The objective of these experiments was to develop an attenuated thermostable Newcastle disease virus(NDV), CBP-1 strain isolated from infected pheasants. Safety, pathogenicity and protective effects against velogenic NDV were also investigated to evaluate if the attenuated NDV, CBP-1 strain could be a candidate for a new NDV vaccine strain. CBP-1 strain was passaged up to the 173 times by nine days old embryonated eggs and chicken embryo fibroblast(CEF) cell cultures. Its sensitivitly to lipid solvents and low pH, thermostability, mean death time(MDT), intracerebral pathogenicity index(ICPI) of one day old chicks and intravenous pathogenicity index(IVPI) of four weeks old chicks were examined. Safety, boosting and protective effects were tested by chicks mortality. CBP-1 NDV strain had significant thermostability at 56$\^{C}$ for 30 minutes. by hemagglutinin activity and egg infectivity test, but was not resistant to lipid solvent. It showed possibility to use as a feed or water vaccine because of the resistance to low pH. MDT, ICPI and IVPI of CBP-1 were attenuated from 51.5, 1.96, 2.60 to 112.4, 1.12, 1.45. These results implied that the 173rd passages in embryonated egg and CEF cell cultures induced a substantial attenuation of the pathogenicity of the parent virus, changing the virulence from velogenic to intermediate between mesogenic and lentogenic. After vaccination with CBP-1 at one day old by drinking water mortality was 17.5%. However, spray vaccination with B1 at one day old, CBP-1 at two weeks ild and challenge with velogenic Kyojeongwon strain at four weeks old showed 93.5% survival rate. Mortality of chicks, vaccination with 173rd passaged CBP-1 strain at one day old, two weeks old and challenge with Kyokeongwon strain at four weeks old, was 20.0%. The results of these studies indicated that partial attenuated CBP-1 strain tended to be a low safety for ND of broiler chicks and would need to be more successive attenuation.
Objective: This study aimed to determine the protective efficacy of Buddha's Temple (BT) extract against tert-butyl hydroperoxide (t-BHP)-induced oxidative stress in Gallus gallus chicken embryo fibroblast cell line (DF-1) and its effects on the cell lipid metabolism. Methods: In this experimental study, Gallus gallus DF-1 fibroblast cells were pretreated with BT 10-7 for 24 hours, followed by their six-hour exposure to t-BHP (100 μM). Water-soluble tetrazolium salt-8 (WST-8) assays were performed, and the growth curve was computed. The intracellular gene expression changes caused by BT extract were confirmed through quantitative polymerase chain reaction (qPCR). Flow cytometry, oil red O staining experiment, and thin-layer chromatography were performed for the detection of intracellular metabolic mechanism changes. Results: The WST-8 assay results showed that the BT pretreatment of Gallus gallus DF-1 fibroblast cell increased their cell survival rate by 1.08%±0.04%, decreased the reactive oxygen species (ROS) level by 0.93%±0.12% even after exposure to oxidants, and stabilized mitochondrial activity by 1.37%±0.36%. In addition, qPCR results confirmed that the gene expression levels of tumor necrosis factor α (TNFα), TIR domain-containing adapter inducing IFN-beta (TICAM1), and glucose-regulated protein 78 (GRP78) were regulated, which contributed to cell stabilization. Thin-layer chromatography and oil red O analyses showed a clear decrease in the contents of lipid metabolites such as triacylglycerol and free fatty acids. Conclusion: In this study, we confirmed that the examined BT extract exerted selective protective effects on Gallus gallus DF-1 fibroblast cells against cell damage caused by t-BHP, which is a strong oxidative inducer. Furthermore, we established that this extract significantly reduced the intracellular ROS accumulation due to oxidative stress, which contributes to an increase in poultry production and higher incomes.
The effect of paternal telomere length on reproductive performance, relationship between paternal and offspring telomere length, and association between paternal telomere length and offspring production performance were investigated in Korean native chickens. Using 22 paternal chickens and 329 offspring, the paternal reproductive performance such as fertility, embryo mortality, and hatchability, as well as the offspring production performance such as survival rate, body weight, and weight gain were analyzed. Telomere length was analyzed through quantitative fluorescence in situ hybridization using lymphocytes. No significant differences were observed in fertility, embryo mortality, and hatchability between paternal chicken telomere lengths (P<0.05). These results indicate that paternal telomere length had a weak negative correlation with fertility and embryonic death rate but a weak positive correlation with hatchability. The correlation coefficient between paternal telomere length and offspring survival rate was r=0.17 (P>0.05). The group of offspring with long paternal telomeres showed relatively poor growth performance. Moreover, a significant negative correlation was estimated between paternal telomere length and offspring growth performance (P<0.05). The correlation coefficient between paternal and offspring telomere lengths was r=0.075 (P>0.05). In conclusion, there was a weak association between paternal telomere length and reproductive performance, offspring survival rate, and offspring telomere length, respectively. However, paternal telomere length and offspring growth performance showed a negative relationship. Such results could be due to the re-extension of telomere length by telomere reprogramming in the early embryonic stage and the different degree of telomere shortening between individuals with increasing age after birth.
Kim, Hyun;Kim, Dong Hun;Han, Jae Yong;Do, Yoon Jung;Kim, Jae Hwan;Kim, Young Sin;Seong, Hwan Hoo;Ko, Yeoung Gyu;Kim, Sung Woo
Korean Journal of Poultry Science
/
v.40
no.3
/
pp.163-169
/
2013
Cryopreservation of poultry semen has been reported, but preservation of female genetic material has not been possible because of the unique anatomical and physiological characteristics of the avian egg. Thus an alternative strategy for conservation of oviparous species of animals must be developed. Recent technological developments for producing germline chimeras by the transfer of primordial germ cells (PGCs) into recipient embryos has enabled the conservation and retrieval of chicken genetic resources in their complete form. In the present study, fertilized eggs were incubated for about 5.5 days to obtain embryos at stage 28. The whole embryo was collected from the germinal gonad using a fine glass micro pipette under a microscope. The PGCs were then purified using MACS method. Two commercially available cryoprotectants (A and B) were used to preserve the PGCs, and EG were used as a control. The average recovery rate of PGCs after thawing was 35.5% and 60.5% with the A and B treatments, respectively. There was no significant difference between B treatments and control, which showed an average recovery rate of 52.8%. However, the recovery rate obtained using A cryoprotectant (35.5%) was significantly lower than using treatment control and B. The average viability of the PGCs after thawing were 77.9% and 77.4% for cryoprotectants A and B, respectively, and the control were was 81.6%. There was no statistically significant difference between the two treatments and control. It was concluded that all of the available cryoprotectants examined in this study could be used for preservation of PGCs from embryos. Further experiments to produce germline chimera from PGCs preserved using this techniques are strongly recommended.
Dental implants have been developed for enhancement of osseointegration. Biocompatibility, bone affinity and surface characteristics of dental implants are very important factors for osseointegration. The aim of the present study was to determine the cytotoxicity and the bone affinity of titanium phosphide(Ti-P) implant material. The Ti-P surface was obtained by vacuum sintering of titanium within compacted hydroxyapatite powder. The composition and the chemical change of the surface were determined by Auger electron spectroscopy. The in vitro cytotoxicity was evaluated by the viability of the bone cells and macrophages obtained from chicken embryo and rat,s peritonium, respectively. For the comparative evaluation, 316L stainless steel, commercially pure titanium and Ti-P materials, prepared in size of 1O.0mm in diameter and 5.0mm in height, were immersed separately in bone cells and macrophages for 10 days. For the evaluation of the in vivo bone affinity, 316L stainless steel, commercially pure titanium and Ti-P materials, prepared in size of 5.0mm in diameter and 10.0mm in length, were implanted after drilling in diameter 5.5mm in femurs of 2 dogs weighing 10Kg more or less. Six weeks after implantation the specimens were prepared for histopathological examination and were observed under light microscope. In comparison of in vitro bone cell viability, Ti-P and commercially pure titanium groups were not significantly different from control group (p>O.1), but 316L stainless steel group was significantly lower than control group(p<0.05). There was no statistical difference in the viability of macrophages between 3 different groups and control group(p>O.l). In comparison of in vivo study, 316L stainless steel and commercially pure titanium showed fibrous encapsulation, but Ti-P showed remarkable new bone formation without any fibrous tissue. The results demonstrate that Ti-P has favorable biocompatibility and bone affinity, and suggest that dental implants with Ti-P surface may enhance osseointegration.
Purpose: The increasing demand for esthetically pleasing results has contributed to the use of ceramics for dental implant abutments. The aim of this study was to compare the biological response of epithelial tissue cultivated on lithium disilicate ($LS_2$) and zirconium oxide ($ZrO_2$) ceramics. Understanding the relevant physicochemical and mechanical properties of these ceramics will help identify the optimal material for facilitating gingival wound closure. Methods: Both biomaterials were prepared with 2 different surface treatments: raw and polished. Their physicochemical characteristics were analyzed by contact angle measurements, scanning white-light interferometry, and scanning electron microscopy. An organotypic culture was then performed using a chicken epithelium model to simulate peri-implant soft tissue. We measured the contact angle, hydrophobicity, and roughness of the materials as well as the tissue behavior at their surfaces (cell migration and cell adhesion). Results: The best cell migration was observed on $ZrO_2$ ceramic. Cell adhesion was also drastically lower on the polished $ZrO_2$ ceramic than on both the raw and polished $LS_2$. Evaluating various surface topographies of $LS_2$ showed that increasing surface roughness improved cell adhesion, leading to an increase of up to 13%. Conclusions: Our results demonstrate that a biomaterial, here $LS_2$, can be modified using simple surface changes in order to finely modulate soft tissue adhesion. Strong adhesion at the abutment associated with weak migration assists in gingival wound healing. On the same material, polishing can reduce cell adhesion without drastically modifying cell migration. A comparison of $LS_2$ and $ZrO_2$ ceramic showed that $LS_2$ was more conducive to creating varying tissue reactions. Our results can help dental surgeons to choose, especially for esthetic implant abutments, the most appropriate biomaterial as well as the most appropriate surface treatment to use in accordance with specific clinical dental applications.
Proceedings of the Korea Society of Poultry Science Conference
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2001.11a
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pp.74-76
/
2001
Comparing to mammals, male bird has the homozygote ZZ and female has the heterozygote n. Therefore, the sex of fertilized eggs is defined by female chromosome constitution. Although this cytological observation had been established, the molecular and cellular mechanism of germ cell differentiation are essentially unknown in aves. Especially, the differentiation of germ cells in mixed-sex chimeras has not yet been clearly elucidated. Primordial germ cells, which are the progenitors of sperm or egg after sexual maturity, firstly arise in the epiblast and migrate to embryonic gonads through the blood vessel. During the embryo development, these PGCs differentiate in the pathway of mate or female, respectively and develop the sperm or egg cells after sexual maturity. In this paper, we confirmed that the female PGCs could migrate into the recipient male gonads after transferring and differentiate into germ cells in the embryonic stages. The primordial germ cells were isolated from the female embryonic gonads of 5.5-day-old incubation and re-injected into the male recipient embryos of 2-day-old incubation, which produced mixed-sex chimera in the germline. The finding in this study demonstrated the ability of migration and differentiation of gonadal primordial germ cells in mixed-sex chicken.
Background: Coinfection with avian leukosis virus subgroup J (ALV-J) and reticuloendotheliosis virus (REV) is common in chickens, and the molecular mechanism of the synergistic pathogenic effects of the coinfection is not clear. Exosomes have been identified as new players in the pathogenesis of retroviruses. The different functions of exosomes depend on their cargo components. Objectives: The aim of this study was to investigate the function of co-regulation differentially expressed proteins in exosomes on coinfection of ALV-J and REV. Methods: Here, viral replication in CEF cells infected with ALV-J, REV or both was detected by immunofluorescence microscopy. Then, we analyzed the exosomes isolated from supernatants of chicken embryo fibroblast (CEF) cells single infected and coinfected with ALV-J and REV by mass spectrometry. KEGG pathway enrichment analyzed the co-regulation differentially expressed proteins in exosomes. Next, we silenced and overexpressed tripartite motif containing 62 (TRIM62) to evaluate the effects of TRIM62 on viral replication and the expression levels of NCK-association proteins 1 (NCKAP1) and actin-related 2/3 complex subunit 5 (ARPC5) determined by quantitative reverse transcription polymerase chain reaction. Results: The results showed that coinfection of ALV-J and REV promoted the replication of each other. Thirty proteins, including TRIM62, NCK-association proteins 1 (NCKAP1, also known as Nap125), and Arp2/3-5, ARPC5, were identified. NCKAP1 and ARPC5 were involved in the actin cytoskeleton pathway. TRIM62 negatively regulated viral replication and that the inhibition of REV was more significant than that on ALV-J in CEF cells coinfected with TRIM62. In addition, TRIM62 decreased the expression of NCKAP1 and increased the expression of ARPC5 in coinfected CEF cells. Conclusions: Collectively, our results indicated that coinfection with ALV-J and REV competitively promoted each other's replication, the actin cytoskeleton played an important role in the coinfection mechanism, and TRIM62 regulated the actin cytoskeleton.
Proceedings of the Korea Society of Poultry Science Conference
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2003.07b
/
pp.37-54
/
2003
It has been recognized that the hen. like its mammalian counterparts. provides young chicks with antibodies as protection against hostile invaders. This system facilitates the transfer of specific antibodies from serum to egg yolk. and provides a supply of antibodies called immunoglobulin Y(IgY) to the developing embryo and the hatched chick. The protection against pathogens that the relatively immuno-incompetent newly hatched chick has. is through transmission of antibodies from the mother via the egg. Egg yolk. therefore. can be loaded with a large amount of IgY against pathogens which can immobilize the existing or invading pathogens during the embryo development or in day-old chicks. Thus. the immunization of laying hens to various pathogens results in production of different antigen-specific IgY in eggs. Egg yolk contains 8~20 mg of immunoglobulins (IgY) per $m\ell$ or 136~340 mg per yolk suggesting that more than 30 g of IgY can be obtained from one immunized hen in a year. By immunizing laying hens with antigens and collecting IgY from egg yolk. low cost antibodies at less than $10 per g compared to more than $20.000 per g of mammalian IgG can be obtained. This IgY technology opens new potential market applications in medicine. public health veterinary medicine and food safety. A broader use of IgY technology could be applied as biological or diagnostic tool. nut-raceutical or functional food development. oral-supplementation for prophylaxis. and as pathogen-specific antimicrobial agents for infectious disease control. This paper has emphasized that when IgY-loaded chicken eggs are produced and consumed. the specific antibody binds. immobilizes and consequently reduces or inhibits the growth or colony forming abilities of microbial pathogens. This concept could serve as an alternative agent to replace the use of antibiotics. since today. more and more antibiotics are less effective in the treatment of infections. due to the emergence of drug-resistant bacteria.
Throughout the studies the following experimental results were obtained and are summarized: 1. Multiplication of agents in primary cell cultures of both GF classical and CR-64 acute strain of Marek's disease infected chicken kidneys was accompanied by the formation of distinct transformed cell foci. This characteristic nature of cell transformation was passaged regularly by addition of dispersed cell from infected cultures to normal chicken kidney cell cultures, and also transferred was the nature of cell transformation to normal chick-embryo liver and neuroglial cell cultures. No cytopathic changes were noticed in inoculated chick-embryo fibroblast cultures. 2. The same cytopathic effects were noticed in normal kidney cell monolayers after the inoculation of whole blood and huffy coat cells derived from both forms of Marek's disease infected chickens. In these cases, however, the number of transformed cell foci appearing was far less than that of uninoculated monolayers prepared directly from the kidneys of Marek's disease infected chickens. 3. The change in cell culture IS regarded as a specific cell transformation focus induced by an oncogenic virus rather than it plaque in slowly progressing cytopathic effect by non-oncogenic viruses, and it is quite similar to RSV focus in chick-embryo fibroblasts in many respects. 4. The infective agent (cell transformable) were extremely cell-associated and could not be separated in an infective state from cells under the experimental conditions. 5. The focus assay of these agents was valid as shown by the high degree of linear correlation (r=0.97 and 0.99) between the relative infected cell concentration (in inoculum) and the transformed cell foci counted. 6. No differences were observed between the GF classical strain and the CR-64 acute strain of Marek's disease as far as cell culture behavior. 7. Characterization of the isolates by physical and chemical treatments, development of internuclear inclusions in Infected cells, and nucleic acid typing by differential stainings and cytochemical treatments indicated that the natures of these cell transformation agents closely resemble to those described fer the group B herpes viruses. 8. Susceptible chicks inoculated with infected kidney tissue culture cells developed specific lesions of Marek's disease, and in a case of prolonged observation after inoculation (5 weeks) the birds developed clinical symptoms and gross lesions of Marek's disease. Kidney cell cultures prepared from those inoculated birds and sacrificed showed a superior recovery of cell transformation property by formation of distinct foci. 9. Electron microscopic study of infected kidney culture cells (GF agent) by negative staining technique revealed virus particles furnishing the properties of herpes viruses. The particle was measured about $100m{\mu}$ and, so far, no herpes virus envelop has been seen from these preparations. 10. No relationship of both isolates to avian leukosis/sarcoma group viruses and PPLO was observed.
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