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Induction of DNA strand scissions in HeLa cells by human polymorphonuclear leucocytes activated by Chlamydia
1Virology Unit, Faculty of Health Sciences, Ben-Gurion University of the Negev, Beer Sheva, Israel.
Abstract:
Incubation of human polymorphonuclear leucocytes (HPMN) with Chlamydia trachomatis elementary bodies (EB) or phorbol 12-myristate 13-acetate (PMA) resulted in the production of superoxide anions (.O2-) and hydrogen peroxide (H2O2). Exposure of HeLa cells to EB- or PMA-activated HPMN and to EB alone, for 2 h, resulted in the formation of DNA strand scissions (nicks) in the HeLa cells. The nicks were visualized by incorporation of biotin 11-dUTP with its detection by streptavidin-peroxidase, and quantified by using [3H]dCTP in the in situ nuclear nick-translation reaction. Catalase, and to a lesser extent superoxide dismutase, reduced the amount of nicks induced by the EB- or PMA-activated HPMN. The possible relationship between the activity of PMN in chlamydial infections and the development of chronic diseases is discussed.
Insights
Human polymorphonuclear leucocytes (HPMN) exposed to Chlamydia trachomatis elementary bodies (EB) produce reactive oxygen species that damage HeLa cell DNA. Antioxidants like catalase reduce this DNA damage, suggesting a link to chronic chlamydial infections.
Area of Science:
- Immunology
- Cell Biology
- Microbiology
Background:
- Chlamydia trachomatis (EB) infections can trigger inflammatory responses.
- Human polymorphonuclear leucocytes (HPMN) are key immune cells involved in combating bacterial infections.
- Reactive oxygen species (ROS) produced by immune cells play a role in host defense but can also cause tissue damage.
Purpose of the Study:
- To investigate the production of reactive oxygen species (ROS) by HPMN upon stimulation with Chlamydia trachomatis elementary bodies (EB).
- To determine if HPMN-derived ROS can induce DNA damage in host cells.
- To explore the potential role of ROS-mediated DNA damage in the pathogenesis of chlamydial infections.
Main Methods:
- Incubation of HPMN with EB or phorbol 12-myristate 13-acetate (PMA) to induce ROS production (superoxide anions and hydrogen peroxide).
- Exposure of HeLa cells to activated HPMN or EB alone, followed by assessment of DNA strand scissions using in situ nick-translation.
- Quantification of DNA damage via biotin-11-dUTP incorporation and detection, and [3H]dCTP labeling.
- Evaluation of the protective effects of antioxidants, catalase and superoxide dismutase, against DNA damage.
Main Results:
- HPMN incubated with EB or PMA produced significant levels of superoxide anions and hydrogen peroxide.
- Exposure of HeLa cells to EB- or PMA-activated HPMN, as well as EB alone, resulted in DNA strand scissions.
- The formation of DNA strand breaks in HeLa cells was significantly reduced by the addition of catalase and, to a lesser extent, superoxide dismutase.
- These findings indicate that ROS generated by HPMN contribute to DNA damage in host cells during Chlamydia trachomatis exposure.
Conclusions:
- HPMN activation by Chlamydia trachomatis leads to the generation of ROS that can directly damage host cell DNA.
- The observed DNA damage suggests a potential mechanism by which chronic inflammation and tissue pathology may develop during chlamydial infections.
- Further research is warranted to elucidate the precise role of PMN activity and ROS in the long-term consequences of chlamydial infections.