BackgroundAlzheimer’s disease (AD) is a complex brain disorder that is greatly affected by genetics. Next-generation sequencing (NGS) has facilitated the discovery of rare variants in new genes that may be linked to AD in different populations. However, we still know very little about the genetic makeup of AD in Saudi Arabia and other Arab populations.ObjectivesThis study aims to explore rare variants that are predicted to be deleterious in a group of 64 Saudi patients diagnosed with sporadic and familial Alzheimer’s disease (AD). These patients previously tested negative for mutations in genes known to cause AD and were genotyped for APOE alleles.MethodsWe performed whole-exome sequencing (WES) on the Ion Proton platform. Then, we used our internal process for filtering, validating, and prioritizing variants.ResultsUsing stringent selection criteria, we identified 107 rare candidate variants with potential functional relevance. Of these, 26 (24.3%) were novel, while the remaining variants had been previously reported in public databases. Among these candidates, 33 were connected to AD, 28 to both AD and other neurodegenerative disorders (OND), 34 to OND-related functions, and 11 to broader processes like aging, inflammation, and neuronal regulation. We found rare missense variants in genes involved in important processes related to Alzheimer’s disease. These processes include mainly Aβ and Tau pathology, kinase signaling, stress response, and neuroinflammation.ConclusionOur analysis reveals diverse genetic contributors to Alzheimer’s disease in a population that remains largely underrepresented in genomic studies. We identified candidate variants in 53% of the patients, highlighting the value of expanding AD genetics research to non-European populations.

Background: Copy number variations (CNVs) play an important role in the genetic etiology of various neurological disorders, including Alzheimer’s disease (AD). Type 2 diabetes mellitus (T2DM) and major depressive disorder (MDD) were shown to have share mechanisms and signaling pathways with AD. Objective: We aimed to assess CNVs regions that may harbor genes contributing to AD, T2DM, and MDD in 67 Saudi familial and sporadic AD patients, with no alterations in the known genes of AD and genotyped previously for APOE. Methods: DNA was analyzed using the CytoScan-HD array. Two layers of filtering criteria were applied. All the identified CNVs were checked in the Database of Genomic Variants (DGV). Results: A total of 1086 CNVs (565 gains and 521 losses) were identified in our study. We found 73 CNVs harboring genes that may be associated with AD, T2DM or MDD. Nineteen CNVs were novel. Most importantly, 42 CNVs were unique in our studied cohort existing only in one patient. Two large gains on chromosomes 1 and 13 harbored genes implicated in the studied disorders. We identified CNVs in genes that encode proteins involved in the metabolism of amyloid-β peptide (AGRN, APBA2, CR1, CR2, IGF2R, KIAA0125, MBP, RER1, RTN4R, VDR and WISPI) or Tau proteins (CACNAIC, CELF2, DUSP22, HTRA1 and SLC2A14). Conclusion: The present work provided information on the presence of CNVs related to AD, T2DM, and MDD in Saudi Alzheimer’s patients.

Background:Alzheimer’s disease (AD) is a chronic neurological disorder associated with mental decline and dementia. Several studies focused on investigating the molecular basis of the disease that led to the identification of several causative genes and risk associated alleles. Replication of these studies and findings from different populations is very important. Objective:Molecular assessment of a cohort of 117 familial and sporadic AD cases from Saudi Arabia. Methods:Comprehensive screening for point mutations was carried out by direct sequencing of coding regions in the three known AD causative genes: PSEN1, PSEN2, APP, as well as the AD associated gene SORL1. All patients were also genotyped for APOE alleles. In silico 3D protein structure analysis was performed for two novel SORL1 variants. Results:We identified a total of eight potential pathogenic missense variants in all studied genes. Five of these variants were not previously reported including four in SORL1 (p.Val297Met, p.Arg1084Cys, p.Asp1100Asn, and p.Pro1213Ser) and one in APP (p.Glu380Lys). The frequency of APOE-ɛ4 allele was 21.37% of total investigated cases. In silico 3D protein structure analysis of two SORL1 novel missense variants (p.Pro1213Ser and p.Arg1084Cys) suggested that these variants may affect the folding of the proteins and disturb their structure. Conclusions:Our comprehensive analysis of the open reading frame of the known genes have identified potential pathogenic rare variants in 18/117 cases. We found that point mutations in AD main genes (PSEN1, PSEN2, and APP) were underrepresented in our cohort of patients. Our results confirm involvement of SORL1 in familial and sporadic AD cases.

Pregnenolone sulfate (PREGS) and dehydroepiandrosterone sulphate (DHEAS) are pro-amnesic, anti-amnesic and neuroprotective steroids in rodents. In Alzheimer’s disease (AD) patient’s brains, their low concentrations are correlated with high levels of Aβ and tau proteins. The unnatural enantiomer ent-PREGS enhanced memory in rodents. We investigated here whether ent-PREGS and ent-DHEAS could be neuroprotective in AD models.

The enantiomers of pregnenolone sulfate (ent -PREGS) and of dehydroepi-androsterone sulfate (ent -DHEAS) are synthetic analogues of the neuroactive neurosteroids PREGS and DHEAS, respectively. In rodents, ent -PREGS improves memory and is 10-fold more potent that PREGS. The effect of ent -DHEAS on memory is unknown. These data lead us to consider the role of neurosteroid enantiomers as neuroprotectants in Alzheimer's disease (AD). The effects of ent- PREGS were investigated in vitro against Aβ25 - 35 peptide induced neuronal death and also on neurite outhgrowth, in comparison to PREGS. ent -PREGS and ent -DHEAS were examined for their block A β 25–35-induced amnesia and oxidative stress in mice. B104 neuroblastoma cell cultures were incubated with ent -PREGS or PREGS at 0.25, 0.5, 2, 5, 10, 20 μM in absence or presence of A β 25–35 (5 μM toxic dose) for 24h. Cell viability and death types were measured by flow cytometry. Neurite outgrowth was analyzed by the Neuron J software. Male Swiss mice were simultaneously administered intracerebroventricularly with A β 25–35 (9 nmol), steroid enantiomers (0.05, 0.2, 0.5 and 2 nmol) or vehicle. Their memory capacities were evaluated by the spontaneous alternation and step passive avoidance tests. Peroxidized lipids quantification in hippocampal extracts was based on Fe(III)xylenol orange complex formation. Results: Neither ent -PREGS nor PREGS were toxic to B104 cells. ent -PREGS pre-treatment (5, 10 μM) attenuated the A b 25–35-induced decreased in cell viability. It lowered the percentage of late apoptotic cells while PREGS (0.25–5μM) decreased that of late apoptotic and necrotic cells. en t-PREGS and PREGS enhanced neurite length as compared to control cells, in a time dependant manner. A β 25–35-induced spontaneous alternation deficits were diminished by en t-PREGS (0.5, 0.2 and 2 nmol). Impaired retention response of step-through passive avoidance was decreased by ent -PREGS (0.5 nmol). ent -DHEAS (0.2, 2 nmol) prevented impaired spontaneous alternation and step down passive avoidance responses. Reduced lipid peroxidation by A β 25–35 was prevented by ent -PREGS (2 nmol) and ent -DHEAS (0.5 nmol). ent -PREGS and en t-DHEAS are neuroprotective steroids that are highly effective against the A β 25–35-associated oxidative damage, in terms of reduction of neuronal cell death and spatial working and contextual long term memory deficits.

SCO-spondin is a newly identified protein that is strongly expressed in the subcommissural organ (SCO), an ependymal differentiation of the brain. When released into the cerebrospinal fluid at the entrance to the Sylvian aqueduct, the glycoproteins condense and form a thread-like structure, Reissner's fiber (RF). To analyze the role of SCO-spondin on neuronal development, we studied the effects induced by an oligopeptide derived from a thrombospondin type 1 repeat (TSR) of SCO-spondin on neuroblastoma B104 cells and compared them with the effects of soluble RF material containing complete SCO-spondin proteins. In low density cell culture, the TSR peptide first induced a notable flattening of cells accompanied by increased neurite outgrowth. Grouping of these differentiated B104 cells, which later formed dense aggregates, was then observed with increasing time in culture. Soluble RF material induced similar morphological changes and neurite-promoting effects on B104 cells, although the cells remained evenly distributed throughout the culture time and no aggregates were visible. In high-density cell culture, both TSR peptide and RF material induced prominent neurite outgrowth and subsequent rapid cell aggregation. Whereas soluble RF material inhibited cell proliferation, no respective effect was observed in the presence of the TSR peptide. A direct interaction of TSR peptide and soluble RF material with a B104 cell binding site was revealed by increased B104 cell metabolic activity by flow cytometry.

In the developing vertebrate nervous system, several proteins of the thrombospondin superfamily act on axonal pathfinding. By successive screening of a SCO-cDNA library, we have characterized a new member of this superfamily, which we call SCO-spondin. This extracellular matrix glycoprotein of 4,560 amino acids is expressed and secreted early in development by the subcommissural organ (SCO), an ependymal differentiation located in the roof of the Sylvian aqueduct. Furthermore, SCO-spondin makes part of Reissner's fiber (RF), a thread-like structure present in the central canal of the spinal cord. This novel protein shows a unique arrangement of several conserved domains, including 26 thrombospondin type 1 repeats (TSR), nine low-density lipoprotein receptor (LDLr) type A domains, two epidermal growth factor (EGF)-like domains, and N- and C-terminal von Willebrand factor (vWF) cysteine-rich domains, all of which are potent sites of protein-protein interaction. Regarding the huge number of TSR, the putative function of SCO-spondin on axonal guidance is discussed in comparison with other developmental molecules of the CNS exhibiting TSR. To correlate SCO-spondin molecular feature and function, we tested the effect of oligopeptides, whose sequences include highly conserved amino acids of the consensus domains on a neuroblastoma cell line B 104. One of these peptides (WSGWSSCSRSCG) markedly increased neurite outgrowth of B 104 cells and this effect was dose dependent. Thus, SCO-spondin is a favorable substrate for neurite outgrowth and may participate in the posterior commissure formation and spinal cord differentiation during ontogenesis of the central nervous system.

. The subcommissural organ (SCO) is a specialized ependymal structure of the brain that secretes glycoproteins into the cerebrospinal fluid (CSF), which condense to form a thread-like structure – Reissner's fiber (RF). The effects of soluble material released by RF were examined on neuroblastoma B104 cells grown in serum-free medium, using "low-density" and "high-density" culture systems. In the presence of soluble RF material, low-density cultures were suitable for analysis of the enhanced neurite outgrowth of B104 cells, while high-density cultures allowed the increased B104 cell aggregation to be examined. RF-induced neuronal aggregation and neuritic outgrowth were restricted to a perimeter around the RF. This standardized cell culture system reproduced in part the effects observed previously with primary cortical and spinal cord cell cultures and may serve the analysis of the mechanisms leading to aggregation and neurite outgrowth. In the present study, we analyzed variations in the rate of neural cell adhesion molecules, such as N-CAM and N-cadherin, induced by soluble RF material in high-density cultures.