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Host epithelium integrity in the female reproductive tract during Neisseria gonorrhoeae infection
Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
2014 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Neisseria gonorrhoeae infections are asymptomatic in approximately fifty percent of the affected women. Left untreated, this can develop into a long-term inflammatory state with detrimental secondary complications such as ectopic pregnancy and sterility. Furthermore, studies have shown that N. gonorrhoeae infection may be a contributing factor to urogenital cancers.

We found that gonococcal infections induce DNA damage in human vaginal and cervical cells. As a consequence of the DNA damage the cell cycle progression is altered. Mitotic checkpoint genes and proteins important in regulation of metaphase were distorted. Partly because of this, the progression of mitosis was hampered. An additional contributing cause of the DNA damage and dysfunctional mitosis is the release of endogenous gonococcal restriction endonucleases.

Most N. gonorrhoeae infection studies are performed on cultured monolayers of cells derived from tumors.  In order to create a system which more resembled in vivo conditions and study N. gonorrhoeae infections, we developed a polarized epithelium of human non-tumorigenic vaginal VK2/E6E7 cells. In the search for an animal model for the human-restricted pathogen we evaluated the CD46 transgenic mouse model in the study of gonococcal infections.

In summary, this thesis aims to increase the understanding of the basic molecular function of how invasive gonococcal infections affect host cell cycle regulation, DNA integrity and potential predisposition to cellular malignancies in the epithelium of the female reproductive tract. Since the female reproductive tract is colonized with Lactobacillus, the impact of lactobacilli to the host cell cycle has also been investigated.

Place, publisher, year, edition, pages
Stockholm: Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University , 2014. , 64 p.
Keyword [en]
Female reproductive tract, Neisseria gonorrhoeae, VK2/E6E7 cells, DNA damage, cell cycle, mitosis, restriction endonucleases, infection models
National Category
Microbiology
Research subject
Molecular Bioscience
Identifiers
URN: urn:nbn:se:su:diva-107713ISBN: 978-91-7649-007-5 (print)OAI: oai:DiVA.org:su-107713DiVA: diva2:750188
Public defence
2014-11-07, Nordenskiöldsalen, Geovetenskapens hus, Svante Arrheniusväg 12, Stockholm, 10:00 (English)
Opponent
Supervisors
Note

At the time of the doctoral defense, the following papers were unpublished and had a status as follows: Paper 3: Manuscript. Paper 4: Manuscript. Paper 5: Manuscript.

Available from: 2014-10-16 Created: 2014-09-24 Last updated: 2014-11-21Bibliographically approved
List of papers
1. Lactobacillus Decelerates Cervical Epithelial Cell Cycle Progression
Open this publication in new window or tab >>Lactobacillus Decelerates Cervical Epithelial Cell Cycle Progression
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2013 (English)In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 8, no 5, e63592- p.Article in journal (Refereed) Published
Abstract [en]

We investigated cell cycle progression in epithelial cervical ME-180 cells during colonization of three different Lactobacillus species utilizing live cell microscopy, bromodeoxyuridine incorporation assays, and flow cytometry. The colonization of these ME-180 cells by L. rhamnosus and L. reuteri, originating from human gastric epithelia and saliva, respectively, was shown to reduce cell cycle progression and to cause host cells to accumulate in the G1 phase of the cell cycle. The G1 phase accumulation in L. rhamnosus-colonized cells was accompanied by the up-regulation and nuclear accumulation of p21. By contrast, the vaginal isolate L. crispatus did not affect cell cycle progression. Furthermore, both the supernatants from the lactic acid-producing L. rhamnosus colonies and lactic acid added to cell culture media were able to reduce the proliferation of ME-180 cells. In this study, we reveal the diversity of the Lactobacillus species to affect host cell cycle progression and demonstrate that L. rhamnosus and L. reuteri exert anti-proliferative effects on human cervical carcinoma cells.

National Category
Microbiology
Research subject
Molecular Bioscience; Molecular Genetics
Identifiers
urn:nbn:se:su:diva-91528 (URN)10.1371/journal.pone.0063592 (DOI)000318852400046 ()
Note

AuthorCount:6;

Available from: 2013-07-01 Created: 2013-06-28 Last updated: 2017-12-06Bibliographically approved
2. Neisseria gonorrhoeae infection causes DNA damage and affects the expression of p21, p27 and p53 in non-tumor epithelial cells
Open this publication in new window or tab >>Neisseria gonorrhoeae infection causes DNA damage and affects the expression of p21, p27 and p53 in non-tumor epithelial cells
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2013 (English)In: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 126, no 1, 339-347 p.Article in journal (Refereed) Published
Abstract [en]

The constant shedding and renewal of epithelial cells maintain the protection of epithelial barriers. Interference with the processes of host cell-cycle regulation and barrier integrity permits the bacterial pathogen Neisseria gonorrhoeae to effectively colonize and invade epithelial cells. Here, we show that a gonococcal infection causes DNA damage in human non-tumor vaginal VK2/E6E7 cells with an increase of 700 DNA strand breaks per cell per hour as detected by an alkaline DNA unwinding assay. Infected cells exhibited elevated levels of DNA double-strand breaks, as indicated by a more than 50% increase in cells expressing DNA damage-response protein 53BP1-positive foci that co-localized with phosphorylated histone H2AX (gamma H2AX). Furthermore, infected cells abolished their expression of the tumor protein p53 and induced an increase in the expression of cyclin-dependent kinase inhibitors p21 and p27 to 2.6-fold and 4.2-fold of controls, respectively. As shown by live-cell microscopy, flow cytometry assays, and BrdU incorporation assays, gonococcal infection slowed the host cell-cycle progression mainly by impairing progression through the G2 phase. Our findings show new cellular players that are involved in the control of the human cell cycle during gonococcal infection and the potential of bacteria to cause cellular abnormalities.

Keyword
DNA damage, N. gonorrhoeae, VK2/E6E7
National Category
Microbiology
Research subject
Molecular Bioscience; Molecular Genetics
Identifiers
urn:nbn:se:su:diva-89534 (URN)10.1242/jcs.117721 (DOI)000316460800032 ()
Note

AuthorCount:6;

Available from: 2013-05-02 Created: 2013-04-29 Last updated: 2017-12-06Bibliographically approved
3. Restriction endonucleases from invasive Neisseria gonorrhoeae cause double-strand breaks and distort mitosis in epithelial cells during infection
Open this publication in new window or tab >>Restriction endonucleases from invasive Neisseria gonorrhoeae cause double-strand breaks and distort mitosis in epithelial cells during infection
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(English)Manuscript (preprint) (Other academic)
Keyword
Restriction endonucleases, N. gonorrhoeae, VK2E6/E7 cells, mitosis
National Category
Microbiology
Research subject
Microbiology; Infectious Diseases
Identifiers
urn:nbn:se:su:diva-107688 (URN)
Available from: 2014-09-24 Created: 2014-09-24 Last updated: 2014-09-29
4. Adherence and transmigration of Neisseria gonorrhoeae to polarized human stratified squamous vaginal epithelium
Open this publication in new window or tab >>Adherence and transmigration of Neisseria gonorrhoeae to polarized human stratified squamous vaginal epithelium
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(English)Manuscript (preprint) (Other academic)
Keyword
Neisseria gonorrhoeae, VK2/E6E7, Transwell, Polarized cells, Epithelium
National Category
Microbiology
Research subject
Microbiology; Infectious Diseases
Identifiers
urn:nbn:se:su:diva-107692 (URN)
Available from: 2014-09-24 Created: 2014-09-24 Last updated: 2014-09-29
5. Gonococcal infection in CD46 transgenic mice – A pilot study
Open this publication in new window or tab >>Gonococcal infection in CD46 transgenic mice – A pilot study
(English)Manuscript (preprint) (Other academic)
Keyword
CD46 transgenic mice, Neisseria gonorrhoeae, animal model
National Category
Microbiology
Research subject
Infectious Diseases; Microbiology
Identifiers
urn:nbn:se:su:diva-107690 (URN)
Available from: 2014-09-24 Created: 2014-09-24 Last updated: 2014-09-29

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