Oxysterols protect bovine endometrial cells against pore‐forming toxins from pathogenic bacteria

R., Thomas J.; Owens, Sian-eleri; Horlock, Anthony D.; Davies, Daphne; Griffiths, William; Wang, Yuqin; Cronin, James; Bromfield, John J.; Sheldon, Martin; Ormsby, Thomas

doi:10.1096/fj.202100036r

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Oxysterols protect bovine endometrial cells against pore‐forming toxins from pathogenic bacteria

Thomas J. R. Ormsby, Sian-eleri Owens

, Anthony D. Horlock, Daphne Davies, William Griffiths

, Yuqin Wang

, James Cronin

, John J. Bromfield, Martin Sheldon

, Thomas Ormsby

The FASEB Journal, Volume: 35, Issue: 10, Pages: 1 - 21

Swansea University Authors: Sian-eleri Owens , William Griffiths , Yuqin Wang , James Cronin , Martin Sheldon , Thomas Ormsby

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DOI (Published version): 10.1096/fj.202100036r

Abstract

Many species of pathogenic bacteria secrete toxins that form pores in mammalian cell membranes. These membrane pores enable the delivery of virulence factors into cells, result in the leakage of molecules that bacteria can use as nutrients, and facilitate pathogen invasion. Inflammatory responses to...

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Published in:	The FASEB Journal
ISSN:	0892-6638 1530-6860
Published:	Wiley 2021
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URI:	https://cronfa.swan.ac.uk/Record/cronfa57632
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These membrane pores enable the delivery of virulence factors into cells, result in the leakage of molecules that bacteria can use as nutrients, and facilitate pathogen invasion. Inflammatory responses to bacteria are regulated by the side-chain-hydroxycholesterols 27-hydroxycholesterol and 25-hydroxycholesterol, but their effect on the intrinsic protection of cells against pore-forming toxins is unclear. Here, we tested the hypothesis that 27-hydroxycholesterol and 25-hydroxycholesterol help protect cells against pore-forming toxins. We treated bovine endometrial epithelial and stromal cells with 27-hydroxycholesterol or 25-hydroxycholesterol, and then challenged the cells with pyolysin, which is a cholesterol-dependent cytolysin from Trueperella pyogenes that targets these endometrial cells. We found that treatment with 27-hydroxycholesterol or 25-hydroxycholesterol protected both epithelial and stomal cells against pore formation and the damage caused by pyolysin. The oxysterols limited pyolysin-induced leakage of potassium and lactate dehydrogenase from cells, and reduced cytoskeletal changes and cytolysis. This oxysterol cytoprotection against pyolysin was partially dependent on reducing cytolysin-accessible cholesterol in the cell membrane and on activating liver X receptors. Treatment with 27-hydroxycholesterol also protected the endometrial cells against Staphylococcus aureus α hemolysin. Using mass spectrometry, we found 27-hydroxycholesterol and 25-hydroxycholesterol in uterine and follicular fluid. Furthermore, epithelial cells released additional 25-hydroxycholesterol in response to pyolysin. In conclusion, both 27-hydroxycholesterol and 25-hydroxycholesterol increased the intrinsic protection of bovine endometrial cells against pore-forming toxins. Our findings imply that side-chain-hydroxycholesterols may help defend the endometrium against pathogenic bacteria.</abstract><type>Journal Article</type><journal>The FASEB Journal</journal><volume>35</volume><journalNumber>10</journalNumber><paginationStart>1</paginationStart><paginationEnd>21</paginationEnd><publisher>Wiley</publisher><placeOfPublication/><isbnPrint/><isbnElectronic/><issnPrint>0892-6638</issnPrint><issnElectronic>1530-6860</issnElectronic><keywords>Cattle, cholesterol, cytoprotection, liver X receptor, uterus</keywords><publishedDay>1</publishedDay><publishedMonth>10</publishedMonth><publishedYear>2021</publishedYear><publishedDate>2021-10-01</publishedDate><doi>10.1096/fj.202100036r</doi><url>http://dx.doi.org/10.1096/fj.202100036r</url><notes/><college>COLLEGE NANME</college><department>Biomedical Sciences</department><CollegeCode>COLLEGE CODE</CollegeCode><DepartmentCode>BMS</DepartmentCode><institution>Swansea University</institution><apcterm>SU Library paid the OA fee (TA Institutional Deal)</apcterm><funders>This study was supported by Eunice Kennedy Shriver National Institute of Child Health & Human Development of the National Institutes of Health under Award Number R01HD084316. Instrumentation for oxysterol analysis was funded by Biotechnology and Biological Sciences Research Council grants BB/I001735/1 and BB/L001942/1, and the European Union via European Structural Funds as part of the Welsh Government funded Academic Expertise for Business project HE09161003.</funders><projectreference>UKRI - BB/K006592/1; BB/L001942/1; BB/I001735/1</projectreference><lastEdited>2022-02-15T08:50:53.2255343</lastEdited><Created>2021-08-18T09:13:20.7290773</Created><path><level id="1">Swansea University Medical School</level><level id="2">Swansea University Medical School</level></path><authors><author><firstname>Thomas J. R.</firstname><surname>Ormsby</surname><order>1</order></author><author><firstname>Sian-eleri</firstname><surname>Owens</surname><orcid>0000-0003-1806-5235</orcid><order>2</order></author><author><firstname>Anthony D.</firstname><surname>Horlock</surname><order>3</order></author><author><firstname>Daphne</firstname><surname>Davies</surname><order>4</order></author><author><firstname>William</firstname><surname>Griffiths</surname><orcid>0000-0002-4129-6616</orcid><order>5</order></author><author><firstname>Yuqin</firstname><surname>Wang</surname><orcid>0000-0002-3063-3066</orcid><order>6</order></author><author><firstname>James</firstname><surname>Cronin</surname><orcid>0000-0002-0590-9462</orcid><order>7</order></author><author><firstname>John J.</firstname><surname>Bromfield</surname><order>8</order></author><author><firstname>Martin</firstname><surname>Sheldon</surname><orcid>0000-0001-7902-5558</orcid><order>9</order></author><author><firstname>Thomas</firstname><surname>Ormsby</surname><order>10</order></author></authors><documents><document><filename>57632__21051__c8145150a083454ca694b73984060628.pdf</filename><originalFilename>ORMSBYfj202100036R.pdf</originalFilename><uploaded>2021-09-29T12:40:44.0642284</uploaded><type>Output</type><contentLength>3779382</contentLength><contentType>application/pdf</contentType><version>Version of Record</version><cronfaStatus>true</cronfaStatus><documentNotes>This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited. © 2021 The Authors.</documentNotes><copyrightCorrect>true</copyrightCorrect><language>eng</language><licence>http://creativecommons.org/licenses/by/4.0/</licence></document></documents><OutputDurs/></rfc1807>
spelling	2022-02-15T08:50:53.2255343 v2 57632 2021-08-18 Oxysterols protect bovine endometrial cells against pore‐forming toxins from pathogenic bacteria 721deb4604d122019244cfdf08820cbe 0000-0003-1806-5235 Sian-eleri Owens Sian-eleri Owens true false 3316b1d1b524be1831790933eed1c26e 0000-0002-4129-6616 William Griffiths William Griffiths true false c92729b58622f9fdf6a0e7d8f4ce5081 0000-0002-3063-3066 Yuqin Wang Yuqin Wang true false 9cfd17551c0d1f7438895121e4fbb6e8 0000-0002-0590-9462 James Cronin James Cronin true false ab0f74b794e59cc270c69e63ee1d9748 0000-0001-7902-5558 Martin Sheldon Martin Sheldon true false c8a0f774d8ff66a2193a035018cc6cda Thomas Ormsby Thomas Ormsby true false 2021-08-18 BMS Many species of pathogenic bacteria secrete toxins that form pores in mammalian cell membranes. These membrane pores enable the delivery of virulence factors into cells, result in the leakage of molecules that bacteria can use as nutrients, and facilitate pathogen invasion. Inflammatory responses to bacteria are regulated by the side-chain-hydroxycholesterols 27-hydroxycholesterol and 25-hydroxycholesterol, but their effect on the intrinsic protection of cells against pore-forming toxins is unclear. Here, we tested the hypothesis that 27-hydroxycholesterol and 25-hydroxycholesterol help protect cells against pore-forming toxins. We treated bovine endometrial epithelial and stromal cells with 27-hydroxycholesterol or 25-hydroxycholesterol, and then challenged the cells with pyolysin, which is a cholesterol-dependent cytolysin from Trueperella pyogenes that targets these endometrial cells. We found that treatment with 27-hydroxycholesterol or 25-hydroxycholesterol protected both epithelial and stomal cells against pore formation and the damage caused by pyolysin. The oxysterols limited pyolysin-induced leakage of potassium and lactate dehydrogenase from cells, and reduced cytoskeletal changes and cytolysis. This oxysterol cytoprotection against pyolysin was partially dependent on reducing cytolysin-accessible cholesterol in the cell membrane and on activating liver X receptors. Treatment with 27-hydroxycholesterol also protected the endometrial cells against Staphylococcus aureus α hemolysin. Using mass spectrometry, we found 27-hydroxycholesterol and 25-hydroxycholesterol in uterine and follicular fluid. Furthermore, epithelial cells released additional 25-hydroxycholesterol in response to pyolysin. In conclusion, both 27-hydroxycholesterol and 25-hydroxycholesterol increased the intrinsic protection of bovine endometrial cells against pore-forming toxins. Our findings imply that side-chain-hydroxycholesterols may help defend the endometrium against pathogenic bacteria. Journal Article The FASEB Journal 35 10 1 21 Wiley 0892-6638 1530-6860 Cattle, cholesterol, cytoprotection, liver X receptor, uterus 1 10 2021 2021-10-01 10.1096/fj.202100036r http://dx.doi.org/10.1096/fj.202100036r COLLEGE NANME Biomedical Sciences COLLEGE CODE BMS Swansea University SU Library paid the OA fee (TA Institutional Deal) This study was supported by Eunice Kennedy Shriver National Institute of Child Health & Human Development of the National Institutes of Health under Award Number R01HD084316. Instrumentation for oxysterol analysis was funded by Biotechnology and Biological Sciences Research Council grants BB/I001735/1 and BB/L001942/1, and the European Union via European Structural Funds as part of the Welsh Government funded Academic Expertise for Business project HE09161003. UKRI - BB/K006592/1; BB/L001942/1; BB/I001735/1 2022-02-15T08:50:53.2255343 2021-08-18T09:13:20.7290773 Swansea University Medical School Swansea University Medical School Thomas J. R. Ormsby 1 Sian-eleri Owens 0000-0003-1806-5235 2 Anthony D. Horlock 3 Daphne Davies 4 William Griffiths 0000-0002-4129-6616 5 Yuqin Wang 0000-0002-3063-3066 6 James Cronin 0000-0002-0590-9462 7 John J. Bromfield 8 Martin Sheldon 0000-0001-7902-5558 9 Thomas Ormsby 10 57632__21051__c8145150a083454ca694b73984060628.pdf ORMSBYfj202100036R.pdf 2021-09-29T12:40:44.0642284 Output 3779382 application/pdf Version of Record true This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited. © 2021 The Authors. true eng http://creativecommons.org/licenses/by/4.0/
title	Oxysterols protect bovine endometrial cells against pore‐forming toxins from pathogenic bacteria
spellingShingle	Oxysterols protect bovine endometrial cells against pore‐forming toxins from pathogenic bacteria Sian-eleri Owens William Griffiths Yuqin Wang James Cronin Martin Sheldon Thomas Ormsby
title_short	Oxysterols protect bovine endometrial cells against pore‐forming toxins from pathogenic bacteria
title_full	Oxysterols protect bovine endometrial cells against pore‐forming toxins from pathogenic bacteria
title_fullStr	Oxysterols protect bovine endometrial cells against pore‐forming toxins from pathogenic bacteria
title_full_unstemmed	Oxysterols protect bovine endometrial cells against pore‐forming toxins from pathogenic bacteria
title_sort	Oxysterols protect bovine endometrial cells against pore‐forming toxins from pathogenic bacteria
author_id_str_mv	721deb4604d122019244cfdf08820cbe 3316b1d1b524be1831790933eed1c26e c92729b58622f9fdf6a0e7d8f4ce5081 9cfd17551c0d1f7438895121e4fbb6e8 ab0f74b794e59cc270c69e63ee1d9748 c8a0f774d8ff66a2193a035018cc6cda
author_id_fullname_str_mv	721deb4604d122019244cfdf08820cbe_*_Sian-eleri Owens 3316b1d1b524be1831790933eed1c26e__William Griffiths c92729b58622f9fdf6a0e7d8f4ce5081__Yuqin Wang 9cfd17551c0d1f7438895121e4fbb6e8__James Cronin ab0f74b794e59cc270c69e63ee1d9748__Martin Sheldon c8a0f774d8ff66a2193a035018cc6cda_*_Thomas Ormsby
author	Sian-eleri Owens William Griffiths Yuqin Wang James Cronin Martin Sheldon Thomas Ormsby
author2	Thomas J. R. Ormsby Sian-eleri Owens Anthony D. Horlock Daphne Davies William Griffiths Yuqin Wang James Cronin John J. Bromfield Martin Sheldon Thomas Ormsby
format	Journal article
container_title	The FASEB Journal
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institution	Swansea University
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doi_str_mv	10.1096/fj.202100036r
publisher	Wiley
college_str	Swansea University Medical School
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department_str	Swansea University Medical School{{{_:::_}}}Swansea University Medical School{{{_:::_}}}Swansea University Medical School
url	http://dx.doi.org/10.1096/fj.202100036r
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description	Many species of pathogenic bacteria secrete toxins that form pores in mammalian cell membranes. These membrane pores enable the delivery of virulence factors into cells, result in the leakage of molecules that bacteria can use as nutrients, and facilitate pathogen invasion. Inflammatory responses to bacteria are regulated by the side-chain-hydroxycholesterols 27-hydroxycholesterol and 25-hydroxycholesterol, but their effect on the intrinsic protection of cells against pore-forming toxins is unclear. Here, we tested the hypothesis that 27-hydroxycholesterol and 25-hydroxycholesterol help protect cells against pore-forming toxins. We treated bovine endometrial epithelial and stromal cells with 27-hydroxycholesterol or 25-hydroxycholesterol, and then challenged the cells with pyolysin, which is a cholesterol-dependent cytolysin from Trueperella pyogenes that targets these endometrial cells. We found that treatment with 27-hydroxycholesterol or 25-hydroxycholesterol protected both epithelial and stomal cells against pore formation and the damage caused by pyolysin. The oxysterols limited pyolysin-induced leakage of potassium and lactate dehydrogenase from cells, and reduced cytoskeletal changes and cytolysis. This oxysterol cytoprotection against pyolysin was partially dependent on reducing cytolysin-accessible cholesterol in the cell membrane and on activating liver X receptors. Treatment with 27-hydroxycholesterol also protected the endometrial cells against Staphylococcus aureus α hemolysin. Using mass spectrometry, we found 27-hydroxycholesterol and 25-hydroxycholesterol in uterine and follicular fluid. Furthermore, epithelial cells released additional 25-hydroxycholesterol in response to pyolysin. In conclusion, both 27-hydroxycholesterol and 25-hydroxycholesterol increased the intrinsic protection of bovine endometrial cells against pore-forming toxins. Our findings imply that side-chain-hydroxycholesterols may help defend the endometrium against pathogenic bacteria.
published_date	2021-10-01T04:13:52Z
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Oxysterols protect bovine endometrial cells against pore‐forming toxins from pathogenic bacteria

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