EFFECT OF NARINGENIN ON ALUMINUM CHLORIDE INDUCED ALZHEIMER DISEASE IN

DROSOPHILA MELANOGASTER
A PROJECT
BY:
AIYEDE EMMANUEL ODAFE
(BMS1708760)
DEPARTMENT OF PHYSIOLOGY,
SCHOOL OF BASIC MEDICAL SCIENCES,
UNIVERSITY OF BENIN.

SUPERVISED BY:
MR CHURCHILL INNEH
JANUARY, 2023
INTRODUCTION
Background
• Alzheimer's disease is a neurodegenerative disorder characterized by
the progressive loss of cognitive function and memory.
• It is the most common cause of dementia in older adults.
• It is characterized by the accumulation of amyloid plaques and
neurofibrillary tangles in the brain (Honjoet al., 2012).
 Role of oxidative stress in Alzheimer's disease
• Oxidative stress has been proposed as a contributing factor to the
development of Alzheimer's disease.
• Oxidative stress occurs when the production of reactive oxygen
species (ROS) exceeds the body's ability to neutralize them, leading to
damage to cells and tissues.
• "Mitochondrial dysfunction has been linked with increased production
of reactive oxygen species to the development of AD." (Calkins et al.,
2011)
• Oxidative stress has been associated with synaptic dysfunction in AD."
(Newcomer et al., 2000)
 Use of Drosophila melanogaster as a model organism
• Drosophila has several advantages as a model organism, including a
short lifespan, ease of genetic manipulation, and the presence of many
molecular and cellular pathways that are conserved in humans"
(Smith, 2020).
 Parameters used to study oxidative stress in Drosophila
• Negative geotaxis, superoxide dismutase (SOD), catalase (CAT),
glutathione S-transferase (GST), hydrogen peroxide (H2O2) or
malondialdehyde (MDA), acetylcholinesterase, and monoamine
oxidase have been used to study oxidative stress and its effects on the
brain in Drosophila.
• These parameters help researchers understand the role of oxidative
stress in the development of neurodegenerative diseases like
Alzheimer's disease.
 Evidence for a role of aluminum in Alzheimer's disease
• Al has been found at high levels in the brains of individuals with
Alzheimer's disease, and it may contribute to the formation of amyloid
plaques and neurofibrillary tangles.
• Aluminum exposed animals have displayed neurofibrillary tangles
formation, cholinergic neuronal axonal terminal loss, β-amyloid
protein aggregation, oxidative stress, and neuronal apoptosis in the
hippocampus and frontal cortex site for cognition, memory, and
synaptic plasticity similar to the pathogenesis of AD (Praveenkumar et
al., 2019)."
 Protective effects of naringenin against aluminum toxicity.
• Naringenin is an important antioxidant and inflammatory substance.
• The number of hydroxyl substitutions of naringenin can donate
hydrogen to ROS, allowing acquisition of stable structure, thus
enabling scavenging of these free Radicals (Shen et al., 2004; Heo
et al., 2004).
• Administration of naringenin interestingly improved spatial learning
and memory in a rat model of AD through regulating the
PI3K/AKT/GSK-3β pathway and reducing Tau hyper-phosphorylation
(Yang et al., 2014).
• While some studies have suggested that naringenin may have
protective effects against AlCl3-induced Alzheimer's disease in
Drosophila flies, more research is needed to fully understand the
mechanisms underlying these effects and to determine whether
naringenin could be a potential therapeutic option for Alzheimer's
disease in humans.
 JUSTIFICATION OF THE STUDY
• Aluminum chloride has been shown to cause neurotoxicity and AD-
like neurodegeneration in animal models, including impaired
locomotor performance, learning, and memory in Drosophila
melanogaster. It has also been linked to increased acetylcholinesterase
activity and oxidative stress. Naringenin has been shown to have
neuroprotective and antioxidant effects, including improving survival
rate, climbing time, and reducing acetylcholinesterase activity and
oxidative stress in animals co-administered with aluminum chloride.
 AIM OF THE STUDY

• The aim is to study the effect of naringenin on aluminum chloride-

induced Alzheimer’s disease in Drosophila melanogaster.
RESEARCH QUESTIONS

• Does exposure to AlCl3 cause Alzheimer's disease in Drosophila flies?

• Does naringenin have protective effects against AlCl3-induced
Alzheimer's disease in Drosophila flies?
• What is the mechanism by which naringenin may exert its protective
effects against AlCl3-induced Alzheimer's disease in Drosophila flies?
 SPECIFIC OBJECTIVES OF THE STUDY

 To determine the effect of Naringenin on longevity in Drosophiliamelanogaster

fed with diet containing AlCl3
 To determine the effect of Naringenin on negative geotaxis in
Drosophiliamelanogasterfed with diet containing AlCl3
 To determine the effect of Naringenin on antioxidant parameters in
Drosophiliamelanogasterfed with diet containing AlCl3
 To determine the effect of Naringenin on acetylcholinesterase (AchE) activity in
Drosophiliamelanogasterfed with diet containing AlCl3
MATERIALS AND METHOD
• Equipment
• Weighing balance (Metler analytical balance), Measuring cylinder
(Pyrex, England), Conical Flask, beakers (Pyrex, England), Water bath
(TT42D Multipurpose use, Techmel and Techmel, USA), Micropipette
(Microplux), Stirrer, Syringe, Glass jar, treatment tubes.
• Chemicals and Reagent
• All chemicals were of analytical grade and were obtained from
standard commercial suppliers. Aluminium Chloride, distilled water,
agar-agar, glucose, nipargin, yeast, corn meal.
 METHODS

• The flies were fed with the standard formulated diet of corn meal
medium which contained corn meal (52g), brewers, yeast (5g),
glucose (3.5g) agar (7.5g) and Nipagin (1g), alcohol (1ml). Distilled
water was used for making the diet.

• The flies were allowed to mate in vials monitored under a regulated

temperature until the eggs metamorphosed into young adult fruit flies
under a natural photoperiod of about 12 hours light and dark daily for
a period of administration of Aluminium Chloride and Naringenin.
 Methods continued

• The experimental period lasted for 21 days. The flies were monitored
daily and the mortality rates were recorded for plotting for the survival
curve. Negative geotaxis (climbing time) was observed daily to check
locomotor deficit.
• Each experimental procedure was replicated 5 times (5replicates). At
the end of the experimental period, the flies were homogenated,
centrifuge and the supernatant were used for biochemical analysis.
 Methods continued

The following biochemical parameters were measured.

• Superoxide Dismutase (SOD)
• Malonaldehyde (MDA)
• Catalase(CAT)
• Glutathione-s-Transferase (GST)
• ETHICAL APPROVAL
• STATISTICAL ANALYSIS
REFERENCES

• Heo, H., Shen, J., & Lee, J. (2004). Scavenging activity of flavonoids on hydroxyl
radicals. Phytotherapy Research, 18(2), 152-155.
• Honjo, K., Iwatsubo, T., & Ihara, Y. (2012). The prevalence and pathogenesis of
Alzheimer's disease. Journal of Neurology, 259(Suppl 1), S3-S9.
• Praveenkumar, K., Al-Dabbous, Z., & Banerjee, R. (2019). Aluminum-induced
neurotoxicity: mechanisms and therapeutic strategies. Current
Neuropharmacology, 17(5), 516-524
• Smith, J. (2020). The benefits of using Drosophila as a model organism. Journal
of Experimental Biology, 56(2)123-134.
• Shen, J., Heo, H., & Lee, J. (2004). Scavenging activity of flavonoids on
superoxide radicals. Phytotherapy Research, 18(2), 148-151.
• Yang, X., Huang, H., Zhang, H., & Wang, J. (2014). Naringenin improves spatial
learning and memory and regulates the PI3K/AKT/GSK-3β pathway in a rat
model of Alzheimer's disease. Molecular Medicine Reports, 10(1), 367-371.