富含半胱氨酸的区域内半胱氨酸的排列方式颇具规律性 ,进而提出了以半胱氨酸排列方式为基础的分类及命名法 ,并阐述了采用这种方法的理由及其可行性。

Lysozyme refolding at high concentration by dilution and size—exclusion chromatography

This study of renaturation by dilution and size exclusion chromatogra phy (SEC) addition of urea to improve yield as well as the initial and final pro tein concentrations showed that although urea decreased the rate of lysozyme ref o lding, it could suppress protein aggregation to sustain the pathway of correct r efolding at high protein concentration; and that there existed an optimum urea c oncentration in renaturation buffer. Under the above conditions, lysozyme was su ccessfully refolded from initial concentration of up to 40 mg/mL by dilution and 100 mg/mL by SEC, with the yield of the former being more than 40% and that of the latter being 34.8%. Especially, under the condition of 30 min interval time, i.e. τ>2(t_R2 -t_R1 ), the efficiency was increased by 25% and the renaturation buffe r could be recycled for SEC refolding in continuous operation of downstream proc ess.

Plasminogen Activator and Plasminogen Activator Inhibitor in Mouse Ovaries during Periovulatory Periods

Changes of ovarian tPA,uPA and PA inhibitor activities were examined in PMSG-and hCG-treatedimmature mice during periovulatory periods.The results show that 15% of the gonadotropin-treatedanimals ovulated 8 hrs after hCG administration,about 6-8 hrs earlier than in rat.It is also shownthat not only tPA activity,but also uPA activity,was regulated by gonadotropins in ovarianhomogenates and granulosa cells,and they reached maximum prior to ovulation.No measurableamount of PAI-1 activity could be detected in mouse granulosa cell conditioned medium andfollicular fluid,but considerable amount of α_2-antiplasmin,a specific inhibitor for plasmin,wasfound in follicular fluid.Cumulus-oocyte complexes contain mainly tPA.Since the ovulated eggsstill have high tPA activity,it is thought that the enzyme in the oocyte may play an important rolein implantation.

Improving the sensitivity of protein microarray by evanescent-field-induced fluorescence

To improve the sensitivity of protein microarray, a prism surface replaces the surface of the common microscope slide.The protein targets arrayed on the surface are hybridized and labelled by fluorescent probes. Evanescent excitation occurs when the convergent laser reaches the surface, and a photomultiplier tube detects the emitted fluorescent signal. A two-dimensional actuator scans the whole surface to achieve planar laser excitation and fluorescence collection. The penetration depth of the evanescent field into the protein targets is only some hundred nanometers and can be controlled by different incident angle of the laser beam, so the undesired background signals are reduced dramatically and the detection sensitivity is improved by a factor of 50 to 100 comparing to confocal excitation. This approach can detect low abundance analytes without signal amplification.

Dendroaspis natriuretic peptide relaxes gastric antral circular smooth muscle of guinea-pig through thecGMP/cGMP-dependent protein kinase pathway

AIM: To systematically investigate if cGMP/cGMP- dependent protein kinase G (PKG) signaling pathway may participate in dendroaspis natriuretic peptide (DNP)-induced relaxation of gastric circular smooth muscle. METHODS: The content of cGMP in guinea pig gastric antral smooth muscle tissue and perfusion solution were measured using radioimmunoassay; spontaneous contraction of gastric antral circular muscles recorded using a 4-channel physiograph; and Ca2+-activated K+ currents (IK(Ca)) and spontaneous transient outward currents (STOCs) in isolated gastric antral myocytes were recorded using the whole-cell patch clamp technique. RESULTS: DNP markedly enhanced cGMP levels in gastric antral smooth muscle tissue and in the perfusion medium. DNP induced relaxation in gastricantral circular smooth muscle, which was inhibited by KT5823, a cGMP-dependent PKG inhibitor. DNP increased IK(Ca). This effect was almost completely blocked by KT5823, and partially blocked by LY83583, an inhibitor of guanylate cyclase to change the production of cGMP. DNP also increased STOCs. The effect of DNP on STOCs was abolished in the presence of KT5823, but not affected by KT-5720, a PKA-specific inhibitor. CONCLUSION: DNP activates IK(Ca) and relaxes guinea-pig gastric antral circular smooth muscle via the cGMP/PKG-dependent singling axis instead of cAMP/ PKA pathway.

Reduced Ammonia Release by Frequently Removal of Manure below Draining Floors in a Floor Housing System for Laying Hens

The hygienic threshold limit values for ammonia （25 ppm） for animal welfare but also for occupational safety and health is often exceeded in floor housing systems for laying hens with long time storage of manure in bins below draining floors. The major reason for high ammonia concentrations is the large amounts of stored and exposed manure. The possibility to reduce ammonia release by reducing the amount of stored manure in bins in floor housing systems for laying hens has therefore been investigated. Investigations were carried out in a climate chamber equipped with a floor housing system with a manure removal system with two parallel motor driven conveyors placed below an elevated draining floor. The conditions when manure is stored in bins below draining floors were simulated by storing manure on the conveyors for several days at constant ventilation rates and temperatures. The investigations clearly showed that storage of manure in the bin caused a rapid increase in ammonia concentrations. After about 7 days storage of manure in the bin the ammonia concentration exceeded the hygienic threshold limit values. It can be concluded that long time storage of manure in storage bins below draining floors should not be recommended. It was possible to maintain the ammonia concentration below the hygienic threshold limit values when manure was removed frequently with conveyors. Floor housing systems for laying hens with elevated draining floors should therefore be equipped with manure removal systems that enable frequent removal of manure in the bins.

Ductal adenocarcinoma of the pancreatic head:A focus on current diagnostic and surgical concepts

Complete surgical resection still remains the only possibility of curing pancreatic cancer,however,only 10% of patients undergo curative surgery.Pancreatic resection currently remains the only method of curing patients,and has a 5-year overall survival rate between 7%-34% compared to a median survival of 3-11 mo for unresected cancer.Pancreatic surgery is a technically demanding procedure requiring highly standardized surgical techniques.Nevertheless,even in experienced hands,perioperative morbidity rates(delayed gastric emptying,pancreatic fistula etc.) are as high as 50%.Different strategies to reduce postoperative morbidity,such as different techniques of gastroenteric reconstruction(pancreatico-jejunostomy vs pancreatico-gastrostomy),intraoperative placement of a pancreatic main duct stent or temporary sealing of the main pancreatic duct with fibrin glue have not led to a significant improvement in clinical outcome.The perioperative application of somatostatin or its analogues may decrease the incidence of pancreatic fistulas in cases with soft pancreatic tissue and a small main pancreatic duct(< 3 mm).The positive effects of external pancreatic main duct drainage and antecolic gastrointestinal reconstruction have been observed to decrease the rate of pancreatic fistulas and delayed gastric emptying,respectively.Currently,the concept of extended radical lymphadenectomy has been found to be associated with higher perioperative morbidity,but without any positive impact on overall survival.However,there is growing evidence that portal vein resections can be performed with acceptable low perioperative morbidity and mortality but does not achieve a cure.

Establishment and characterization of an oxaliplatin-resistant hepatic cancer cell line

Objective The aim of the current study was to establish an oxaliplatin-resistant hepatoma cell line(HepG2/OXA) and investigate the potential mechanisms of its drug resistance.Methods The hepatoma cell subline, HepG2/OXA, resistant to oxaliplatin(OXA), was established from a parent cell line HepG2, by stepwise exposure to gradually increasing concentrations of OXA over a half-year period. Chemosenstivity of the cytotoxic drugs, OXA, cisplatin(CDDP), adriamycin(ADM), and 5-fuorouracil(5-FU), was determined in HepG2 and HepG2/OXA cells, by the Cell counting kit-8(CCK8) assay. Cell cycle distribution of HepG2 and HepG2/OXA cells was analyzed by Flow cytometry(FCM). The expression levels of several drug resistance-related proteins, such as P-glycoprotein(P-gp), multidrug resistant protein 1(MRP1), and excision repair-cross complementing 1(ERCC1) protein in the two cell lines were tested by the western blot assay.Results The IC50 of OXA in HepG2/OXA and HepG2 were 136.84 μmol/L and 23.86 μmol/L, respectively. The resistance index(RI) was 5.34. HepG2 was also demonstrated to be cross-resistant to other antitumor agents, such as 5-FU, ADM, and CDDP. The percentage of HepG2/OXA cells in the S phase was significantly decreased compared to HepG2 cells(25.58% ± 2.36% vs 14.37% ± 2.54%, P < 0.05), while the percentage of cells in the G0/G1 and G2/M phases showed no statistical difference(respectively 55.29% ± 4.98% vs 56.73% ± 4.56%, P > 0.05, and 24.63% ± 4.81% vs 28.26% ± 3.82%, P > 0.05). The ERCC1 was found to be over expressed in HepG2/OXA cells, while there was no difference in the expressions of P-gp and MRP1 between the multiple drug resistance(MDR) phenotype cell line and its parental cell line.Conclusion HepG2/OXA showed an MDR ability; the over expression of ERCC1 might be associated with the platinum resistance of the cells, but P-gp and MRP1 are not.

Design of Manure Gas Exhaustion to Reduce Ammonia Concentrations in Loose-Housing Systems for Laying Hens

Uniformity of air flow in extraction openings in exhaust air channels for manure gas exhaustion is determined by the distribution of pressure. The areas required in extraction vents and in extraction ducts are determined by the uniformity of air flow desired along the duct and by the loss of pressure that can be accepted. The area ratio between the vents and the cross section of the exhaust air duct will have a strong influence on both uniformity of flow and loss of pressure. The following ventilation properties were studied： Uniformity of air flow; Variations in static pressure along a duct; Air velocity at different distances from the vents. The area ratio should be about 1 for uniform exhaustion. The studies showed that the relative variation in air velocity is independent of the level of the ventilation rate. The uniformity of the exhaust distance is influenced in about the same way by the area ratio as the air velocity in the exhaust vents. Thus, it is important that the area ratio is not too high if a good exhaust function should be guaranteed. The studies also demonstrated that the uniformity of the exhaust distance is independent of the ventilation flow rate. The exhaust ventilation range is, maximally 0.3 m from the vents. The static friction coefficient was on average 0.80.

Effect of Prolactin on Gonadotropin Regulation of Mouse Ovarian Function

We have demonstrated that prolactin inhibits gonadotropin-induced ovulation in PMSG-primed mice.The number of ova in oviducts considerably decreased in the group of hCG plus prolactin (PRL)(19.7 + 4.9) as compared with that of hCG alone (31.7 + 6.7). PRL inhibition of hCG-inducedovulation in mice may be through decreasing the ovarian plasminogen activator (PA) activity on onehand, and inhibiting the preovulatory increase in estrogen (E) secretion on the other.

Ligation of Caeca Improves Nitrogen Utilization and Decreases Urinary Uric Acid Excretion in Chickens

This study was carried out to clarify role of ceca in nitrogen nutrition of the chicken. Exp. 1： The effect of cecal ligation on nitrogen utilization and excretion was investigated in chickens fed 5% to 14% protein diet. Irrespective of dietary protein level and different protein sources, the ligation of cecal decreased uric acid excretion and tended to increase nitrogen utilization and balance with the exception of urea-added 10% protein diet. Exp. 2： The effect of cecal ligation on nitrogen utilization and excretion was investigated in conventional and colostomized chickens fed a 5% protein diet or 5% protein diet plus urea. Total nitrogen excretion and uric acid excretion increased by colostomy were depressed by cecal ligation in chickens. Therefore, nitrogen utilization and balance decreased by colostomy were increased by cecal ligation. Urinary nitrogen excretion was significantly decreased by cecal ligation in colostomized chickens, but the cecal ligation did not change fecal nitrogen excretion in chickens fed either diet. Exp. 3： This experiment was carried out to examine the effects of removal of cecal contents on nitrogen utilization, balance and nitrogen excretion in cecally ligated chickens. Total nitrogen excretion was significantly decreased by washing out the cecal contents with saline or antibiotics. The cecal ligation and the removal of cecal contents significantly decreased uric acid excretion in the excreta. There was a highly inverse relationship between microbial counts in the ceca and an excretory amount of uric acid. Exp. 4： In order to examine effects of cecal ligation on microbial activity, microbes were counted and products of microbial fermentation were determined. The ligation of caeca decreased microbial counts, concentrations of acetic, propionic and butyric acids and ammonia concentration of cecal contents. These results suggest that nitrogen metabolism in chickens is affected by possible changes in cecal fermentation caused by preventing the substances from urine and digesta from entering into the ceca.

The Synergistic Effect of HT and Complement Regulatory Proteins in Resisting the Immunological Rejection of Heterogenic Transplantation

Objective：To establish the polytransgenic mice expressing the human HT and complement regulatory proteins （CRPs） and discuss their ability to resist the hyperacute rejection （HAR） and delayed xenograft rejection （DXR） of heterogenic transplantation. Methods ：Transgenic mice were produced by microinjection to construct gene for human HT, delay acceleration factor （DAF） and/or CD59 into the male pronucleus of zygote. PCR and Southern blot were used to screen the positive trarisgenic mice. Flow cytometry （FCM） was used to detect the expression of HT, ct-Gal and DAF or CD59 on the PBMCs of transgenic mice. The survival time and function of the heart of transgenic mice were determined by a modified Langendorff cardiac perfusion apparatus： The change of proteinosis on IgM,IgG, C3c and C9 from different cardiac vascular iendothelial cells of transgenic mice were detected by immunohistochemistry. Results：HT, DAF or CD59 were highly expressed on the positive transgenic mice by FCM. The deposition of IgM,IgG,C3c or C9 in the cardiac vascular endothelial cells of the positive transgenic mice were de- creased. The survival time and function of the heart of the co-transgenic mice with AB serum perfusion were significantly longer and higher than that of the single HT positive transgenic mice（P 〈0.05）. Conclusion ：The mice co-expressing HT/DAF or HT/CD59 could resist the HAR,which was better than those expressing HT alone. It is feasible to use HT and CRPs co-transgenic methods to resist the HAR and DXR.

Biased Immunoglobulin Genes Rearrangement in Mantle Cell Lymphoma： Hints to Identify the Normal B-cell Counterpart

Mantle cell lymphoma （MCL） is an aggressive nonHodgkin＇s lymphoma, originating from naive B-cells. The blastoid MCL tumors often show complex cytogenetic aberrations. In this review, we summarized the data available on immunoglobulin heavy-chain （IgH） genes rearrangement for their importance in suggesting the MCL normal counterpart B-cell. Some new data suggesting an antigen selection process were also presented in this review.

Polymerase chain reaction-single strand conformational polymorphism analysis of rearranged during transfection proto-oncogene in Chinese familial hirschsprung's disease

AIM: To investigate the relationship between mutations of rearranged during transfection (RET) proto-oncogene and Chinese patients with Hirschsprung's disease (HD), and to elucidate the genetic mechanism of familial HD patient at the molecular level.METHODS: Genomic DNA was extracted from venous blood of probands and their relatives in two genealogies.Polymerase chain reaction (PCR) products, which were amplified using specific primers (RET, exons 11, 13, 15and 17), were electrophoresed to analyze the single-strand conformational polymorphism (SSCP) patterns. The positive amplified products were sequenced. Forty-eight sporadic HD patients and 30 normal children were screened for mutations of RET proto-oncogene simultaneously.RESULTS: Three cases with HD in one family were found to have a G heterozygous insertion at nucleotide 18 974 in exon 13 of RET cDNA (18 974insG), which resulted in a frameshift mutation. In another family, a heterozygosity for T to G transition at nucleotide 18 888 in the same exon which resulted in a synonymous mutation of Leu at codon 745 was detected in the proband and his father. Eight RET mutations were confirmed in 48 sporadic HD patients.CONCLUSION: Mutations of RET proto-oncogene may play an important role in the pathogenesis of Chinese patients with HD. Detection of mutated RET proto-oncogene carriers may be used for genetic counseling of potential risk for HD in the affected families.

Characterization of the Mycobacterium Tuberculosis Protein Rv2302

The Mycobacterium tuberculosis protein Rv2302 has been characterized using nuclear magnetic resonance （NMR） and circular dichroism （CD） spectroscopy. While the biochemical function of Rv2302 is still unknown, recent microarray analyses show that Rv2302 is upregulated in response to starvation and overexpression of heat shock proteins and, consequently, may play a role in the biochemical processes associated with these events. Rv2302 is a monomer in solution as shown by size exclusion chromatography and NMR spectroscopy. CD spectroscopy suggests that Rv2302 partially unfolds upon heating and that this unfolding is reversible. Using NMR-based methods, the solution structure of Rv2302 was determined. The protein contains a five-strand, antiparalle β-sheet core with one C-terminal α-helix （A61 to A75） nestled against its side. Hydrophobic interactions between residues in the α-helix and β-strands 3 and 4 hold the α-helix near the β-sheet core. The electrostatic potential on the solvent-accessible surface is primarily negative with the exception of a positive arginine pocket composed of residues R18, R70, and R74. Steady-state {^1H}-^15N heteronuclear nuclear Overhauser effects indicate that the protein＇s core is rigid on the picosecond timescale. The absence of amide cross-peaks for residues G 13 to H 19 in the ^1H-^15N heteronuclear single quantum correlation spectrum suggests that this region, a loop between β-strands 1 and 2, undergoes motion on the millisecond to microsecond timescale. Dali searches using the structure closest to the average structure do not identify any high similarities to any other known protein structure, suggesting that the structure of Rv2302 may represent a novel protein fold.

Accumulation of ciprofloxacin and lomefloxacinin fluoroquinolone-resistant strains of Escherichia coli

Objective To evaluate the role of outer membrane protein (Omp) F-deficiency and active efflux in the accumulation of hydrophilic fluoroquinolones ciprofloxacin (CPLX) and lomefloxacin (LMLX) in resistant E. coli strains. Methods Fluoroquinolone accumulation in bacteria and the effect of active efflux were measured by a fluorescence method. The outer membrane proteins of the bacteria were analysed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). E. coli strains in this study included control strains JF701 and JF703 that are OmpC- or OmpF-deficient mutants of E. coli K-12, respectively, and the fluoroquinolone susceptible strain the fluoroquinolone susceptible strain of Escherichia coli (Ecs) and its in vitroselected resistant strains R2 and R256, and the clinical resistant isolates R5 and R6. Results The steady-state accumulation concentration of each drug in Ecs appeared to be the same as in JF701, while in the OmpF- deficient strain JF703, it was 1/5 CPLX or 1/2 LMLX lower than that in JF701, but JF703 was still susceptible to fluoroquinolones. On the other hand, compared with susceptible strains, a 2- to 10-fold decrease in the accumulation of each drug was found in the resistant strains except R2, in which the accumulation was slightly higher than in JF703. After the addition of 2,4-dinitrophenol (DNP), accumulation of each drug increased, especially in resistant strains, indicating that the function of the active efflux (pump) system in these bacteria had been enhanced dramatically. Furthermore, both OmpF and OmpC in Ecs, OmpF-deficiency in R2 and R256 and OmpC-deficiency in R5 and R6 were observed.Conclusion The decreased accumulation of hydrophilic fluoroquinolones in E. coli involved OmpF-deficiency and active efflux (pump), and the latter may be an important factor.

Adaptor protein APPL1 interacts with EGFR to orchestrate EGF-stimulated signaling

Epidermal growth factor receptor （EGFR） mediates multiple signaling pathways that regulate cell migration, proliferation, and differentiation. Adaptor pro- tein APPL1 has been reported to function as a downstream effector of EGFR signaling pathway. However, molecular mechanisms underlying the role of APPL1 downstream of EGFR signaling remains elusive. Here, we identified APPL1 as a critical molecule that interacts with EGFR. Suppression of APPL 1 by siRNA inhibited EGF-stimulated Akt phosphorylation. Functionally, EGF stimulation of cells caused phosphorylation of APPL1 at Ser636, which subsequently promoted the interaction between APPL1 and EGFR, indicating that APPL1 sensitizes EGF stimulation by acting at a site downstream of the EGFR signaling. Importantly, non-phosphorylatable mutant of APPL1 reduced cell migration compared with wild-type APPL1 in an Akt-dependent manner. Our study reveals a novel function of APPL1 in EGF signaling and defines a novel molecular mechanism by which phosphorylation of APPL1 upon EGF stimulation regulates cell migration underlying EGF-stimulated Akt pathway.

RLKs orchestrate the signaling in plant male-female interaction

Different from animals, sessile plants are equipped with a large receptor-like kinase(RLK) superfamily. RLKs are a family of single trans-membrane proteins with divergent N-terminal extracellular domains capped by a signal peptide and C-terminal intracellular kinase. Researches in the last two decades have uncovered an increasing number of RLKs that regulate plant development, stress response and sexual reproduction, highlighting a dominant role of RLK signaling in cell-to-cell communications. Sexual reproduction in flowering plants is featured by interactions between the male gametophyte and the female tissues to facilitate sperm delivery and fertilization. Emerging evidences suggest that RLKs regulate almost every aspect of plant reproductive process, especially during pollination. Therefore, in this review we will focus mainly on the function and signaling of RLKs in plant male-female interaction and discuss the future prospects on these topics.