Table1_The Role of Ion Channel-Related Genes in Autism Spectrum Disorder: A Study Using Next-Generation Sequencing.DOCX

The clinical heterogeneity of autism spectrum disorder (ASD) is closely associated with the diversity of genes related to ASD pathogenesis. With their low effect size, it has been hard to define the role of common variants of genes in ASD phenotype. In this study, we reviewed genetic results and clinical scores widely used for ASD diagnosis to investigate the role of genes in ASD phenotype considering their functions in molecular pathways. Genetic data from next-generation sequencing (NGS) were collected from 94 participants with ASD. We analyzed enrichment of cellular processes and gene ontology using the Database for Annotation, Visualization, and Integrated Discovery (DAVID). We compared clinical characteristics according to genetic functional characteristics. We found 266 genes containing nonsense, frame shift, missense, and splice site mutations. Results from DAVID revealed significant enrichment for “ion channel” with an enrichment score of 8.84. Moreover, ASD participants carrying mutations in ion channel-related genes showed higher total IQ (p = 0.013) and lower repetitive, restricted behavior (RRB)-related scores (p = 0.003) and mannerism subscale of social responsiveness scale scores, compared to other participants. Individuals with variants in ion channel genes showed lower RRB scores, suggesting that ion channel genes might be relatively less associated with RRB pathogenesis. These results contribute to understanding of the role of common variants in ASD and could be important in the development of precision medicine of ASD.</p

Table_1_Effects of a Dehydroevodiamine-Derivative on Synaptic Destabilization and Memory Impairment in the 5xFAD, Alzheimer's Disease Mouse Model.docx

Carboxy-dehydroevodiamine·HCl (cx-DHED) is a derivative of DHED, which improves memory impairment. Carboxyl modification increases solubility in water, indicating that its bioavailability is higher than that of DHED. Cx-DHED is expected to have better therapeutic effects on Alzheimer's disease (AD) than DHED. In this study, we investigated the therapeutic effects of cx-DHED and the underlying mechanism in 5xFAD mice, transgenic (Tg) mouse model of AD model mice. In several behavioral tests, such as Y-maze, passive avoidance, and Morris water maze test, memory deficits improved significantly in cx-DHED-treated transgenic (Tg) mice compared with vehicle-treated Tg mice. We also found that AD-related pathologies, including amyloid plaque deposition and tau phosphorylation, were reduced after the treatment of Tg mice with cx-DHED. We determined the levels of synaptic proteins, such as GluN1, GluN2A, GluN2B, PSD-95 and Rabphilin3A, and Rab3A in the brains of mice of each group and found that GluN2A and PSD-95 were significantly increased in the brains of cx-DHED-treated Tg mice when compared with the brains of Tg-vehicle mice. These results suggest that cx-DHED has therapeutic effects on 5xFAD, AD model mice through the improvement of synaptic stabilization.</p

Image_1_Effects of a Dehydroevodiamine-Derivative on Synaptic Destabilization and Memory Impairment in the 5xFAD, Alzheimer's Disease Mouse Model.tiff

Carboxy-dehydroevodiamine·HCl (cx-DHED) is a derivative of DHED, which improves memory impairment. Carboxyl modification increases solubility in water, indicating that its bioavailability is higher than that of DHED. Cx-DHED is expected to have better therapeutic effects on Alzheimer's disease (AD) than DHED. In this study, we investigated the therapeutic effects of cx-DHED and the underlying mechanism in 5xFAD mice, transgenic (Tg) mouse model of AD model mice. In several behavioral tests, such as Y-maze, passive avoidance, and Morris water maze test, memory deficits improved significantly in cx-DHED-treated transgenic (Tg) mice compared with vehicle-treated Tg mice. We also found that AD-related pathologies, including amyloid plaque deposition and tau phosphorylation, were reduced after the treatment of Tg mice with cx-DHED. We determined the levels of synaptic proteins, such as GluN1, GluN2A, GluN2B, PSD-95 and Rabphilin3A, and Rab3A in the brains of mice of each group and found that GluN2A and PSD-95 were significantly increased in the brains of cx-DHED-treated Tg mice when compared with the brains of Tg-vehicle mice. These results suggest that cx-DHED has therapeutic effects on 5xFAD, AD model mice through the improvement of synaptic stabilization.</p

Image_3_Effects of a Dehydroevodiamine-Derivative on Synaptic Destabilization and Memory Impairment in the 5xFAD, Alzheimer's Disease Mouse Model.tiff

Carboxy-dehydroevodiamine·HCl (cx-DHED) is a derivative of DHED, which improves memory impairment. Carboxyl modification increases solubility in water, indicating that its bioavailability is higher than that of DHED. Cx-DHED is expected to have better therapeutic effects on Alzheimer's disease (AD) than DHED. In this study, we investigated the therapeutic effects of cx-DHED and the underlying mechanism in 5xFAD mice, transgenic (Tg) mouse model of AD model mice. In several behavioral tests, such as Y-maze, passive avoidance, and Morris water maze test, memory deficits improved significantly in cx-DHED-treated transgenic (Tg) mice compared with vehicle-treated Tg mice. We also found that AD-related pathologies, including amyloid plaque deposition and tau phosphorylation, were reduced after the treatment of Tg mice with cx-DHED. We determined the levels of synaptic proteins, such as GluN1, GluN2A, GluN2B, PSD-95 and Rabphilin3A, and Rab3A in the brains of mice of each group and found that GluN2A and PSD-95 were significantly increased in the brains of cx-DHED-treated Tg mice when compared with the brains of Tg-vehicle mice. These results suggest that cx-DHED has therapeutic effects on 5xFAD, AD model mice through the improvement of synaptic stabilization.</p

Image_2_Effects of a Dehydroevodiamine-Derivative on Synaptic Destabilization and Memory Impairment in the 5xFAD, Alzheimer's Disease Mouse Model.tiff

Carboxy-dehydroevodiamine·HCl (cx-DHED) is a derivative of DHED, which improves memory impairment. Carboxyl modification increases solubility in water, indicating that its bioavailability is higher than that of DHED. Cx-DHED is expected to have better therapeutic effects on Alzheimer's disease (AD) than DHED. In this study, we investigated the therapeutic effects of cx-DHED and the underlying mechanism in 5xFAD mice, transgenic (Tg) mouse model of AD model mice. In several behavioral tests, such as Y-maze, passive avoidance, and Morris water maze test, memory deficits improved significantly in cx-DHED-treated transgenic (Tg) mice compared with vehicle-treated Tg mice. We also found that AD-related pathologies, including amyloid plaque deposition and tau phosphorylation, were reduced after the treatment of Tg mice with cx-DHED. We determined the levels of synaptic proteins, such as GluN1, GluN2A, GluN2B, PSD-95 and Rabphilin3A, and Rab3A in the brains of mice of each group and found that GluN2A and PSD-95 were significantly increased in the brains of cx-DHED-treated Tg mice when compared with the brains of Tg-vehicle mice. These results suggest that cx-DHED has therapeutic effects on 5xFAD, AD model mice through the improvement of synaptic stabilization.</p

Image_4_Effects of a Dehydroevodiamine-Derivative on Synaptic Destabilization and Memory Impairment in the 5xFAD, Alzheimer's Disease Mouse Model.tiff

Carboxy-dehydroevodiamine·HCl (cx-DHED) is a derivative of DHED, which improves memory impairment. Carboxyl modification increases solubility in water, indicating that its bioavailability is higher than that of DHED. Cx-DHED is expected to have better therapeutic effects on Alzheimer's disease (AD) than DHED. In this study, we investigated the therapeutic effects of cx-DHED and the underlying mechanism in 5xFAD mice, transgenic (Tg) mouse model of AD model mice. In several behavioral tests, such as Y-maze, passive avoidance, and Morris water maze test, memory deficits improved significantly in cx-DHED-treated transgenic (Tg) mice compared with vehicle-treated Tg mice. We also found that AD-related pathologies, including amyloid plaque deposition and tau phosphorylation, were reduced after the treatment of Tg mice with cx-DHED. We determined the levels of synaptic proteins, such as GluN1, GluN2A, GluN2B, PSD-95 and Rabphilin3A, and Rab3A in the brains of mice of each group and found that GluN2A and PSD-95 were significantly increased in the brains of cx-DHED-treated Tg mice when compared with the brains of Tg-vehicle mice. These results suggest that cx-DHED has therapeutic effects on 5xFAD, AD model mice through the improvement of synaptic stabilization.</p

S100A9KOxTg crossbred mice showed significant improvement in spatial reference memory.

<p>We performed memory tests at the age of 13-months. (A) The Morris water maze test was performed. Training trials were conducted for 5 consecutive days. From the 5^th day of training trials, escape latency was significantly increased in the WT/Tg group. However, the latency was decreased in the KO/Tg group compared to the WT/Tg group.<i>*p</i><0.05 by one-way ANOVA. (B) The probe test was performed 48 h after the final training session. The times that the mice of each group stayed in zones 1, 2, 3 and 4 were compared. The time spent in the platform quadrant (zone 4) was significantly decreased in the WT/Tg group. However, the KO/Tg group showed memory improvement compared to the WT/Tg group in zone 4. (C) In the Y-maze, the WT/Tg group showed a significant decrease in the alternation rate of arm entries. In the passive avoidance test, the latency time of the KO/Tg group was greatly increased. Behavior task groups were as follows: n = 7–11 per group. These results were analyzed by ANOVA followed by the LSD test <i>*p</i><0.05, <i>**p<</i>0.01.</p

Accumulation of phosphorylated tau was detected in neurites surrounding amyloid plaques in the brain of S100A9 crossbred mice at 14 months old.

<p>Phosphorylated tau (P-tau) was detected in the hippocampus and cortex of WT/Tg and KO/Tg mice brains by immunohistochemistry. The amount of P-tau was significantly reduced in KO/Tg mice brains compared with WT/Tg mice brains. (h) – (j) are P-tau stained region of (c) and (g). Sections are 4 µm thick. ((a) – (h) Scale bar; 200 µm, (h) – (k) scale bar; 50 µm).</p

The number of amyloid plaques and amount of Aβ_1-42 were reduced in S100A9 KO/Tg mice.

<p>After the tests, the mice brains were isolated and the brain slices were stained with Congo red for the detection of amyloid plaques. (A) Histological analysis was performed out at the age of 14-months. In the cortex, hippocampus and entorhinal cortex, amyloid plaques were detected using Congo red staining. (i)-(l) The Congo red-stained region of (c), (g) and (h). Thio-S stained region of (m). Sections are 4 µm thick. ((a) – (h) Scale bar; 200 µm, (i)-(l) scale bar; 50 µm). (B) Quantitative analysis of Congo red-stained plaque number. The number of amyloid plaques was counted in brain slices containing the hippocampal region of each group, and the average number of plaques per brain slice was calculated. In brains from the KO/Tg group, Aβ deposition was significantly reduced in the cortex and hippocampus, compared to the WT/Tg group. The total numbers of mice per group were as follows: n = 3–6 per group.<i>*p</i><0.05 by one-way ANOVA. (C, D) Western blot analysis was performed with the total lysates from the cortical region and hippocampal region of brains in each group using the 6E10 antibody. Aβ and CT bands were detected and normalized by the amount of APP and GAPDH. In the WT/Tg mice brain, many Aβ and CT were produced compared to KO/Tg mice. (n = 10). (E, F) Aβ<b>_1-42</b> levels in cortical or hippocampal brain regions in all groups were analyzed by Aβ ELISA. The levels of Aβ<b>_1-42</b> were highly increased in the cortex and hippocampus of WT/Tg mice compared with age-matched WT/WT or WT/KO mice. Note that the levels of Aβ<b>_1-42</b> were decreased in the cortex and hippocampus of KO/Tg mice brains compared with WT/Tg mice brains (n = 4). (G, H) Western blot analysis was performed with total lysates from the cortical region of the brains in each group using an antibody against oligomeric Aβ. In the cortex of KO/Tg mice brains, the relative quantity of oligomeric Aβ was decreased compared with WT/Tg mice. (n = 3) <i>*p</i><0.05 by one-way ANOVA.</p

The anti-inflammatory cytokine IL-10 was significantly increased and pro-inflammatory cytokines IL-6 and TNF-α were decreased in the cortex of KO/Tg mice brains compared to WT/Tg mice brains at 14 months old.

<p>(A) The level of IL-10 was detected in the tissue lysates from the cortical region of the brain from each group by sandwich ELISA. IL-10, which is a representative anti-inflammatory cytokine, was increased in the cortex of KO/Tg mice brains compared to WT/Tg mice brains. (B) The level of IL-6 was detected in the tissue lysates from the cortical region of the brain from each group by sandwich ELISA. IL-6, which is a representative pro-inflammatory cytokine, was decreased in the cortex of KO/Tg mice brains compared to WT/Tg mice brains. (C) The level of TNF-α was detected in the tissue lysates from the cortical region of the brain from each group by sandwich ELISA. TNF-α, which is a representative pro-inflammatory cytokine, was decreased in the cortex of KO/Tg mice brains compared to WT/Tg mice brains. (n = 9–10), *<i>p</i><0.05 by one-way ANOVA.</p