Abstract
Objectives: To investigate the fundamental mechanisms of the neuroprotective impact of Astaxanthin (AST) in a mouse model of Alzheimer’s disease (AD) induced by scopolamine.
Methods: This research constituted an in vivo animal study encompassing 36 adult male mice, divided into 6 groups: Control, 100 mg/kg AST, 2 mg/kg scopolamine (AD group), 100 mg/kg AST+2 mg/kg scopolamine, 3 mg/kg galantamine+2 mg/kg scopolamine, and 100 mg/kg AST+3 mg/kg galantamine+2 mg/kg scopolamine. After 14 days, the mice’s short-term memory, hippocampus tissue, oxidative and inflammatory markers were evaluated.
Results: The AST demonstrated a beneficial influence on short-term memory and a reduction in acetylcholinesterase activity in the brain. It exhibited neuroprotective and anti-amyloidogenic properties, significantly decreased pro-inflammatory markers and oxidative stress, and reversed the decline of the Akt-1 and phosphorylated Akt pathway, a crucial regulator of abnormal tau. Furthermore, AST enhanced the effect of galantamine in reducing inflammation and oxidative stress.
Conclusion: The findings indicate that AST may offer therapeutic benefits against cognitive dysfunction in AD. This is attributed to its ability to reduce oxidative stress, control neuroinflammation, and enhance Akt-1 and pAkt levels, thereby underscoring its potential in AD treatment strategies.
Dementia is a clinical condition characterized by persistent deterioration in two or more cognitive aspects, including memory, vocabulary, visual/ spatial activity, personality, and actions, resulting in a loss of ability to perform everyday life’s instrumental and/or fundamental activities. The most common cause of dementia is Alzheimer’s disease (AD).1 AD is a condition that progressively deteriorates the neurons, leading to their degeneration. It is well known for two definitive features: the build-up of β-amyloid plaques outside the cells and the excessive phosphorylation of tau proteins inside the cells.2 The AD is recognized as one of the most worldwide neurodegenerative disorders, and it is recorded as a cause of more than eighty percent of dementia cases in elderly patients. It is estimated that by 2050, every half a minute, one case of AD will develop, or approximately 1,000,000 newly diagnosed patients yearly.1
The AD pathophysiology includes different molecular pathways of oxidative stress mechanisms, inflammatory mediators, and neuronal destructions.3 For this reason, compounds with antioxidant and anti-inflammatory features have been of specific interest as prophylactic and therapeutic strategies in neurodegenerative disorders.
While there are several drugs approved by the US Food and Drug Administration for treating AD, the most commonly used medications to address the cognitive impairments experienced by AD patients include three acetylcholinesterase (AchE) inhibitors (galantamine, rivastigmine, donepezil) and one N-Methyl-D-Aspartate (NMDA) receptor antagonist (memantine).2 All these medications work by delaying the aggressive deterioration of symptomatic cognitive impairment, but at least 50% of AD patients who are on these medications do not respond to them. Therefore, the development of new disease-modifying therapeutic strategies to inhibit the progression of AD is an important goal in the management of this devastating disease.
Astaxanthin (AST) is considered one of the most effective natural carotenoid compounds, with recent studies showing its ability to inhibit oxidative stress and inflammation and protect against chronic neurodegenerative disorders. Furthermore, AST’s unique molecular structure allows it to penetrate the blood-brain barrier, making it highly effective in preventing human neurodegenerative disorders.3 In this way, the brain is one of the most significant target organs of AST. Thus, this study was conducted to investigate the potential neuroprotective effect of AST and its mechanism of action on scopolamine-induced Alzheimer model in mice.
Methods
Animals
The experimental procedure was conducted at the King Abdulaziz University’s College of Pharmacy animal facility in Saudi Arabia from February through June 2021. 36 adult male mice of the C57BL6/J strain (6 to 8 weeks old, weighing 20-30g) were used. The animals were housed for one week prior to the experimental research and were divided into groups. During their stay, they were provided with regular pellets and unlimited access to water. The mice were maintained at a temperature of 22±3°C, on a 12-hour light/dark cycle, and with a constant relative humidity. The experiment was conducted in accordance with the international guidelines for the treatment and use of laboratory animals and was approved by the biomedical ethics research committee at King Abdulaziz University’s Faculty of Medicine.
Chemicals
Scopolamine hydrobromide, AST and galantamine were purchased from (Medchem, CA, USA) in powder form. Scopolamine hydrobromide powder was dissolved in normal saline to reach a final concentration of 0.2 mg/ml (purity: 99.3%). An injection of 5mg/ml AST was prepared by dissolving the powder in 5% Dimethyl Sulfoxide (DMSO) (purity≥ 98%). An oral solution of galantamine 0.3mg/ml was prepared by mixing the powder with normal saline (purity:99.32%). Reagent enzyme-linked immunosorbent assay (ELISA) kits for tumor necrosis factor-alpha (TNF-α; Catalog #: K1051), AchE (Catalog #: E4453), Malondialdehyde (MDA; Catalog #: E4601), superoxide dismutase (SOD; Catalog #: E4583) and nitric oxide (NO; Catalog #: ab65328) were purchased from (BioVision, CA, USA). Interleukin-6 (IL-6, Catalog #: M6000B), Akt1 (Catalog #: DYC887B-2), phosphorylated Akt antirat antibodies (pAkt, Catalog #: NBP1-69923) (Novus Biologicals, USA).
Study design and treatment groups
In this in vivo study, thirty-six adult male C57BL6/J mice (8 weeks old, 20–30 g) were divided into 6 groups of 6 mice each. Group 1 (control group) received saline (10 ml/kg, i.p.) for 14 successive days. Group 2 (control + AST) was treated with AST (100 mg/kg, i.p) for 14 days. Group 3 (AD group) received an injected dose of scopolamine hydrobromide (2 mg/kg, i.p.) on days 4-14 for 10 days. Group 4 (AD/AST) was administered AST (100 mg/kg, i.p.) on day 1-14 and scopolamine (2 mg/kg, i.p.) on days 4-14 for 10 days. Group 5 (AD/galantamine) received oral galantamine (3 mg/kg, orally) on days 1-14 and scopolamine (2 mg/kg, i.p.) on days 4-14 for 10 days. Group 6 (AD/ AST /galantamine) was treated with AST (100 mg/kg, i.p.) and galantamine (3 mg/kg, orally) on days 1-14 and scopolamine hydrobromide (2 mg/kg, i.p.) on days (4-14) for 10 days. The dose of AST was chosen based on a previous study that examines the neuroprotective effect of AST on the brain traumatic mice model.4
Short-memory test
Y-maze spontaneous alternation test: The Y-maze test was used to assess short-term memory in the mice.5 The maze consisted of three identical arms (labelled A, B and C), each 40 cm long, 35 cm high, and 12 cm wide at equal angles. The mice were allowed to navigate the maze for multiple 5-minute sessions. The test measured the mouse’s ability to enter in a different arm each time, which is called an alteration choice. Entering the same arm twice was considered an error. An arm entry was considered to be complete when the mouse’s hind paws were entirely placed in the arm. Alternation was described as consecutive entries into the three arms of a triplet set overlapped (i.e., ABC, BCA….). The spontaneous alternation percentage (SAP%), the total arm entries (TAE) and the spontaneous alternation efficiency (SAE) score were determined. The spontaneous alternation test measures a mouse’s spatial working memory by taking advantage of their natural curiosity to explore previously unvisited arms of a Y-maze. The mouse is then considered to remember the previously explored area. A random alternation happens when the animal reaches a particular arm of the maze in each of 3 successive arm entries. The following formula is used to measure the percent of spontaneous alternation: SAP% = [(number of alternations)/(total arm entries)] × 100.
Estimation of brain content of AchE activity: The brain content of AchE activity was measured using mice ELISA kits (BioVision, USA, CA) based on the Ellman method.6
Estimation of brain biomarkers: All brain biomarkers measured in the current study (MDA, SOD enzyme activity, TNF-α and NO, IL-6, Akt1 and pAkt) were measured using ELISA techniques.
Histological examination of brain tissues
To identify amyloid plaque deposits, brain samples were stained with 0.2% Congo red stain and incubated for one hour. The samples were then counterstained with hematoxylin solution. Amyloid plaques were detected and photographed using a fluorescent microscope at a magnification of 400X.
Paraffin brain blocks were used for immunohistochemical detection of glial fibrillary acidic protein (GFAP) in the hippocampus tissue to clarify astrocytes response to the neural degeneration in different experimental groups. Briefly, 4µm thick paraffin sections were cut and mounted on slides, deparaffinized in xylene, and hydrated in descending grades of ethanol. A primary rabbit monoclonal anti-GFAP antibody (Abcam, Cambridge, UK) was added, followed by a secondary antibody and then conjugated with streptavidin horseradish peroxidase. Finally, the sections were counterstained with Mayer’s hematoxylin. The GFAP-positive astrocytes were stained brown. In negative control sections, the primary antibody was omitted.
Statistical analysis
GraphPad Prism software version 5 (GraphPad, USA) was used to analyze data and generate figures. Data were represented as means ± standard deviation (M±SD). Statistical significance was determined by using one-way analysis of variance (ANOVA) followed by Dunnett’s test to compare the means of each individual group to the control group. A probability value less than 0.05 (p<0.05) was considered statistically significant.
Results
Effect of AST on short-term memory in scopolamine-induced Alzheimer’s model in mice: Figure 1 shows that the scopolamine-induced AD group showed a significant decrease in SAP compared to the control mice. However, treatment with AST significantly increased SAP compared to control group and the GLA treated group. Additionally, co-administration of AST and galantamine potentiated the effect on SAP in the AD+AST+GLA group compared to the AD+AST group.
- Spontaneous arm alternation percentage (SAP). AST - Astaxanthin, AD - Alzheimer’s disease, GLA - galantamine. Data are presented as mean±SD. Statistical analysis was carried out using ONE-WAY ANOVA test. ***statistically significant from the corresponding control group at p<0.001
Effect of AST on brain AchE activity in scopolamine-induced Alzheimer’s model in mice: As a cholinergic blocker, scopolamine significantly increased AchE activity in the brain of the AD group compared to the control group, as shown in Figure 2. However, administration of AST to scopolamine-induced AD group (AD+AST group) significantly decreased AchE activity by more than 5-fold. On the other hand, the effect of galantamine on AchE activity in the (AD+GLA) group was significantly higher than that of AST. Co-administration of AST and galantamine decreased AchE activity compared to the scopolamine-induced Alzheimer’s group. It is worth mentioning that administration of AST to control mice did not affect AchE activity levels.
- Effect of AST on brain AchE activity in scopolamine-induced Alzheimer’s model in mice. AST - Astaxanthin, AD - Alzheimer’s disease, GLA - galantamine. Data are presented as mean ±SD. Statistical analysis was carried out using ONE-WAY ANOVA test followed by Dunnett’s test. ***statistically significant from the corresponding AD group at p<0.001.
Effect of AST on brain histology of mice with scopolamine-induced Alzheimer’s model: Immunohistochemical staining of the hippocampus using the anti-GFAP antibody was performed to assess astrocytes’ response to neural degeneration in the different experimental groups. In the control group (Figure 3A, 3B), a few GFAP-positive immunoreactive astrocytes were detected among the pyramidal cells in CA3 and dentate gyrus, with many dispersed among the molecular layers. These astrocytes had ramifying processes passing between the neuronal cells. In the AST-treated group, GFAP immunostaining showed similar histological features as in the control group in the CA3 and DG regions (Figure 3C, 3D).
- CA3 region (A,C,E,G,I,K) & the dentate gyrus (DG) (B,D,F,H,J,L) stained by GFAP. (↑) GFAP immunostaining. A photomicrograph of the hippocampus proper of negative control group (A, B), Astaxanthin (AST) group (C, D), Scopolamine-induced Alzheimer’s (AD) model (E, F), Alzheimer’s mice treated with AST (AD+AST) (G, H), Alzheimer’s treated with galantamine (AD+GLA) (I, J), Alzheimer’s mice treated with AST and galantamine (AD+AST+GLA) (K, L). CA3 region (M,O,Q,S,U,W) & the dentate gyrus (DG) (N,P,R,T,V,X) stained by Congo red. (↑) represents neurofibrillary tangles within the cytoplasm of nerve cells. Inset: Higher magnification of the NFT in the subgranular zone. A photomicrograph of the hippocampus proper of negative control group (M,N), Astaxanthin (AST) group (O,P), Scopolamine-induced Alzheimer’s (AD) model (Q,R), Alzheimer’s mice treated with AST (AD+AST) (S,T), Alzheimer’s treated with galantamine (AD+GLA) (U,V), Alzheimer’s mice treated with AST and galantamine (AD+AST+GLA) (W, X). Scale bar 50µm.
In the scopolamine-induced AD group (AD) (Figure 3E, 3F), there was a marked increase in GFAP-positive staining of the cytoplasm and astrocytes’ processes, which were increased in number and size with multiple long thick processes.
In the scopolamine-AD treated with AST (AD+AST), a mild decrease in the number of GFAP-positive astrocytes with thin processes was observed in the CA3 and DG regions, particularly in the molecular and polymorphic layers. (Figure 3G, 3H). Brain sections from the scopolamine-AD group treated with galantamine (AD + GLA) (Figure 3I, 3J) showed astrocytes with large and thin ramified processes in-between the pyramidal cells CA3 region and granular cells in DG. Also, a moderated decrease of astrocytes dispersed among the molecular and the polymorphic layers of the CAS and DG. Interestingly, in the scopolamine-AD group with both AST and galantamine (AD+AST+GLA), there was a marked decrease in the GFAP-positive astrocytes in CA3 and DG regions (Figure 3K, 3L).
Congo red-stained sections of the control group showed the CA3 and DG regions of the hippocampus proper displaying pyramidal cells in the CA3 and granular cells of the DG with no obvious immature neurons with neurofibrillary tangles (NFTs) (Figure 3M, 3N). The AST-treated group showed a few orange colorations of the finely NFT within the cytoplasm of nerve cells of the subgranular zone and pyramidal cell layer (Figure 3O, 3P). The CA3 and DG region of the scopolamine-induced AD group (AD) showed an increase in the deposition of amyloid plaque. Also, there was a marked increase in the orange colorations of the NFT within the cytoplasm of immature cells of the subgranular zone and the granular cell layer in DG and pyramidal cell layer of the CA3 (Figure 3Q, 3R). However, the CA3 and DG region of the scopolamine-induced AD group treated with AST group (AD+AST) (Figure 3S, 3T) exhibited a mild decrease in the orange colorations of the NFT within the cytoplasm of immature cells in the subgranular zone of DG area and the pyramidal cell layer of the CA3 as compared to the scopolamine-induced AD group.
Similarly, the scopolamine-AD group treated with galantamine group (AD+GLA) showed a moderate decrease in the orange colorations of the NFT within the cytoplasm of immature cells in the granular cell layer and the pyramidal cells of the CA3 area as compared to the AD group (Figure 3U, 3V). Interestingly, simultaneous treatment with AST and galantamine (AD+AST+GLA) (Figure 3W, 3X) caused a marked decrease in the orange coloration of the NFT in immature cells in the subgranular zone of the DG compared to the AD group. However, the pyramidal cell layers of the CA3 area appeared nearly similar to the control group.
Effect of AST on brain oxidative stress biomarkers in the brain of mice with scopolamine-induced Alzheimer’s model: In the AD group, scopolamine induced a significant increase in MDA levels in the brain compared with the control group, as shown in (Figure 4A). Treatment with AST in the AD+AST group significantly reduced MDA levels by 72% compared to the AD group. The reduction in MDA levels induced by AST was significant compared to the effect of galantamine. Treatment with galantamine in the AD+GLA group significantly reduced MDA levels by only 36% compared to AD group. Co-administration of AST and galantamine in the AD+AST+GLA group significantly reduced MDA levels by 65% compared to the AD group.
- Effect of AST on brain oxidative stress biomarkers in scopolamine-induced Alzheimer’s model in mice. AST - Astaxanthin, AD - Alzheimer’s disease, GLA - galantamine. Data are presented as mean±SD. Statistical analysis was carried out using One-Way ANOVA followed by Dunnett’s test.***statistically significant from the corresponding AD group at p<0.001. #statistically significant from the AD+AST group at p<0.05.
In the scopolamine-induced AD group (AD), SOD activity was markedly reduced by approximately 50% compared with the control group, as shown in (Figure 4B). In contrast, administration of AST normalized the level of SOD activity compared to the AD group. Similarly, galantamine alone or with AST significantly prevented the scopolamine-induced reduction in SOD activity in the AD group.
Treatment with AST in the AD+AST group dramatically inhibited the scopolamine-induced elevation in NO levels in the brain of the AD group, as shown in (Figure 4C). Similarly, galantamine significantly decreased NO levels in the AD+GLA group compared to the AD group. However, the reduction in NO levels induced by AST was significant compared to the effect of galantamine. Co-administration of AST and galantamine in the AD+AST+GLA group significantly reduced NO levels compared to the AD group.
Effect of AST on proinflammatory biomarkers in the brain of scopolamine-induced Alzheimer’s model in mice: Scopolamine administration significantly increased the proinflammatory TNF-α level by 7-fold in the AD group compared to the control group, as shown in (Figure 5 A). However, there was a dramatic decrease in brain TNF-α levels in the AD+AST group by more than 70% compared to the AD group. Also, treatment with galantamine caused a significant reduction in TNF-α levels, but this reduction was potentially enhanced with AST in the AD+AST+GLA group.
- Effect of AST on brain inflammatory biomarkers in scopolamine-induced Alzheimer’s model in mice. AST: Astaxanthin, AD: Alzheimer’s disease, GLA: galantamine. Data are presented as mean ±SD. Statistical analysis was carried out using One-Way ANOVA followed by Dunnett’s test. ***: statistically significant from the corresponding AD group at p<0.001. #: statistically significant from the AD+AST group at p<0.05.
Additionally, scopolamine administration significantly increased IL-6 levels by 3.5-fold in the AD group compared to the control group, as shown in (Figure 5B). Administration of AST alone or galantamine alone induced a significant decrease in the IL-6 level comparable to the AD group. However, the reduction in IL-6 induced by AST was significant.
Effect of AST on brain levels of Akt-1, pAkt of scopolamine-induced Alzheimer’s model in mice: Figure 6 shows that Akt -1 and pAkt levels were dramatically decreased with scopolamine administration in the AD group. AST restored the elevated Akt -1 and pAkt levels compared to the AD group. Similarly, pAkt was significantly increased in the AD+AST group compared to the AD group. On the other hand, galantamine showed a significant increase in pAkt, but not in Akt-1compared to the AD group. Remarkably, the co-administration of AST and galantamine showed a significant increase in Akt-1 and pAkt, more than AST or galantamine alone.
- Effect of AST on brain levels of Akt-1 (A), pAkt (B) in scopolamine-induced Alzheimer’s model in mice. AST: Astaxanthin, AD: Alzheimer’s disease, GLA: galantamine. Data are presented as mean±SD. Statistical analysis was carried out using One-Way ANOVA followed by Dunnett’s test. **statistically significant from the corresponding AD group at p<0.01, and ***at p<0.001.
Discussion
Globally, AD is one of the most common neurodegenerative conditions, accounting for over 80% of dementia cases in the elderly population.1 The pathology of AD involves several specific and complicated processes, including neuronal death, extracellular amyloid beta plaques, intracellular tau protein deposition, and free radical generation. Therefore, developing novel pathways to create new medications capable of altering other processes is a worldwide goal for AD treatment.
In the current study, the AD-like mice model was induced by i.p injection of scopolamine. Scopolamine is known to induce cholinergic suppression, which in turn increases AchE activity in the brain and stimulate inflammatory responses and oxidative stress, leading to amnesia and a dementia-like state that simulates AD in animals.7 Furthermore, scopolamine alters cholinergic receptors in neurons and the brain’s memory system. Additionally, a previous study has shown that scopolamine has a unique effect in that it can cause the formation of amyloid plaques and hyperphosphorylation of tau, both of which are strongly associated with AD pathogenesis.8 These pathological confounders result in a loss of memory in rodents that mimics the impairments seen in AD. These behavioral impairments occurred in conjunction with increased cholinesterase activity, as well as increased brain oxidative stress and inflammation.9 Taken together, these exciting characteristics of the scopolamine-induced AD group make it a reliable and time-effective in vivo model to investigate the pharmacological effects of different therapeutic agents.
The current study demonstrates the efficacy of AST in protection against impairments in learning and memory, as well as its adjuvant with galantamine, which is one of the established therapeutic drugs for managing AD. AST is a pioneer among the current best natural carotenoid compounds, as multiple recent studies have demonstrated its suppressive activities on inflammation and oxidative stress, which are major contributors to neurodegenerative diseases such as Parkinson’s disease,10 in a mouse model of cerebral ischemia11 and chemotherapy-induced cognitive impairment.12 Indeed, AST can work against oxidative process in various ways, including by inhibiting lipid peroxidation, neutralizing radicals, scavenging singlet oxygen, and regulating gene expression associated with oxidative harm.13,14
The present study implicated a protective effect of AST against memory deficits in an AD mice model. Notably, the SAP was significantly improved in AD mice after administration of AST and achieved more than the control group. This finding is in accordance with a previous study in which AST improved cognitive behavior in a transgenic AD mouse.15 Furthermore, AST reversed cognitive and memory dysfunction in an Aβ-induced AD group in a substantial and dose-dependent manner.9 On the other hand, in this study, galantamine has minimal significant effect on improving the cognitive activity of AD mice. However, remarkably, AST showed a potentiating effect when administered with galantamine in the AD+AST+GLA group.
The cholinergic hypothesis proposed about 30 years ago that the degeneration of cholinergic neurons in elderly groups played a significant role in the deterioration of perception, attention, learning, memory, and other tasks.16 Amnesia in AD patients is associated with loss of cholinergic neurons and an increases in AchE levels in the brain.17 In alignment with the above findings, the current study showed that scopolamine accounts for the observed rise in AchE activity.18 Furthermore, the study found that AST significantly reduces AchE activity compared to the group treated with galantamine and enhances the effect of galantamine when used in combination. Additionally, recent research has indicated that AST has an inhibitory effect on AchE activity in mice with genetically induced AD15 and the Aβ-induced AD group.9
Moreover, histopathological analysis in the current study of the scopolamine-induced AD mice showed an obvious decline in the number of neurons and a rise in immature neurons. However, the administration of AST in the current study significantly reduced the hippocampal neuronal loss. In concomitant with the above observation, AST significantly decreased the number of astrocytes in the scopolamine-induced AD mice model. This outcome supports a recent study that demonstrated that a single dose of 10 mg/kg of AST administered intraperitoneally significantly reduced the activation of astrocytes in a rat model of AD induced by ferrous amyloid buthionine infusion.19
Aβ accumulation is the prominent pathology in the development of neurodegeneration in AD patients. Neuronal damage in the hippocampal areas of AD mice could be a result of Aβ accumulation.9 Congo red staining was used as a histological marker for Aβ accumulation in the current study, which revealed a significant rise in Aβ deposition in the brain of the scopolamine-treated AD group. The AST therapy protected the brain from Aβ-mediated neuronal damage and plaque formation in scopolamine-induced AD mice. These results were in accordance with recent studies that reported the effectiveness of AST to reducing Aβ in other in vivo models of AD.9,20
In the current study, hippocampus oxidative stress biomarkers were studied as a proposed mechanism of AST neuroprotective effect in the scopolamine-induced AD group. AST dramatically decreased the NO content. This result supports a previous study in which AST dose-dependently decreased NO levels in rats’ hippocampus following Aβ peptide infusion.9 Additionally, another study showed that AST improves cognitive abilities by inhibiting hippocampal NO expression in an aluminum chloride-induced memory impairment model.21
In AD, endogenous antioxidant enzymes exhibit decreased activity and performance. The findings of the current study support AST’s beneficial impact on antioxidant pathways by increasing the activity levels of SOD. Notably, AST has a synergistic effect when combined with galantamine in the current research. It enhances galantamine’s ability to activate antioxidant enzymes and lower the MDA concentration. This result is in agreement with a previous in vitro study.22
Additionally, the results of the current research showed that AST demonstrated potent anti-inflammatory properties. It inhibited the development of pro-inflammatory mediators such as TNF-α and IL-6, which were increased by scopolamine injection. This finding was consistent with previous research on other disease models. AST was reported to have a potent effect in reducing the inflammation induced by lipopolysaccharide in vitro and in vivo models.23 Also, AST has marked effectiveness in reducing TNF-α release in the uveitis model in rats.24 More importantly, the anti-inflammatory properties of AST in AD group were demonstrated in a recent study in which Aβ-infused hippocampal tissue showed a significant rise in TNF-α levels. AST treatment substantially decreased TNF-α release caused by Aβ peptides.9 In addition, AST demonstrated a potent anti-inflammatory role by inhibiting the expression of proinflammatory mediators such as IL-6. Previous research has demonstrated similar anti-inflammatory effects of AST using a variety of different experimental models. In inactivated microglial cells, AST (50 µM) significantly reduced the release of IL-6.25 Interestingly, the findings of this study indicated that AST was more effective than galantamine in suppressing oxidative stress biomarkers and the inflammatory cytokines in an AD mice model. Furthermore, AST enhanced the effects of galantamine in that regard.
Although oxidative stress and chronic pro-inflammatory cytokines could explain neuronal death in AD pathogenesis,26 other pathways such as Akt-1 and PAkt pathways could also be involved. The current study showed that scopolamine injection in the AD group led to decreased Akt1 and its active phosphorylated form (pAkt), which represent a critical pathway for neuronal survival. In alignment with this, a previous study demonstrated a decrease in pAkt activity in a scopolamine-induced AD group.8 In line with this, studies have shown that scopolamine increases tau protein gene expression and decreases Akt activation.27,28 The current study showed that AST effectively corrected the decline caused by scopolamine in the levels of active forms of Akt and PAkt. This is the first known investigation to evaluate the role of Akt-1 and PAkt pathways in vivo as a new mechanism for AST’s neuroprotective effects at a molecular level.
Conclusion
The current study demonstrated that AST has a potent protective effect against the scopolamine-induced Alzheimer’s disease model in mice. The reduction of oxidative stress and chronic proinflammatory cytokines might be the main reasons for AST’s improvement of memory. However, the role of Akt1 and pAkt pathways in this process is not yet clear, and the study is the first to examine this mechanism in a live Alzheimer’s disease model. Therefore, it can be suggested that AST’s modulation of Akt may provide a new explanation for its protective effect against memory impairment in Alzheimer’s disease.
Acknowledgments
The authors, therefore, acknowledge with thanks DSR for technical and financial support.
Footnotes
Disclosure. This project was funded by the Deanship of Scientific Research (DSR) at King Abdulaziz University, Jeddah, under grant no. G. 502-248-1442. The authors, therefore, acknowledge with thanks DSR for technical and financial support.
- Received July 12, 2023.
- Accepted December 28, 2023.
- Copyright: © Neurosciences
Neurosciences is an Open Access journal and articles published are distributed under the terms of the Creative Commons Attribution-NonCommercial License (CC BY-NC). Readers may copy, distribute, and display the work for non-commercial purposes with the proper citation of the original work.
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