Toro Carlos A, Johnson Kaitlin, Hansen Jens, Siddiq Mustafa M, Vásquez Walter, Zhao Wei, Graham Zachary A, Sáez Juan C, Iyengar Ravi, Cardozo Christopher P
Spinal Cord Damage Research Center, James J. Peters VA Medical Center, Bronx, NY, United States.
Department of Medicine, Icahn School of Medicine at Mount Sinai, New York, NY, United States.
Front Cell Neurosci. 2023 Jun 21;17:1163436. doi: 10.3389/fncel.2023.1163436. eCollection 2023.
Membrane channels such as those formed by connexins (Cx) and P2X receptors (P2XR) are permeable to calcium ions and other small molecules such as adenosine triphosphate (ATP) and glutamate. Release of ATP and glutamate through these channels is a key mechanism driving tissue response to traumas such as spinal cord injury (SCI). Boldine, an alkaloid isolated from the Chilean boldo tree, blocks both Cx and Panx1 hemichannels (HCs). To test if boldine could improve function after SCI, boldine or vehicle was administered to treat mice with a moderate severity contusion-induced SCI. Boldine led to greater spared white matter and increased locomotor function as determined by the Basso Mouse Scale and horizontal ladder rung walk tests. Boldine treatment reduced immunostaining for markers of activated microglia (Iba1) and astrocytic (GFAP) markers while increasing that for axon growth and neuroplasticity (GAP-43). Cell culture studies demonstrated that boldine blocked glial HC, specifically Cx26 and Cx30, in cultured astrocytes and blocked calcium entry through activated P2XR. RT-qPCR studies showed that boldine treatment reduced expression of the chemokine Ccl2, cytokine IL-6 and microglial gene CD68, while increasing expression of the neurotransmission genes Snap25 and Grin2b, and Gap-43. Bulk RNA sequencing revealed that boldine modulated a large number of genes involved in neurotransmission in spinal cord tissue just caudal from the lesion epicenter at 14 days after SCI. Numbers of genes regulated by boldine was much lower at 28 days after injury. These results indicate that boldine treatment ameliorates injury and spares tissue to increase locomotor function.
膜通道，如由连接蛋白（Cx）和P2X受体（P2XR）形成的通道，对钙离子以及其他小分子如三磷酸腺苷（ATP）和谷氨酸具有通透性。通过这些通道释放ATP和谷氨酸是驱动组织对诸如脊髓损伤（SCI）等创伤作出反应的关键机制。波尔定碱是一种从智利波尔多树中分离出的生物碱，可阻断Cx和Panx1半通道（HCs）。为了测试波尔定碱是否能改善脊髓损伤后的功能，将波尔定碱或赋形剂给予中度严重挫伤诱导的脊髓损伤小鼠进行治疗。根据巴索小鼠量表和水平梯级行走测试，波尔定碱可使脊髓白质保留更多，并增强运动功能。波尔定碱治疗减少了活化小胶质细胞标志物（Iba1）和星形胶质细胞标志物（GFAP）的免疫染色，同时增加了轴突生长和神经可塑性标志物（GAP-43）的免疫染色。细胞培养研究表明，波尔定碱可阻断培养的星形胶质细胞中的胶质半通道，特别是Cx26和Cx30，并阻断通过活化的P2XR的钙内流。RT-qPCR研究表明，波尔定碱治疗降低了趋化因子Ccl2、细胞因子IL-6和小胶质细胞基因CD68的表达，同时增加了神经传递基因Snap25、Grin2b和Gap-43的表达。批量RNA测序显示，在脊髓损伤后14天，波尔定碱调节了损伤震中尾侧脊髓组织中大量参与神经传递的基因。损伤后28天，受波尔定碱调节的基因数量要少得多。这些结果表明，波尔定碱治疗可减轻损伤并保留组织以增强运动功能。
