CAG-GFP AAV (Serotype BR1)

Adeno-associated viruses (AAV) are a common tool in gene therapy approaches and have been engineered to specifically target different cells. There is interest in targeting endothelial cells (ECs) at the blood-brain barrier, and the AAV2 capsid variant BR1 has been found to transduce ECs with high selectivity in various mouse models. However, this has not been tested in rat models. Here, researchers show that systemic injection of the AAV-BR1-CAG-GFP virus in Sprague-Dawley rats does not transduce ECs, but rather transduces brain parenchymal cells with neuronal morphology. These findings highlight the importance of species differences in the use of AAV.

The researchers injected AAV-BR1-CAG-GFP into three three-week-old Sprague-Dawley rats via tail injection. Due to differences in size and metabolism between mice and rats, they decided to inject three different doses: one dose that was the same as the usual dose for mice (1.5 x 10¹¹ vg/rat), one dose that was twice the usual dose for mice (3 x 10¹¹ vg/rat), and one dose that was three times the original dose for mice (4.5 x 10¹¹ vg/rat). The AAV stock was diluted in sterile PBS and then injected into the tail vein under anesthesia. Three weeks after the injection, the brains were perfused, harvested, and immunostained for isolectin B4 (a blood vessel marker), ERG (an endothelial nuclear marker), and GFP (the fluorescence the AAV-BR1 was tagged with). In Sprague-Dawley rats, unlike mice, GFP expression by AAV-BR1 was not seen in the ECs of any cerebral blood vessels across any brain regions observed. In contrast, brain parenchymal cells with neuronal morphology had high levels of GFP expression, and GFP expression was higher in rats that received the higher dose.

Figure 1. AAV-BR1-CAG-GFP fails to transduce endothelial cells in Sprague-Dawley rats. (Kremer R, Williams A., 2024)