# Improving Nonviral Gene Delivery by Activating Mechanosensing-Dependent Endocytic Pathways

## TL;DR

This study shows that applying mechanical stretching to cells can boost the efficiency of gene delivery without changing the delivery vectors themselves.

## Contribution

A novel mechanobiological approach using mechanical stimulation to enhance non-viral gene delivery is introduced.

## Key findings

- Cyclic mechanical stimulation increased nuclear translocation of YAP in HeLa cells and human myoblasts.
- Mechanical conditioning upregulated clathrin-mediated endocytosis and macropinocytosis pathways.
- Transfection efficiency of bPEI-based DNA and mRNA complexes improved significantly under mechanical stimulation.

## Abstract

The delivery of nucleic
acids into host cells has emerged
as an
innovative and promising therapeutic approach for various diseases.
Despite significant advances in nanoparticle delivery systems, persistent
cellular barriers limit the clinical application of most existing
technologies. In this study, we developed a programmable device that
applies precise uniaxial cyclic stretching to cells cultured on custom
polydimethylsiloxane chambers to investigate whether mechanical stimulation
can enhance the transfection efficiency (TE) of gold-standard non-viral
gene delivery vectors. Applying cyclic mechanical stimulation (f = 0.1 Hz, ε = 10% strain, t = 30
min) to HeLa cells and human myoblasts (hMyo) significantly increased
nuclear translocation of the mechanosensitive transcription factor
Yes-Associated Protein (YAP). Gene expression analysis revealed that
this mechanical conditioning orchestrated a coordinated modulation
of endocytic machinery, upregulating clathrin-mediated endocytosis
(FCHO1) and macropinocytosis (STX1B) pathways while downregulating
endocytic inhibitors (DLC1, EHD2). These mechanically induced cellular
adaptations resulted in significantly enhanced TE of both plasmid
DNA (pDNA)- and mRNA (mRNA)-carrying gold-standard branched polyethylenimine
(bPEI)-based complexes in both HeLa cells and hMyo,
compared to static conditions. Our findings demonstrate that mechanical
stimulation is an effective complementary strategy for improving non-viral
gene delivery by leveraging endogenous cellular mechanotransduction
pathways. Rather than modifying vector chemistry, this mechanobiological
approach enhances the performance of existing delivery systems by
transiently modulating cellular uptake capacity and nuclear accessibility.
This work offers mechanistic insights into how mechanotransduction
regulates cellular uptake and highlights opportunities for leveraging
controlled mechanical environments in applications such as ex vivo
cell engineering.