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Review
, 168, 69-85

In Vivo Methods for Acute Modulation of Gene Expression in the Central Nervous System

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Review

In Vivo Methods for Acute Modulation of Gene Expression in the Central Nervous System

Andrzej W Cwetsch et al. Prog Neurobiol.

Abstract

Accurate and timely expression of specific genes guarantees the healthy development and function of the brain. Indeed, variations in the correct amount or timing of gene expression lead to improper development and/or pathological conditions. Almost forty years after the first successful gene transfection in in vitro cell cultures, it is currently possible to regulate gene expression in an area-specific manner at any step of central nervous system development and in adulthood in experimental animals in vivo, even overcoming the very poor accessibility of the brain. Here, we will review the diverse approaches for acute gene transfer in vivo, highlighting their advantages and disadvantages with respect to the efficiency and specificity of transfection as well as to brain accessibility. In particular, we will present well-established chemical, physical and virus-based approaches suitable for different animal models, pointing out their current and future possible applications in basic and translational research as well as in gene therapy.

Keywords: Electroporation; In vivo genetic manipulations; Nanoparticles; Polymers; Sonoporation; Viruses.

Figures

Fig. 1
Fig. 1. Chemical methods of gene delivery.
Lipid-mediated gene transfer (left) occurs by the interaction between the positively charged surface of the carriers and the negatively charged cell membrane. This interaction promotes endocytosis, creating an endosome lipocomplex. The complex is then degraded in the cell cytoplasm, and the released DNA is transported to the nucleus. Nanoparticle-mediated gene transfer (middle) occurs by the nanoparticles binding to receptors on the cell surface, followed by endocytosis. During endosome degradation in the cell cytoplasm, the DNA attached to the core of the nanoparticles is released and transported to the nucleus. Similar to lipid-mediated gene transfer, polymer-mediated gene transfer (right) occurs via charge differences between the carriers and the cell membrane, which promotes binding and endocytosis. During endosome degradation, the DNA is released from the polymer structure and is transported to the nucleus. After DNA transcription, the mRNA exits the nucleus, and it is translated into a protein in the cytoplasm.
Fig. 2
Fig. 2. Physical methods of gene delivery.
Electroporation-mediated gene transfer (left) enables the directional guidance of plasmids carrying the negatively charged DNA of interest towards the positive pole of the electrode. At the same time, electroporation causes destabilization of the structure of the cell membrane, creating temporary pores in its surface and allowing the plasmid to enter the cell. The plasmid is then transported to the nucleus. Sonoporation-mediated gene transfer (middle) occurs by ultrasound application, which promotes destabilization of the cell membrane in the presence of oscillating microbubbles. During this process, the plasmid contained in the microbubble mixture enters the cell and is then transported to the nucleus. Magnet-assisted transfection (right) is mediated by magnetic field oscillations that guide and promote endocytosis of the magnetic carriers with attached DNA. In the cytoplasm of the cell, the endosome undergoes degradation and the released DNA enters the cell nucleus. After DNA transcription, the mRNA exits the nucleus, and it is translated into a protein in the cytoplasm.
Fig. 3
Fig. 3. Virus-mediated transduction.
A lentivirus (left) can carry RNA inside its capsid. Contact with cell-specific receptors induces the fusion of the capsid with the cell membrane, followed by RNA release. In the cell cytoplasm, the released RNA undergoes reverse transcription, and upon transport to the nucleus, the RNA is integrated with the host DNA. Adenovirus infection (middle left) is mediated by specific receptor binding on the cell membrane, followed by endocytosis. Endosome degradation then results in the release of the virus capsid and lysosomal degradation of the endosome. The released virus capsid binds to the nuclear pore of the infected-cell nucleus, allowing the introduction of the double-stranded DNA inside the nucleus. Adeno-associated virus infection (middle right) is mediated by receptor binding on the host-cell surface and endocytosis. Endosome degradation results in the release of single-stranded DNA. The single-stranded DNA enters the nucleus and undergoes a conversion to double-stranded DNA, which can be either integrated in the host DNA (and later transcribed) or can remain in the host cell nucleus as nonintegrated viral DNA. Herpes simplex virus-mediated gene delivery (right) is the result of receptor binding and fusion of the virus with the cell membrane of the host cell. Inside the cell, the released capsid binds to the nuclear pore and introduces double-stranded DNA into the nucleus to be transcribed. The viral DNA in the host cell nucleus is transcribed into mRNA. The transcribed viral mRNA exits the nucleus, and it is translated into a protein in the cytoplasm of the host cell.

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