Departamento de Bioquímica y Biología Molecular, Instituto Universitario de Investigación en Neuroquímica, Facultad de Medicina, Universidad Complutense, Madrid 28040, Spain.
Centro de Investigación Biomédica en Red de Enfermedades Neurodegenerativas (CIBERNED), Madrid 28040, Spain.
Dis Model Mech. 2017 May 1;10(5):551-558. doi: 10.1242/dmm.028373. Epub 2017 Jan 9.
Targeting of the CB receptor results in neuroprotection in the SOD1 mutant mouse model of amyotrophic lateral sclerosis (ALS). The neuroprotective effects of CB receptors are facilitated by their upregulation in the spinal cord of the mutant mice. Here, we investigated whether similar CB receptor upregulation, as well as parallel changes in other endocannabinoid elements, is evident in the spinal cord of dogs with degenerative myelopathy (DM), caused by mutations in the superoxide dismutase 1 gene (). We used well-characterized post-mortem spinal cords from unaffected and DM-affected dogs. Tissues were used first to confirm the loss of motor neurons using Nissl staining, which was accompanied by glial reactivity (elevated GFAP and Iba-1 immunoreactivity). Next, we investigated possible differences in the expression of endocannabinoid genes measured by qPCR between DM-affected and control dogs. We found no changes in expression of the CB receptor (confirmed with CB receptor immunostaining) or NAPE-PLD, DAGL, FAAH and MAGL enzymes. In contrast, CB receptor levels were significantly elevated in DM-affected dogs determined by qPCR and western blotting, which was confirmed in the grey matter using CB receptor immunostaining. Using double-labelling immunofluorescence, CB receptor immunolabelling colocalized with GFAP but not Iba-1, indicating upregulation of CB receptors on astrocytes in DM-affected dogs. Our results demonstrate a marked upregulation of CB receptors in the spinal cord in canine DM, which is concentrated in activated astrocytes. Such receptors could be used as a potential target to enhance the neuroprotective effects exerted by these glial cells.
靶向 CB 受体可导致肌萎缩侧索硬化症 (ALS) 的 SOD1 突变小鼠模型中的神经保护。CB 受体的神经保护作用是通过其在突变小鼠脊髓中的上调来促进的。在这里，我们研究了在由超氧化物歧化酶 1 基因 (SOD1) 突变引起的退行性脊髓病 (DM) 的犬的脊髓中是否存在类似的 CB 受体上调以及其他内源性大麻素成分的平行变化。我们使用了经过充分表征的未受影响和受 DM 影响的犬的死后脊髓。首先使用尼氏染色来确认运动神经元的丧失，这伴随着神经胶质反应（升高的 GFAP 和 Iba-1 免疫反应性）。接下来，我们通过 qPCR 研究了 DM 影响和对照犬之间内源性大麻素基因表达的可能差异。我们没有发现 CB 受体（通过 CB 受体免疫染色证实）或 NAPE-PLD、DAGL、FAAH 和 MAGL 酶的表达变化。相比之下，通过 qPCR 和 Western blot 确定 CB 受体在 DM 影响的犬中显著升高，并用 CB 受体免疫染色在灰质中得到证实。使用双标记免疫荧光，CB 受体免疫标记与 GFAP 共定位，但不与 Iba-1 共定位，表明在 DM 影响的犬的星形胶质细胞中 CB 受体上调。我们的结果表明，犬 DM 脊髓中 CB 受体明显上调，主要集中在激活的星形胶质细胞中。这种受体可以用作增强这些神经胶质细胞发挥神经保护作用的潜在靶标。
