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11 pages, 1819 KiB

Open AccessArticle

Delivery of miRNA-Targeted Oligonucleotides in the Rat Striatum by Magnetofection with Neuromag^®

by Simoneide Souza Titze de Almeida, Camila Hillesheim Horst, Cristina Soto-Sánchez, Eduardo Fernandez and Ricardo Titze de Almeida

Molecules 2018, 23(7), 1825; https://doi.org/10.3390/molecules23071825 - 23 Jul 2018

Cited by 31 | Viewed by 5358

Abstract

MicroRNAs (miRNAs) regulate gene expression at posttranscriptional level by triggering RNA interference. In such a sense, aberrant expressions of miRNAs play critical roles in the pathogenesis of many disorders, including Parkinson’s disease (PD). Controlling the level of specific miRNAs in the brain is [...] Read more.

MicroRNAs (miRNAs) regulate gene expression at posttranscriptional level by triggering RNA interference. In such a sense, aberrant expressions of miRNAs play critical roles in the pathogenesis of many disorders, including Parkinson’s disease (PD). Controlling the level of specific miRNAs in the brain is thus a promising therapeutic strategy for neuroprotection. A fundamental need for miRNA regulation (either replacing or inhibition) is a carrier capable of delivering oligonucleotides into brain cells. This study aimed to examine a polymeric magnetic particle, Neuromag^®, for delivery of synthetic miRNA inhibitors in the rat central nervous system. We injected the miRNA inhibitor complexed with Neuromag^® into the lateral ventricles next to the striatum, by stereotaxic surgery. Neuromag efficiently delivered oligonucleotides in the striatum and septum areas, as shown by microscopy imaging of fluorescein isothiocyanate (FITC)-labeled oligos in astrocytes and neurons. Transfected oligos showed efficacy concerning miRNA inhibition. Neuromag^®-structured miR-134 antimiR (0.36 nmol) caused a significant 0.35 fold decrease of striatal miR-134, as revealed by real-time quantitative polymerase chain reaction (RT-qPCR). In conclusion, the polymeric magnetic particle Neuromag^® efficiently delivered functional miRNA inhibitors in brain regions surrounding lateral ventricles, particularly the striatum. This delivery system holds potential as a promising miRNA-based disease-modifying drug and merits further pre-clinical studies using animal models of PD. Full article

(This article belongs to the Special Issue Gene Delivery)

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14 pages, 3860 KiB

Open AccessArticle

DNA Interaction with Head-to-Tail Associates of Cationic Surfactants Prevents Formation of Compact Particles

by Nina Kasyanenko, Ivan Unksov, Vladimir Bakulev and Svetlana Santer

Molecules 2018, 23(7), 1576; https://doi.org/10.3390/molecules23071576 - 28 Jun 2018

Cited by 9 | Viewed by 4395

Abstract

Cationic azobenzene-containing surfactants are capable of condensing DNA in solution with formation of nanosized particles that can be employed in gene delivery. The ratio of surfactant/DNA concentration and solution ionic strength determines the result of DNA-surfactant interaction: Complexes with a micelle-like surfactant associates [...] Read more.

Cationic azobenzene-containing surfactants are capable of condensing DNA in solution with formation of nanosized particles that can be employed in gene delivery. The ratio of surfactant/DNA concentration and solution ionic strength determines the result of DNA-surfactant interaction: Complexes with a micelle-like surfactant associates on DNA, which induces DNA shrinkage, DNA precipitation or DNA condensation with the emergence of nanosized particles. UV and fluorescence spectroscopy, low gradient viscometry and flow birefringence methods were employed to investigate DNA-surfactant and surfactant-surfactant interaction at different NaCl concentrations, [NaCl]. It was observed that [NaCl] (or the Debye screening radius) determines the surfactant-surfactant interaction in solutions without DNA. Monomers, micelles and non-micellar associates of azobenzene-containing surfactants with head-to-tail orientation of molecules were distinguished due to the features of their absorption spectra. The novel data enabled us to conclude that exactly the type of associates (together with the concentration of components) determines the result of DNA-surfactant interaction. Predomination of head-to-tail associates at 0.01 M < [NaCl] < 0.5 M induces DNA aggregation and in some cases DNA precipitation. High NaCl concentration (higher than 0.8 M) prevents electrostatic attraction of surfactants to DNA phosphates for complex formation. DAPI dye luminescence in solutions with DNA-surfactant complexes shows that surfactant tails overlap the DNA minor groove. The addition of di- and trivalent metal ions before and after the surfactant binding to DNA indicate that the bound surfactant molecules are located on DNA in islets. Full article

(This article belongs to the Special Issue Gene Delivery)

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18 pages, 6843 KiB

Open AccessArticle

Synthesis and Comparative Evaluation of Novel Cationic Amphiphile C12-Man-Q as an Efficient DNA Delivery Agent In Vitro

by Gunita Apsite, Irena Timofejeva, Aleksandra Vezane, Brigita Vigante, Martins Rucins, Arkadij Sobolev, Mara Plotniece, Karlis Pajuste, Tatjana Kozlovska and Aiva Plotniece

Molecules 2018, 23(7), 1540; https://doi.org/10.3390/molecules23071540 - 26 Jun 2018

Cited by 9 | Viewed by 4194

Abstract

New amphiphilic 1,4-DHP derivative C12-Man-Q with remoted cationic moieties at positions 2 and 6 was synthesised to study DNA delivery activity. The results were compared with data obtained for cationic 1,4-DHP derivative D19, which is known to be the most efficient one [...] Read more.

New amphiphilic 1,4-DHP derivative C12-Man-Q with remoted cationic moieties at positions 2 and 6 was synthesised to study DNA delivery activity. The results were compared with data obtained for cationic 1,4-DHP derivative D19, which is known to be the most efficient one among the previously tested 1,4-DHP amphiphiles. We analysed the effects of C12-Man-Q concentration, complexation media, and complex/cell contact time on the gene delivery effectiveness and cell viability. Transmission electron microscopy data confirms that lipoplexes formed by the compound C12-Man-Q were quite uniform, vesicular-like structures with sizes of about 50 nm, and lipoplexes produced by compound D19 were of irregular shapes, varied in size in the range of 25–80 nm. Additionally, confocal microscopy results revealed that both amphiphiles effectively delivered green fluorescent protein expression plasmid into BHK-21 cells and produced a fluorescent signal with satisfactory efficiency, although compound C12-Man-Q was more cytotoxic to the BHK-21 cells with an increase of concentration. It can be concluded that optimal conditions for C12-Man-Q lipoplexes delivery in BHK-21 cells were the serum free media without 0.15 M NaCl, at an N/P ratio of 0.9. Compound D19 showed higher transfection efficiency to transfect BHK-21 and Cos-7 cell lines, when transfecting active proliferating cells. Although D19 was not able to transfect all studied cell lines we propose that it could be cell type specific. The compound C12-Man-Q showed modest delivery activity in all used cell lines, and higher activity was obtained in the case of H2-35 and B16 cells. The transfection efficiency in cell lines MCF-7, HeLa, and Huh-7 appears to be comparable to the reference compound D19 and minimal in the HepG2 cell line. Full article

(This article belongs to the Special Issue Gene Delivery)

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10 pages, 1772 KiB

Open AccessArticle

Development of a Mouse Model of Prostate Cancer Using the Sleeping Beauty Transposon and Electroporation

by Hyun-Ji Choi, Han-Byul Lee, Sunyoung Jung, Hyun-Kyu Park, Woori Jo, Sung-Min Cho, Woo-Jin Kim and Woo-Chan Son

Molecules 2018, 23(6), 1360; https://doi.org/10.3390/molecules23061360 - 5 Jun 2018

Cited by 5 | Viewed by 4868

Abstract

The Sleeping Beauty (SB) transposon system is non-viral and uses insertional mutagenesis, resulting in the permanent expression of transferred genes. Although the SB transposon is a useful method for establishing a mouse tumor model, there has been difficulty in using this method to [...] Read more.

The Sleeping Beauty (SB) transposon system is non-viral and uses insertional mutagenesis, resulting in the permanent expression of transferred genes. Although the SB transposon is a useful method for establishing a mouse tumor model, there has been difficulty in using this method to generate tumors in the prostate. In the present study, electroporation was used to enhance the transfection efficiency of the SB transposon. To generate tumors, three constructs (a c-Myc expression cassette, a HRAS (HRas proto-oncogene, GTPase) expression cassette and a shRNA against p53) contained within the SB transposon plasmids were directly injected into the prostate. Electroporation was conducted on the injection site after the injection of the DNA plasmid. Following the tumorigenesis, the tumors were monitored by animal PET imaging and identified by gross observation. After this, the tumors were characterized by using histological and immunohistochemical techniques. The expression of the targeted genes was analyzed by Real-Time qRT-PCR. All mice subjected to the injection were found to have prostate tumors, which was supported by PSA immunohistochemistry. To our knowledge, this is the first demonstration of tumor induction in the mouse prostate using the electroporation-enhanced SB transposon system in combination with c-Myc, HRAS and p53. This model serves as a valuable resource for the future development of SB-induced mouse models of cancer. Full article

(This article belongs to the Special Issue Gene Delivery)

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Review

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24 pages, 7275 KiB

Open AccessFeature PaperReview

Biodegradable Polymers for Gene Delivery

by T. J. Thomas, Heidar-Ali Tajmir-Riahi and C. K. S. Pillai

Molecules 2019, 24(20), 3744; https://doi.org/10.3390/molecules24203744 - 17 Oct 2019

Cited by 122 | Viewed by 9219

Abstract

The cellular transport process of DNA is hampered by cell membrane barriers, and hence, a delivery vehicle is essential for realizing the potential benefits of gene therapy to combat a variety of genetic diseases. Virus-based vehicles are effective, although immunogenicity, toxicity and cancer [...] Read more.

The cellular transport process of DNA is hampered by cell membrane barriers, and hence, a delivery vehicle is essential for realizing the potential benefits of gene therapy to combat a variety of genetic diseases. Virus-based vehicles are effective, although immunogenicity, toxicity and cancer formation are among the major limitations of this approach. Cationic polymers, such as polyethyleneimine are capable of condensing DNA to nanoparticles and facilitate gene delivery. Lack of biodegradation of polymeric gene delivery vehicles poses significant toxicity because of the accumulation of polymers in the tissue. Many attempts have been made to develop biodegradable polymers for gene delivery by modifying existing polymers and/or using natural biodegradable polymers. This review summarizes mechanistic aspects of gene delivery and the development of biodegradable polymers for gene delivery. Full article

(This article belongs to the Special Issue Gene Delivery)

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10 pages, 4434 KiB

Open AccessReview

Gene Delivery into the Inner Ear and Its Clinical Implications for Hearing and Balance

by Sho Kanzaki

Molecules 2018, 23(10), 2507; https://doi.org/10.3390/molecules23102507 - 30 Sep 2018

Cited by 14 | Viewed by 5243

Abstract

The inner ear contains many types of cell, including sensory hair cells and neurons. If these cells are damaged, they do not regenerate. Inner ear disorders have various etiologies. Some are related to aging or are idiopathic, as in sudden deafness. Others occur [...] Read more.

The inner ear contains many types of cell, including sensory hair cells and neurons. If these cells are damaged, they do not regenerate. Inner ear disorders have various etiologies. Some are related to aging or are idiopathic, as in sudden deafness. Others occur due to acoustic trauma, exposure to ototoxic drugs, viral infections, immune responses, or endolymphatic hydrops (Meniere’s disease). For these disorders, inner ear regeneration therapy is expected to be a feasible alternative to cochlear implants for hearing recovery. Recently, the mechanisms underlying inner ear regeneration have been gradually clarified. Inner ear cell progenitors or stem cells have been identified. Factors necessary for regeneration have also been elucidated from the mechanism of hair cell generation. Inducing differentiation of endogenous stem cells or inner ear stem cell transplantation is expected. In this paper, we discuss recent approaches to hair cell proliferation and differentiation for inner ear regeneration. We discuss the future road map for clinical application. The therapies mentioned above require topical administration of transgenes or drug onto progenitors of sensory cells. Developing efficient and safe modes of administration is clinically important. In this regard, we also discuss our development of an inner ear endoscope to facilitate topical administration. Full article

(This article belongs to the Special Issue Gene Delivery)

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16 pages, 2266 KiB

Open AccessReview

Recent Advances in Mitochondria-Targeted Gene Delivery

by Yoon-ha Jang and Kwang-il Lim

Molecules 2018, 23(9), 2316; https://doi.org/10.3390/molecules23092316 - 11 Sep 2018

Cited by 39 | Viewed by 9278

Abstract

Mitochondria are the energy-producing organelles of cells. Mitochondrial dysfunctions link to various syndromes and diseases including myoclonic epilepsy and ragged-red fiber disease (MERRF), Leigh syndrome (LS), and Leber hereditary optic neuropathy (LHON). Primary mitochondrial diseases often result from mutations of mitochondrial genomes and [...] Read more.

Mitochondria are the energy-producing organelles of cells. Mitochondrial dysfunctions link to various syndromes and diseases including myoclonic epilepsy and ragged-red fiber disease (MERRF), Leigh syndrome (LS), and Leber hereditary optic neuropathy (LHON). Primary mitochondrial diseases often result from mutations of mitochondrial genomes and nuclear genes that encode the mitochondrial components. However, complete intracellular correction of the mutated genetic parts relevant to mitochondrial structures and functions is technically challenging. Instead, there have been diverse attempts to provide corrected genetic materials with cells. In this review, we discuss recent novel physical, chemical and biological strategies, and methods to introduce genetic cargos into mitochondria of eukaryotic cells. Effective mitochondria-targeting gene delivery systems can reverse multiple mitochondrial disorders by enabling cells to produce functional mitochondrial components. Full article

(This article belongs to the Special Issue Gene Delivery)

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13 pages, 259 KiB

Open AccessReview

Turning Stem Cells Bad: Generation of Clinically Relevant Models of Human Acute Myeloid Leukemia through Gene Delivery- or Genome Editing-Based Approaches

by Maria Mesuraca, Nicola Amodio, Emanuela Chiarella, Stefania Scicchitano, Annamaria Aloisio, Bruna Codispoti, Valeria Lucchino, Ylenia Montalcini, Heather M. Bond and Giovanni Morrone

Molecules 2018, 23(8), 2060; https://doi.org/10.3390/molecules23082060 - 17 Aug 2018

Cited by 6 | Viewed by 4327

Abstract

Acute myeloid leukemia (AML), the most common acute leukemia in the adult, is believed to arise as a consequence of multiple molecular events that confer on primitive hematopoietic progenitors unlimited self-renewal potential and cause defective differentiation. A number of genetic aberrations, among which [...] Read more.

Acute myeloid leukemia (AML), the most common acute leukemia in the adult, is believed to arise as a consequence of multiple molecular events that confer on primitive hematopoietic progenitors unlimited self-renewal potential and cause defective differentiation. A number of genetic aberrations, among which a variety of gene fusions, have been implicated in the development of a transformed phenotype through the generation of dysfunctional molecules that disrupt key regulatory mechanisms controlling survival, proliferation, and differentiation in normal stem and progenitor cells. Such genetic aberrations can be recreated experimentally to a large extent, to render normal hematopoietic stem cells “bad”, analogous to the leukemic stem cells. Here, we wish to provide a brief outline of the complementary experimental approaches, largely based on gene delivery and more recently on gene editing, employed over the last two decades to gain insights into the molecular mechanisms underlying AML development and progression and on the prospects that their applications offer for the discovery and validation of innovative therapies. Full article

(This article belongs to the Special Issue Gene Delivery)

14 pages, 781 KiB

Open AccessReview

MicroRNA-Regulated Gene Delivery Systems for Research and Therapeutic Purposes

by Bijay Dhungel, Charmaine A. Ramlogan-Steel and Jason C. Steel

Molecules 2018, 23(7), 1500; https://doi.org/10.3390/molecules23071500 - 21 Jun 2018

Cited by 22 | Viewed by 5441

Abstract

Targeted gene delivery relies on the ability to limit the expression of a transgene within a defined cell/tissue population. MicroRNAs represent a class of highly powerful and effective regulators of gene expression that act by binding to a specific sequence present in the [...] Read more.

Targeted gene delivery relies on the ability to limit the expression of a transgene within a defined cell/tissue population. MicroRNAs represent a class of highly powerful and effective regulators of gene expression that act by binding to a specific sequence present in the corresponding messenger RNA. Involved in almost every aspect of cellular function, many miRNAs have been discovered with expression patterns specific to developmental stage, lineage, cell-type, or disease stage. Exploiting the binding sites of these miRNAs allows for construction of targeted gene delivery platforms with a diverse range of applications. Here, we summarize studies that have utilized miRNA-regulated systems to achieve targeted gene delivery for both research and therapeutic purposes. Additionally, we identify criteria that are important for the effectiveness of a particular miRNA for such applications and we also discuss factors that have to be taken into consideration when designing miRNA-regulated expression cassettes. Full article

(This article belongs to the Special Issue Gene Delivery)

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