Tissue nanotransfection

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Tissue nanotransfection (TNT) is an electroporation-based technique capable of gene and drug cargo delivery or transfection at the nanoscale. Furthermore, TNT is a scaffold-less tissue engineering (TE) technique that can be considered cell-only or tissue inducing depending on cellular or tissue level applications. The transfection method makes use of nanochannels to deliver cargo to tissues topically.  

Contents

History

Cargo delivery methods rely on carriers, for example nanoparticles, viral vectors, or physical approaches such as gene guns, microinjection, or electroporation. [1] [2] [3] [4] [5] [6] [7] [8] [9] The various methods can be limited by size constraints or their ability to efficiently deliver cargo without damaging tissue. Electroporation is a physical method which harnesses an electric field to open pores in the normally semi-permeable cell membrane through which cargo can enter. In this process, the charges can be used to drive cargo in a specific direction.

Bulk electroporation (BEP) is the most conventional electroporation method. Benefits come in the form of high throughput and minimal set-up times. [7] The downside of BEP is that the cell membrane experiences an uneven distribution of the electric field and many membranes receive irreversible damage from which they can no longer close, thus leading to low cell viability.

Attempts have been made to miniaturize electroporation such as microelectroporation (MEP) [10] and nanochannel electroporation (NEP) [11] which uses electroporation approached to deliver cargo through micro/nanochannels respectively. These techniques have shown to have higher efficiency of delivery, increased uniform transfection, and increased cell viability compared to BEP. [12]

Technique

Tissue nanotransfection uses custom fabricated nanochannel arrays for nanoscale delivery of genetic cargo directly onto the surface of the skin. The postage stamp-sized chip is placed directly on the skin and an electric current is induced lasting for milliseconds to deliver the gene cargo with precise control. This approach delivers ample amounts of reprogramming factors to single-cells, creating potential for a powerful gene transfection and reprogramming method. [11] [12] The delivered cargo then transforms the affected cells into a desired cell type without first transforming them to stem cells. TNT is a novel technique and has been used on mice models to successfully transfect fibroblasts into neuron-like cells along with rescue of ischemia in mice models with induced vasculature and perfusion. [13] Current methods require the fabricated TNT chip to be placed on the skin and the loading reservoir filled with a gene solution. An electrode (cathode) is placed into the well with a counter electrode (anode) placed under the chip intradermally (into the skin). The electric field generated delivers the genes. [13]

Initial TNT experiments showed that genes could be delivered to the skin of mice. [13] Once this was confirmed, a cocktail of gene factors (ABM) used by Vierbuchen [14] and collaborators to reprogram fibroblast into neurons was used. [12] [13] Delivery of these factors demonstrated successful reprogramming in-vivo and signals propagated from the epidermis to the dermis skin layers. This phenomenon is believed to be mediated by extracellular vesicles [15] and potentially other factors [18]. Successful reprogramming was determined by performing histology and electrophysiological tests to confirm the tissue behaved as functional neurons. [13]

Beyond inducing neurons, Gallego-Perez et al. also set out to induce endothelial cells in an ischemic mouse limb that, without proper blood flow, becomes necrotic and decays. Using a patented cocktail of plasmids (Etv2, Fli1, Foxc2, or EFF), these factors were delivered to the tissue above the surgery site. Using various methods, including histology and laser speckle imaging, perfusion and the establishment of new vasculature was verified as early as 7 days post-treatment. [13]

The technique was developed to combat the limitations of current approaches, such as a shortage in donors to supply cell sources and the need to induce pluripotency. [14] [15] [16] [17] [18] [19] Reprogramming cells in vivo takes advantage of readily available cells, bypassing the need for pre-processing. [20] [21] Most reprogramming methods have a heavy reliance on viral transfection. [22] [23] TNT allows for implementation of a non-viral approach which is able to overcome issues of capsid size, increase safety, and increase deterministic reprogramming. [13]

Development

The tissue nanotransfection technique was developed as a method to efficiently and benignly deliver cargo to living tissues. This technique builds on the high-throughput nanoelectroporation methods developed for cell reprogramming applications by Lee and Gallego-Perez of Ohio State's Chemical and Biomolecular Engineering department. Sen (Surgery/Regenerative Medicine) adapted this technology, in collaboration with Lee in Engineering, for in vivo tissue reprogramming applications with Gallego-Perez serving the role of a shared fellow between the two programs. Development was a joint effort between OSU's College of Engineering and College of Medicine led by Gallego-Perez (Ph.D), Lee (Ph.D), and Sen (Ph.D).

This technology was fabricated using cleanroom techniques and photolithography and deep reactive ion etching (DRIE) of silicon wafers to create nanochannels with backside etching of a reservoir for loading desired factors as described in Gallego-Perez et al 2017. [13] This chip is then connected to an electrical source capable of delivering an electrical field to drive the factors from the reservoir into the nanochannels, and onto the contacted tissue. Later, with support from Xuan, Sen developed the current version of the tissue nanotransfection chip. [24]

Related Research Articles

<span class="mw-page-title-main">Electroporation</span> Method in molecular biology to make pores in cell membranes

Electroporation, or electropermeabilization, is a technique in which an electrical field is applied to cells in order to increase the permeability of the cell membrane. This may allow chemicals, drugs, electrode arrays or DNA to be introduced into the cell.

Transdifferentiation, also known as lineage reprogramming, is the process in which one mature somatic cell is transformed into another mature somatic cell without undergoing an intermediate pluripotent state or progenitor cell type. It is a type of metaplasia, which includes all cell fate switches, including the interconversion of stem cells. Current uses of transdifferentiation include disease modeling and drug discovery and in the future may include gene therapy and regenerative medicine. The term 'transdifferentiation' was originally coined by Selman and Kafatos in 1974 to describe a change in cell properties as cuticle producing cells became salt-secreting cells in silk moths undergoing metamorphosis.

<span class="mw-page-title-main">Small interfering RNA</span> Biomolecule

Small interfering RNA (siRNA), sometimes known as short interfering RNA or silencing RNA, is a class of double-stranded non-coding RNA molecules, typically 20–24 base pairs in length, similar to microRNA (miRNA), and operating within the RNA interference (RNAi) pathway. It interferes with the expression of specific genes with complementary nucleotide sequences by degrading messenger RNA (mRNA) after transcription, preventing translation. It was discovered in 1998 by Andrew Fire at the Carnegie Institution for Science in Washington, D.C. and Craig Mello at the University of Massachusetts in Worcester.

Transfection is the process of deliberately introducing naked or purified nucleic acids into eukaryotic cells. It may also refer to other methods and cell types, although other terms are often preferred: "transformation" is typically used to describe non-viral DNA transfer in bacteria and non-animal eukaryotic cells, including plant cells. In animal cells, transfection is the preferred term, as the term "transformation" is also used to refer to a cell's progression to a cancerous state (carcinogenesis). Transduction is often used to describe virus-mediated gene transfer into prokaryotic cells.

<span class="mw-page-title-main">Morpholino</span> Chemical compound

A Morpholino, also known as a Morpholino oligomer and as a phosphorodiamidate Morpholino oligomer (PMO), is a type of oligomer molecule used in molecular biology to modify gene expression. Its molecular structure contains DNA bases attached to a backbone of methylenemorpholine rings linked through phosphorodiamidate groups. Morpholinos block access of other molecules to small specific sequences of the base-pairing surfaces of ribonucleic acid (RNA). Morpholinos are used as research tools for reverse genetics by knocking down gene function.

<span class="mw-page-title-main">Gene delivery</span> Introduction of foreign genetic material into host cells

Gene delivery is the process of introducing foreign genetic material, such as DNA or RNA, into host cells. Gene delivery must reach the genome of the host cell to induce gene expression. Successful gene delivery requires the foreign gene delivery to remain stable within the host cell and can either integrate into the genome or replicate independently of it. This requires foreign DNA to be synthesized as part of a vector, which is designed to enter the desired host cell and deliver the transgene to that cell's genome. Vectors utilized as the method for gene delivery can be divided into two categories, recombinant viruses and synthetic vectors.

Cell-penetrating peptides (CPPs) are short peptides that facilitate cellular intake and uptake of molecules ranging from nanosize particles to small chemical compounds to large fragments of DNA. The "cargo" is associated with the peptides either through chemical linkage via covalent bonds or through non-covalent interactions.

Magnetofection is a transfection method that uses magnetic fields to concentrate particles containing vectors to target cells in the body. Magnetofection has been adapted to a variety of vectors, including nucleic acids, non-viral transfection systems, and viruses. This method offers advantages such as high transfection efficiency and biocompatibility which are balanced with limitations.

<span class="mw-page-title-main">Induced pluripotent stem cell</span> Pluripotent stem cell generated directly from a somatic cell

Induced pluripotent stem cells are a type of pluripotent stem cell that can be generated directly from a somatic cell. The iPSC technology was pioneered by Shinya Yamanaka and Kazutoshi Takahashi in Kyoto, Japan, who together showed in 2006 that the introduction of four specific genes, collectively known as Yamanaka factors, encoding transcription factors could convert somatic cells into pluripotent stem cells. Shinya Yamanaka was awarded the 2012 Nobel Prize along with Sir John Gurdon "for the discovery that mature cells can be reprogrammed to become pluripotent."

Magnet-assisted transfection is a transfection method which uses magnetic interactions to deliver DNA into target cells. Nucleic acids are associated with magnetic nanoparticles, and magnetic fields drive the nucleic acid-particle complexes into target cells, where the nucleic acids are released.

Heart nanotechnology is the "Engineering of functional systems at the molecular scale".

<span class="mw-page-title-main">Vectors in gene therapy</span>

Gene therapy utilizes the delivery of DNA into cells, which can be accomplished by several methods, summarized below. The two major classes of methods are those that use recombinant viruses and those that use naked DNA or DNA complexes.

A list of examples of in vivo transdifferentiation through transfection:

Gene therapy for osteoarthritis is the application of gene therapy to treat osteoarthritis (OA). Unlike pharmacological treatments which are administered locally or systemically as a series of interventions, gene therapy aims to establish sustained therapeutic effect after a single, local injection.

DNA-directed RNA interference (ddRNAi) is a gene-silencing technique that utilizes DNA constructs to activate a cell's endogenous RNA interference (RNAi) pathways. DNA constructs are designed to express self-complementary double-stranded RNAs, typically short-hairpin RNAs (shRNA), that bring about the silencing of a target gene or genes once processed. Any RNA, including endogenous messenger RNA (mRNAs) or viral RNAs, can be silenced by designing constructs to express double-stranded RNA complementary to the desired mRNA target.

William Mark Saltzman was named the Goizueta Foundation Professor of Biomedical and Chemical Engineering at Yale University on July 1, 2002 and became the founding chair of Yale's Department of Biomedical Engineering in 2003. Saltzman's research aims to promote new methods for drug delivery and develop new biotechnologies to combat human disease. A pioneer in the fields of biomaterials, nanobiotechnology, and tissue engineering, Saltzman has contributed to the design and implementation of a number of clinical technologies that have become essential to medical practice today. His popular course Frontiers of Biomedical Engineering is available to everyone through Open Yale Courses.

Quantum dots (QDs) are semiconductor nanoparticles with a size less than 10 nm. They exhibited size-dependent properties especially in the optical absorption and the photoluminescence (PL). Typically, the fluorescence emission peak of the QDs can be tuned by changing their diameters. So far, QDs were consisted of different group elements such as CdTe, CdSe, CdS in the II-VI category, InP or InAs in the III-V category, CuInS2 or AgInS2 in the I–III–VI2 category, and PbSe/PbS in the IV-VI category. These QDs are promising candidates as fluorescent labels in various biological applications such as bioimaging, biosensing and drug delivery.

RNA therapeutics are a new class of medications based on ribonucleic acid (RNA). Research has been working on clinical use since the 1990s, with significant success in cancer therapy in the early 2010s. In 2020 and 2021, mRNA vaccines have been developed globally for use in combating the coronavirus disease. The Pfizer–BioNTech COVID-19 vaccine was the first mRNA vaccine approved by a medicines regulator, followed by the Moderna COVID-19 vaccine, and others.

<span class="mw-page-title-main">Intracellular delivery</span> Scientific research area

Intracellular delivery is the process of introducing external materials into living cells. Materials that are delivered into cells include nucleic acids, proteins, peptides, impermeable small molecules, synthetic nanomaterials, organelles, and micron-scale tracers, devices and objects. Such molecules and materials can be used to investigate cellular behavior, engineer cell operations or correct a pathological function.

<span class="mw-page-title-main">Hydrodynamic delivery</span> Gene Transfer Method

Hydrodynamic Delivery (HD) is a method of DNA insertion in rodent models. Genes are delivered via injection into the bloodstream of the animal, and are expressed in the liver. This protocol is helpful to determine gene function, regulate gene expression, and develop pharmaceuticals in vivo.

References

  1. ↑ Chen Z, Zhang A, Wang X, Zhu J, Fan Y, Yu H, Yang Z (2017). "The Advances of Carbon Nanotubes in Cancer Diagnostics and Therapeutics". Journal of Nanomaterials. 2017: 1–13. doi: 10.1155/2017/3418932 .
  2. ↑ Kang C, Sun Y, Zhu J, Li W, Zhang A, Kuang T, Xie J, Yang Z (2016-09-30). "Delivery of Nanoparticles for Treatment of Brain Tumor". Current Drug Metabolism. 17 (8): 745–754. doi:10.2174/1389200217666160728152939. PMID   27469219.
  3. ↑ Xie J, Yang Z, Zhou C, Zhu J, Lee RJ, Teng L (July 2016). "Nanotechnology for the delivery of phytochemicals in cancer therapy". Biotechnology Advances. 34 (4): 343–353. doi:10.1016/j.biotechadv.2016.04.002. PMID   27071534.
  4. ↑ Chen Z, Chen Z, Zhang A, Hu J, Wang X, Yang Z (June 2016). "Electrospun nanofibers for cancer diagnosis and therapy". Biomaterials Science. 4 (6): 922–32. doi:10.1039/C6BM00070C. PMID   27048889.
  5. ↑ Sha L, Chen Z, Chen Z, Zhang A, Yang Z (2016). "Polylactic Acid Based Nanocomposites: Promising Safe and Biodegradable Materials in Biomedical Field". International Journal of Polymer Science. 2016: 1–11. doi: 10.1155/2016/6869154 .
  6. ↑ Xie J, Teng L, Yang Z, Zhou C, Liu Y, Yung BC, Lee RJ (2013). "A polyethylenimine-linoleic acid conjugate for antisense oligonucleotide delivery". BioMed Research International. 2013: 710502. doi: 10.1155/2013/710502 . PMC   3683435 . PMID   23862153.
  7. 1 2 Shi J, Ma Y, Zhu J, Chen Y, Sun Y, Yao Y, Yang Z, Xie J (November 2018). "A Review on Electroporation-Based Intracellular Delivery". Molecules. 23 (11): 3044. doi: 10.3390/molecules23113044 . PMC   6278265 . PMID   30469344.
  8. ↑ Sun J, Wang X, Wu J, Jiang C, Shen J, Cooper MA, Zheng X, Liu Y, Yang Z, Wu D (April 2018). "Biomimetic Moth-eye Nanofabrication: Enhanced Antireflection with Superior Self-cleaning Characteristic". Scientific Reports. 8 (1): 5438. Bibcode:2018NatSR...8.5438S. doi:10.1038/s41598-018-23771-y. PMC   5883013 . PMID   29615712.
  9. ↑ Sun J, Kormakov S, Liu Y, Huang Y, Wu D, Yang Z (July 2018). "Recent Progress in Metal-Based Nanoparticles Mediated Photodynamic Therapy". Molecules. 23 (7): 1704. doi: 10.3390/molecules23071704 . PMC   6099795 . PMID   30002333.
  10. ↑ Kurosawa O, Oana H, Matsuoka S, Noma A, Kotera H, Washizu M (2006-12-01). "Electroporation through a micro-fabricated orifice and its application to the measurement of cell response to external stimuli". Measurement Science and Technology. 17 (12): 3127–3133. doi:10.1088/0957-0233/17/12/S02. S2CID   121182614.
  11. 1 2 Boukany PE, Morss A, Liao WC, Henslee B, Jung H, Zhang X, Yu B, Wang X, Wu Y, Li L, Gao K, Hu X, Zhao X, Hemminger O, Lu W, Lafyatis GP, Lee LJ (October 2011). "Nanochannel electroporation delivers precise amounts of biomolecules into living cells". Nature Nanotechnology. 6 (11): 747–54. Bibcode:2011NatNa...6..747B. doi:10.1038/nnano.2011.164. PMID   22002097.
  12. 1 2 3 Gallego-Perez D, Otero JJ, Czeisler C, Ma J, Ortiz C, Gygli P, et al. (February 2016). "Deterministic transfection drives efficient nonviral reprogramming and uncovers reprogramming barriers". Nanomedicine. 12 (2): 399–409. doi:10.1016/j.nano.2015.11.015. PMC   5161095 . PMID   26711960.
  13. 1 2 3 4 5 6 7 8 Gallego-Perez D, Pal D, Ghatak S, Malkoc V, Higuita-Castro N, Gnyawali S, Chang L, Liao WC, Shi J, Sinha M, Singh K, Steen E, Sunyecz A, Stewart R, Moore J, Ziebro T, Northcutt RG, Homsy M, Bertani P, Lu W, Roy S, Khanna S, Rink C, Sundaresan VB, Otero JJ, Lee LJ, Sen CK (October 2017). "Topical tissue nano-transfection mediates non-viral stroma reprogramming and rescue". Nature Nanotechnology. 12 (10): 974–979. Bibcode:2017NatNa..12..974G. doi:10.1038/nnano.2017.134. PMC   5814120 . PMID   28785092.
  14. 1 2 Vierbuchen T, Ostermeier A, Pang ZP, Kokubu Y, Südhof TC, Wernig M (February 2010). "Direct conversion of fibroblasts to functional neurons by defined factors". Nature. 463 (7284): 1035–41. Bibcode:2010Natur.463.1035V. doi:10.1038/nature08797. PMC   2829121 . PMID   20107439.
  15. 1 2 Valadi H, Ekström K, Bossios A, Sjöstrand M, Lee JJ, Lötvall JO (June 2007). "Exosome-mediated transfer of mRNAs and microRNAs is a novel mechanism of genetic exchange between cells". Nature Cell Biology. 9 (6): 654–9. doi:10.1038/ncb1596. PMID   17486113. S2CID   8599814.
  16. ↑ Davis DM, Sowinski S (June 2008). "Membrane nanotubes: dynamic long-distance connections between animal cells". Nature Reviews. Molecular Cell Biology. 9 (6): 431–6. doi:10.1038/nrm2399. PMID   18431401. S2CID   8136865.
  17. ↑ Rosová I, Dao M, Capoccia B, Link D, Nolta JA (August 2008). "Hypoxic preconditioning results in increased motility and improved therapeutic potential of human mesenchymal stem cells". Stem Cells. 26 (8): 2173–82. doi:10.1634/stemcells.2007-1104. PMC   3017477 . PMID   18511601.
  18. ↑ Kinoshita M, Fujita Y, Katayama M, Baba R, Shibakawa M, Yoshikawa K, Katakami N, Furukawa Y, Tsukie T, Nagano T, Kurimoto Y, Yamasaki K, Handa N, Okada Y, Kuronaka K, Nagata Y, Matsubara Y, Fukushima M, Asahara T, Kawamoto A (October 2012). "Long-term clinical outcome after intramuscular transplantation of granulocyte colony stimulating factor-mobilized CD34 positive cells in patients with critical limb ischemia". Atherosclerosis. 224 (2): 440–5. doi:10.1016/j.atherosclerosis.2012.07.031. PMID   22877866.
  19. ↑ Losordo DW, Dimmeler S (June 2004). "Therapeutic angiogenesis and vasculogenesis for ischemic disease: part II: cell-based therapies". Circulation. 109 (22): 2692–7. doi: 10.1161/01.CIR.0000128596.49339.05 . PMID   15184293.
  20. ↑ Lee AS, Tang C, Rao MS, Weissman IL, Wu JC (August 2013). "Tumorigenicity as a clinical hurdle for pluripotent stem cell therapies". Nature Medicine. 19 (8): 998–1004. doi:10.1038/nm.3267. PMC   3967018 . PMID   23921754.
  21. ↑ Cunningham JJ, Ulbright TM, Pera MF, Looijenga LH (September 2012). "Lessons from human teratomas to guide development of safe stem cell therapies". Nature Biotechnology. 30 (9): 849–57. doi:10.1038/nbt.2329. PMID   22965062. S2CID   20383770.
  22. ↑ Leduc PR, Wong MS, Ferreira PM, Groff RE, Haslinger K, Koonce MP, et al. (January 2007). "Towards an in vivo biologically inspired nanofactory". Nature Nanotechnology. 2 (1): 3–7. Bibcode:2007NatNa...2....3L. doi:10.1038/nnano.2006.180. PMID   18654192.
  23. ↑ Heinrich C, Spagnoli FM, Berninger B (March 2015). "In vivo reprogramming for tissue repair". Nature Cell Biology. 17 (3): 204–11. doi:10.1038/ncb3108. PMID   25720960. S2CID   32061267.
  24. ↑ Xuan, Yi; Ghatak, Subhadip; Clark, Andrew; Li, Zhigang; Khanna, Savita; Pak, Dongmin; Agarwal, Mangilal; Roy, Sashwati; Duda, Peter; Sen, Chandan K. (December 2021). "Fabrication and use of silicon hollow-needle arrays to achieve tissue nanotransfection in mouse tissue in vivo". Nature Protocols. 16 (12): 5707–5738. doi:10.1038/s41596-021-00631-0. PMC   9104164 . PMID   34837085.
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