The Experts below are selected from a list of 585 Experts worldwide ranked by ideXlab platform
Shou-long Deng - One of the best experts on this subject based on the ideXlab platform.
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overexpression of toll like receptor 4 affects autophagy oxidative stress and inflammatory responses in monocytes of Transgenic Sheep
Frontiers in Cell and Developmental Biology, 2020Co-Authors: Sutian Wang, Shou-long Deng, Xuting Song, Kunli Zhang, Peixin Jiao, Meiyu Qi, Zheng Xing LianAbstract:Toll-like receptor 4 (TLR4) is a critical pattern recognition receptor that plays a critical role in the host innate immune system’s recognition of Gram-negative bacteria. Since it is the lipopolysaccharide (LPS) receptor, it links the activated inflammatory response with autophagy and oxidative stress. Autophagy, or type II programmed cell death, was reported to have defensive functions in response to the production of inflammatory cytokines and oxidative stress. To explore the relationship between autophagy, inflammation, and oxidative stress, a TLR4-enriched Transgenic animal (Tg) model (Sheep) was generated. Autophagy activity in the Tg blood monocytes was significantly higher than in the wild-type animal under LPS stress and it returned to normal after transfection of TLR4 siRNA. Pretreatment with 3-methyladenine (3-MA) inhibited autophagy and enhanced oxidative stress and the production of TNF-α. The LPS-induced reactive oxygen species (ROS) level was markedly increased in the Tg group at an early stage, before quickly returning to normal values. In addition, suppressing ROS production by N-acetyl-L-cysteine (NAC) down-regulated the number of intracellular autophagosomes, and the expression of Beclin-1, ATG5 and cytokines IL-1β, IL-6, and TNF-α. Further mechanistic investigation suggested that the TLR4-associated p38 mitogen-activated protein kinase (MAPK) signaling pathway was involved in autophagy and oxidative stress. P38 MAPK promotes intracellular autophagy, ROS production and the inflammatory response. Moreover, TLR4 over-expression suppressed oxidative stress and the production of inflammatory cytokines and increased autophagy activity in vivo. Taken together, our results showed that LPS induced autophagy, which was related to TLR4-mediated ROS production through the p38 MAPK signaling pathway. In addition, our study also provided a novel Transgenic animal model to analyze the effects of TLR4 on autophagy, oxidative stress and inflammatory responses.
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Overexpression of Toll-Like Receptor 4 Contributes to Phagocytosis of Salmonella Enterica Serovar Typhimurium via Phosphoinositide 3-Kinase Signaling in Sheep
Cellular Physiology and Biochemistry, 2018Co-Authors: Sutian Wang, Shou-long Deng, Xiaosheng Zhang, Zhixian Wang, Rui Zhang, Xiaojing Jiang, Jiahao Wang, Jinlong ZhangAbstract:Phagocytosis of bacteria by monocytes/macrophages can trigger the immune response and the clearance of bacteria. This innate immune response involves Toll-like receptor 4 (TLR4). However, much remains unknown about the mechanism of TLR4-regulated phagocytosis of Salmonella enterica serovar Typhimurium (S. typhimurium) within Sheep monocytes/macrophages. Here, we aimed to address these knowledge gaps by infecting Transgenic Sheep overexpressing TLR4 with S. typhimurium and examining the phagocytic mechanisms involved. Transgenic Sheep were generated by microinjection of the constructed plasmids into fertilized eggs. Monocytes/macrophages isolated from Sheep blood were stimulated with LPS and S. typhimurium. Phagocytosis-related factor expression, phagocytic ability, and adhesion were then determined. TLR4/phosphatidylinositide 3-kinase (PI3K) signaling was inhibited to investigate if this pathway is involved in changes in bacterial internalization in Sheep. We found that TLR4 overexpression effectively activated the PI3K signaling pathway and upregulated the expression of scavenger receptors. Additionally, actin polymerization and adhesive capacity were both enhanced in TLR4-overexpressing Sheep monocytes/macrophages. TLR4 inhibition decreased S. typhimurium phagocytosis by reducing the actin polymerization and adhesive capacity of cells. Furthermore, inhibition of PI3K markedly impaired TLR4-dependent phagocytosis by restraining actin polymerization and scavenger receptor expression and reduced the adhesive capacity of the monocytes/macrophages. Our findings indicate that overexpression of TLR4 enhances phagocytosis through PI3K signaling and the subsequent activation of actin polymerization and scavenger receptors in Sheep monocytes/macrophages infected with S. typhimurium. © 2018 The Author(s). Published by S. Karger AG, Basel.
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aanat Transgenic Sheep generated via ops vitrified microinjected pronuclear embryos and reproduction efficiency of the Transgenic offspring
PeerJ, 2018Co-Authors: Xiuzhi Tian, Shou-long Deng, Xiaosheng Zhang, Juncai Fu, Dongying Lv, Minghui Yang, Yukun Song, Jinglong Zhang, Zheng Xing LianAbstract:Background: The open pulled straw (OPS) vitrification method has been successfully applied in mouse, pig, and goat embryos as well as in buffalo oocytes, but it has not yet been applied to the microinjected embryos. This study examined the effects of OPS vitrification on embryo development and the reproductive capacity of the Transgenic offspring in order to establish a method for preservation of microinjected embryos. Methods: Ovine pronuclear embryos were microinjected with the exogenous aralkylamine N-acetyltransferase gene (AANAT), frozen by the OPS method, and subsequently thawed for embryo transplantation. Pregnancy rate, lambing rate, survival rate, average birth weight and Transgenic positive rate as well as reproduction efficiency and hormone level of the Transgenic offspring were investigated to analyze the effect of OPS vitrification on microinjectd pronuclear embryos. Results: No significant differences were observed in the birth rate, lamb survival rate and Transgenic positive rate between the frozen and non-frozen AANAT-microinjected pronuclear embryos. The average birth weight of the frozen embryos offspring was greater than that of the non-frozen embryos. Importantly, the Transgenic offspring that overexpressed the AANAT gene showed improved ovulation efficiency and lambing rate by regulating their hormone levels. Conclusions: The OPS vitrification approach may be a valuable method in microinjected- embryo transfer technology, which could reserve embryos and result in fewer unnecessary animal sacrifices. In addition, the AANAT+ Transgenic offspring exhibited improved reproductive capacity on account of regulation effect of melatonin on reproductive hormone. These data may provide available references for human-assisted reproduction.
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AANAT Transgenic Sheep generated via OPS vitrified-microinjected pronuclear embryos and reproduction efficiency of the Transgenic offspring
PeerJ, 2018Co-Authors: Xiuzhi Tian, Shou-long Deng, Xiaosheng Zhang, Juncai Fu, Dongying Lv, Minghui Yang, Yukun Song, Jinglong Zhang, Teng Ma, Zheng Xing LianAbstract:The open pulled straw (OPS) vitrification method has been successfully applied in mouse, pig, and goat embryos as well as in buffalo oocytes, but it has not yet been applied to the microinjected embryos. This study examined the effects of OPS vitrification on embryo development and the reproductive capacity of the Transgenic offspring in order to establish a method for preservation of microinjected embryos. Ovine pronuclear embryos were microinjected with the exogenous aralkylamine N-acetyltransferase gene (AANAT), frozen by the OPS method, and subsequently thawed for embryo transplantation. Pregnancy rate, lambing rate, survival rate, average birth weight and Transgenic positive rate as well as reproduction efficiency and hormone level of the Transgenic offspring were investigated to analyze the effect of OPS vitrification on microinjectd pronuclear embryos. No significant differences were observed in the birth rate, lamb survival rate and Transgenic positive rate between the frozen and non-frozen AANAT-microinjected pronuclear embryos. The average birth weight of the frozen embryos offspring was greater than that of the non-frozen embryos. Importantly, the Transgenic offspring that overexpressed the AANAT gene showed improved ovulation efficiency and lambing rate by regulating their hormone levels. The OPS vitrification approach may be a valuable method in microinjected- embryo transfer technology, which could reserve embryos and result in fewer unnecessary animal sacrifices. In addition, the AANAT+ Transgenic offspring exhibited improved reproductive capacity on account of regulation effect of melatonin on reproductive hormone. These data may provide available references for human-assisted reproduction.
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Overexpression of Toll-like Receptor 4-linked Mitogen-activated Protein Kinase Signaling Contributes to Internalization of Escherichia coli in Sheep.
International Journal of Biological Sciences, 2018Co-Authors: Sutian Wang, Shou-long Deng, Xiaosheng Zhang, Jinlong Zhang, Xiaojing Jiang, Jiahao Wang, Zheng Xing LianAbstract:Escherichia coli is one of the most common causal pathogens of mastitis in milk-producing mammals. Toll-like receptor 4 (TLR4) is important for host recognition of this bacteria. Increased activation of TLR4 can markedly enhance the internalization of E. coli. In this study, the relationship between TLR4 and mitogen-activated protein kinase (MAPK) signaling pathways in mediating E. coli internalization was evaluated in Sheep monocytes. Using a TLR4-overexpressing Transgenic (Tg) Sheep model, we explored the bacterial internalization mechanism in Sheep. We found that monocytes of Tg Sheep could phagocytize more bacteria and exhibited higher adhesive capacity. The specific inhibition of p38 MAPK or c-Jun N-terminal kinase (JNK) or extracellular signal-regulated kinases (ERKs) reduced TLR4-dependent internalization of bacteria into Sheep monocytes. Furthermore, the inhibition of MAPK signaling down-regulated the adhesive capacity of monocytes and the expression of scavenger receptors and adhesion molecules. Taken together, the overexpression of TLR4 in Transgenic Sheep enhanced the internalization of E. coli via MAPK signaling.
Juncai Fu - One of the best experts on this subject based on the ideXlab platform.
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aanat Transgenic Sheep generated via ops vitrified microinjected pronuclear embryos and reproduction efficiency of the Transgenic offspring
PeerJ, 2018Co-Authors: Xiuzhi Tian, Shou-long Deng, Xiaosheng Zhang, Juncai Fu, Dongying Lv, Minghui Yang, Yukun Song, Jinglong Zhang, Zheng Xing LianAbstract:Background: The open pulled straw (OPS) vitrification method has been successfully applied in mouse, pig, and goat embryos as well as in buffalo oocytes, but it has not yet been applied to the microinjected embryos. This study examined the effects of OPS vitrification on embryo development and the reproductive capacity of the Transgenic offspring in order to establish a method for preservation of microinjected embryos. Methods: Ovine pronuclear embryos were microinjected with the exogenous aralkylamine N-acetyltransferase gene (AANAT), frozen by the OPS method, and subsequently thawed for embryo transplantation. Pregnancy rate, lambing rate, survival rate, average birth weight and Transgenic positive rate as well as reproduction efficiency and hormone level of the Transgenic offspring were investigated to analyze the effect of OPS vitrification on microinjectd pronuclear embryos. Results: No significant differences were observed in the birth rate, lamb survival rate and Transgenic positive rate between the frozen and non-frozen AANAT-microinjected pronuclear embryos. The average birth weight of the frozen embryos offspring was greater than that of the non-frozen embryos. Importantly, the Transgenic offspring that overexpressed the AANAT gene showed improved ovulation efficiency and lambing rate by regulating their hormone levels. Conclusions: The OPS vitrification approach may be a valuable method in microinjected- embryo transfer technology, which could reserve embryos and result in fewer unnecessary animal sacrifices. In addition, the AANAT+ Transgenic offspring exhibited improved reproductive capacity on account of regulation effect of melatonin on reproductive hormone. These data may provide available references for human-assisted reproduction.
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AANAT Transgenic Sheep generated via OPS vitrified-microinjected pronuclear embryos and reproduction efficiency of the Transgenic offspring
PeerJ, 2018Co-Authors: Xiuzhi Tian, Shou-long Deng, Xiaosheng Zhang, Juncai Fu, Dongying Lv, Minghui Yang, Yukun Song, Jinglong Zhang, Teng Ma, Zheng Xing LianAbstract:The open pulled straw (OPS) vitrification method has been successfully applied in mouse, pig, and goat embryos as well as in buffalo oocytes, but it has not yet been applied to the microinjected embryos. This study examined the effects of OPS vitrification on embryo development and the reproductive capacity of the Transgenic offspring in order to establish a method for preservation of microinjected embryos. Ovine pronuclear embryos were microinjected with the exogenous aralkylamine N-acetyltransferase gene (AANAT), frozen by the OPS method, and subsequently thawed for embryo transplantation. Pregnancy rate, lambing rate, survival rate, average birth weight and Transgenic positive rate as well as reproduction efficiency and hormone level of the Transgenic offspring were investigated to analyze the effect of OPS vitrification on microinjectd pronuclear embryos. No significant differences were observed in the birth rate, lamb survival rate and Transgenic positive rate between the frozen and non-frozen AANAT-microinjected pronuclear embryos. The average birth weight of the frozen embryos offspring was greater than that of the non-frozen embryos. Importantly, the Transgenic offspring that overexpressed the AANAT gene showed improved ovulation efficiency and lambing rate by regulating their hormone levels. The OPS vitrification approach may be a valuable method in microinjected- embryo transfer technology, which could reserve embryos and result in fewer unnecessary animal sacrifices. In addition, the AANAT+ Transgenic offspring exhibited improved reproductive capacity on account of regulation effect of melatonin on reproductive hormone. These data may provide available references for human-assisted reproduction.
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rnai combining sleeping beauty transposon system inhibits ex vivo expression of foot and mouth disease virus vp1 in Transgenic Sheep cells
Scientific Reports, 2017Co-Authors: Shou-long Deng, Wenting Li, Guangdong Li, Kun Yu, Xiuzhi Tian, Feng Wang, Wuqi Jiang, Pengyun Ji, Juncai FuAbstract:Foot and mouth disease, which is induced by the foot and mouth disease virus (FMDV), takes its toll on the cloven-hoofed domestic animals. The VP1 gene in FMDV genome encodes the viral capsid, a vital element for FMDV replication. Sleeping Beauty (SB) is an active DNA-transposon system responsible for genetic transformation and insertional mutagenesis in vertebrates. In this study, a conserved VP1-shRNA which specifically targets the ovine FMDV-VP1 gene was constructed and combined with SB transposase and transposon. Then, they were microinjected into pronuclear embryos to breed Transgenic Sheep. Ninety-two lambs were born and the VP1-shRNA was positively integrated into eight of them. The rate of Transgenic Sheep production in SB transposon system was significantly higher than that in controls (13.04% vs. 3.57% and 7.14%, P < 0.05). The ear fibroblasts of the Transgenic lambs transfected with the PsiCheck2-VP1 vector had a significant inhibitory effect on the VP1 gene of the FMDV. In conclusion, the VP1-shRNA Transgenic Sheep were successfully generated by the current new method. The ear fibroblasts from these Transgenic Sheep possess a great resistance to FMDV. The result indicated that RNAi technology combining the “Sleeping Beauty” transposon system is an efficient method to produce Transgenic animals.
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RNAi combining Sleeping Beauty transposon system inhibits ex vivo expression of foot-and-mouth disease virus VP1 in Transgenic Sheep cells
Scientific Reports, 2017Co-Authors: Shou-long Deng, Wenting Li, Guangdong Li, Kun Yu, Xiuzhi Tian, Feng Wang, Wuqi Jiang, Pengyun Ji, Juncai FuAbstract:Foot and mouth disease, which is induced by the foot and mouth disease virus (FMDV), takes its toll on the cloven-hoofed domestic animals. The VP1 gene in FMDV genome encodes the viral capsid, a vital element for FMDV replication. Sleeping Beauty (SB) is an active DNA-transposon system responsible for genetic transformation and insertional mutagenesis in vertebrates. In this study, a conserved VP1-shRNA which specifically targets the ovine FMDV-VP1 gene was constructed and combined with SB transposase and transposon. Then, they were microinjected into pronuclear embryos to breed Transgenic Sheep. Ninety-two lambs were born and the VP1-shRNA was positively integrated into eight of them. The rate of Transgenic Sheep production in SB transposon system was significantly higher than that in controls (13.04% vs. 3.57% and 7.14%, P
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Efficient production of pronuclear embryos in breeding and nonbreeding season for generating Transgenic Sheep overexpressing TLR4
Journal of animal science and biotechnology, 2016Co-Authors: Yan Li, Di Lian, Shou-long Deng, Xiaosheng Zhang, Jinlong Zhang, Wenting Li, Zhixian Wang, Hongping Wu, Juncai FuAbstract:Brucella is a zoonotic Gram-negative pathogen that causes abortion and infertility in ruminants and humans. TLR4 is the receptor for LPS which can recognize Brucella and initiate antigen-presenting cell activities that affect both innate and adaptive immunity. Consequently, Transgenic Sheep over-expressing TLR4 are an suitable model to investigate the effects of TLR4 on preventing Brucellosis. In this study, we generated Transgenic Sheep overexpressing TLR4 and aimed to evaluate the effects of different seasons (breeding and non-breeding season) on superovulation and the imported exogenous gene on growth. In total of 43 donor ewes and 166 recipient ewes in breeding season, 37 donor ewes and 144 recipient ewes in non-breeding season were selected for super-ovulation and injected embryo transfer to generate Transgenic Sheep. Our results indicated the no. of embryos recovered of donors and the rate of pronuclear embryos did not show any significant difference between breeding and non-breeding seasons (P > 0.05). The positive rate of exogenous TLR4 tested were 21.21 % and 22.58 % in breeding and non-breeding season by Southern blot. The expression level of TLR4 in the Transgenic Sheep was 1.5 times higher than in the non-Transgenic group (P
Zheng Xing Lian - One of the best experts on this subject based on the ideXlab platform.
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The Application of PCR-DGGE Technique in Sheep Intestinal Microbial Diversity Research
Advanced Materials Research, 2020Co-Authors: Sen Yuan, Hongxing Zhang, Zheng Xing LianAbstract:To identify the intestinal microflora diversity of Transgenic Sheep and non-Transgenic one with overexpression of foreign antiviral gene (VP1 & SB-SW), DNA of total bacteria were extracted from 16 Sheep fecal samples (4 non-Transgenic Sheep, 6 VP1 Transgenic Sheep and 6 SB-SW Transgenic Sheep). And then PCR amplification with bacterial universal primers of V3 variable region of 16S rRNA were taken to get the fingerprint profile by denaturing gradient gel electrophoresis (DGGE) technology. The results showed that the DGGE profiles of the 16 fecal samples were highly polymorphic. The number of DGGE bands, considered indicative of the total species richness, did not vary predictably among the three different samples. The DNA sequences were analyzed and the dominant bacteria in Sheep fecal were found to be Bacteroides, Clostridium, and Ruminococcus. Specially, non-Transgenic Sheep had more Alistipes finegoldii and Clostridium lentocellum, Transgenic Sheep with VP1 had more Clostridium drakei and Clostridium populeti and Transgenic Sheep with SB-SW had more Barnesiella intestinihominis and Clostridium ljungdahlii. So the PCR-DGGE technique can be applied to evaluate genetically modified (GM) animal potential risks to the environment.
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overexpression of toll like receptor 4 affects autophagy oxidative stress and inflammatory responses in monocytes of Transgenic Sheep
Frontiers in Cell and Developmental Biology, 2020Co-Authors: Sutian Wang, Shou-long Deng, Xuting Song, Kunli Zhang, Peixin Jiao, Meiyu Qi, Zheng Xing LianAbstract:Toll-like receptor 4 (TLR4) is a critical pattern recognition receptor that plays a critical role in the host innate immune system’s recognition of Gram-negative bacteria. Since it is the lipopolysaccharide (LPS) receptor, it links the activated inflammatory response with autophagy and oxidative stress. Autophagy, or type II programmed cell death, was reported to have defensive functions in response to the production of inflammatory cytokines and oxidative stress. To explore the relationship between autophagy, inflammation, and oxidative stress, a TLR4-enriched Transgenic animal (Tg) model (Sheep) was generated. Autophagy activity in the Tg blood monocytes was significantly higher than in the wild-type animal under LPS stress and it returned to normal after transfection of TLR4 siRNA. Pretreatment with 3-methyladenine (3-MA) inhibited autophagy and enhanced oxidative stress and the production of TNF-α. The LPS-induced reactive oxygen species (ROS) level was markedly increased in the Tg group at an early stage, before quickly returning to normal values. In addition, suppressing ROS production by N-acetyl-L-cysteine (NAC) down-regulated the number of intracellular autophagosomes, and the expression of Beclin-1, ATG5 and cytokines IL-1β, IL-6, and TNF-α. Further mechanistic investigation suggested that the TLR4-associated p38 mitogen-activated protein kinase (MAPK) signaling pathway was involved in autophagy and oxidative stress. P38 MAPK promotes intracellular autophagy, ROS production and the inflammatory response. Moreover, TLR4 over-expression suppressed oxidative stress and the production of inflammatory cytokines and increased autophagy activity in vivo. Taken together, our results showed that LPS induced autophagy, which was related to TLR4-mediated ROS production through the p38 MAPK signaling pathway. In addition, our study also provided a novel Transgenic animal model to analyze the effects of TLR4 on autophagy, oxidative stress and inflammatory responses.
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aanat Transgenic Sheep generated via ops vitrified microinjected pronuclear embryos and reproduction efficiency of the Transgenic offspring
PeerJ, 2018Co-Authors: Xiuzhi Tian, Shou-long Deng, Xiaosheng Zhang, Juncai Fu, Dongying Lv, Minghui Yang, Yukun Song, Jinglong Zhang, Zheng Xing LianAbstract:Background: The open pulled straw (OPS) vitrification method has been successfully applied in mouse, pig, and goat embryos as well as in buffalo oocytes, but it has not yet been applied to the microinjected embryos. This study examined the effects of OPS vitrification on embryo development and the reproductive capacity of the Transgenic offspring in order to establish a method for preservation of microinjected embryos. Methods: Ovine pronuclear embryos were microinjected with the exogenous aralkylamine N-acetyltransferase gene (AANAT), frozen by the OPS method, and subsequently thawed for embryo transplantation. Pregnancy rate, lambing rate, survival rate, average birth weight and Transgenic positive rate as well as reproduction efficiency and hormone level of the Transgenic offspring were investigated to analyze the effect of OPS vitrification on microinjectd pronuclear embryos. Results: No significant differences were observed in the birth rate, lamb survival rate and Transgenic positive rate between the frozen and non-frozen AANAT-microinjected pronuclear embryos. The average birth weight of the frozen embryos offspring was greater than that of the non-frozen embryos. Importantly, the Transgenic offspring that overexpressed the AANAT gene showed improved ovulation efficiency and lambing rate by regulating their hormone levels. Conclusions: The OPS vitrification approach may be a valuable method in microinjected- embryo transfer technology, which could reserve embryos and result in fewer unnecessary animal sacrifices. In addition, the AANAT+ Transgenic offspring exhibited improved reproductive capacity on account of regulation effect of melatonin on reproductive hormone. These data may provide available references for human-assisted reproduction.
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AANAT Transgenic Sheep generated via OPS vitrified-microinjected pronuclear embryos and reproduction efficiency of the Transgenic offspring
PeerJ, 2018Co-Authors: Xiuzhi Tian, Shou-long Deng, Xiaosheng Zhang, Juncai Fu, Dongying Lv, Minghui Yang, Yukun Song, Jinglong Zhang, Teng Ma, Zheng Xing LianAbstract:The open pulled straw (OPS) vitrification method has been successfully applied in mouse, pig, and goat embryos as well as in buffalo oocytes, but it has not yet been applied to the microinjected embryos. This study examined the effects of OPS vitrification on embryo development and the reproductive capacity of the Transgenic offspring in order to establish a method for preservation of microinjected embryos. Ovine pronuclear embryos were microinjected with the exogenous aralkylamine N-acetyltransferase gene (AANAT), frozen by the OPS method, and subsequently thawed for embryo transplantation. Pregnancy rate, lambing rate, survival rate, average birth weight and Transgenic positive rate as well as reproduction efficiency and hormone level of the Transgenic offspring were investigated to analyze the effect of OPS vitrification on microinjectd pronuclear embryos. No significant differences were observed in the birth rate, lamb survival rate and Transgenic positive rate between the frozen and non-frozen AANAT-microinjected pronuclear embryos. The average birth weight of the frozen embryos offspring was greater than that of the non-frozen embryos. Importantly, the Transgenic offspring that overexpressed the AANAT gene showed improved ovulation efficiency and lambing rate by regulating their hormone levels. The OPS vitrification approach may be a valuable method in microinjected- embryo transfer technology, which could reserve embryos and result in fewer unnecessary animal sacrifices. In addition, the AANAT+ Transgenic offspring exhibited improved reproductive capacity on account of regulation effect of melatonin on reproductive hormone. These data may provide available references for human-assisted reproduction.
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Overexpression of Toll-like Receptor 4-linked Mitogen-activated Protein Kinase Signaling Contributes to Internalization of Escherichia coli in Sheep.
International Journal of Biological Sciences, 2018Co-Authors: Sutian Wang, Shou-long Deng, Xiaosheng Zhang, Jinlong Zhang, Xiaojing Jiang, Jiahao Wang, Zheng Xing LianAbstract:Escherichia coli is one of the most common causal pathogens of mastitis in milk-producing mammals. Toll-like receptor 4 (TLR4) is important for host recognition of this bacteria. Increased activation of TLR4 can markedly enhance the internalization of E. coli. In this study, the relationship between TLR4 and mitogen-activated protein kinase (MAPK) signaling pathways in mediating E. coli internalization was evaluated in Sheep monocytes. Using a TLR4-overexpressing Transgenic (Tg) Sheep model, we explored the bacterial internalization mechanism in Sheep. We found that monocytes of Tg Sheep could phagocytize more bacteria and exhibited higher adhesive capacity. The specific inhibition of p38 MAPK or c-Jun N-terminal kinase (JNK) or extracellular signal-regulated kinases (ERKs) reduced TLR4-dependent internalization of bacteria into Sheep monocytes. Furthermore, the inhibition of MAPK signaling down-regulated the adhesive capacity of monocytes and the expression of scavenger receptors and adhesion molecules. Taken together, the overexpression of TLR4 in Transgenic Sheep enhanced the internalization of E. coli via MAPK signaling.
Caird E Rexroad - One of the best experts on this subject based on the ideXlab platform.
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DNA preparation method can influence outcome of Transgenic animal experiments
Animal Biotechnology, 2000Co-Authors: Robert Wall, Caird E Rexroad, Anne M Powell, R.k. Paleyanda, J.a. Foster, H. LubonAbstract:Abstract In our continuing quest to improve the efficiency of producing Transgenic animals, we have compared the influence of two transgene purification techniques on the efficiency of creating Transgenic Sheep and mice. Three hundred eighty‐seven Sheep zygotes and 2,737 mouse zygotes were microinjected with one of four transgenes. Transgenes were isolated from plasmid sequences either by agarose gel electrophoresis followed by gel extraction or by a single step sodium chloride gradient fractionation technique. Four Transgenic Sheep and 61 Transgenic mice were produced. Both Sheep and mice embryos responded similarly to transgene preparation methods. Overall, pregnancy rate was higher for recipients that received embryos injected with NaCl purified DNA (mean + SEM: 64 ± 7% vs. 38 ± 7%). Furthermore, offspring per zygote transferred (NaCl, 22 + 3% vs. Gel, 12 + 3%) and Transgenics born per zygote transferred (NaCl, 3.9 ± 0.6% vs. Gel, 1.5 ± 0.6%) were higher when the NaCl purified DNA was used. However, th...
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Synthesis and secretion of the mouse whey acidic protein in Transgenic Sheep
Transgenic Research, 1996Co-Authors: Robert J. Wall, Caird E Rexroad, Anne Powell, Avi Shamay, Robert Mcknight, Lothar HennighausenAbstract:The synthesis of foreign proteins can be targeted to the mammary gland of Transgenic animals, thus permitting commercial purification of otherwise unavailable proteins from milk. Genetic regulatory elements from the mouse whey acidic protein (WAP) gene have been used successfully to direct expression of transgenes to the mammary gland of mice, goats and pigs. To extend the practical usefulness of WAP promoter-driven fusion genes and further characterize WAP expression in heterologous species, we introduced a 6.8 kb DNA fragment containing the genomic form of the mouse WAP gene into Sheep zygotes. Two lines of Transgenic Sheep were produced. The transgene was expressed in mammary tissue of both lines and intact WAP was secreted into milk at concentrations estimated to range from 100 to 500 mg/litre. Ectopic WAP gene expression was found in salivary gland, spleen, liver, lung, heart muscle, kidney and bone marrow of one founder ewe. WAP RNA was not detected in skeletal muscle and intestine. These data suggest that unlike pigs, Sheep may possess nuclear factors in a variety of tissues that interact with WAP regulatory sequences. Though the data presented are based on only two lines, these findings suggest WAP regulatory sequences may not be suitable as control elements for transgenes in Sheep bioreactors.
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development of Transgenic Sheep that express the visna virus envelope gene
Virology, 1994Co-Authors: Janice E Clements, R J Wall, Opendra Narayan, Debra Hauer, R Schoborg, Darlene Sheffer, Anne M Powell, Lucy M Carruth, M C Zink, Caird E RexroadAbstract:Abstract The ovine lentiviruses cause encephalitis, pneumonia, and arthritis in Sheep worldwide. Visna virus is a prototype of this family and the pathogenesis and molecular biology of the virus has been well characterized. The envelope proteins of visna virus are responsible for binding of virus to host cells and for causing cell fusion. The surface glycoprotein also elicits cellular and humoral immune responses to the virus, the former being thought to be responsible for eliminating infected cells as well as causing inflammatory lesions. In this study, Transgenic Sheep were constructed that expressed the envelope genes of visna virus under the control of the visna LTR to investigate the role of the env gene in the pathogenesis of lentiviral disease in its natural host. Three Transgenic lambs were identified that contain the env transgene and express the envelope glycoproteins. These Transgenic animals have remained healthy and expression of the viral gene has had no obvious deleterious effect. Expression of the visna envelope protein was demonstrated by cell fusion mediated by the envelope gene as well as by immunoprecipitation of the envelope proteins with monoclonal antibodies and immunofluorescence analyses of Env protein in cells. The target cell for visna virus replication in infected animals is the monocyte/macrophage. In natural infection, the level of viral gene expression in these cells increases with cell maturation. In the Transgenic Sheep, monocytes did not express the envelope glycoproteins until they differentiated into macrophages in vitro . Expression of the env mRNA in macrophages was quantitated by an RNase protection assay. In addition to expression in macrophages, the transgene was expressed by fibroblasts isolated from skin of the Transgenic Sheep. Expression of both the Env and Rev proteins was detected by immunoprecipitation and immunofluorescence. Two of the three lambs responded immunologically to the expression of the transgene by producing binding antibodies to the envelope glycoproteins. Thus, these Transgenic Sheep provide a model to study whether a lentivirus glycoprotein will prevent infection or modulate disease in its natural host after virus challenge.
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recent progress in the Transgenic modification of swine and Sheep
Molecular Reproduction and Development, 1993Co-Authors: V G Pursel, Caird E RexroadAbstract:INTRODUCTION For centuries the genetic composition of domestic animals has been manipulated to enhance their usefulness to man. In the past decade, development of recombinant DNA technology has enabled scientists to isolate single genes, analyze and modify their nucleotide structures, make copies of these isolated genes, and transfer copies into the genome. Animals that integrate this recombinant DNA in their genome are called “Transgenic.” Recently, medically important human proteins have been produced in large quantities in milk of Transgenic Sheep. Unless unforeseen complications arise during extraction and purification of these proteins, we can expect to see such products to begin clinical evaluation very soon. Use of Transgenic animals for food and fiber remains more distant in the future. Few agriculturally useful genes have thus far been isolated, sequenced, and cloned. In addition, insufficient information on gene regulation is available so that these integrated transgenes can be regulated sufficiently to prevent overexpression that has been shown in some instances to adversely impact the health status. The purpose of this paper will be to briefly review the progress that has been achieved since Transgenic modification of swine and Sheep was first reported in 1985 (Hammer et al., 1985), with emphasis on developments during the past year.
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expression of mouse iga by Transgenic mice pigs and Sheep
European Journal of Immunology, 1991Co-Authors: Caird E Rexroad, Richard R Behringer, Richard D Palmiter, David Lo, V G Pursel, Phyllis J Linton, Eric P Sandgren, Ralph L BrinsterAbstract:: The development of Transgenic animal technology allows the introduction of desired traits into the germ line of mice and other animals. Since the production of antibody to various polysaccharide antigens can be protective against pathogenic bacteria, we generated Transgenic mice, Sheep and pigs carrying genes encoding the mouse alpha and kappa chains for antibodies against phosphorylcholine (PC) to determine whether transgene antibody might be used to influence susceptibility to disease. High serum levels of mouse IgA were detected in Transgenic mice and pigs but not in Transgenic Sheep. It has been noted that transgene immunoglobulin expression can suppress endogenous immunoglobulin gene rearrangement and expression, and indeed, in one mouse line, expression of the transgene resulted in less than 10% of spleen B cells expressing endogenous IgM. Despite this suppression, significant levels of endogenous IgM were still secreted into the serum. Suppression of endogenous IgM expression was not seen in other mouse lines, nor was it seen in Transgenic pigs. In the Transgenic pigs, the mouse IgA was detected in the serum despite the absence of an intact mouse kappa transgene, so the secreted antibody presumably included pig light chains. Little if any of the mouse IgA in these sera showed binding specificity for PC. In one of the founder Sheep, mouse IgA was detectable in peripheral lymphocytes but not in serum. Mouse kappa expression was not detected in the Transgenic Sheep harboring an intact kappa transgene. These results illustrate the potential of introducing beneficial traits such as germ-line-encoded immunity into large mammalian species.
Wei Li - One of the best experts on this subject based on the ideXlab platform.
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cloning of the xuhuai goat pparγ gene and the preparation of Transgenic Sheep
Biochemical Genetics, 2013Co-Authors: Hao Chen, Yani Zhang, Wei Li, Mengmeng Zheng, Qi Xu, Jiuzhou Song, Bichun LiAbstract:This study aimed to clone the peroxisome proliferator-activated receptor γ (PPARγ) gene of the Xuhuai goat and to make Transgenic Sheep using intratesticular injection, so as to improve the meat quality and flavor by increasing the intramuscular fat content. The coding sequence of the goat PPARγ gene was 1,428 bp, encoding 475 amino acids. Its similarity with other species was 81 (chicken), 89 (mouse), 92 (pig), 98 (cow), and 99% (Sheep). The similarity of the corresponding amino acid sequences was 92.9, 97.3, 98.3, 99.6, and 99.8%, respectively. The signal peptide region of the PPARγ protein was not found in this study, demonstrating that the protein is not secreted. RT-PCR and western blot revealed that PPARγ was expressed in vitro, and the protein was localized in the cytoplasm. The PPARγ gene was expressed in F1 Transgenic Sheep at both the mRNA and the protein levels; the positive ratio was 13.7%.
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production of brown yellow patches in the slc7a11 Transgenic Sheep via testicular injection of transgene
Journal of Genetics and Genomics, 2012Co-Authors: Xin He, Hongtao Li, Zhiyong Zhou, Zongsheng Zhao, Wei LiAbstract:Abstract The gene, SLC7A11, which encodes the solute carrier family 7 member 11 (anionic amino acid transporter light chain, xCT), has been reported to be implicated in multiple processes such as in pheomelanin production, cell proliferation and migration, Kaposi's sarcoma herpesvirus (KSHV) entry into the host cells, learning and memory. Its involvement in cancer cell proliferation and metastasis has been widely studied. Its role in pheomelanogenesis is likely conserved in Sheep. The full-length cDNA of Sheep SLC7A11 was cloned from Sheep skin fibroblasts for evaluating its role in regulating Sheep coat color. The complete open reading frame of Sheep xCT (sxCT) is 1512 bp in length, encoding a 503 amino acid polypeptide. We explored its function on pheomelanogenesis in vitro and in vivo. In the melan-a non-agouti mouse melanocytes that mainly produce eumelanin, overexpressed sxCT reduced the content of eumelanin. Using a testicular injection Transgenic method, sxCT-Transgenic Sheep were generated and exhibited patches of brown/yellow coat, suggesting that sxCT can be selectively expressed to increase the pheomelanin production in wool. Our studies suggest that testicular injection of transgene can be used to genetically modify Sheep coat color.
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cloning of xuhuai goat lipoprotein lipase gene and the preparation of Transgenic Sheep
Molecular Biology Reports, 2012Co-Authors: Yani Zhang, Wei Li, Feng Xu, Bichun LiAbstract:This paper presents cloning of cDNA of lipoprotein lipase (LPL) gene from Xuhuai goat, and the sub-cellular localization analysis through enhanced green fluorescent (EGFP) fusion protein. cDNA was cloned by reverse transcription polymerase chain reaction (RT-PCR). Fusion expression vector named pEGFP–LPL was constructed successfully. Then NIH-3T3 cells were transfected with pEGFP–LPL through polyethylene imine and observed under inverted microscope after 48 h transfection. The RT-PCR was performed to analysis the level of expression of mRNA. The complete coding sequence (1,530 bp) of LPL was acquired, and the open reading frame size was 1,437 bp with a capacity to encode 478 amino acids. The prediction of signal peptide region showed that LPL protein contained a short signal peptide with a probability of 100 %, and the signal peptidase cleavage site located between the 23rd and the 24th amino acid with a probability of 65.9 %. RT-PCR results showed the LPL mRNA expressed successfully in vitro. Sub-cellullar localization analysis showed that pEGFP–LPL fusion protein located at the cytoplasm. LPL gene of Xuhuai goat was transfer into Sheep by testicular injection. According to detection from different level, the LPL gene was expressed successfully in F1 generation.
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Production of brown/yellow patches in the SLC7A11 Transgenic Sheep via testicular injection of transgene.
Journal of Genetics and Genomics, 2012Co-Authors: Xin He, Hongtao Li, Zhiyong Zhou, Zongsheng Zhao, Wei LiAbstract:Abstract The gene, SLC7A11, which encodes the solute carrier family 7 member 11 (anionic amino acid transporter light chain, xCT), has been reported to be implicated in multiple processes such as in pheomelanin production, cell proliferation and migration, Kaposi's sarcoma herpesvirus (KSHV) entry into the host cells, learning and memory. Its involvement in cancer cell proliferation and metastasis has been widely studied. Its role in pheomelanogenesis is likely conserved in Sheep. The full-length cDNA of Sheep SLC7A11 was cloned from Sheep skin fibroblasts for evaluating its role in regulating Sheep coat color. The complete open reading frame of Sheep xCT (sxCT) is 1512 bp in length, encoding a 503 amino acid polypeptide. We explored its function on pheomelanogenesis in vitro and in vivo. In the melan-a non-agouti mouse melanocytes that mainly produce eumelanin, overexpressed sxCT reduced the content of eumelanin. Using a testicular injection Transgenic method, sxCT-Transgenic Sheep were generated and exhibited patches of brown/yellow coat, suggesting that sxCT can be selectively expressed to increase the pheomelanin production in wool. Our studies suggest that testicular injection of transgene can be used to genetically modify Sheep coat color.
