romv tmao in vivo  (MedChemExpress)

Journal: Gut Microbes

Article Title: Blautia coccoides -derived metabolite trimethylamine-N-oxide exacerbates Alzheimer's disease progression via targeting HIF1α signaling

doi: 10.1080/19490976.2025.2605768

Figure Lengend Snippet: B. coccoides supplementation promoted the production of TMAO in both AD mouse models and in vitro . (A) and (B) α-diversity measurement of the Shannon index and Simpson index between P301s and B. coccoides group ( n = 5). (C) PCoA showed variations in the gut microbiota composition between the two groups ( n = 5). (D) KEGG pathway enrichment analysis of B. coccoides functional genes. (E) Volcano plot displayed of differential metabolites between the two groups ( n = 5), FDR < 0.05. (F) VIP analysis revealed that TMAO was an important differential metabolite ( n = 5). (G) Bubble chart displayed the significant Spearman correlation between Blautia abundance and differential metabolites from the VIP analysis. The gradient colors and the size of the circle represent the correlation coefficient. (H) Functional sequence of C utC indicated the bacterial biology of B. coccoides . (I) Linear regression analysis of B. coccoides and the fecal TMAO levels. (J) Illustration of the experimental setup in which the fermentation broth from a 24-h co-culture of B. coccoides or B. producta and 50 mg/L Choline was applied to LM3 cells. (K) Quantitative analysis of TMAO level among Con, B. coccoides, B. coccoides + Choline, B. producta and B. producta + Choline groups ( n = 3). (L) Representative Western blot images and quantitative analysis of Occludin in P301s and B. coccoides group ( n = 4). (M) Quantitative analysis of Occludin and ZO-1 levels in the P301s group and B. coccoides group ( n = 4). ( N ) Representative immunohistochemical images of Occludin and ZO-1 in the P301s group and B. coccoides group. The image above: magnification 200×. Scale bar = 100 μm. (O) Quantitative analysis of TMAO level in plasma between two groups ( n = 5). (P) Quantitative analysis of TMAO levels in the brain between the two groups ( n = 5). (Q) Linear regression analysis of B. coccoides and brain's TMAO levels. The error bars indicate the SEM, * p < 0.05 and **** p < 0.0001.

Article Snippet: The cells were pretreated with TMAO (MCE, USA) for 16 h. After pretreatment, the cells were exposed to 1% O2 hypoxic conditions for an additional 6 h to stabilize HIF1α expression.

Techniques: In Vitro, Functional Assay, Sequencing, Co-Culture Assay, Western Blot, Immunohistochemical staining, Clinical Proteomics

Journal: Gut Microbes

Article Title: Blautia coccoides -derived metabolite trimethylamine-N-oxide exacerbates Alzheimer's disease progression via targeting HIF1α signaling

doi: 10.1080/19490976.2025.2605768

Figure Lengend Snippet: The TMAO level was correlated with dementia severity in AD patients, and TMAO treatment promoted Tau phosphorylation and oxidative stress in vitro . (A) Forest plot of MR analyses for the associations between TMAO and Dementia in AD, points represent beta estimates and 95% CI. (B) P301L-N2a cells were treated with 0, 1, 10, 20 and 50 μM TMAO, and cell viability was assessed using the CCK-8 method ( n = 5). (C) Representative Western blot images of p-Tau181 and p-Tau396 of Con group and TMAO group in P301L-N2a cells. (D) Quantitative analysis of p-Tau181 and p-Tau396 levels ( n = 3). The Con group was used as a reference value. (E) P301L-293T cells were treated with 0, 1, 10, 20 and 50 μM TMAO, and cell viability was assessed using the CCK-8 method ( n = 5). (F) Representative Western blot images of p-Tau181 and p-Tau396 of Con group and TMAO group in P301L-293T cells. (G) Quantitative analysis of p-Tau181 and p-Tau396 levels ( n = 3). The Con group was used as a reference value. (H) Representative immunofluorescence images of p-Tau396 of Con group and TMAO group in P301L-N2a cells. (I) Representative immunofluorescence images of p-Tau396 of Con group and TMAO group in P301L-293T cells. (J) Flow cytometry of ROS levels in Con and TMAO treated group of P301L-N2a. (K) Quantitative analysis of ROS in the two groups of P301L-N2a ( n = 3). (L) Flow cytometry of the ROS levels in the Con and TMAO treated group of P301L-293T. (M) Quantitative analysis of ROS in the two groups of P301L-293T ( n = 3). The image above: magnification 200×. Scale bar = 100 μm. The error bars indicate the SEM, * p < 0.05, ** p < 0.01, *** p < 0.01, and **** p < 0.0001.

Article Snippet: The cells were pretreated with TMAO (MCE, USA) for 16 h. After pretreatment, the cells were exposed to 1% O2 hypoxic conditions for an additional 6 h to stabilize HIF1α expression.

Techniques: Phospho-proteomics, In Vitro, CCK-8 Assay, Western Blot, Immunofluorescence, Flow Cytometry

Journal: Gut Microbes

Article Title: Blautia coccoides -derived metabolite trimethylamine-N-oxide exacerbates Alzheimer's disease progression via targeting HIF1α signaling

doi: 10.1080/19490976.2025.2605768

Figure Lengend Snippet: TMAO promoted oxidative stress by suppressing the HIF1α signaling pathway. (A) Chemical structure of TMAO. (B) Venn diagram showed overlap between TMAO binding targets and AD-related genes. (C) PPI network diagram of TMAO and AD intersection genes (interaction score > 0.7). (D) MCC screening of overlap targets identified the top 10 core targets with the shortest interaction paths in the PPI network. (E) KEGG pathway enrichment showing that the HIF1 and Alzheimer disease pathways were significantly enriched in overlap genes, FDR < 0.05 was considered statistically significant. (F) Volcano plot showing the distribution of gene expression level differences between the Con and TMAO group. FDR < 0.05 were screened as differentially expressed genes. (G) KEGG pathway enrichment showing that the HIF1 and Alzheimer's disease pathways were significantly enriched in DEGs between the Con and TMAO groups; FDR < 0.05 was considered statistically significant. (H) Representative Western blot analysis of HIF1 pathway in the Con and TMAO group, including HIF1α, HO-1, SOD2 and GPX4. (I) Quantification of HIF pathway between two groups ( n = 3); the Con group was used as a reference value. (J) Representative Western blot of HIF1 pathway in P301s and B. coccoides group, including HIF1α, HO-1, SOD2 and GPX4. (K) Quantification of HIF pathway between P301s and B. coccoides group ( n = 4); the P301s group was used as a reference value. (L) Representative Western blot of HIF1 pathway in the TMAO and TMAO + OE group, including HIF1α, HO-1, SOD2 and GPX4. (M) Quantification of HIF pathway between TMAO and TMAO + OE groups ( n = 3); the TMAO group was used as a reference. (N) Representative Western blot images of AT8, p-Tau181 and p-Tau396 in the two groups. (O) Quantitative analysis of AT8, p-Tau181 and p-Tau396 levels between TMAO group and TMAO + OE group ( n = 3); TMAO group was used as a reference. The error bars indicate the SEM, * p < 0.05 and ** p < 0.01.

Article Snippet: The cells were pretreated with TMAO (MCE, USA) for 16 h. After pretreatment, the cells were exposed to 1% O2 hypoxic conditions for an additional 6 h to stabilize HIF1α expression.

Techniques: Binding Assay, Gene Expression, Western Blot

Journal: Gut Microbes

Article Title: Blautia coccoides -derived metabolite trimethylamine-N-oxide exacerbates Alzheimer's disease progression via targeting HIF1α signaling

doi: 10.1080/19490976.2025.2605768

Figure Lengend Snippet: TMAO promoted Tau phosphorylation via the HIF1α interaction. (A) AutoDock simulation of the TMAO-HIF1α interaction. (B) Diagram of the HIF1α deletion mutant at hydrogen bond binding sites 235–237 with TMAO. (C) Flow cytometry of the ROS level of P301L cells in HIF1α-WT, HIF1α-mut, TMAO + HIF1α-WT and TMAO + HIF1α-mut group. (D) Quantitative analysis of ROS among the four groups ( n = 3). (E) Representative Western blot of the HIF1 pathway in the four groups. (F) Quantification of the HIF pathway; the HIF1α-WT group was used as a reference value ( n = 3). (G) Representative immunofluorescence images of p-Tau396 in the four groups. The image above: magnification 200×. Scale bar = 100 μm. (H) Quantification of p-Tau396 levels ( n = 3). (I) Representative Western blot images of p-Tau181 in the four groups. (J) Quantitative analysis of p-Tau181 levels; the HIF1α-WT group was used as a reference value ( n = 3). Error bars indicate the SEM, * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001.

Article Snippet: The cells were pretreated with TMAO (MCE, USA) for 16 h. After pretreatment, the cells were exposed to 1% O2 hypoxic conditions for an additional 6 h to stabilize HIF1α expression.

Techniques: Phospho-proteomics, Mutagenesis, Binding Assay, Flow Cytometry, Western Blot, Immunofluorescence