Please wait a minute...

Frontiers of Agricultural Science and Engineering

Front. Agr. Sci. Eng.    2014, Vol. 1 Issue (4) : 314-320     https://doi.org/10.15302/J-FASE-2014038
RESEARCH ARTICLE |
Effects of the TLR4 transgene on reproductive traits and DNA methylation pattern of oocytes in ewes
Yi FANG1,Xiangwei FU1,Junjie LI2,Ming DU1,Baoyu JIA1,Jinlong ZHANG3,Xiaosheng ZHANG3,Shien ZHU1,*()
1. National Engineering Laboratory for Animal Breeding and Key Laboratory of Animal Genetics, Breeding and Reproduction of Ministry of Agriculture, College of Animal Science and Technology, China Agricultural University, Beijing 100193, China
2. College of Animal Science and Technology, Agricultural University of Hebei, Baoding 071000, China
3. Animal Husbandry and Veterinary Research Institute of Tianjin, Tianjin 300412, China
Download: PDF(378 KB)   HTML
Export: BibTeX | EndNote | Reference Manager | ProCite | RefWorks
Abstract

This study was conducted to systematically assess the reproductive performance of transgenic TLR4 ewes. In the TLR4 transgenic founders (F0) and their positive offspring (F1), hematological and reproductive parameters and the global DNA methylation level in oocytes at various stages were analyzed. The values of the physiological and biochemical parameters determined from the blood samples did not differ significantly between the transgenic and wild-type ewes. Moreover, the transgenic ewes showed reproductive traits similar to the wild-type ewes. These traits included characteristics of puberty, the estrus cycle, estrus duration, gestation, the pregnancy rate and the superovulation response. Additionally, no significant differences were found between transgenic and wild-type ewes in the DNA methylation level of the oocytes at various stages. In summary, the preliminary evidence presented in this paper demonstrates that the presence of the TLR4 transgene did not affect the reproductive performance in sheep.

Keywords TLR4 transgenic ewe      safety assessment      reproductive trait      oocyte      DNA methylation     
Corresponding Authors: Shien ZHU   
Online First Date: 28 January 2015    Issue Date: 10 March 2015
 Cite this article:   
Yi FANG,Xiangwei FU,Junjie LI, et al. Effects of the TLR4 transgene on reproductive traits and DNA methylation pattern of oocytes in ewes[J]. Front. Agr. Sci. Eng. , 2014, 1(4): 314-320.
 URL:  
http://journal.hep.com.cn/fase/EN/10.15302/J-FASE-2014038
http://journal.hep.com.cn/fase/EN/Y2014/V1/I4/314
Service
E-mail this article
E-mail Alert
RSS
Articles by authors
Yi FANG
Xiangwei FU
Junjie LI
Ming DU
Baoyu JIA
Jinlong ZHANG
Xiaosheng ZHANG
Shien ZHU
Physiological Parameters F0 F1
TG (n = 5) WT (n = 5) TG (n = 7) WT (n = 6)
RBC/(1012·L-1) 12.01±0.79 12.32±0.45 12.22±0.48 12.15±0.65
WBC/(109·L-1) 18.45±3.44 20.06±3.51 17.44±2.98 20.75±4.05
HGB/(g·L-1) 112.79±8.88 107.58±5.19 110.40±4.17 110.00±12.06
HCT/% 35.15±1.28 32.77±2.48 34.34±1.15 34.95±4.15
Tab.1  Physiological parameters of TG and WT ewes in F0 and F1
Biochemical Parameters F0 F1
TG (n = 5) WT (n = 5) TG (n = 7) WT (n = 6)
TP/(g·L-1) 75.19±4.41 77.04±5.39 78.98±6.58 68.10±4.81
ALB/(g·L-1) 34.42±6.11 37.40±2.22 31.68±1.19 35.90±3.34
GLO/(g·L-1) 35.38±1.25 34.62±3.41 35.90±3.30 32.20±1.55
ALT/μL 15.29±3.29 14.38±2.77 16.67±4.12 14.50±1.52
AST/μL 151.49±10.01 139.47±7.09 148.00±12.48 133.50±7.59
BUN/(mmol·L-1) 6.66±0.42 6.01±0.29 6.60±0.23 5.85±0.35
GLU/(mmol·L-1) 3.44±0.14 3.51±0.17 3.32±0.13 3.46±0.14
TG/(mmol·L-1) 0.27±0.03 0.27±0.04 0.26±0.04 0.30±0.06
Tab.2  Biochemical parameters of TG and WT ewes in F0 and F1
Traits F0 F1
TG (n = 5) WT (n = 5) TG (n = 7) WT (n = 6)
Puberty(days) 143.91±6.14(132-151) 145.05±5.28(128-149) 145.44±6.14(131-153) 144.05±4.28(135-149)
Estrus cycle(days) 17.05±1.31(15-19) 17.87±0.36(16-18) 17.42±0.94(16-20) 17.55±0.99(16-18)
Estrus duration(hours) 33.65±2.45(24-39) 32.9±3.53(19-40) 32.65±3.31(18-41) 34.01±4.13(17-38)
Gestation(days) 148.47±1.15(145-153) 151.95±1.29(146-155) 148.41±1.67(144-152) 150.95±1.22(144-153)
Total pregnancy rate/% 100% (5/5) 100% (5/5) 100% (7/7) 100% (6/6)
Tab.3  The basic reproductive parameters of TG and WT ewes in F0 and F1
Superovulation parameters F0 F1
TG (n = 4) WT (n = 4) TG (n = 4) WT (n = 4)
Average no. of corpora lutea 12.33±2.80 15.26±1.48 16.37±3.42 14.91±1.77
Average no. of oocytes recovered 8.50±1.12 11.31±1.83 14.84±2.15 13.44±0.91
Tab.4  Superovulation response of TG and WT ewes in F0 and F1
Fig.1  Typical picture of 5-MeC patterns in TG and WT oocytes. (a)-(d) Immunofluorescence results on stage GV, GVBD, MI and MII oocytes, respectively (left, blue), PI-stained nuclei (middle), merged (right). Bar= 20 μm
Fig.2  Effect of TLR4 transgene on global DNA methylation levels in oocytes at various stages in F0 and F1. Note: The average fluorescent intensity of GV oocyte from TG group was set as 1 in (a) and (b).
1 Doblhoff-Dier O, Collins C H. Biosafety: future priorities for research in health care. Journal of Biotechnology, 2001, 85(2): 227-239
https://doi.org/10.1016/S0168-1656(00)00362-X pmid: 11165365
2 Clark J, Whitelaw B. A future for transgenic livestock. Nature Reviews. Genetics, 2003, 4(10): 825-833
https://doi.org/10.1038/nrg1183 pmid: 14526378
3 Einsiedel E F. Public perceptions of transgenic animals. Revue Scientifique et Technique (International Office of Epizootics), 2005, 24(1): 149-157
pmid: 16110885
4 Van Reenen C G, Meuwissen T H, Hopster H, Oldenbroek K, Kruip T H, Blokhuis H J. Transgenesis may affect farm animal welfare: a case for systematic risk assessment. Journal of Animal Science, 2001, 79(7): 1763-1779
pmid: 11465364
5 van der Meer M, Rolls A, Baumans V, Olivier B, van Zutphen L F. Use of score sheets for welfare assessment of transgenic mice. Laboratory Animals, 2001, 35(4): 379-389
https://doi.org/10.1258/0023677011911859 pmid: 11669323
6 Webster J. The assessment and implementation of animal welfare: theory into practice. Revue Scientifique et Technique (International Office of Epizootics), 2005, 24(2): 723-734
pmid: 16358522
7 Food and Agriculture Organization (FAO). Safety assessment of foods derived from genetically modified animals, including fish. Rome: FAO Food and Nutrition Paper, 2004, 79: 1-36
8 Jackson K A, Berg J M, Murray J D, Maga E A. Evaluating the fitness of human lysozyme transgenic dairy goats: growth and reproductive traits. Transgenic Research, 2010, 19(6): 977-986
https://doi.org/10.1007/s11248-010-9371-z pmid: 20135222
9 Brundige D R, Maga E A, Klasing K C, Murray J D. Lysozyme transgenic goats’ milk influences gastrointestinal morphology in young pigs. The Journal of Nutrition, 2008, 138(5): 921-926 18424602
https://doi.org/18424602
10 Brundige D R, Maga E A, Klasing K C, Murray J D. Consumption of pasteurized human lysozyme transgenic goats’ milk alters serum metabolite profile in young pigs. Transgenic Research, 2010, 19(4): 563-574
https://doi.org/10.1007/s11248-009-9334-4 pmid: 19847666
11 Tang M, Zheng X, Cheng W, Jin E, Chen H, Yang S, Cui W, Li K. Safety assessment of sFat-1 transgenic pigs by detecting their co-habitant microbe in intestinal tract. Transgenic Research, 2011, 20(4): 749-758
https://doi.org/10.1007/s11248-010-9457-7 pmid: 21082244
12 Xu J, Zhao J, Wang J, Zhao Y, Zhang L, Chu M, Li N. Molecular-based environmental risk assessment of three varieties of genetically engineered cows. Transgenic Research, 2011, 20(5): 1043-1054
https://doi.org/10.1007/s11248-010-9477-3 pmid: 21221780
13 Zhao J, Xu J, Wang J, Zhao Y, Zhang L, He J, Chu M, Li N. Impacts of human lysozyme transgene on the microflora of pig feces and the surrounding soil. Journal of Biotechnology, 2012, 161(4): 437-444
https://doi.org/10.1016/j.jbiotec.2012.05.018 pmid: 22750647
14 Huber R C, Remuge L, Carlisle A, Lillico S, Sand?e P, S?rensen D B, Whitelaw C B, Olsson I A. Welfare assessment in transgenic pigs expressing green fluorescent protein (GFP). Transgenic Research, 2012, 21(4): 773-784
https://doi.org/10.1007/s11248-011-9571-1 pmid: 22173943
15 Deppenmeier S, Bock O, Mengel M, Niemann H, Kues W, Lemme E, Wirth D, Wonigeit K, Kreipe H. Health status of transgenic pigs expressing the human complement regulatory protein CD59. Xenotransplantation, 2006, 13(4): 345-356
https://doi.org/10.1111/j.1399-3089.2006.00317.x pmid: 16768728
16 Maga E A, Murray J D. Welfare applications of genetically engineered animals for use in agriculture. Journal of Animal Science, 2010, 88(4): 1588-1591
https://doi.org/10.2527/jas.2010-2828 pmid: 20154173
17 Cao Z, Li Y, Wen X, Li Z, Mi C, Zhang Z, Li N, Li Q. Recloned transgenic pigs possess normal reproductive performance and stable genetic transmission capacity. Zygote, 2014, 22(1): 18-24
https://doi.org/10.1017/S0967199412000238 pmid: 22784554
18 Merlino G T, Stahle C, Jhappan C, Linton R, Mahon K A, Willingham M C. Inactivation of a sperm motility gene by insertion of an epidermal growth factor receptor transgene whose product is overexpressed and compartmentalized during spermatogenesis. Genes & Development, 1991, 5(8): 1395-1406
https://doi.org/10.1101/gad.5.8.1395 pmid: 1714416
19 Pellas T C, Ramachandran B, Duncan M, Pan S S, Marone M, Chada K. Germ-cell deficient (gcd), an insertional mutation manifested as infertility in transgenic mice. Proceedings of the National Academy of Sciences of the United States of America, 1991, 88(19): 8787-8791
https://doi.org/10.1073/pnas.88.19.8787 pmid: 1924340
20 Soriano P, Gridley T, Jaenisch R. Retroviruses and insertional mutagenesis in mice: proviral integration at the Mov 34 locus leads to early embryonic death. Genes & Development, 1987, 1(4): 366-375
https://doi.org/10.1101/gad.1.4.366 pmid: 2824282
21 Oliveri R S, Kalisz M, Schjerling C K, Andersen C Y, Borup R, Byskov A G. Evaluation in mammalian oocytes of gene transcripts linked to epigenetic reprogramming. Reproduction, 2007, 134(4): 549-558
https://doi.org/10.1530/REP-06-0315 pmid: 17890290
22 Kues W A, Schwinzer R, Wirth D, Verhoeyen E, Lemme E, Herrmann D, Barg-Kues B, Hauser H, Wonigeit K, Niemann H. Epigenetic silencing and tissue independent expression of a novel tetracycline inducible system in double-transgenic pigs. The FASEB Journal, 2006, 20(8): 1200-1202
https://doi.org/10.1096/fj.05-5415fje pmid: 16684801
23 Hofmann A, Kessler B, Ewerling S, Kabermann A, Brem G, Wolf E, Pfeifer A. Epigenetic regulation of lentiviral transgene vectors in a large animal model. Molecular Therapy, 2006, 13(1): 59-66
https://doi.org/10.1016/j.ymthe.2005.07.685 pmid: 16140581
24 Reik W, R?mer I, Barton S C, Surani M A, Howlett S K, Klose J. Adult phenotype in the mouse can be affected by epigenetic events in the early embryo. Development, 1993, 119(3): 933-942
pmid: 8187648
25 Reik W, Dean W, Walter J. Epigenetic reprogramming in mammalian development. Science, 2001, 293(5532): 1089-1093
https://doi.org/10.1126/science.1063443 pmid: 11498579
26 Yue M, Fu X, Zhou G, Hou Y, Du M, Wang L, Zhu S. Abnormal DNA methylation in oocytes could be associated with a decrease in reproductive potential in old mice. Journal of Assisted Reproduction and Genetics, 2012, 29(7): 643-650
https://doi.org/10.1007/s10815-012-9780-4 pmid: 22618193
27 Takeda K, Akira S. Toll-like receptors in innate immunity. International Immunology, 2005, 17(1): 1-14
https://doi.org/10.1093/intimm/dxh186 pmid: 15585605
28 Borjesson D L, Christopher M M, Boyce W M. Biochemical and hematologic reference intervals for free-ranging desert bighorn sheep. Journal of Wildlife Diseases, 2000, 36(2): 294-300
https://doi.org/10.7589/0090-3558-36.2.294 pmid: 10813611
29 Deng S, Yu K, Zhang B, Yao Y, Liu Y, He H, Zhang H, Cui M, Fu J, Lian Z, Li N. Effects of over-expression of TLR2 in transgenic goats on pathogen clearance and role of up-regulation of lysozyme secretion and infiltration of inflammatory cells. BMC Veterinary Research, 2012a, 8(1): 196
https://doi.org/10.1186/1746-6148-8-196 pmid: 23082910
30 Garrels W, Holler S, Cleve N, Niemann H, Ivics Z, Kues W A. Assessment of fecundity and germ line transmission in two transgenic pig lines produced by sleeping beauty transposition. Genes, 2012, 3(4): 615-633
https://doi.org/10.3390/genes3040615 pmid: 24705079
31 Jungi T W, Farhat K, Burgener I A, Werling D. Toll-like receptors in domestic animals. Cell and Tissue Research, 2011, 343(1): 107-120
https://doi.org/10.1007/s00441-010-1047-8 pmid: 20927536
32 Kannaki T R, Shanmugam M, Verma P C. Toll-like receptors and their role in animal reproduction. Animal Reproduction Science, 2011, 125(1-4): 1-12
https://doi.org/10.1016/j.anireprosci.2011.03.008 pmid: 21497464
33 Müller M, Brem G. Transgenic approaches to the increase of disease resistance in farm animals. Revue Scientifique et Technique (International Office of Epizootics), 1998, 17(1): 365-378
pmid: 9638824
34 Girling J E, Hedger M P. Toll-like receptors in the gonads and reproductive tract: emerging roles in reproductive physiology and pathology. Immunology and Cell Biology, 2007, 85(6): 481-489
https://doi.org/10.1038/sj.icb.7100086 pmid: 17592495
35 Deng S, Wu Q, Yu K, Zhang Y, Yao Y, Li W, Deng Z, Liu G, Li W, Lian Z. Changes in the relative inflammatory responses in sheep cells overexpressing of toll-like receptor 4 when stimulated with LPS. PLoS ONE, 2012a, 7(10): e47118
https://doi.org/10.1371/journal.pone.0047118 pmid: 23056598
Related articles from Frontiers Journals
[1] Shen YIN,Junyu MA,Wei SHEN. Effects of DNA damage on oocyte meiotic maturation and early embryonic development[J]. Front. Agr. Sci. Eng. , 2014, 1(3): 185-190.
Viewed
Full text


Abstract

Cited

  Shared   
  Discussed