الفهرس 
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Abstract
The most common type of dementia is Alzheimer’s disease (AD). It is an irreversible, progressive neurodegenerative illness marked by synapse malfunction and neuronal cell death, resulting in memory impairment and cognitive decline. As a result, therapeutic alternatives to currently existing treatments appear to be required. Exosomes are microvesicles with a diameter of 30-100 nm and small lipid vesicles that are secreted by all cell types. They performed their roles by fusing with the target cell and releasing active tiny molecules such as messenger RNA, microRNA, lipids, and proteins from their cell of origin into the target cell’s membranes and cytoplasm. As a result, the surface receptors are stimulated, and the target cell is regulated.
Exosomes produced from mesenchymal stem cells have recently been shown to have the ability to regenerate the functioning of numerous tissues via their paracrine action via the production of growth factors, cytokines, or chemokines.
In many inflammatory illnesses, conditioned medium (CM) generated from stem cells has an anti-inflammatory impact. CM derived from mesenchymal stem cells contains a variety of cytokines, growth factors, and microRNAs that play essential roles in inflammation modulation and can, in certain situations, substitute stem cell therapy.
So, the purpose of this study was to compare the therapeutic effect of different novel therapies; stem cell derived exosomes and their conditioned
medium on the hippocampus of a rat model of Alzheimer’s disease, as well as to explain the various mechanisms involved in those effects using histological, immunohistochemical, and biochemical methods.
42 adult male albino rats were utilized to demonstrate those roles, together with 10 male albino rats aged 4-6 weeks and weighing 70-80 gm for the generation of mesenchymal stem cell-derived conditioned media and exosomes. Furthermore, 32 rats aged 6-8 weeks with weights ranging from 150-200 gm were divided into four equal groups of eight rats each: group I received the vehicle (distilled water) orally via gastric tube for 42 days at a dose of 0.5 ml/100g body weight. For 42 days, group II Alzheimer’s group (AD group) received daily administration of Alcl3 100 mg/kg body weight diluted in distilled water at a dose of 0.5 ml/100g body weight via a gastric tube. group III (CM group) received daily oral administration of Alcl3 at the same dose and duration as the AD-group, followed by a single injection of CM into the rats’ tail vein at a dose of 0.5 ml/rat under ether inhalation anesthesia. group IV Exosomes treated group (Exo group) received daily oral administration of Alcl3 at the same dose and duration as the AD-group, followed by a single injection of BM-MSC-derived exosomes at a dose of 100g/rat suspended in 1ml phosphate buffered saline via the rats’ tail vein under ether inhalation anesthesia. The effect of ALCL3 treatment, conditioned medium, and BM-MSC derived exosomes injection on rats’ activity and memory was assessed using behavioral assays. Each group had five rats. These experiments were carried out at three different times: following 42 days of ALCL3 administration, as well as 2 and 4 weeks of conditioned medium and BM-MSC generated exosomes injection.
Four weeks after being injected with conditioned medium and exosomes, the animals were slaughtered.
• Behavioral results: AD-group animals had significantly lower mean levels of SAP and DI than control rats. At the end of the second week, the CM-treated and Exo-treated groups had significantly lower SAP than the control group but not the AD group. At the conclusion of the trial (4th week), the CM-treated group was considerably lower than the control group but insignificant when compared to the AD-group. The SAP of the Exo-treated group, on the other hand, was considerably lower than that of the AD-group.
The AD group had severe hypoactivity and anxiety-like behaviour. At the second week, the CM-treated group had a considerably higher number of crossed peripheral squares and rearing frequency than the control group. The number of crossed peripheral squares decreased significantly in the exo- treated group while rearing frequency decreased non-significantly. Furthermore, when compared to the control group, the Exo-treated group spent a substantial amount of time freezing and a non-significant amount of time grooming. The number of crossed peripheral squares and rearing frequency in the CM-treated group increased considerably by the fourth week. In addition, it greatly increased freezing time and had no effect on grooming duration as compared to the control. The exo-treated group greatly increased the number of crossing peripheral squares and rearing frequency while having no effect on anxiety-like behaviors.
• Biochemical results: When compared to the C-group, the AD-group had significantly higher levels of MDA, TNF-, and AchE. However, administration of CM and exosomes in CM and Exo-treated groups dramatically lowered MDA levels in the hippocampus when compared to the AD- group. When compared to the C-group, the TAC level in the AD group was much lower. When compared to the AD- group, the treatment of CM and exosomes enhanced the level of TAC in the hippocampus considerably.
• Hematoxylin and eosin staining revealed significant histological abnormalities in the AD-group, including degeneration of most small and big pyramidal neurons, congested blood capillaries, and numerous shrunken granular cells surrounded by pericellular haloes with vacuolated cytoplasm. In this group, glial cells were seen on a pink neuropil in all regions of the hippocampus. CA3 and CA4 were found to be the most affected locations. While the CM-group showed some improvement in histopathological alterations, it was more localized to the CA1, CA3, and DG areas. The majority of pyramidal cells in CA1 and CA3 have normal shape. The Exo- treated group improved more than the CM-group in all of the previously reported histopathological alterations that occurred in the AD-group.
• Modified Bielschowsky’s silver stain results: Intra-neuronal aggregations of argyrophilic neurofibrillary tangles (NFTs) were identified in cells of the CA1, CA4, and DG regions in the AD group. There were also two types of plaques found. While the CM-group demonstrated that the majority of CA1 and CA4 pyramidal neurons had
lightly stained nuclei and undamaged faintly stained axons. Other neurons seemed smaller, with intra-neuronal argyrophilic neurofibrillary tangle buildup. Most granular neurons were preserved in the DG. When compared to the CM-treated group, the Exo-group showed higher preservation of normal structure and less degeneration of pyramidal neurons in CA1 and CA4.
• The Immunohistochemical and morphometrical The results demonstrated a substantial decrease in the mean number of anti-CD68 immune-reactive cells and mean area fraction of GFAP in the treated groups compared to the AD group, but an increase in the mean area fraction of nestin and CD163 in the treated groups compared to the AD group.
• The electron microscope results։ The prior observations were confirmed. Elongated microglia showed electron dense cytoplasm with tiny cytoplasmic vacuoles, while others appeared as dark dystrophic microglia in the AD group. This group of astrocytes had an uneven nucleus with less electron dense nuclear chromatin, vacuolated cytoplasm with engulfed myelin sheath, dilated endoplasmic reticulum cisternae, and damaged mitochondria. Some degenerative neurons, dark elongated microglia, and astrocytes in the CM-group had an irregular nucleus with electron rich nuclear chromatin. While the Exo- treated group showed improvement with more or less typical pyramidal neurons.