Ultramicroscopy as a novel tool to unravel the tropism of AAV gene therapy vectors in the brain.
Recombinant adeno-associated viral (AAV) vectors have advanced to the vanguard of gene therapy. Numerous naturally occurring serotypes have been used to target cells in various tissues. There is a strong need for fast and dynamic methods which efficiently unravel viral tropism in whole organs. Ultramicroscopy (UM) is a novel fluorescence microscopy technique that images optically cleared undissected specimens, achieving good resolutions at high penetration depths while being non-destructive. UM was applied to obtain high-resolution 3D analysis of AAV transduction in adult mouse brains, especially in the hippocampus, a region of interest for Alzheimer's disease therapy. We separately or simultaneously compared transduction efficacies for commonly used serotypes (AAV9 and AAVrh10) using fluorescent reporter expression. We provide a detailed comparative and quantitative analysis of the transduction profiles. UM allowed a rapid analysis of marker fluorescence expression in neurons with intact projections deep inside the brain, in defined anatomical structures. Major hippocampal neuronal transduction was observed with both vectors, with slightly better efficacy for AAV9 in UM. Glial response and synaptic marker expression did not change post transduction.We propose UM as a novel valuable complementary tool to efficiently and simultaneously unravel tropism of different viruses in a single non-dissected adult rodent brain.
INSERM U1169/MIRCen CEA, Fontenay aux Roses 92265, France, Université Paris-Sud, Université Paris-Saclay, Orsay 91400, France.
Sandro
S
Alves
Schaller Research Group at the University of Heidelberg and the German Cancer Research Center (DKFZ), Im Neuenheimer Feld 581, 69120 Heidelberg, Germany.|Molecular Mechanisms of Tumor Invasion (V077), DKFZ, Im Neuenheimer Feld 581, 69120 Heidelberg, Germany.
Julia
J
Bode
Commissariat à l´Energie Atomique et aux Energies Alternatives (CEA), Départment de la Recherche Fondamentale (DRF), Institut d´Imagerie Biomédicale (I2BM), Molecular Imaging Research Center (MIRCen), Fontenay-aux-Roses, France.|Centre National de la Recherche Scientifique (CNRS), Université Paris-Sud, Université Paris-Saclay, UMR 9199, Neurodegenerative Diseases Laboratory, Fontenay-aux Roses, France.
Alexis-Pierre
AP
Bemelmans
Department of Translational Oncology, National Center for Tumor Diseases (NCT) and German Cancer Research Center (DKFZ), Im Neuenheimer Feld 280, 69120 Heidelberg, Germany.
Christof
C
von Kalle
INSERM U1169/MIRCen CEA, Fontenay aux Roses 92265, France, Université Paris-Sud, Université Paris-Saclay, Orsay 91400, France.
Nathalie
N
Cartier
Schaller Research Group at the University of Heidelberg and the German Cancer Research Center (DKFZ), Im Neuenheimer Feld 581, 69120 Heidelberg, Germany.|Molecular Mechanisms of Tumor Invasion (V077), DKFZ, Im Neuenheimer Feld 581, 69120 Heidelberg, Germany.
Björn
B
Tews
D000818:Animals; D001921:Brain / Q000378:metabolism / Q000473:pathology / Q000821:virology*; D000229:Dependovirus / Q000502:physiology*; D015316:Genetic Therapy; D008297:Male; D051379:Mice; D008856:Microscopy, Fluorescence; D009474:Neurons / Q000378:metabolism / Q000473:pathology / Q000821:virology*; D014161:Transduction, Genetic; D056189:Viral Tropism