Establishment and evaluation of a PRRSV-sensitive porcine endometrial epithelial cell line by transfecting SV40 large T antigen
Abstract Background PRRSV is an infectious illness causing lung injury and abortion in sows. Cells apoptosis in the interface between the endometrium and fetal placenta is a crucial factor causing abortion. Previous study confirmed PRRSV could cause apoptosis of macrophages but rarely produced an ob...
Main Authors: | , , , , , , , , |
---|---|
Format: | Article |
Language: | English |
Published: |
BMC
2019-08-01
|
Series: | BMC Veterinary Research |
Subjects: | |
Online Access: | http://link.springer.com/article/10.1186/s12917-019-2051-1 |
id |
doaj-c67161e2e87d4b398bbd842f2cd22558 |
---|---|
record_format |
Article |
spelling |
doaj-c67161e2e87d4b398bbd842f2cd225582020-11-25T03:18:29ZengBMCBMC Veterinary Research1746-61482019-08-0115111010.1186/s12917-019-2051-1Establishment and evaluation of a PRRSV-sensitive porcine endometrial epithelial cell line by transfecting SV40 large T antigenKang Zhang0Houshen Li1Shasha Dong2Ying Liu3Dong Wang4Haichang Liu5Feng Su6Lijiang Ge7Yunliang Jiang8Shandong Provincial Key Laboratory of Animal Biotechnology and Disease Control and Prevention, College of Animal Science and Veterinary Medicine, Shandong Agricultural UniversityCollege of Chemistry and Material Science, Shandong Agricultural UniversityDepartment of Cardiology, Shandong First Medical University and Shandong Academy of Medical ScienceShandong Provincial Key Laboratory of Animal Biotechnology and Disease Control and Prevention, College of Animal Science and Veterinary Medicine, Shandong Agricultural UniversityShandong Provincial Key Laboratory of Animal Biotechnology and Disease Control and Prevention, College of Animal Science and Veterinary Medicine, Shandong Agricultural UniversityNingbo Defangyuan Biotech Co. Ltd.Shandong Provincial Key Laboratory of Animal Biotechnology and Disease Control and Prevention, College of Animal Science and Veterinary Medicine, Shandong Agricultural UniversityShandong Provincial Key Laboratory of Animal Biotechnology and Disease Control and Prevention, College of Animal Science and Veterinary Medicine, Shandong Agricultural UniversityShandong Provincial Key Laboratory of Animal Biotechnology and Disease Control and Prevention, College of Animal Science and Veterinary Medicine, Shandong Agricultural UniversityAbstract Background PRRSV is an infectious illness causing lung injury and abortion in sows. Cells apoptosis in the interface between the endometrium and fetal placenta is a crucial factor causing abortion. Previous study confirmed PRRSV could cause apoptosis of macrophages but rarely produced an obvious change in porcine endometrial epithelial cells (PECs). Recently, PRRSV-induced abortion was attributed to fetal placental and endometrium epithelial cells (Sn+ and CD163+) apoptosis. However, the mechanism of abortion is still unrevealed because of the limit of porcine endometrium epithelial cells (PEC). The aim of this study was to establish a stable immortalized PECs lines and use it to reveal the abortion mechanism. Results In this study, highly purified primary PECs were harvested through differential digestion, and their characteristics were confirmed by CK18, ERɑ and PR staining. Cells were then immortalized by transfecting a lentiviral vector that expressed SV40 large T antigen. PECs lines were obtained after puromycin screening. Proliferation of cell line was evaluated by cell growth curve and cell cycle assays. Cell lines exhibited faster proliferation capacity than primary cells. Biological characteristics of cell line were assessed by Western blot, karyotype analysis and staining, which confirmed that the cell line retained the endometrium characteristics. Finally, PRRSV sensitivity was assessed; expression of Sn and CD163 indicated that primary PECs and cell lines were all potentially sensitive to PRRSV. PRRSV infection tests showed an obvious increase in apoptotic rate in the infected PEC cell line, which suggested its susceptibility. Conclusion The newly constructed cell line is a useful tool for studying the mechanism of abortion caused by PRRSV.http://link.springer.com/article/10.1186/s12917-019-2051-1Porcine endometrial epithelial cell lineSV40T antigenPRRSV-sensitivityLentrival vectorReplication efficiency |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Kang Zhang Houshen Li Shasha Dong Ying Liu Dong Wang Haichang Liu Feng Su Lijiang Ge Yunliang Jiang |
spellingShingle |
Kang Zhang Houshen Li Shasha Dong Ying Liu Dong Wang Haichang Liu Feng Su Lijiang Ge Yunliang Jiang Establishment and evaluation of a PRRSV-sensitive porcine endometrial epithelial cell line by transfecting SV40 large T antigen BMC Veterinary Research Porcine endometrial epithelial cell line SV40T antigen PRRSV-sensitivity Lentrival vector Replication efficiency |
author_facet |
Kang Zhang Houshen Li Shasha Dong Ying Liu Dong Wang Haichang Liu Feng Su Lijiang Ge Yunliang Jiang |
author_sort |
Kang Zhang |
title |
Establishment and evaluation of a PRRSV-sensitive porcine endometrial epithelial cell line by transfecting SV40 large T antigen |
title_short |
Establishment and evaluation of a PRRSV-sensitive porcine endometrial epithelial cell line by transfecting SV40 large T antigen |
title_full |
Establishment and evaluation of a PRRSV-sensitive porcine endometrial epithelial cell line by transfecting SV40 large T antigen |
title_fullStr |
Establishment and evaluation of a PRRSV-sensitive porcine endometrial epithelial cell line by transfecting SV40 large T antigen |
title_full_unstemmed |
Establishment and evaluation of a PRRSV-sensitive porcine endometrial epithelial cell line by transfecting SV40 large T antigen |
title_sort |
establishment and evaluation of a prrsv-sensitive porcine endometrial epithelial cell line by transfecting sv40 large t antigen |
publisher |
BMC |
series |
BMC Veterinary Research |
issn |
1746-6148 |
publishDate |
2019-08-01 |
description |
Abstract Background PRRSV is an infectious illness causing lung injury and abortion in sows. Cells apoptosis in the interface between the endometrium and fetal placenta is a crucial factor causing abortion. Previous study confirmed PRRSV could cause apoptosis of macrophages but rarely produced an obvious change in porcine endometrial epithelial cells (PECs). Recently, PRRSV-induced abortion was attributed to fetal placental and endometrium epithelial cells (Sn+ and CD163+) apoptosis. However, the mechanism of abortion is still unrevealed because of the limit of porcine endometrium epithelial cells (PEC). The aim of this study was to establish a stable immortalized PECs lines and use it to reveal the abortion mechanism. Results In this study, highly purified primary PECs were harvested through differential digestion, and their characteristics were confirmed by CK18, ERɑ and PR staining. Cells were then immortalized by transfecting a lentiviral vector that expressed SV40 large T antigen. PECs lines were obtained after puromycin screening. Proliferation of cell line was evaluated by cell growth curve and cell cycle assays. Cell lines exhibited faster proliferation capacity than primary cells. Biological characteristics of cell line were assessed by Western blot, karyotype analysis and staining, which confirmed that the cell line retained the endometrium characteristics. Finally, PRRSV sensitivity was assessed; expression of Sn and CD163 indicated that primary PECs and cell lines were all potentially sensitive to PRRSV. PRRSV infection tests showed an obvious increase in apoptotic rate in the infected PEC cell line, which suggested its susceptibility. Conclusion The newly constructed cell line is a useful tool for studying the mechanism of abortion caused by PRRSV. |
topic |
Porcine endometrial epithelial cell line SV40T antigen PRRSV-sensitivity Lentrival vector Replication efficiency |
url |
http://link.springer.com/article/10.1186/s12917-019-2051-1 |
work_keys_str_mv |
AT kangzhang establishmentandevaluationofaprrsvsensitiveporcineendometrialepithelialcelllinebytransfectingsv40largetantigen AT houshenli establishmentandevaluationofaprrsvsensitiveporcineendometrialepithelialcelllinebytransfectingsv40largetantigen AT shashadong establishmentandevaluationofaprrsvsensitiveporcineendometrialepithelialcelllinebytransfectingsv40largetantigen AT yingliu establishmentandevaluationofaprrsvsensitiveporcineendometrialepithelialcelllinebytransfectingsv40largetantigen AT dongwang establishmentandevaluationofaprrsvsensitiveporcineendometrialepithelialcelllinebytransfectingsv40largetantigen AT haichangliu establishmentandevaluationofaprrsvsensitiveporcineendometrialepithelialcelllinebytransfectingsv40largetantigen AT fengsu establishmentandevaluationofaprrsvsensitiveporcineendometrialepithelialcelllinebytransfectingsv40largetantigen AT lijiangge establishmentandevaluationofaprrsvsensitiveporcineendometrialepithelialcelllinebytransfectingsv40largetantigen AT yunliangjiang establishmentandevaluationofaprrsvsensitiveporcineendometrialepithelialcelllinebytransfectingsv40largetantigen |
_version_ |
1724626496449937408 |