Acta Veterinaria et Zootechnica Sinica

Relationship between Polymorphism of HSD11B1 and MyoG Gene and Gestation Length in Pigs

MA Zhi-gang;LI Jia-lian;GUO Wan-hu;PAN Wan-ping;WANG Jia-sheng;HAN An-qin;DING Shan-he;JIANG Si-wen

2008, 39(6): 689-695. doi:

Abstract ( 1570 )

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11βhydroxysteroid dehydrogenase-1（HSD11B1）and myogenin（MyoG）were selected as candidate genes influencing on gestation length in pigs. The present study analyzed relationship between HSD11B1 and MyoG genes polymorphisms by PCR-RFLP and gestation length in 7 pig populations. Results showed that the frequencies of the A allele of HSD11B1 gene based on PCR-Bsh1236Ⅰ-RFLP analysis were 0.538, 0.752, 0.522, 0.941, 1.000, 0.889 and 0.594, respectively in Qingping, New Qingping line, Large White, Landrace, Duroc, DIV1 and DIV2 of New line of Chinese lean-type. Except for DIV2 line, there was no any significant differences between genotypes of HSD11B1 and gestation length (P>0.05). The frequencies of the M allele of MyoG gene based on PCR-MspⅠ-RFLP analysis were 1.000, 0.534, 0.370, 0.115, 0.094, 0.382 and 0.243, respectively in Qingping, New Qingping line, Large White, Landrace, Duroc, DIV1 and DIV2 of New line of Chinese leantype. The Qingping pig breeds only presented genotype MM, the genetic polymorphisms of the locus were detected in the other populations. The gestation length of genotype MM is less than that of genotype NN and the difference is significant in multiple parity Landrace（P<0.01）.

Association Analysis of Bovine ACAA1 and ACAA2 Gene Polymorphisms with Production Traits

LI Heng-de;WANG Shu-hui;XU Shang-zhong;GAO Xue;REN Hong-yan;CHEN Jin-bao

2008, 39(6): 696-700. doi:

Abstract ( 1278 )

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178 cattle from five breeds, Luxi, Qinchuan, Angus, Hereford and crossbreed of Simmental and Menggolian were used in the experiment. Through allelespecific polymerase chain reaction (AS-PCR), two single nucleotide polymorphisms (SNP): A1066G (GenBank Accession No. EF576938) and A1452G were found on ACAA1 gene. A1066G SNP is located at exon 9, but it is a silent mutation, while A1452G SNP is located at 3′-untranslated region (UTR). C33119T SNP (GenBank Accession No. EF583005) was found at intron 8 of ACAA2 gene. No haplotype was found between these two SNPs of ACAA1 gene. Results of association analysis of these SNPs with production traits showed that A1066G substitution of ACAA1 is associated significantly (P<005) with loin-eye area, the mean value of genotype AA is the highest, genotype AG second highest and genotype GG the lowest. A1452G substitution is associated significantly (P<0.05) with loin-eye area, the mean of genotype GG is the highest, genotype AG second highest and genotype AA the lowest. The C33119T substitution is associated significantly with daily gain (P<0.01) and loineye area (P<0.05), for daily gain, the mean of genotype CT is the highest, genotype TT second highest, and genotype CC the lowest; for loineye area, the mean of genotype CC is the highest, genotype TT second highest, and genotype CT the lowest.

Genetic Variation of Myostatin Gene in 4 Bos Species in China

JI De-jun;CHANG Hong;CHANG Chun-fang;GENG Rong-qing;LI Yong-hong

2008, 39(6): 701-704. doi:

Abstract ( 1018 )

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Through designing the primers to the three exons of the myostatin (MSTN) gene and using the methods of PCR amplification and sequencing, variations of the MSTN gene were detected in 4 Chinese Bos species, normal cattle (Mongolia cattle), zebu(Leiqiong cattle), yak (Bayingolin), and gayal (Dulong gayal). The results showed that 375，372，381 base pairs were detected respectively in the exon 1, 2 and 3 of the MSTN gene, and 7 nucleotide polymorphic sites were observed with 17 haplotypes in the 4 Bovinae populations. The nucleotide diversities within the 4 species (Pi) ranged from 0.000 36 to 0.001 03, and between the 4 species （Dxy）ranged from 0.000 76 to 0.003 22, indicating a low intrapopulation genetic diversity. Construction of the phylogenic tree based on these MSTN haplotypes indicated that yak was diverged earlier than yellow cattle (normal cattle and zebu); gayals were clustered into two groups, probably due to hybridization.

Polymorphisms of Y Chromosome Microsatellites of Seven Cattle Breeds from Southwest China

CAI Xin

2008, 39(6): 705-708. doi:

Abstract ( 888 )

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Polymorphisms of four Y chromosome microsatellites were analyzed among 81 bulls from 7 indigenous cattle breeds from southwest region of China. Only two out of the five microsatellites exhibited polymorphisms in the bulls analyzed. UMN 2404 showed specific Bos taurus (taurine) (104, 91 bp) and Bos indicus (indicine) (120, 110, 85 bp) haplotypes in these breeds, similarly, UMN 0103 also displayed specific taurine (155, 140 bp) and indicine (136, 125 bp) haplotypes consistent with UMN2404 in each bull examined. Based on the investigation of UMN2404 and UMN0103, we revealed the distribution and genetic introgression of male indicine and taurine in southwest region of China. The frequency of indicine Y specific haplotype (72.8%) was significantly higher than that of taurine haplotype (27.2%). The taurine haplotype dominated in Tibetan (100%) and Diqing cattle (81.8%), while the indicine dominated in the other breeds (76.5%-100%).

Genetic Structure and Diversity Study on Five Goat Breeds

WU Yan-ping;MA Yue-hui;HAN Jun-wen;ZHAO Qian-jun;HE Xiao-hong;PU Ya-bin;GUAN Wei-jun;CHU Ming-xing;LI Kui

2008, 39(6): 709-714. doi:

Abstract ( 864 )

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The genetic diversity of five goat breeds was surveyed using twenty-ive microsatellite loci. 199 alleles were gotten in all the loci，and the HardyWeiberg equilibrium analysis revealed that almost the 5 breeds in the 25 loci showed a remarkable genetic disequilibrium. The PIC value (0.93) were less than 0.5 in BMS1943, the other loci were high diversity. The mean heterozygosity of different locus varied from 0.436 to 0.820,and of different population was from 0.592 to 0.738.The F-statistic analysis indicated that the Fst of different locus varied from 0.011(in BMC1206) to 0.078 (in BMS1943). The Nm was from 2.972 to 23.115,and the mean value was 8.884. The result of the cofficient of gene differentiation showed it was not remarkable differentiation in all the breeds.Adopted the method of UPGMA with the Dcoa genetic distance in the 5 goat breeds ，the dendrograms were drawed，Hanshan White Cashmere goat and Wuzhumuqin Cashmere goat were in the close relationship each other.

The Statistical Analysis on the Effect of the SNP in PGC-1α on Meat Quality in White Plymouth Rock

AN Jian-yong;CHEN Liao;HOU Zhuo-cheng;WU Gui-qin;XU Gui-yun

2008, 39(6): 715-720. doi:

Abstract ( 1471 )

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The peroxisome proliferators-activated receptor-γ coactivator-1α (PGC-1α) plays a prominent role in energy metabolism and adipogenesis. A single nucleotide polymorphism (SNP) from G to A was found at position 646 in its open reading frame which causes an Asp216Asn amino acid substitution. A Mendelian segregated population of White Plymouth Rock was designed to study its effect on meat quality. Breast muscle was collected from these birds at 8 wks old and the meat quality was measured by the TA-XT2 texture analyzer. There after, least square model was established for the sake of statistical analysis. The significant differences were found among the three genotypes and the AG genotype had the better meat quality. Hardness, cohesiveness, gumminess, chewiness, resilience had significant differences among the groups (P<0.05). And there were significant correlations between rigidity and flexibility, viscosity and tenderness. Principal components analysis showed six components reflected the meat quality thoroughly. Hardness, freshness and adhesiveness were the first three characteristics in the components.

Impacting of Insulin and Prolactin on Mammary Gland Epithelial Cell Line

TONG Hui-li;GAO Xue-jun;LI Qing-zhang;YAN Yun-qin

2008, 39(6): 721-725. doi:

Abstract ( 945 )

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Mammary gland epithelial cells which have lactating function can be a cellular mode of research of mammary gland development and lactating regulation mechanism. In this paper we studied the lactating function of dairy goat mammary gland epithelial cell line. SDS-PAGE, testing of triglyceride content and HPLC were used for detection of total proteins, milk fat and lactose respectively reflected lactating function of both insulin and prolactin impacting on dairy goat mammary gland epithelial cells. Results show that dairy goat mammary gland epithelial cells can excrete milk fat, milk proteins and lactose cultured in DF12 fluid after adding EGF, IGF-1, ITS and FBS. Lactating function of mammary gland epithelial cells has no obvious change after 48 h treated by insluin. However, treatment of prolactin can obviously raise the level of secretion of milk proteins and lactose. A mammary gland epithelial cell line which has lactating function has been established. This result is convenient for the study of lactation mechanism of dairy goats, dairy cows and other economic animals.

Establishment and Characteristics of Tianzhu White Yaks Kidney Explant Fibroblast

FENG Ruo-fei;MA Zhong-ren;GUAN Wei-jun;LI Ming-sheng;QIAO Zi-lin;FENG Yu-ping;ZHOU Xue-yan

2008, 39(6): 726-732. doi:

Abstract ( 1001 )

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The Tianzhu White yaks kidney explant was cultured with trypsinization and then subcultured．Observations on morphology，dynamic growth，fluorescin plasmid transfection and expresstion，and analysis of karyotype，isoenzyme of lactate dehydrogenase were carried out．The results showed that population doubling time of cells (PDT) was 27.2 h；the chromosomes were 60 and diploid cells were dominant of 74%．The banding patterns of the isoenzyme of lactate dehydrogenase had significant difference.As follow，the LDH5 was higher and stronger；the exogenous plasmid could copy and express in the cell，tests for bacteria，fungi and yeasts，virus and mycoplasma were negative．Those showed that the cell line was successful，and it make the Tianzhu White yaks，a national important genetic resource preserved in cell level，as well as will provide an ideal experimental material for genetic studies．

Effect of Thiram on Growth Performance and Histopathologic Changes of Tibial Dyschondroplasia in Broiler

TIAN Wen-xia;LI Jia-kui;BI Ding-ren;ZHANG Yan-hong;QIN Ping

2008, 39(6): 733-738. doi:

Abstract ( 961 )

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The experiment was conducted to research the effect of thiram on growth performance and histopathologic changes of tibial dyschondroplasia in broilers. 160 one-day-old AA broilers were randomly allotted into two treatments after raised seven days, they were given a regular diet (control group), or the same diet containing thiram 100 mg/kg for 96 h, then control and experimental group broilers were given the same regular diet. The feeding experiment was ended at 23rd days. ADG, F/G, TD score and TD incidence were studied at 4th,9th,16th, 23rd day. Pathological and histopathologic changes of growth plate were also observed at different stages. The results showed that TD incidence was 94.44% at 4th day. Body weight was decreased at 4th and 9th day, and feed to gain ratio (F/G) was increased at 4th day. The pathological symptoms were typical but also more severe than those of spontaneous TD. Through every stage observing with microscope, we found that the thiram promoted chondrocyte proliferation, resulted in large numbers of death cells, compressed blood vessels which led to necrosis of blood vessel, hindered extracellular matrix synthesis, deferred endochondral calcification, formed cartilage plug finally because of accumulated prehypertrophic chondrocytes. As a result, it was demonstrated that thiram could decline growth performance and elevate TD incidence significantly, lead to pathological and histopathologic changes in growth plate, which were similar to those of spontaneously occurring TD. So broiler TD induced by thiram is an ideal animal model for studying TD development and selecting effective different micronutrients against TD.

Effect of Lactic Acid Bacteria Inoculants Applying to Mixed Gramineous Grass on Fermentation Characteristics

LIU Han-lu;GUI Rong;TA Na

2008, 39(6): 739-745. doi:

Abstract ( 1473 )

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The study was conducted to investigate the effect of applying Lactobacillus plantarum and Enterococcus mundtti that were isolated from ensilaging materials on fermentation characteristic of mixed gramineous grass Elymus excelsus and Elymus sirbiricus. There were four treatments as followed: the control (no additives) (Con), the E.mundtii (Em), the L.plantarum (Lp) and the E.mundtii+L.plantarum (EL). All the strains were diluted with distilled water, and then sprayed on ensilaging materials at 1×105cfu/g fresh matter(FM). Then about 530 g grasses was packed into a plastic bottle and sealed. Each treatment was carried out 8 repetitions. The bottle silos were kept at room temperature. After 60 days, the bottles were opened and the fermentation quality of silages was determined. The results showed: (1) All LAB inoculants could significantly (P<0.05) reduce pH value compared with the control.(2) All LAB inoculants could evidently (P<0.05) increase the content of Lactic acid(LA) and the ratio of Lactic acid to acetic acid(AA) in silage, especially the EL; and distinctly (P<0.05) decrease the content of propionic acid(PA) and butyric acid(BA), compared with control. (3) The silage inoculated with EL had the lowest concentration of ammoniaN (20.85 μmol/g FM). (4) All LAB inoculants could reduce dry matter loss , Em，Lp，EL, respectively , 38.4%，43.4%，23.4%. As a result, applying LAB can indeed improve silage fermentation quality. Synthetically thought, EL treatment was the best one.

Detection of the Interference to the Reproduction of CSFV by PK-15 Cell Strains Transcribing shRNA Targeted to the 3′-UTR

GUO Kang-kang;XU Hao;ZHANG Yan-ming;ZHANG Wei-min;YE Gui-sheng;XIONG Kui-zhou;XIE Lin-hong;DANG Ru-yi

2008, 39(6): 746-751. doi:

Abstract ( 980 )

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The P-1(114-132 sites), P-2(136-154 sites) and P-3(209-227 sites) PK-15 cell strains transcribing shRNA targeted to the classical swine fever virus( CSFV) 3′-UTR were infected with CSFV shimen strain respectively.The NS3 and β-actin gene of infected cells were detected by semi-quantitive reverse transcription polymerase chain reaction(RT-PCR) at the 72nd and 96th hour post-infection, to study the influence of transcribing shRNA on the reproduction of CSFV at RNA level. At the same time ,the NS3 protein of cells was detected by enzyme linked immunosorbent assay(ELISA) at the 72nd, 96th and 148th hour post-infection to estimate the influence at protein level by comparing the optical density at 490 nm.Results showed that:(1) The NS3 genes of P-1,P-2 and P-3 cell strains decreased obviously than that of PK-15 cells(positive control) at the 72nd and 96th hour post-infection, respectively(P<0.05) ,it showed the transcribed shRNA could interfere the reproduction of CSFV at RNA level.(2)The expressing quantity of NS3 protein of P-1,P-2 and P-3 cell strains were obvious less than that of PK-15 cells(positive control) at the 72nd and 96th hour postinfection, respectively(P<0.05), indicating that the transcribed shRNA could interfere the reproduction of CSFV at protein level, but the discrepancy were not obvious at the 148th hour postinfection.

Development of Rea1-Time Fluorescent Quantitative PCR Assay to Detect H1N1 Subtype Swine Influenza Virus

DUAN Ting-yun;CHEN Hong-ying;CUI Bao-an;WEI Zhan-yong;LI Xin-sheng;WANG Xue-bin

2008, 39(6): 752-756. doi:

Abstract ( 1486 )

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A pair of primers and a TaqMan probe were designed according to NP gene nucleotide sequence (DQ889686) of H1N1 subtype swine influenza virus (SIV) in GenBank. NP gene was amplified by RT-PCR from RNA extracted from allantoic fluid infected by HN407 isolate of SIV. The PCR product was cloned into pGEM-T Easy vector and sequenced. The positive recombined plasmid was used as a positive quantitative template to establish a standard curve. By optimizing the probe’s concentration, Mg²⁺concentration, primers concentration and the annealing temperature, rea1-time fluorescent quantitative PCR was established. Clinical detection of H1N1 subtype SIV by rea1-time fluorescent quantitative PCR showed that the constructed method paved the way for the early and rapid detection of SIV and quantitative analysis for the infect degree of SIV.

Localization of Linear B-cell Epitopes on Goose Parvovirus Non-structural Protein and Structural Protein

YU Tian-fei;;QIU Zheng;MA Bo;YUAN Shuai-zhen;;LI Li;WANG Jun-wei

2008, 39(6): 757-763. doi:

Abstract ( 1574 )

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The nonstructural protein and structural protein of goose parvovirus were dissected into 15 or 19 fragments, respectively, and expressed in Escherichia coli. Then Western blot reactivity of these short peptide fused proteins were surveyed by using viral infected sera. The results showed that the nonstructural protein linear B-cell epitopes are located on the C-terminal (453-627 aa) and the structural protein linear Bcell epitopes are located on 35-198, 423-491, 531-595, 616-669, 678-732 aa.

Studies on the Pathomorphological Development of Experimental Duck Viral Swollen Head Hemorrhagic Disease

LI Chuan-feng;CHENG An-chun;WANG Ming-shu;SHEN Chan-juan;ZHANG She-yu;ZHOU Yi;CHEN Xiao-yue;

2008, 39(6): 764-770. doi:

Abstract ( 890 )

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72 Ying-Tao-Gu ducks of 28-day-old were infected with duck viral swollen head hemorrhagic disease virus (DSHDV) by muscle injection. The sick ducks and the dead ducks were dissected at different time to study the changes of histology and ultrastructure of tissues by using light microscopic techniques and electron microscopic techniques. The results of light microscopic observation: At the early stage of infection (2-72 h), the congestion, hemorrhage and the infiltration of inflammatory cells in different number were found in tissues. At the last course of the disease (119-170 h), necrotic lesions of different sizes were seen in immune organs (spleen, thymus, bursa of Fabricius and gland of Harder), digestive organs (liver and pancreas), kidney and cerebrum tissues, digestive tract (oesophagus, stomachus glandularis and all intestinal canals) and windpipes epithelium mucosaes completely suffered with necrosis and desquamation, lamina propria showed severe congestion and hemorrhage; Diffuse hemorrhage of heart as well as engorgement of the small vessels of lung and pectoralis were seen clearly. The results of electron microscope observation: The main ultrastructural changes were the cavitation of cell nucleus, swelling and expanding of all kinds of organelles, even some dissolved and disappeared. The Occlusionbody was formed in cytoplasm and a large amount of virions filled in it. The results maked it clear that bursa of Fabricius, thymus, spleen, liver, tractus alimentarius and windpipes were the target organs that DSHDV attacked mainly. The main target cells that DSHDV attacked were hepatocyte, lymphocyte, collagenoblast, bile duct endothelial cell and digestive tube endothelial cells.

Research of Dissociation Culture and Subculture of Epithelium Cells from Swine Bronchia

SUN Pei;ZHANG Yan-ming;YANG Xiao-yun;WEN Yuan-peng;ZHANG Hao;HE Jian-feng;GUO Kang-kang;DANG Ru-yi

2008, 39(6): 771-777. doi:

Abstract ( 945 )

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By the method of douche, trachea-bronchia epithelium was digested overnight and separated. The digestion products of the two enzymes, Trypsin and Pronase, were compared. Different cell culture mediums which including different cell factors and serum concentration were set up in order to select optimal trachea-bronchia epithelium culture technology which is economical and easy to be repeated. Through immunohistochemistry of cell No.8/18 keratins and testing cell program death with flow cytometer, the maximal number of possible cell generation was measured. The research results indicated that by the method of Trypsin digestion only a small quantity of cells were gotten and the acquired cells were very difficult to conglutinate and grow, and by Pronase digestion a number of cells were obtained which is pure and highly active. To the system of basic trachea-bronchia epithelium culture the use of low serum concentration，transferrin and epidermal growth factor are enough to assure primary trachea-bronchia epithelium and its subculture to grow well. The trachea-bronchia epithelium separated by us could be successively propagated six times and keep active well and pure highly, which may provide ideal research material and basis on technology for study of immortalisation of swine trachea-bronchia epithelium.

Apoptosis and Relative Factors Expression of Endothelial CellsInduced by Deoxynivalenol

LIU Xiu-fang;ZHANG Ai-hua;ZHANG Hong-ying;HE Cheng-hua;FAN Yan-hong;ZHANG Hai-bin

2008, 39(6): 778-783. doi:

Abstract ( 902 )

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Deoxynivalenol (DON), a.k.a. vomitoxin, is a kind of mycotoxin mainly produced by the fungi Fusariu culmorum and F. graminearum while growing on crops. It is a trichothecene with relatively low toxicity. In the present experiment, five different concentrations of deoxynivalenol (DON) were added to culture PIEC and HUVEC cell lines for 24 h. Then the morphological changes of the cell lines were observed under microscope. The MTT method was performed to determine the inhibition ratio on proliferation of PIEC and HUVEC cell lines. And TUNEL method was used to assay the apoptotic rate of these cell lines. Immunohistochemical method was employed to measure the expression of Caspase-9 and Bax in apoptototic PIEC and HUVEC cell lines. The results showed that different concentration of DON can inhibit the proliferation of these cell lines and higher concentration can alter the morphology of PIEC and HUVEC cell lines. Different concentration of DON induced apoptosis of the two cell lines, for PIEC cell line, the apoptotic ratio reached the highest level at 0.5 μg/mL (71.74%), and HUVEC at 5 μg/mL (26.61%). Then the apoptotic ratio decreased with increase of DON concentration, which may result from the collapse of apoptotic cells and their falling from the cell slice. The results implied that the Caspases pathway may contribute to the apoptosis of PIEC induced by DON, and the Bax may promote the process of the apoptosis. The changes of Caspase-9 and Bax were undetectable in the process of apoptosis of HUVEC cell line, which implied that the process did not relate to the Caspases pathway and Bax.

Development of Real-time RTPCR for Detecting the Expression Level of CD4 and CD8 mRNA in Chicken

YUE Hua;HUANG Xing;YANG Fa-long;LI Ming-yi;FAN Gen-cheng;MA Li;TANG Cheng

2008, 39(6): 784-790. doi:

Abstract ( 956 )

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Leukocyte differentiation antigens CD4 and CD8 employed important role in avian immune response and signal transduction. In present study, a SYBR Green I based real-time reverse-transcription PCR (RRT-PCR) for quantitative analysis of chicken CD4 and CD8 mRNA was developed and the expression level of CD4 and CD8 mRNA were evaluated in 2650 days old commercial chickens by using this method. The following results were obtained: The amplification efficiency of RRT-PCRs for CD4 and CD8 were 93% and 91%, respectively; The linear range of relative quantitative standard curve was 10-4 to 10-9 and 10-3 to 10-9 with good correlation coefficient of 0.998 0(y=-3.49x+27.29) and 0.999 9(y=-3.54x+31.48), respectively; The detection limit was 105 copies per reaction for CD4 and 120 copies per reaction for CD8; The melting curve presented a single peak at (78.05±0.15)℃ and (86.50±0.20)℃ for CD4 and CD8 gene respectively. The intra-assay variability were 1.13%-2.15% for CD4 and 1.17%-3.68% for CD8, the inter-assay variability were 1.16%-3.25% and 1.66%-2.86%.CD4 and CD8 mRNA levels in PBL from 26.50 days old commercial chickens were slightly fluctuated, which is accord to the results obtained by using flow cytometry. The results in this study indicated that the RRT-PCR have advantages of high sensitivity, repeatability and reproducibility, and provided a novel way for accurate quantification of CD4 and CD8 mRNA expression in chickens.

Effect of Sodium Fluoride on the Expression of Osteoprotegerin to Receptor Activator of NF-kappa B Ligand mRNA in Mouse Osteoblasts in vitro

WANG Jian-fang;ZHANG Guo-zhong;WU Pei-fu;QU Wei-jie;ZHAO Ji-xun;HAN Bo;ZHAO De-ming

2008, 39(6): 791-796. doi:

Abstract ( 912 )

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The receptor activator of nuclear factor kappa B ligand (RANKL) and its decoy receptor, osteoprotegerin (OPG), play an important role in bone metabolism, which directly control osteoclastogenesis. Here we investigate the expression of OPG and RANKL mRNA using an in vitro osteoblast culture assay. The osteoblasts were derived from mouse calvarial bone and isolated by complete trypsinization and collagenase digestion and cultured in DMEM supplemented with 15% FBS at 37℃ containing 5% CO₂, and the osteoblasts were exposed to the different concentrations of sodium fluoride (0, 10^-12, 10^-11, 10^-10, 10^-9 and 10^-8 mol/L) for 40 h at 37℃ containing 5% CO₂, and the total RNA was extracted and reverse transcribed into firststrand cDNA. The cDNA was amplified using OPG, RANKL and β-actin primers by real-time fluorescent quantitative PCR (FQ-PCR) in DNA Engine Opticon^TM2 continuous fluorescence detection system. The results indicated that OPG mRNA expression of mouse osteoblasts in sodium fluoride treated groups were upregulated and the RANKL mRNA expression also increased excepting the group of 10^-12 mol/L sodium fluoride. When exposed to sodium fluoride of 1.0×10^-12 to 1.0×10^-8 mol/L, the relative expression level of OPG/RANKL mRNA was also up-regulated. Particularly the ratio of OPG/RANKL increased significantly from 1.0×10^-12 to 1.0×10^-10 mol/L sodium fluoride(P<0.01). Thereafter, it was decreased gradually from 1.0×10^-10 to 1.0×10^-8 mol/L sodium fluoride(P<0.05). In conclusion, these in vitro data demonstrated that traces of sodium fluoride could decrease osteoclastogenesis.

Pathological Study of Effect of Dietary High Copper on the Thymus in Ducklings

YANG Fan;ZHAO Li;PENG Xi;DENG Jun-liang;CUI Heng-min

2008, 39(6): 797-803. doi:

Abstract ( 986 )

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The experiment was conducted with the objective of examining the effects of high copper on the thymus in the duckling by the methods of experimental pathology and flow cytometry (FCM). 360 one-day-old Tianfu ducklings were randomly divided into six groups, and fed on diets as follows: Controls (Cu 8 mg/kg) and high copper (Cu 100 mg/kg, high copper group Ⅰ; Cu 200 mg/kg, high copper group Ⅱ; Cu 400 mg/kg, high copper group Ⅲ; Cu 600 mg/kg, high copper group Ⅳ; Cu 800 mg/kg, high copper group Ⅴ) for six weeks. The results showed that histological changes were observed in high copper copper groups Ⅲ, Ⅳ and Ⅴ in comparison with those of control group. The weight and the growth index of the thymus were lower in high copper groups Ⅲ, Ⅳ and Ⅴ than that in control group (P<0.05 or P<0.01). The G0/G1 phase of cell cycle of the thymus was increased, and proliferating index (PI) was decreased in group Ⅳ and Ⅴcompared with those of control group（P<0.05 or P<0.01）. Percentage of apoptotic lymphocytes in the thymus was higher in high copper groups Ⅲ, Ⅳ and Ⅴ than that in control group (P<0.05 or P<0.01). It was concluded that dietary copper in excess of 400 mg/kg inhibited the development of the thymus, caused pathological changes in the thymus and impaired the function of cellular immunity in ducklings.

The Dynamic Changes of Serum TNF-α, IL-2 and β-EP Concentrations after Penetrating Keratoplasty in Dogs

ZHOU Qing-guo;LIU Wei-min;HOU Jia-fa

2008, 39(6): 804-809. doi:

Abstract ( 919 )

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In order to determine the postoperative immune rejection, inflammatory and pain reactions in dogs with penetrating keratoplasty, 11 dogs of Shih Tzu, Pekingese and small type of mongrels with ages from 6 to 12 months and weights from 4.0 to 6.5 kg were given penetrating keratoplasty. Following the surgery, the recipient dogs were closely monitored everyday for the general and ocular inflammatory symptoms and the samples of peripheral blood were obtained every five days during initial 30 days and every ten days afterwards to determine serum TNF-α, IL-2 and β-EP levels by radioimmunoassay. The results showed that the serum TNF-α, IL-2 and β-EP levels increased by varying degrees in all of the recipient dogs following the surgery compared with those preoperatively, and that the serum TNF-α, IL-2 and β-EP levels varied owing to many different factors from the fifteenth day postoperation. These findings suggested that subacute immune rejection occurred seldom in the homografts treated with local dexamethasone; If the rejection did occur, it was mild. Dynamic changes of serum TNF-α contents were not suitable for early detection and diagnosis of postoperative immune rejection. The serous IL-2 levels varied with the extent of postoperative ocular inflammation, which might be an important indicator of the inflammative extent and its transformation following homologous corneal transplantation. The dogs showed painful response to corneal wound and suture-knot irritation, and notably increased serum β-EP content, determination of whose dynamic changes could accurately reflect the stress response of the recipient dogs to pain inflicted upon cornea following penetrating keratoplasty.

Effects of Chinese Herbal Medicinal Prescription on Splenic

CHEN Han-ing;CHEN Wen-yun;LIU Li-mei;CHEN Lin;WANG Rui-hai;XU Jian-qin;LIU Feng-hua

2008, 39(6): 810-813. doi:

Abstract ( 888 )

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Splenic IFN-γ and IL-4 contents in mice under heat stimulation were detected with ELISA to evaluate the effects of Chinese herbal medicine on them. The results showed that all of IFN-γ and IL-4 contents and IL-4/IFN-γ ratio increased in high temperature control group (HTCG); In Chinese herbal medicine group (CHMG), IFN-γ content began to upgrade on the first day after heat stimulation, raised at peak on the 3rd day, and then downgrade to the content of normal temperature control group (NTCG) on the 6th day without significant difference; IL-4 content downgraded significantly on the first day, then upgraded gradually and dramatically on the 6th day, and then downgraded to the content of NTCG; IL-4/IFN-γ ratio decreased significantly on the first day, and then increased gradually. In conclusion, the heat stimulation may cause the disorder of IFN-γ and IL-4 levels in mice spleen; Chinese herbal medicine have the significant effects of regulation on IFN-γ and IL-4 levels to improve and recover the immune function.

Immunomodulation and Antitumor Activity of Fermented Decoction of Sijunzi in S180-bearing Mice

HAN Chun-yang;LIN De-gui;LIU Cui-yan;GUO Shun-xing

2008, 39(6): 814-818. doi:

Abstract ( 884 )

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The immunomodulation and antitumor activity of fermented decoction of Sijunzi (FDSJZ) in S180-bearing mice was investigated. The mice inoculated with S180 cell suspension were treated with FDSJZ for 10 days. The homolysin assays, MTT colorimetric method and flow cytometry were used to study the humoral immunity and cellular immunity of S180-bearing mice. The results showed that FDSJZ could significantly inhibit the growth of transplantable sarcoma S180 and it could enhance the anti-tumor activity of CTX, and lower the side effect of CTX. FDSJZ could increase organ indices, spleen lymphocyte proliferation, the form of antibody secreted by spleen cells, and the percent of CD3⁺ and CD8⁺ of S180-bearing mice. And it could lower the ratio of CD4⁺/CD8⁺ which was elevated abnormally in S180-bearing mice. The results showed that there was some relation between the antitumor activity of FDSJZ and the immunomodulation.

Cloning and Tissue Specific Expression of Duck A-FABP Gene

ZHANG Jun;DONG Biao;ZHANG Hong;CHU Dong-sheng;XU Guo-qing;GONG Dao-qing;DUAN Xiu-jun;ZHAO Xu-ting;GU Zhi-liang

2008, 39(6): 819-822. doi:

Abstract ( 935 )

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A 422 bp cDNA of A-FABP gene was cloned and sequenced from the adipose tissue of a 35-week-old female duck, it contains a complete 399 bp CDS, encoding 132 amino acids. The duck A-FABP gene shared about 94% homology with that of poultry(goose and chicken), and 73.4%-75.7% homology with that of mammal (human, mouse,pig and cattle), while the corresponding protein shared more than 92.4% and 72.0%-77.3％ homology with the above animals’ A-FABP protein, respectively. The results of the semi-quantity RT-PCR indicated that duck A-FABP mRNA did not expressed in diencephalons, lung and kidney, and its mRNA expression level of adipose tissue was higher than that of other tissues (P<0.5). A-FABP gene is of high conservation in the course of animal evolution. The expression level of A-FABP mRNA is particular to tissues.

Cloning of the Prolactin Receptor Gene and Studying on Its Expression Characteristics in Eastern Zhejiang White Geese

CHU Xiao-hong;HU Jin-ping;LU Li-zhi;WANG Ya-qin;CHEN Wei-hu;XU Ning-ying

2008, 39(6): 823-826. doi:

Abstract ( 887 )

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This study was conducted to clone partial sequence of 1 305 nucleotides of the prolactin receptor (PRLR) gene in Eastern Zhejiang White geese from genome DNA. Meanwhile, the PRLR gene expression characteristics were investigated during different reproductive periods by real time PCR. Comparisons of concentration of prolactin receptor mRNA in the hypothalamus, pituitary gland and ovary of adult female geese at egglaying, out-of-lay and incubating periods were made, respectively. The results indicated that there were significant difference (P<0.05) in PRLR mRNA expression between different reproduction periods of the geese. The lowest level of PRLR expression was found in out-of-lay geese, higher in the egglaying geese, and the greatest in incubating geese. Furthermore, the analysis of PRLR expression in different tissues indicated that the highest levels of PRLR was expressed in the pituitary gland, followed in hypothalamus, and the least in ovary of the geese. There were significant difference（P<0.01）of PRLR expression between the pituitary gland/hypothalamus and ovary of the geese, whereas no any difference was observed between the pituitary gland and hypothalamus.

Effects of High Ambient Temperature on Performance, Carcass Traits and Fat Deposition in Two Breeds of Broiler

LU Qing-ping;WEN Jie;ZHANG Hong-fu

2008, 39(6): 827-831. doi:

Abstract ( 1624 )

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The effects of chronic heat stress on growth, carcass traits and fat deposition were investigated in two breeds of broilers. Male chickens (108, 5 wk old) from Arbor Acres (AA) and Beijing-You (BJY) respectively were equally distributed to 3 treatments: constant high ambient temperature at 34℃ with ad libitum feeding (34AL), constant optimal ambient temperature at 21℃ with ad libitum feeding (21AL), and constant optimal ambient temperature 21℃ but pairfed to the amount consumed by the 34AL group (21PF). The results showed that under hot condition, for AA broilers, the feed intakes were decreased by 45%（P<0.001）and average daily gain (ADG) were decreased from 61.45 g/d to 22.29 g/d, the feed efficiency decreased significantly. While for BJY chickens, their growth were not affected by high temperature even though the feed intake were decreased. The mortality of heatstressed AA broilers was 36％ but all of the BJY groups survived the experimental feeding period. The breast muscle rate and fat deposition of heatstressed AA broilers were decreased significantly. While the breast muscle rate of BJY chickens were not affected by high temperature and abdominal fat deposition rate of BJY chickens was enhanced by heat stress (P<0.05). The data indicated that BJY chickens had higher resistance to extreme heat comparing with AA broilers which got bad growth performance under high temperature. The higher feed efficiency and enhanced abdominal fat deposition rate would account for the high adaptability to extreme heat in BJY chickens.

Construction of Eukaryotic Expression Plasmid of TGF-β₁ Gene and Its Expression in Swine Umbilical Vein Endothelial Cells

WANG Wen-xiu;DENG Wen;ZHANG Yan-ming;DAI Chen;XIONG Kui-zhou;XIE Lin-hong;WEN Yuan-peng

2008, 39(6): 832-836. doi:

Abstract ( 831 )

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In this study, transforming growth factor-β₁ (TGF-β₁ ) full length cDNA was cloned from swine peripheral blood mononuclear cells by RT-PCR ，the eukaryotic expression plasmid of TGF-β₁ gene containing green fluorescent protein (GFP) report gene was constructed. The recombinant plasmid was transfected by lipofectin to prepared swine umbilical vein vascular endothelial cells(SUVECs). The fluorescence expression was directly detected with fluorescence microscope, and the expression of TGF-β₁ were tested by RT-PCR and indirect immunofluorescence assay (IFA), respectively. One week after transfection，green fluorescent can be seen by fluorescent microscope；RT-PCR and indirect immunofluorescence assay showed that the expression of TGF-β₁ are positive.The results indicated the successful construction of the eukaryotic expression plasmid pEGFP-C1-TGF-β₁ , and showed that TGF-β₁ gene was expressed efficiently in transfected SUVECs.

A Multiplex PCR Assay for Simultaneous Detection of Six Serotypes of Streptococcus suis

WANG Hua-ru;;WANG Chang-jun;TANG Jia-qi;LU Cheng-ping;PAN Xiu-zhen;Guo Heng-bin

2008, 39(6): 837-841. doi:

Abstract ( 905 )

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A multiplex PCR assay for detection and identification of various streptococcus suis strains in tonsillar specimens from pigs was developed and evaluated. In this research five distinct DNA targets were chose: One target, based on the gdh gene, encoded the glutamate dehydrogenase of streptococcus suis, can be applied to the detection of streptococcus suis at species level. The other targets, based on SS capsular polysaccharide (cps) genes specific for serotypes 1(and 14),2(and 1/2),7,9 were amplified by multiplex PCR. 73 isolates，which included 49 strains of streptococcus suis and 24 strains of negative control，and 94 clinical specimens （45 clinical strains in Sichuan）were detected by the multiplex PCR assay. In 73 streptococcus suis strains, 64 (87.5%)strains were gdh+ .There were no PCR products in strains of serotypes 13，19，30，32，33，34.The multiplex PCR could detect all strains of serotypes 1(and 14),2(and 1/2),7,9. In 45 clinical strains in Sichuan, the PCR results of 41 strains were gdh⁺cps2⁺. These results indicated that the PCR can be used as a reliable speciesspecific molecular diagnostic reagent and a serotype-specific method for the detection of streptococcus suis and six main serotypes 1,2,1/2,7,9 and 14, respectively. The PCR method has potential value for clinical and epidemiological applications.

Molecular Cloning, Sequence Analysis and Prokaryotic Expression of Ovine Mucosal Chemokine CCL28

QIAO Xin-an;LI Hong-ji;WANG Yue-ying;ZHU Yan-cai;HAN Li-qiang;YANG Guo-yu;WANG Yan-ling

2008, 39(6): 842-847. doi:

Abstract ( 1052 )

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A pair of cloning primers were designed according to the cloning principle of homologous sequence CCL28. CCL28 mRNA extracted from mucosal tissue of ovine colon was amplified by RT-PCR, then the PCR product was cloned, sequenced and analyzed. Then, a pair of expression primers were designed according to the cloned sequence. A fragment of ovine CCL28 cDNA containing BamHⅠ/XhoⅠ were amplified from recombinant vector by PCR. The fragment cleaved by BamHⅠ/XhoⅠ was subcloned into pGEX-4T-1 vector to construct a recombinant expression vector, pGEX-CCL28. Subsequently, E.coli BL2l competent cells were transformed by the recombinant vector. The fusion protein, which was induced by IPTG was analyzed by SDS-PAGE. The cloned sequence containing the open reading frame (ORF) of ovine CCL28 consists of 444 bp, the ORF is 387 bp and encodes 129 amino acids. Analysis showed that the ovine CCL28 nucleotide sequence shared 76.4％, 61.4%, 84.3% and 92.9% homology with that of human, mouse, pig and cattle, the predicted amino acid shared 76％, 63%, 84% and 93% homology with that of human, mouse, pig and cattle. The fusion protein expressed in E.coli BL21(DE3) was about 39 ku and mainly existed as inclusion bodies. Our research provides the experiment basis for further researching biological function of CCL28.

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