Transient Expression of Proteins by Hydrodynamic Gene Delivery in Mice

Abstract

Efficient expression of transgenes in vivo is of critical importance in studying gene function and developing treatments for diseases. Over the past years, hydrodynamic gene delivery (HGD) has emerged as a simple, fast, safe and effective method for delivering transgenes into rodents. This technique relies on the force generated by the rapid injection of a large volume of physiological solution to increase the permeability of cell membranes of perfused organs and thus deliver DNA into cells. One of the main advantages of HGD is the ability to introduce transgenes into mammalian cells using naked plasmid DNA (pDNA). Introducing an exogenous gene using a plasmid is minimally laborious, highly efficient and, contrary to viral carriers, remarkably safe. HGD was initially used to deliver genes into mice, it is now used to deliver a wide range of substances, including oligonucleotides, artificial chromosomes, RNA, proteins and small molecules into mice, rats and, to a limited degree, other animals. This protocol describes HGD in mice and focuses on three key aspects of the method that are critical to performing the procedure successfully: correct insertion of the needle into the vein, the volume of injection and the speed of delivery. Examples are given to show the application of this method to the transient expression of two genes that encode secreted, primate-specific proteins, apolipoprotein L-I (APOL-I) and haptoglobinrelated protein (HPR). Video LinkThe video component of this article can be found at

Advances and potential of gene therapy for type 2 diabetes mellitus

“…However, viruses have wild tropism to transduce multiple cell types, and genetic manipulation of viruses is needed before the virus genome with the gene target is packaged [52]. Direct injection of naked DNA is of low immunity and low cost and is convenient for preparation, but the main barrier for its wide use is poor delivery efficiency and transient gene expression [53]. Administration of (nano-scale) DNAmaterial conjugates significantly improved delivery efficiency and stability but its expression and localization are still uncontrollable [54,55].…”

Prime-and-Trap Malaria Vaccination To Generate Protective CD8+ Liver-Resident Memory T Cells

“…Hydrodynamic transfection (HDT) of DNA by large-volume tail vein injection was performed under isoflurane anesthetic and adapted from published protocols (31,32). Briefly, mice were weighed, and 10% v/w PBS containing 20 mg of endotoxin-free plasmid DNA (Qiagen) was injected into the tail vein in 5-10 s using a 27.5-gauge needle.…”

“…To determine if CSP-specific CD8 + T cells produced by ggCSP vaccination were capable of giving rise to liver Trm cells, we performed HDT on ggCSP-vaccinated mice to directly express the P. yoelii CSP epitope in hepatocytes and to assay in vivo CTL responses against Ag-expressing hepatocytes and subsequent liver Trm cell formation. In HDT, mice are anesthetized and injected in the tail vein with a volume of DNA-containing saline equal to 10% body weight over 5-10 s; the mice tolerate the procedure without incident, recover within minutes, and suffer no apparent long-term side effects (31,43). The sudden increase in the oncotic pressure gradient drives injected plasmid DNA into hepatocytes, where plasmid-encoded genes are expressed.…”

Mouse Models of Hepatocellular Carcinoma

“…A simple and inexpensive alternative method to transfect and gene-edit hepatocytes in vivo is represented by hydrodynamic tail vein injection (HTVI) of "naked" plasmid DNA (non-viral vector, not associated with protective proteins or lipids) directly into the liver of adult mice (136,137). The technique consists of rapid injection of a large amount of plasmid DNA (about 50 μg in a volume of saline that signifies 10% of the body weight of the injected mouse) into the mouse lateral tail vein (138). HTVI results in transient heart dysfunction and fluid accumulation in the inferior vena cava.…”

“…The enormous hydrodynamic pressure then forces the fluid into the liver in a retrograde movement, enlarges the liver, and pushes the plasmid DNA into hepatocytes via enlarged sinusoidal fenestrae and transient membrane pores (139), with transfection efficiencies ranging from around 10 to 40% of all hepatocytes (136,140). While the liver is primarily targeted, other organs including heart, kidney, lung, and spleen are also affected by HTVI; however, to only 0.1% of the levels achieved in the liver (138,141). One caveat of HTVI is that the expression of transfected genes is transient, peaking within 24 h, but dropping dramatically thereafter (142).…”