Cavanagh, M., Amabebe, E. orcid.org/0000-0002-3924-5270 and Anumba, D.O.C. orcid.org/0000-0003-2502-3033 (2020) Differential cytokine and metabolite production by cervicovaginal epithelial cells infected with Lactobacillus crispatus and Ureaplasma urealyticum. Anaerobe, 62. 102101. ISSN 1075-9964
Abstract
Introduction
We sought to quantify targeted metabolites (d-lactate, pyruvate, urea, ammonia) and the cytokine IL-8 produced by human cervicovaginal epithelial cells co-cultured with Ureaplasma urealyticum (a preterm birth-associated bacterium) or Lactobacillus crispatus (a healthy vaginal commensal associated with term birth).
Methods
Concentrations of d-lactate, pyruvate, urea and ammonia measured by enzyme-based spectrophotometry and IL-8 by ELISA were determined and compared between monolayer-cultured HeLa cells (ATCC 35241) infected with strains of U. urealyticum (ATCC 27618, 0.5 mL = 3640 CFU/mL, U. urealyticum) or L. crispatus (ATCC 33820, MOI = 10,000, 1000 and 100, L. crispatus) and incubated in 5% CO2 at 37 °C for 24 h. Uninfected HeLa cells (Hc) were used as controls and cytotoxicity was determined by the amount (optical density) of lactate dehydrogenase (LDH) released by the dead HeLa cells.
Results
The amount of LDH released by untreated Hc (P = 0.002) and U. urealyticum-infected cells (P < 0.0001) was higher than those of L. crispatus-infected cells, with U. urealyticum-infected cells recording the highest % cytotoxicity and L. crispatus-infected cells MOI 10,000 (Lc10,000) the least (P < 0.0001). Though there was no significant difference in the concentration of urea between the samples, U. urealyticum-infected cells showed higher ammonia compared to other samples (p = 0.03). In contrast, all L. crispatus samples had higher d-lactate than untreated Hc (p = 0.01) and U. urealyticum-infected cells (P = 0.01). Also, Lc10,000 had the highest d-lactate (p = 0.001) and lowest pyruvate (P = 0.04, excluding UU) compared to other samples. Furthermore, U. urealyticum-infected cells produced the highest IL-8 (P = 0.01) compared to other samples, with Lc10,000 producing undetectable levels.
Conclusion
Infection of cervicovaginal epithelial cells by U. urealyticum stimulates production of ammonia from urea and induces elevated IL-8 production possibly leading to significantly higher cytotoxicity. In contrast, L. crispatus appeared protective against HeLa cell inflammation and death, producing more d-lactate and less IL-8, consistent with a role for L. crispatus in promoting vaginal floral health and reducing infection/inflammation-associated preterm birth.
Metadata
Item Type: | Article |
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Authors/Creators: |
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Copyright, Publisher and Additional Information: | © 2019 Published by Elsevier Ltd. |
Keywords: | Ureaplasma; Lactobacillus; HeLa cells; Preterm birth; IL-8; D-lactate |
Dates: |
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Institution: | The University of Sheffield |
Academic Units: | The University of Sheffield > Sheffield Teaching Hospitals |
Depositing User: | Symplectic Sheffield |
Date Deposited: | 22 Sep 2020 12:47 |
Last Modified: | 22 Sep 2020 12:47 |
Status: | Published |
Publisher: | Elsevier BV |
Refereed: | Yes |
Identification Number: | 10.1016/j.anaerobe.2019.102101 |
Related URLs: | |
Open Archives Initiative ID (OAI ID): | oai:eprints.whiterose.ac.uk:165570 |