Δ8-THC Protects against Amyloid Beta Toxicity Modulating ER Stress In Vitro: A Transcriptomic Analysis

Abstract

Alzheimer's disease (AD) represents the most common form of dementia, characterized by amyloid β (Aβ) plaques and neurofibrillary tangles (NFTs). It is characterized by neuroinflammation, the accumulation of misfolded protein, ER stress and neuronal apoptosis. It is of main importance to find new therapeutic strategies because AD prevalence is increasing worldwide. Cannabinoids are arising as promising neuroprotective phytocompounds. In this study, we evaluated the neuroprotective potential of Δ⁸-THC pretreatment in an in vitro model of AD through transcriptomic analysis. We found that Δ⁸-THC pretreatment restored the loss of cell viability in retinoic acid-differentiated neuroblastoma SH-SY5Y cells treated with Aβ_1-42. Moreover, the transcriptomic analysis provided evidence that the enriched biological processes of gene ontology were related to ER functions and proteostasis. In particular, Aβ_1-42 upregulated genes involved in ER stress and unfolded protein response, leading to apoptosis as demonstrated by the increase in Bax and the decrease in Bcl-2 both at gene and protein expression levels. Moreover, genes involved in protein folding and degradation were also deregulated. On the contrary, Δ⁸-THC pretreatment reduced ER stress and, as a consequence, neuronal apoptosis. Then, the results demonstrated that Δ⁸-THC might represent a new neuroprotective agent in AD.

Keywords: Alzheimer’s disease; neuronal apoptosis; unfolded protein response; Δ8-THC.

Figure 1

Cell viability after Aβ_1-42 and Δ⁸-THC treatment. The treatment with 10 µM Aβ_1-42 reduced cell viability of RA-differentiated SH-SY5Y cells, but Δ⁸-THC pretreatment at the concentrations 10 and 20 µM was able to restore cell viability. N = 4 independent experiments. The results are expressed by mean ± standard deviation (SD). ** p < 0.01; **** p < 0.0001.

Figure 2

DEGs distribution between CTRL vs. Aβ_1-42, CTRL vs. Δ⁸-THC groups or Aβ_1-42 vs. Δ⁸-THC + Aβ_1-42. Venn diagram in the center of the plot highlights the amount of DEGs found exclusively in each comparison (outer circles), how many DEGs found in two groups but not in the other (intersection of two circles) or how many DEGs were found in each comparison (center of the diagram). Each donut plot highlights, in turn, the intersection of two comparisons showing in dark red the upregulated and in the light red the downregulated DEGs.

Figure 3

Bubbleplot of biological process terms enriched in gene ontology between CTRL vs. Aβ_1-42 and Aβ_1-42 vs. Δ⁸-THC + Aβ_1-42 groups. For each ontology reported on the y axis, a bubble for the CTRL vs. Aβ_1-42 (orange) and one for the Aβ_1-42 vs. Δ⁸-THC + Aβ_1-42 (light blue) groups were plotted. The position of the bubble in the x axis shows the number of DEGs in the ontology (the more on the right, the higher the number of DEGs). The number of DEGs was normalized over the number of genes included in the ontology term itself so that the terms were comparable to each other. The size of the bubble is a score given by −log(q-value).

Figure 4

Western blot for Bax and Bcl-2. Aβ_1-42 treatment caused an increase in Bax and a reduction in Bcl-2 protein levels. Δ⁸-THC treatment restored protein levels of Bax and Bcl-2. N = 3 independent experiments. The results are expressed by mean ± standard deviation (SD). * p < 0.05; *** p < 0.001.

Figure 5

Proteins encoded by DEGs modulated in Aβ_1-42 and Δ⁸-THC treated cells in ER pathway. The name of the shown proteins was obtained by KEGG. The figure was drawn using the vector image bank of Servier Medical Art by Servier (http://smart.servier.com/, accessed on 10 February 2023). Licensed under a Creative Commons Attribution 3.0 Unported License (https://creativecommons.org/licenses/by/3.0/, accessed on 10 February 2023).

Figure 6

Δ⁸-THC chemical structure.