Abstract
Simple SummaryAims and objectives: For this study, we hypothesized that the two TSPO ligands CB86 and CB204 can inhibit cellular apoptosis and necrosis by in in vitro cellular PD model of undifferentiated PC12 cells exposed to 6-hydroxydopamine (6-OHDA, 80 µM). The two TSPO ligands CB86 and CB204 seem to suppress cell death of PC12 induced by 6-OHDA. The results may be relevant to the use of these two TSPO ligands as therapeutic options for neurodegenerative diseases like Parkinson disease (PD). Results: The two ligands normalized significantly (57% and 52%, respectively, from 44%; whereas the control was 68%) cell proliferation at different time points from 0–24 h. As compared to control, the red count was increased up to 57-fold whereas CB86 and CB204 inhibited to 2.7-fold and 3.2-fold, respectively. CB86 and CB204 inhibited also normalized the cell viability up to 1.8-fold after the exposure to 6-OHDA, as assessed by XTT assay. The two TSPO ligands also inhibited apoptosis significantly (1.3-fold for both) as assessed by apopxin green staining. Conclusion: It appears that CB86, CB204, and maybe other TSPO ligands are able to slow the progression of neurodegenerative diseases like PD.Parkinson’s disease (PD) is a progressive neurodegenerative disorder which is characterized by the degeneration of dopaminergic neurons in substantia nigra (SN). Oxidative stress or reactive oxygen species (ROS) generation was suggested to play a role in this specific type of neurodegeneration. Therapeutic options which can target and counteract ROS generation may be of benefit. TSPO ligands are known to counteract with neuro-inflammation, ROS generation, apoptosis, and necrosis. In the current study, we investigated an in vitro cellular PD model by the assessment of 6-hydroxydopamine (6-OHDA, 80 µM)-induced PC12 neurotoxicity. Simultaneously to the exposure of the cells to 6-OHDA, we added the TSPO ligands CB86 and CB204 (25 µM each) and assessed the impact on several markers of cell death. The two ligands normalized significantly (57% and 52% respectively, from 44%; whereas the control was 68%) cell proliferation at different time points from 0–24 h. Additionally, we evaluated the effect of these two TSPO ligands on necrosis using propidium iodide (PI) staining and found that the ligands inhibited significantly the 6-OHDA-induced necrosis. As compared to control, the red count was increased up to 57-fold whereas CB86 and CB204 inhibited to 2.7-fold and 3.2-fold respectively. Necrosis was also analyzed by LDH assay which showed significant effect. Both assays demonstrated similar potent anti-necrotic effect of the two TSPO ligands. Reactive oxygen species (ROS) generation induced by 6-OHDA was also inhibited by the two TSPO ligand up to 1.3 and 1.5-fold respectively, as compared to 6-OHDA group. CB86 and CB204 inhibited also normalized the cell viability up to 1.8-fold after the exposure to 6-OHDA, as assessed by XTT assay. The two TSPO ligands also inhibited apoptosis significantly (1.3-fold for both) as assessed by apopxin green staining. In summary, it appears that the two TSPO ligands CB86 and CB204 can suppress cell death of PC12 induced by 6-OHDA. The results may be relevant to the use of these two TSPO ligands as therapeutic option neurodegenerative diseases like PD.
Highlights
Parkinson’s Disease (PD) is one of the most common progressive neurodegenerative disorders, movement disorder associated most often with advanced age and is related to dopaminergic neurons loss in substantia nigra (SN) [1,2]
2.2. 6-Hydroxidopamine Exposure and translocator protein (TSPO) Ligands Treatment 2.5 × 104 undifferentiated PC12 cells were seeded in 24-well plate and incubated for 48 h until the confluency of 50–60% was reached and subsequently exposed to 6-OHDA (80 μM) as well as treated simultaneously with the TSPO ligands CB86 and CB204 (25 μM each) for 24 h
PC12 cells were seeded in 24-well plate and exposed to 6-OHDA (80 μM) with or without CB86 or CB204 (25 μM each) for 24 h and stained with propidium iodide (PI)
Summary
Parkinson’s Disease (PD) is one of the most common progressive neurodegenerative disorders, movement disorder associated most often with advanced age and is related to dopaminergic neurons loss in substantia nigra (SN) [1,2]. Based on a previous study, PC12 cells were used as an in vitro model of PD, by exposing them to the neurotoxic agent 6-OHDA [4]. In the present study, based on the neuroprotective effect of TSPO ligands [5], we investigated the such an effect in PC12 cellular model of PD. Several factors as oxidative stress, inflammation, toxicity of reactive oxygen species (ROS) products are thought to be involved in the cell death in SN [6,7]. Both neuroinflammation-related apoptotic and necrotic neuronal cell death were reported in various neurodegenerative disorders [6,8]
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