Liver Tissue-Specific Expression Regulation of AAV

Background

The liver is the largest solid organ in the human body, a frequent site of organ-specific and systemic diseases, and a common site of tumor metastasis. In liver biology, it is particularly important to study hepatocytes, which are composed of several cell types of different embryonic origins, including hepatocytes, biliary epithelial cells (cholangiocytes), stellate cells, Kupffer cells, and sinusoidal endothelial cells. Hepatocytes are the most common cell type in the liver, accounting for approximately 60% to 80% of the total number of liver cells. They have a variety of functions, including synthesizing, decomposing, and secreting a variety of substances, such as bile, cholesterol, fatty acids, glucose, and ammonia. It can also store and release metal ions such as iron, copper, and zinc, as well as fat-soluble vitamins such as vitamins A, D, and K. Bile duct cells are the second most abundant epithelial cell group in the liver. As cells in the bile duct lumen, they have traditional epithelial functions and bile secretion functions. Hepatic stellate cells are a special type of liver cells that are mainly distributed in the outer layer of the liver. Hepatic stellate cells have secretion and phagocytosis functions, and are involved in liver repair and extracellular matrix accumulation. Kupffer cells are the resident macrophage population of the liver. These cells recognize many pathogenic stimuli and exert pro/anti-inflammatory effects, depending on many contributing factors. Liver sinusoidal endothelial cells are a special group of endothelial cells. These cells form a porous sieve plate in the sinusoidal cavity, which plays a screening role in material exchange between plasma and liver cell types, while maintaining a certain barrier function.

AAV Serotypes

Different serotypes have a great influence on the infection tendency of AAV. Choosing the appropriate serotype according to your own experimental needs is an important factor in determining the experimental effect. After the first discovery, research and modification of AAV2, researchers have successively discovered multiple wild serotypes, and based on them, modified the capsid protein Cap gene to develop new serotypes to improve targeting to different tissues. Among them Types 8 and 9 both have a strong tendency towards the liver.

Figure 1. AAV Gene Transfer with Tandem Promoter Design Prevents Anti-transgene Immunity and Provides Persistent Efficacy in Neonate Pompe Mice. (Source: Colella P, et al.; 2018)

AAV8 was a serotype developed to improve the low transfection efficiency of existing serotypes into the liver at that time. The infection efficiency of AAV8 was much higher than that of AAV2. In animal experiments, compared with portal vein injection, which is more difficult to operate, AAV8 can also achieve similar effects through tail vein injection. Comparing the liver infection effect, AAV9 can also achieve effects similar to AAV8, and the infection effect of AAV9 is better than AAV8.

AAV Injection Method

There are many injection methods for AAV-infected liver. Those that require surgical participation include hepatic parenchymal injection, hepatic portal vein injection and intracapsular injection. Non-surgical injection methods include intraperitoneal injection and tail vein injection:

• For liver parenchyma injection, the left outer lobe of the liver of the mouse needs to be surgically exposed before injection. Use a 29G needle insulin syringe to inject the virus into the liver parenchyma. Select 3-5 different sites for injection, with a total volume of 100-200 μL
• For hepatic portal vein injection, you need to use a scalpel to cut the area between the second rib and the fourth nipple of the mouse, with a length of about 3cm. Use a sterile cotton swab to carefully pull out the large and small intestines and other internal organs. Find the hepatic portal vein and use a 32G needle syringe. Inhale 50-100 μL of viral fluid, and slowly insert and inject the viral fluid along the hepatic portal vein at a distance of about 1 cm below the liver at an angle of less than 5°
• For intracapsular injection of the spleen, first use a scalpel to make an incision of about 1cm in length on the left side of the mouse's abdomen, expose the spleen, use a 30G needle syringe to inhale 100 μL of viral fluid, and inject it under the spleen capsule

AAV8 Related Antibodies

AAV8 Related Antigen

AAV8 Related ELISA