Eliminating blood oncogenic exosomes into the small intestine with aptamer-functionalized nanoparticles

There are disease-causing biohazards in the blood that cannot be treated with modern medicines. Here we show that an intelligently designed safe biomaterial can precisely identify, tow and dump a targeted biohazard from the blood into the small intestine. Positively charged mesoporous silica nanoparticles (MSNs) functionalized with EGFR-targeting aptamers (MSN-AP) specifically recognize and bind blood-borne negatively charged oncogenic exosomes (A-Exo), and tow A-Exo across hepatobiliary layers and Oddi’s sphincter into the small intestine. MSN-AP specifically distinguish and bind A-Exo from interfering exosomes in cell culture and rat and patient blood to form MSN-AP and A-Exo conjugates (MSN-Exo) that transverse hepatocytes, cholangiocytes, and endothelial monolayers via endocytosis and exocytosis mechanisms, although Kupffer cells have been shown to engulf some MSN-Exo. Blood MSN-AP significantly decreased circulating A-Exo levels, sequentially increased intestinal A-Exo and attenuated A-Exo-induced lung metastasis in mice. This study opens an innovative avenue to relocate blood-borne life-threatening biohazards to the intestine.


Statistics
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Software and code
Policy information about availability of computer code Data collection BioRadCFXManager3.0, Image Lab, Leica Microsystems CMS GmbH, NanoScopeAnalysis, Flow Jo 7.6.1, NanoApplication and office 2010 were used to collect data.

Data analysis
GraphPad 5.0 and SPSS statistics 17 were used to analyze data.
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Life sciences
Behavioural & social sciences Ecological, evolutionary & environmental sciences nature research | reporting summary

October 2018
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Life sciences study design
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Sample size
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Flow Cytometry
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Methodology
Sample preparation Exosomes were attached to 2.7-μm aldehyde/sulphate latex beads (Invitrogen) by mixing 10 μg of exosomes with 10 μl of beads for 15 min at room temperature with continuous rotation. This suspension was diluted to 1 ml with PBS and rotated for another 30 min. The reaction was stopped with 100 mM glycine and 2% BSA in PBS for 30 min rotation. The exosome-bound beads were washed with 2% BSA in PBS and centrifuged at 15,000 g for 1 min. The beads were blocked with 10% BSA for 30 min with rotation and washed again in 2% BSA and centrifuged for 1 min at 15,000g, and incubated with MSN-AP-Cy-or MSN-AP-Cy (100

October 2018
μg/mL) for 30 min rotation at 4°C. The beads were centrifuged at 15,000 g for 1 min and the supernatant was discarded. The beads were washed in 2% BSA and centrifuged at 15,000 g for 1 min. The blank beads alone were used as a control. Instrument BD FACSAria III flow cytometry Software FlowJo 7.6.1 Cell population abundance The beads incubated with secondary antibody alone were used as control, and the preliminary FSC/SSC gates were established according to the characteristic population of control. The exosome-beads conjugation abundance were obtained by the gate of control group.

Gating strategy
The beads incubated with secondary antibody alone were used as control, and the preliminary FSC/SSC gates were established according to the characteristic population of control. The control group as negative population, the fluorescence exceeded the control group as positive population.
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