Adeno-associated virus (AAV)-mediated gene delivery has been proposed to be an essential tool of gene therapy for various brain diseases. Among several cell types in the brain, astrocyte has become a promising therapeutic target for brain diseases, as more and more contribution of astrocytes in pathophysiology has been revealed. Until now, genetically targeting astrocytes has been possible by utilizing the glial fibrillary acidic protein (GFAP) promoter. In some brain areas including thalamus, however, the GFAP expression in astrocytes is reported to be low, making it difficult to genetically target astrocytes using GFAP promoter. To study the function of astrocytes in thalamus, which serves as a relay station, there is a great need for identifying an alternative astrocyte-specific promoter in thalamus. Recently, a new astrocyte-specific promoter of ALDH1L1 has been identified. However, it has not been examined in thalamus. Here we developed and characterized an AAV vector expressing Cre recombinase under the human ALDH1L1 promoter, AAV-hALDH1L1-Cre. To test the cell-type specific expression of AAV-hALDH1L1-Cre, AAV virus was injected into several brain regions of Ai14 (RCL-tdTomato) mouse, which reports Cre activity by tdTomato expression. In thalamus, we observed that tdTomato was found mostly in astrocytes (91.71%), with minimal occurrence in neurons (2.67%). In contrast, tdTomato signal was observed in both neurons and astrocytes of the amygdala (neuron: 68.13%, astrocyte: 28.35%) and hippocampus (neuron: 76.25%, astrocyte: 18.00%), which is consistent with the previous report showing neuronal gene expression under rat ALDH1L1 promoter. Unexpectedly, tdTomato was found mostly in neurons (91.98%) with minimal occurrence in astrocytes (6.66%) of the medial prefrontal cortex. In conclusion, hALDH1L1 promoter shows astrocyte-specificity in thalamus and may prove to be useful for targeting thalamic astrocytes in mouse.

中文翻译：

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小鼠丘脑中人ALDH1L1启动子下AAV介导的星形胶质细胞特异性基因表达。

腺相关病毒（AAV）介导的基因传递已被提议为各种脑疾病的基因治疗的重要工具。在大脑中的几种细胞类型中，星形胶质细胞已成为脑疾病的有希望的治疗靶标，因为越来越多的星形胶质细胞在病理生理中的作用已被发现。到目前为止，通过利用神经胶质原纤维酸性蛋白（GFAP）启动子，遗传靶向星形胶质细胞成为可能。然而，据报道，在包括丘脑在内的某些大脑区域，星形胶质细胞中GFAP的表达很低，因此难以使用GFAP遗传靶向星形胶质细胞启动子。为了研究充当中继站的丘脑中星形胶质细胞的功能，非常需要鉴定丘脑中另一种星形胶质细胞特异性启动子。最近，已经鉴定了ALDH1L1的新的星形胶质细胞特异性启动子。但是，尚未在丘脑中对其进行检查。在这里，我们开发并表征了在人ALDH1L1下表达Cre重组酶的AAV载体启动子，AAV-hALDH1L1-Cre。为了测试AAV-hALDH1L1-Cre的细胞类型特异性表达，将AAV病毒注射到Ai14（RCL-tdTomato）小鼠的几个大脑区域，该小鼠通过tdTomato表达报告Cre活性。在丘脑中，我们观察到tdTomato主要在星形胶质细胞中发现（占91.71％），在神经元中的发生率最低（占2.67％）。相反，在杏仁核（神经元：68.13％，星形胶质细胞：28.35％）和海马（神经元：76.25％，星形胶质细胞：18.00％）的神经元和星形胶质细胞中均观察到tdTomato信号，这与先前报道的神经元基因的报道一致在大鼠ALDH1L1启动子下的表达。出乎意料的是，tdTomato主要在神经元（91.98％）中发现，而在额前内侧皮层的星形胶质细胞（6.66％）中发生的可能性最小。总之，h ALDH1L1 启动子在丘脑中显示星形胶质细胞特异性，并且可能被证明可用于靶向小鼠丘脑星形胶质细胞。