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Author Spotlight: Understanding DNA Damage Response in Mammalian Oocytes and Preimplantation Embryos
Published on: June 23, 2023
Escherichia Coli Outer Membrane Vesicles Induced DNA Double-Strand Breaks in Intestinal Epithelial Caco-2 Cells
Zhou Ling1, Chen Dayong1, Yu Denggao1
1State Key Laboratory of Ultrasound Engineering in Medicine Co-Founded by Chongqing and the Ministry of Science and Technology, College of Biomedical Engineering, Chongqing Medical University, Chongqing, China (mainland).
Abstract:
BACKGROUND Recent studies have shown that Escherichia coli induced digestive tract diseases may be related to outer membrane vesicles (OMVs) induced intestinal double-strand breaks (DSBs) in intestinal epithelial cells. This study aimed to compare the impact of OMVs forces on DSBs in intestinal epithelial Caco-2 cells, and provide a new treatment for digestive diseases caused by E. coli. MATERIAL AND METHODS E.coli OMVs were prepared and co-cultured with Caco-2 cells. The uptake of OMVs by Caco-2 cells was observed by confocal microscopy. The γ-H2AX protein was detected by western-blots. The DSBs caused by OMVs was detected by single cell gel electrophoresis. RESULTS The particle size analyzer showed that the average diameters of OMVs centrifuged at 20 000×g and 50 000×g were 217.5±7.29 nm and 186.3±6.59 nm (P<0.05), respectively. Transmission electron microscopy of the OMVs revealed a lipid bilayer structure with a variety of different sizes. Confocal fluorescence microscopy revealed that OMVs almost completely entered Caco-2 cells after 24 hours. The ratio of γ-H2AX protein band gray value normalized data in the OMVs centrifuged at 20 000×g and 50 000×g, and the control group (without OMVs) were 2.23±0.18, 1.58±0.20, 1±0.30 (P<0.05), respectively, while DNA levels of the comet tail (TailDNA%, TDNA%) were 72.21±14.61%, 23.11±4.98%, and 1.02±1.41% (P<0.05), respectively. The corresponding DNA damage was categorized as high (grade 3), moderate (grade 2), and no damage (grade 0). CONCLUSIONS Different sizes of OMVs induced different degrees of DNA damage in intestinal epithelial Caco-2 cells.
Insights
Escherichia coli outer membrane vesicles (OMVs) cause varying degrees of DNA double-strand breaks (DSBs) in intestinal cells. Smaller OMVs induced less DNA damage, offering potential therapeutic insights for E. coli-related digestive diseases.
Area of Science:
- Microbiology
- Cell Biology
- Gastroenterology
Background:
- Escherichia coli outer membrane vesicles (OMVs) are implicated in digestive tract diseases.
- OMVs may induce double-strand breaks (DSBs) in intestinal epithelial cells.
Purpose of the Study:
- To compare the impact of different sized OMVs on DSBs in Caco-2 cells.
- To explore potential new treatments for E. coli-induced digestive diseases.
Main Methods:
- Preparation and characterization of E. coli OMVs.
- Co-culture of OMVs with Caco-2 cells and observation of OMV uptake.
- Detection of DSBs using γ-H2AX protein levels and single cell gel electrophoresis.
Main Results:
- OMVs of different sizes (217.5±7.29 nm and 186.3±6.59 nm) were prepared.
- OMVs were efficiently internalized by Caco-2 cells within 24 hours.
- Larger OMVs (217.5 nm) induced significantly higher levels of DNA damage (72.21% TailDNA) compared to smaller OMVs (186.3 nm, 23.11% TailDNA).
Conclusions:
- OMV size is a critical factor influencing the degree of DNA damage in intestinal epithelial cells.
- Different sizes of OMVs induce distinct levels of DSBs, categorized as high, moderate, or no damage.
- Findings suggest size-dependent OMV pathogenicity and potential therapeutic targets for digestive diseases.
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