# Nanomaterials Covered with Cell Membranes for Intracellular Delivery Without Lysosomal Degradation and Innate Immunity Induction

## TL;DR

This paper explores how cell membrane-covered nanomaterials can enter cells without triggering immune responses or being broken down in lysosomes.

## Contribution

The study introduces a novel method for isolating extracellular nanomaterials and demonstrates their non-immunogenic intracellular delivery.

## Key findings

- Cell membrane-covered nanomaterials accumulate in cells over seven days without lysosomal degradation.
- Extracellular vesicles do not induce significant interferon production, indicating minimal immune activation.
- EVs localize differently from lysosomes, suggesting membrane fusion as a delivery mechanism.

## Abstract

Cellular uptake of nanomaterials is based on endocytosis with their endosomal–lysosomal entrapment resulting in enzymatic hydrolysis. Besides biodegradation, the antigen presentation induces innate and adaptive immunity. Our goal was isolation of extracellular particles to study their structures, penetration into cells, stability, intracellular distribution, and interferon (IFN) production. Extracellular nanomaterials were isolated from conditioned culture media of human embryonic and cancer cells by two-stage differential centrifugation. Cellular uptake of Cy5-labeled particles was evaluated using spectrofluorimetry and confocal fluorescent microscopy. IFN gene expression was analyzed by reverse transcription with real-time PCR and ELISA. Vesicles of 10–200 nm were isolated by centrifugation at 20,800× g at +4 °C for 30 min. The fluorescent vesicles were gradually accumulated inside cells for seven days. Intracellular distribution patterns of the Cy5-labeled vesicles differed from lysosomes stained with LysoRed tracker. IFNs α, β and γ were not detected after treatment with the vesicles. IFN λ was found in cells in the presence of allogenic but not autologous particles. The gradual cellular uptake occurred without significant differences between autologous and heterologous vesicles. Different localization of the extracellular vesicles (EV) and lysosomes along with weak innate immune response (if any) suggested membrane fusion.