102. AAV9 Exclusively Targets the Juxtaglomerular Apparatus in the Adult Mouse Kidney After Intravenous Injection

Abstract

Aim: Gene delivery to the kidney by adeno-associated virus (AAV) remains a relatively unexplored field. We found that the cells in renal tissue infected by AAV9 vectors were very few but interestingly distributed in some specific regions adjacent to the glomerulus. Therefore we aim to identify the type of the cells infected by the AAV9 vectors, and compare it with other AAVs for the specific targeting by different routes of vector injection.Material and Methods: We injected AAV9-CMV-GFP, AAVH43-CMV-GFP and AAVH48-CMV-GFP (5×1011 vg/mouse) into the adult C57BL/6 mice (n=4 /group) via the tail vein. For the renal parenchyma injection (n=3/group) the vectors (10μl, 1 × 1011vg) were slowly injected into the renal parenchyma by a 33-gauge needle, which was connected to a 50μl Hamilton syringe with PE10 tubing. Two weeks later, kidneys were frozen and sectioned to slices of 10 μm.Results: We found that the AAV9 vector specifically infected juxtaglomerular apparatus in the kidney by intravenous injection. The infection efficiency reached up to 58.2±3.2% of total juxtaglomerular apparatus. On the other hand, The AAVH43 and AAVH48 vectors were negative in the kidney yet achieved GFP expression similarly to AAV9 in the heart and liver. Immunostaining showed the cells infected by AAV9-CMV-GFP vectors were exclusively adjacent to juxtaglomerular (JG) cells. These JG cells synthesize, store and secrete renin, and were identified by anti-renin antibody. Neuronal nitric oxide synthase (nNOS) and prostaglandin E receptor 3 (EP3) immunostaining further indicated that GFP-positive cells were peri-macular cells, including the cells opposite the macula densa plaque. These were epithelial cells around the macula densa and one of four contact sites between the distal nephron and vasculature. The peri-macular cells, not including the macula densa, can generate spontaneous [Ca2+ ]i oscillations, and signal to the connected arterioles, regulating renal blood flow and tubular flow. Interestingly, specific infection in the juxtaglomerular apparatus by AAV9 vectors was dependent on the route of vector injection. When injected directly into the renal parenchyma, the peri-macular cells could no longer be infected, whereas the AAVH43 and AAVH48 vectors infected renal tubular epithelial cells.Conclusion: We have identified for the first time that a peri-macular cell type of the kidney is selectively targeted by intravenous AAV9 injection. Considering the importance role of those cells in regulation of renal blood flow and tubular flow, specific targeting by AAV9 may present a novel therapeutic method for renal hypertension and renal dysfunction. On the other hand, the data provide novel proof that the peri-macular cells are a group of functional epithelial cells, which differ from other renal tubular cells and may highly express AAV9 receptors. Additionally, the specificity of AAV9 infection in peri-macular cells was via the blood vessel, suggesting unique communication routes between peri-macular cells and the arterioles.