article published yes Margaret E Maes author 3838F452-F248-11E8-B48F-1D18A9856A870000-0001-9642-1085 Gabriele M. Wögenstein author Gloria Colombo author 3483CF6C-F248-11E8-B48F-1D18A9856A870000-0001-9434-8902 Raquel Casado Polanco author 15240fc1-dbcd-11ea-9d1d-ac5a786425fd0000-0001-8293-4568 Sandra Siegert author 36ACD32E-F248-11E8-B48F-1D18A9856A870000-0001-8635-0877 SaSi department SiHi department Microglia action towards neuronal circuit formation and function in health and disease project Adeno-associated viruses (AAVs) are widely used to deliver genetic material in vivo to distinct cell types such as neurons or glial cells, allowing for targeted manipulation. Transduction of microglia is mostly excluded from this strategy, likely due to the cells’ heterogeneous state upon environmental changes, which makes AAV design challenging. Here, we established the retina as a model system for microglial AAV validation and optimization. First, we show that AAV2/6 transduced microglia in both synaptic layers, where layer preference corresponds to the intravitreal or subretinal delivery method. Surprisingly, we observed significantly enhanced microglial transduction during photoreceptor degeneration. Thus, we modified the AAV6 capsid to reduce heparin binding by introducing four point mutations (K531E, R576Q, K493S, and K459S), resulting in increased microglial transduction in the outer plexiform layer. Finally, to improve microglial-specific transduction, we validated a Cre-dependent transgene delivery cassette for use in combination with the Cx3cr1CreERT2 mouse line. Together, our results provide a foundation for future studies optimizing AAV-mediated microglia transduction and highlight that environmental conditions influence microglial transduction efficiency. https://research-explorer.ista.ac.at/download/10655/10657/2021_MolTherMethodsClinDev_Maes.pdf application/pdfno Elsevier2021 eng 2329-0501 00074874850001910.1016/j.omtm.2021.09.006 23210-224 Maes ME, Wögenstein GM, Colombo G, Casado Polanco R, Siegert S. Optimizing AAV2/6 microglial targeting identified enhanced efficiency in the photoreceptor degenerative environment. <i>Molecular Therapy - Methods and Clinical Development</i>. 2021;23:210-224. doi:<a href="https://doi.org/10.1016/j.omtm.2021.09.006">10.1016/j.omtm.2021.09.006</a> M.E. Maes, G.M. Wögenstein, G. Colombo, R. Casado Polanco, S. Siegert, Molecular Therapy - Methods and Clinical Development 23 (2021) 210–224. M. E. Maes, G. M. Wögenstein, G. Colombo, R. Casado Polanco, and S. Siegert, “Optimizing AAV2/6 microglial targeting identified enhanced efficiency in the photoreceptor degenerative environment,” <i>Molecular Therapy - Methods and Clinical Development</i>, vol. 23. Elsevier, pp. 210–224, 2021. Maes, Margaret E, Gabriele M. Wögenstein, Gloria Colombo, Raquel Casado Polanco, and Sandra Siegert. “Optimizing AAV2/6 Microglial Targeting Identified Enhanced Efficiency in the Photoreceptor Degenerative Environment.” <i>Molecular Therapy - Methods and Clinical Development</i>. Elsevier, 2021. <a href="https://doi.org/10.1016/j.omtm.2021.09.006">https://doi.org/10.1016/j.omtm.2021.09.006</a>. Maes ME, Wögenstein GM, Colombo G, Casado Polanco R, Siegert S. 2021. Optimizing AAV2/6 microglial targeting identified enhanced efficiency in the photoreceptor degenerative environment. Molecular Therapy - Methods and Clinical Development. 23, 210–224. Maes, Margaret E., et al. “Optimizing AAV2/6 Microglial Targeting Identified Enhanced Efficiency in the Photoreceptor Degenerative Environment.” <i>Molecular Therapy - Methods and Clinical Development</i>, vol. 23, Elsevier, 2021, pp. 210–24, doi:<a href="https://doi.org/10.1016/j.omtm.2021.09.006">10.1016/j.omtm.2021.09.006</a>. Maes, M. E., Wögenstein, G. M., Colombo, G., Casado Polanco, R., &#38; Siegert, S. (2021). Optimizing AAV2/6 microglial targeting identified enhanced efficiency in the photoreceptor degenerative environment. <i>Molecular Therapy - Methods and Clinical Development</i>. Elsevier. <a href="https://doi.org/10.1016/j.omtm.2021.09.006">https://doi.org/10.1016/j.omtm.2021.09.006</a> 106552022-01-23T23:01:28Z2025-04-14T07:41:46Z