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Production, Purification, and Quality Control for Adeno-associated Virus-based Vectors
Published on: January 29, 2019
Optimal different adeno-associated virus capsid/promoter combinations to target specific cell types in the common
Yasunori Matsuzaki1,2, Yuuki Fukai1, Ayumu Konno1,2
1Department of Neurophysiology & Neural Repair, Gunma University Graduate School of Medicine, Maebashi, Gunma 371-8511, Japan.
Abstract:
To achieve cell-type-specific gene expression, using target cell-type-tropic different adeno-associated virus (AAV) capsids is advantageous. However, their tropism across brain cell types in nonhuman primates has not been fully elucidated. We assessed the tropism of nine AAV serotype capsids (AAV1, 2, 5, 6, 7, 8, 9, rh10, and DJ) expressing EGFP by chicken β-actin hybrid (CBh) promoter in marmoset cerebral cortical cells. All nine AAV capsid vectors, especially AAV9 and AAVrh10, caused highly neuron-selective EGFP expression. Some AAV capsids, including AAV5, induced EGFP expression to a lesser extent in oligodendrocytes. Different ubiquitous cytomegalovirus (CMV) and CMV early enhancer/chicken β-actin (CAG) promoters exhibited similar neuron-predominant transgene expression. Conversely, all nine AAV capsid vectors with the astrocyte-specific hGFA(ABC1D) promoter selectively expressed EGFP in astrocytes, except AAV5, which modestly expressed EGFP in oligodendrocytes. Oligodendrocyte-specific mouse myelin basic protein (mMBP) promoter in AAV5 vectors expressed EGFP in oligodendrocytes specifically and efficiently. The following are optimal combinations of capsids and promoters for cell-type-specific expression: AAV9 or AAVrh10 and ubiquitous CBh or CMV promoter for neuron-specific transgene expression, AAV2 or AAV7 and hGFA(ABC1D) promoters for astrocyte-specific transgene expression, and AAV5 and mMBP promoters for oligodendrocyte-specific transgene expression.
Insights
This study identifies optimal adeno-associated virus (AAV) capsid and promoter combinations for cell-type-specific gene expression in marmoset brains. Researchers found specific AAVs for targeting neurons, astrocytes, and oligodendrocytes, crucial for neuroscience research.
Area of Science:
- Neuroscience
- Molecular Biology
- Gene Therapy
Background:
- Adeno-associated virus (AAV) vectors are crucial for gene delivery in neuroscience research.
- Understanding AAV tropism in nonhuman primate brains is essential for targeted gene expression.
- Previous studies have limited data on AAV tropism across diverse brain cell types in primates.
Purpose of the Study:
- To elucidate the tropism of nine adeno-associated virus (AAV) serotype capsids in marmoset cerebral cortical cells.
- To identify optimal AAV capsid and promoter combinations for cell-type-specific gene expression in the primate brain.
- To guide the development of targeted gene therapies and neuroscience research tools.
Main Methods:
- In vivo assessment of nine AAV serotype capsids (AAV1, 2, 5, 6, 7, 8, 9, rh10, DJ) expressing EGFP in marmoset cerebral cortex.
- Utilized ubiquitous promoters (CBh, CMV, CAG) and cell-type-specific promoters (hGFA, mMBP).
- Analyzed EGFP expression patterns in neurons, astrocytes, and oligodendrocytes.
Main Results:
- AAV9 and AAVrh10 capsids demonstrated high neuron-selective EGFP expression with ubiquitous promoters.
- AAV2 and AAV7 capsids combined with the hGFA promoter showed selective EGFP expression in astrocytes.
- AAV5 capsids with the mMBP promoter efficiently targeted oligodendrocytes for EGFP expression.
Conclusions:
- Established optimal AAV capsid-promoter pairings for neuron-, astrocyte-, and oligodendrocyte-specific gene expression in marmosets.
- AAV9/rh10 with CBh/CMV for neurons, AAV2/7 with hGFA for astrocytes, and AAV5 with mMBP for oligodendrocytes are recommended.
- Findings provide critical insights for advancing targeted gene therapy and functional studies in the primate brain.
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