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Antibiotic-induced release of small extracellular vesicles (exosomes) with surface-associated DNA.

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.

SUBMITTER: Nemeth A

PROVIDER: S-EPMC5557920 | biostudies-literature | 2017 Aug

REPOSITORIES: biostudies-literature

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Antibiotic-induced release of small extracellular vesicles (exosomes) with surface-associated DNA.

Németh Andrea A Orgovan Norbert N Sódar Barbara W BW Osteikoetxea Xabier X Pálóczi Krisztina K Szabó-Taylor Katalin É KÉ Vukman Krisztina V KV Kittel Ágnes Á Turiák Lilla L Wiener Zoltán Z Tóth Sára S Drahos László L Vékey Károly K Horvath Robert R Buzás Edit I EI

Scientific reports 20170815 1

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, ciprofloxaci ...[more]

PMID: 28811610

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Project description:Purpose: Smoking is a strong relative risk factor for lung cancer. Extracellular vesicles (EVs), particularly exosomes, have been implicated in cancers. In this study, we characterized smoking induced extracellular vesicles in smokers with non-small cell lung cancer (NSCLC). Methods: EVs were isolated from bronchoalveolar lavage (BAL) from smokers and NSCLC patients. EV microRNAs (miRNAs) were analyzed by using a TaqMan microRNA assays. Vesicle mRNAs and long non-coding RNAs (lncRNAs) were measured with quantitative RT-PCR. Tumor associated antigens were examined by Western Blot. Results: Higher levels of local site EVs are found in the lung of smokers and NSCLC patients. Further, over 90% of lung EVs are round vesicles of approximately 50-200 nm, ie., exosomes. There are 21 EV miRNAs up regulated, while 10 miRNAs under regulated, in smokers when compared to controls (relative fold > 2, p < 0.05). These miRNAs were further observed to be dysregulated in NSCLC patients when compared to smokers. Bioinformatic analysis demonstrated that Proteoglycans, Fatty acid biosynthesis, ErbB, Hippo, TGF-beta, Wnt, Rap1, AMPK and Ras pathways were the most prominent pathways enriched in NSCLC EV miRNA signatures. In addition, messenger RNA transcripts including EGFR, KRAS, ALK, MET, LKB1, BRAF, PIK3CA, RET, and ROS1 were significantly higher expressed in lung EVs in smokers and NSCLC patients compared to controls. Long non-coding RNAs, including MALAT1, HOTAIR, HOTTIP, AGAP2-AS1, ATB, TCF7, FOXD2-AS1, HOXA11-AS, PCAF1, and BCAR4, were over expressed in EVs from smokers and NSCLC patients. Furthermore, protein levels of tumor associated antigens including BAGE, PD-L1, MAGE-3, and AKAP4 were significantly dysregulated in EVs of smokers and NSCLC patients compared to healthy controls. Conclusions: In conclusion, these data demonstrated an intrinsic relationship of smoking dysregulated EVs and EVs contained RNA, proteins which may involve in the development of NSCLC.

Project description:Abstract Small extracellular vesicles (EVs) have emerged as a focal point of EV research due to their significant role in a wide range of physiological and pathological processes within living systems. However, uncertainties about the nature of these vesicles have added considerable complexity to the already difficult task of developing EV‐based diagnostics and therapeutics. Whereas small EVs have been shown to be negatively charged, their surface charge has not yet been properly quantified. This gap in knowledge has made it challenging to fully understand the nature of these particles and the way they interact with one another, and with other biological structures like cells. Most published studies have evaluated EV charge by focusing on zeta potential calculated using classical theoretical approaches. However, these approaches tend to underestimate zeta potential at the nanoscale. Moreover, zeta potential alone cannot provide a complete picture of the electrical properties of small EVs since it ignores the effect of ions that bind tightly to the surface of these particles. The absence of validated methods to accurately estimate the actual surface charge (electrical valence) and determine the zeta potential of EVs is a significant knowledge gap, as it limits the development of effective label‐free methods for EV isolation and detection. In this study, for the first time, we show how the electrical charge of small EVs can be more accurately determined by accounting for the impact of tightly bound ions. This was accomplished by measuring the electrophoretic mobility of EVs, and then analytically correlating the measured values to their charge in the form of zeta potential and electrical valence. In contrast to the currently used theoretical expressions, the employed analytical method in this study enabled a more accurate estimation of EV surface charge, which will facilitate the development of EV‐based diagnostic and therapeutic applications.

Dataset Information

Antibiotic-induced release of small extracellular vesicles (exosomes) with surface-associated DNA.

Publications

Antibiotic-induced release of small extracellular vesicles (exosomes) with surface-associated DNA.

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