Abstract
Recently, biological roles of extracellular vesicles (which include among others exosomes, microvesicles and apoptotic bodies) have attracted substantial attention in various fields of biomedicine. Here we investigated the impact of sustained exposure of cells to the fluoroquinolone antibiotic ciprofloxacin on the released extracellular vesicles. Ciprofloxacin is widely used in humans against bacterial infections as well as in cell cultures against Mycoplasma contamination. However, ciprofloxacin is an inducer of oxidative stress and mitochondrial dysfunction of mammalian cells. Unexpectedly, here we found that ciprofloxacin induced the release of both DNA (mitochondrial and chromosomal sequences) and DNA-binding proteins on the exofacial surfaces of small extracellular vesicles referred to in this paper as exosomes. Furthermore, a label-free optical biosensor analysis revealed DNA-dependent binding of exosomes to fibronectin. DNA release on the surface of exosomes was not affected any further by cellular activation or apoptosis induction. Our results reveal for the first time that prolonged low-dose ciprofloxacin exposure leads to the release of DNA associated with the external surface of exosomes.
Highlights
Extracellular vesicles (EVs) play key roles in intercellular communication by which they may impact a wide range of biological functions of cells
In line with previous observations by others[27, 30], we found that the presence of this low-dose (10 μg/mL) antibiotic did not have a significant effect on cell viability (Fig. 1a and b)
In agreement with previous published findings[27,28,29, 31], our mass spectrometry (MS) analysis of cells showed that the presence of ciprofloxacin resulted in a slightly elevated percentage of cellular proteins associated for example with oxidative stress and defense responses, mitochondrial degradation, and in a somewhat reduced percentage of respiratory electron transport chain-associated proteins (Supplementary Fig. S1, Supplementary Dataset S1)
Summary
Extracellular vesicles (EVs) play key roles in intercellular communication by which they may impact a wide range of biological functions of cells. EVs have been shown to deliver specific mRNAs and various small RNAs8–10 as well as DNA11–15 to healthy cells They modify the genetic composition of recipient cells and alter their functions[12, 16,17,18,19]. In a recent short report, a human mast cell line was shown to carry DNAse I-sensitive nucleic acid on the surface of EXOs25. In yet another publication DNAse sensitive chromatin was shown on the surface of EVs secreted into blood[26] This EV-associated chromatin as a potential self-antigen could induce an anti-DNA antibody response. We report for the first time that ciprofloxacin induced the release of both mitochondrial and chromosomal DNA associated with the surface of EXOs. We demonstrate that this exofacial DNA facilitates EXO binding to the extracellular matrix protein fibronectin
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