In lab experiments, the researchers have found that this device, a vector, is able to deliver DNA deeply enough into a cell to allow genetic material to be activated – a critical step in gene therapy. This vector is between 2 ½ and 10 times more effective than other experimental materials, according to the research.
In this case, Ohio State University scientists combined two ingredients – calcium phosphate and a lipid shell – to create a nanoparticle that protects DNA during its journey to the cell and then dissolves to allow for gene activation in the target cell. Nano refers to the tiny size of the particle in question – its general structure can be detected only by an atomic force microscope.
Calcium phosphate is a mineral found in bones and teeth. Lipids are fatty molecules that help maintain the structure of cell membranes. Alone, calcium phosphate is toxic and lipids get absorbed by cells. Together, they form a protective and inflexible structure that, thanks to complex chemical reactions, self-destructs once inside a cell.
“Our nanoparticle is a foreign body just like a viral vector is, but it has a self-destructive mechanism so it does not generate a strong response from the immune system,” said Chenguang Zhou, a graduate student in pharmaceutics at Ohio State and lead author of the study. “The material we use is also biocompatible. Calcium phosphate is in our bones and the lipids we use are synthetic, but can be biologically degraded. That’s why there is low toxicity.”
The research is published in a recent issue of the International Journal of Pharmaceutics.
Zhou noted that other researchers have tried to use liposomes – nanometer-sized bubbles made out of the same material as a cell membrane – to create nonviral vectors for gene delivery. While the material did a good job of protecting the DNA, it did not do a good job of releasing the gene into a cell.
“The liposome gets internalized into cells. It’s sort of like eating food that gets stuck in the stomach or intestines, but never gets to the rest of the body,” he said.
Similarly, calcium phosphate alone has been considered as a gene delivery vehicle. But because of its salty properties, it becomes unstable and expands in size, which makes it too big to penetrate some cell and vascular walls, and which can cause the immune system to reject it.
“So what we do is encapsulate a calcium phosphate core inside the liposome,” Zhou said. “And when this calcium phosphate gets inside a cell and that environment becomes acidic, it gets dissolved and then the gene can be very effectively released into the cytoplasm and transported to the nucleus. That is the theory.”
Zhou and colleagues have developed what they consider an easy method to manufacture this particle. They create a synthetic lipid and place it in a solution that contains calcium and phosphate, which becomes integrated with the lipid. As the acidic properties in the solution change, the calcium phosphate forms a core.
The scientists then mix a solution containing plasmid DNA with their newly formed particle, and all the materials become bundled together. Plasmid DNA is a circular DNA molecule that is able to turn on gene activity that starts a protein-building process without altering an entire genome, or the complete hereditary information of an organism.
Because this particular vector is intended for injection into the bloodstream as a cancer treatment, the particle is designed to protect the DNA from being digested by enzymes as it travels to target cells. A test that exposed this solution to serum, a component of blood, showed that the hybrid particle provided this protection, while unprotected DNA was digested by enzymes.
The researchers next applied this DNA-infused particle solution to mouse cells. The DNA contained the gene code for green fluorescent protein that would be turned on only after it entered the cell. They observed that the particle penetrated the cell membrane and, after a series of interactions occurred, the green fluorescent protein lit up inside the cell, indicating the DNA had reached its target.
For comparison, the group also monitored DNA movement on its own and in other types of vectors. Their hybrid vector was 24 times more effective at delivering genetic material to the cell than was DNA on its own, and 10 times more effective than calcium phosphate preparations.
“We know the particle gets to where it needs to go and what happens to the particle,” Zhou said. “Do we know that the DNA reaches the nucleus? That is something we still need to find out. But because we saw the green fluorescent protein expressed, we think it got to the nucleus or at least as far as the cytoplasm. What’s important is that the protein got inside the cell.”
The study also showed that this hybrid particle maintained its structure for at least 21 days and, when compared with a variety of other potential vector substances, did very little damage to cells, meaning it is not as toxic as most other materials.
With viral vectors, gene therapy is considered a one-time treatment because when the virus carrying new genes infects a cell, that interaction changes the recipient’s entire genome, effectively canceling the activity of the defective gene. Zhou said that with this nonviral vector, treatment would be designed as an intravenous injection on a regular basis until cells are “infected enough to make a change.”
The researchers next plan to test the particle’s ability to travel through the bloodstream and enter target cells in animals.
This work was supported by the National Science Foundation (NSF) and the National Institutes of Health. Zhou has a fellowship in Ohio State’s NSF Nanoscale Science and Engineering Center (NSEC) for Affordable Nanoengineering of Polymeric Biomedical Devices.
He completed the research with Ohio State co-authors Bo Yu and L. James Lee of the Department of Chemical and Biomolecular Engineering; Xiaojuan Yang and Robert Lee of the Division of Pharmaceutics; and Tianyao Huo and Rolf Barth of the Department of Pathology. Yu, Yang, L. James Lee and Robert Lee are also affiliated with the NSEC.Contact: Chenguang Zhou, (614) 292-5870; firstname.lastname@example.org
Chenguang Zhou | EurekAlert!
Making fuel out of thick air
08.12.2017 | DOE/Argonne National Laboratory
‘Spying’ on the hidden geometry of complex networks through machine intelligence
08.12.2017 | Technische Universität Dresden
The miniaturization of the current technology of storage media is hindered by fundamental limits of quantum mechanics. A new approach consists in using so-called spin-crossover molecules as the smallest possible storage unit. Similar to normal hard drives, these special molecules can save information via their magnetic state. A research team from Kiel University has now managed to successfully place a new class of spin-crossover molecules onto a surface and to improve the molecule’s storage capacity. The storage density of conventional hard drives could therefore theoretically be increased by more than one hundred fold. The study has been published in the scientific journal Nano Letters.
Over the past few years, the building blocks of storage media have gotten ever smaller. But further miniaturization of the current technology is hindered by...
With innovative experiments, researchers at the Helmholtz-Zentrums Geesthacht and the Technical University Hamburg unravel why tiny metallic structures are extremely strong
Light-weight and simultaneously strong – porous metallic nanomaterials promise interesting applications as, for instance, for future aeroplanes with enhanced...
An interdisciplinary group of researchers interfaced individual bacteria with a computer to build a hybrid bio-digital circuit - Study published in Nature Communications
Scientists at the Institute of Science and Technology Austria (IST Austria) have managed to control the behavior of individual bacteria by connecting them to a...
Physicists in the Laboratory for Attosecond Physics (run jointly by LMU Munich and the Max Planck Institute for Quantum Optics) have developed an attosecond electron microscope that allows them to visualize the dispersion of light in time and space, and observe the motions of electrons in atoms.
The most basic of all physical interactions in nature is that between light and matter. This interaction takes place in attosecond times (i.e. billionths of a...
Transistors based on carbon nanostructures: what sounds like a futuristic dream could be reality in just a few years' time. An international research team working with Empa has now succeeded in producing nanotransistors from graphene ribbons that are only a few atoms wide, as reported in the current issue of the trade journal "Nature Communications."
Graphene ribbons that are only a few atoms wide, so-called graphene nanoribbons, have special electrical properties that make them promising candidates for the...
08.12.2017 | Event News
07.12.2017 | Event News
05.12.2017 | Event News
08.12.2017 | Life Sciences
08.12.2017 | Information Technology
08.12.2017 | Information Technology