Accurate Delivery

Highly efficient intracellular delivery strategies are essential for developing therapeutic, diagnostic, biological, and biomedical applications. These strategies can be used in cell-based therapy, genome editing, regenerative medicine, personalized medicine and different biological and biomedical analyses.

The main focus while developing such intracellular delivery strategies include:

1. Ensuring high throughput (indicates the analysis of a large number of cells uniformly within a small time frame).
2. High transfection efficiency (indicates delivery of desired biomolecules into different cell-types).
3. Increased cell viability (indicates the number of delivered cells that remain alive after post-delivery).

There are two types of intracellular delivery strategies, namely:

1. Career-mediated.
2. Membrane disruption-based.

Although career-mediated methods are useful for gene transfer and therapy, they are specific to cell-types, and are often restricted by cytotoxicity and unwanted immune and inflammatory responses, leading to lower cell viabilities.

On the other hand, membrane disruption methods involve the permeabilization of the cellular membrane to generate transient nanopores, which then permit the delivery of external molecules through diffusion and/or fluid convection. These methods have the advantage of lesser chance of inducing harsh cellular responses such as unwanted immunogenic and chemical reactions, thus leading to better delivery efficiencies and cell viabilities.  

The most popular membrane disruption techniques include electroporation, optoporation, magnetoporation, accoustoporation, and mechanoporation. Although most of these techniques require some form of energy to facilitate membrane permeabilization (electric, optical, magnetic, etc.), mechanoporation is the only one that doesn’t need an external energy to create transient nanopores.

Mechanoporation imparts physical forces on the cell membrane to generate the transient nanopores that are required for biomolecular delivery. Because of the elimination of an external carrier or energy source, the mechanoporation techniques are simple to use, induce minimal cell damage and avoid unwanted cellular toxicity and death.

Initially, high cost and low throughput were the drawbacks of mechanoporation. However, with the advent of micro/nanotechnologies, mechanoporation integrated with microfluidics have gained significant attention amongst researchers, especially in the past one-and-a-half decades. Microfluidic-based mechanoporation devices have low fabrication costs, lightweight, compact, easy to use, biocompatible, and thereby enable high throughput intracellular delivery studies at the single-cell level with high transfection efficiency and viability.

The different microfluidic-based mechanoporation techniques have been categorized into four types, namely:

1. Microfluidic-based microinjection
2. Micro/nanoneedle arrays
3. Microfluidic device employing mechanical confinement
4. Microfluidic device employing hydrodynamic manipulation

In this study, for the first time as per the authors’ knowledge, a brief overview of the microfluidic-based mechanoporation techniques has been given and summarized. The authors of this paper include:

1. Mr. Pulasta Chakrabarty, Dr. Pallavi Gupta, Ms. Kavitha Illath, and Dr. Tuhin Subhra Santra, from the Department of Engineering Design, Indian Institute of Technology Madras, Chennai, India
2. Dr. Srabani Kar, from the Department of Electrical Engineering, University of Cambridge, Cambridge, UK
3. Dr. Moeto Nagai, from the Department of Mechanical Engineering, Toyohashi University of Technology, Aichi, Japan
4. Dr. Fang-Gang Tseng, from the Department of Engineering and System Science, National Tsing Hua University, Hsinchu, Taiwan.

Of the methods, the microinjection and microneedle arrays-based techniques have high delivery efficiency and cell viability but are low throughput and expensive. Cell squeezing and hydroporation techniques have obtained increased throughputs but at lower efficiency as compared to the former. However, for different biological and biomedical applications, the transfection efficiency for larger molecules needs to be increased for the high throughput platforms.

Combining mechanoporation with other techniques such as electroporation and magnetoporation has significantly improved the macromolecular transfection efficiency. However, the transfection efficiency for ultra-large biomolecules and pathogens into hard-to-transfect cells still remains a significant challenge. Nevertheless, the currently available microfluidic mechanoporation platforms are a very economical and effective for intracellular delivery and analysis

To conclude, the challenges for intracellular delivery and cellular analysis are highly interdisciplinary. A better understanding of the biomolecule-cell interaction is required. A collaborative effort between doctors, medical professionals, engineers, and pharmacologists is necessary to better understand proper drug delivery into cells. The objective should be to develop low-cost, versatile, GMP (good manufacturing practice)-compliant platforms for drug delivery, thereby enabling rapid clinical applications. Effective drug delivery should be the aim of all researchers.

Prof. Ki-Taek Lim from the Department of Biosystems Engineering, Kangwon National University, Chuncheon, South Korea, points out the importance of this study giving the following comments: “This paper demonstrated highly efficient intracellular delivery strategies that can be applicable for developing therapeutic, diagnostic, biological, and various biomedical research purposes. The manuscript covers different mechanical stress-induced membrane disruptions for intracellular delivery, where membrane disruption methods find popularity due to reduced toxicity, enhanced delivery efficiency, and cell viability.

This method is advantageous compared to other techniques because no external energy source is required for membrane deformation, so I think it values high impact to develop alternative delivery strategies and enhance delivery efficiencies and cell viability into different cell types with negligible toxicity.”

Article by Akshay Anantharaman
Here is the original link to the paper:
https://www.sciencedirect.com/science/article/pii/S2590006421001010