Journal: International Journal of Biological Sciences

Article Title: Metabolic Adaptation of CD8⁺ T Cells Limits the Efficacy of Fatty Acid Oxidation Inhibition in Type 1 Diabetes

doi: 10.7150/ijbs.125649

Figure Lengend Snippet: Trimetazidine modulates mitochondrial function, reduces activation, and impairs proliferation in human T cells. (A) Total human T cells were stimulated with anti-CD3, CD28, and CD2 with or without TMZ for 24 hours and analyzed for mitochondrial membrane potential using JC-1 dye. Gates indicate populations with low membrane potential (monomers), high membrane potential (aggregates), and intermediate states. (B) Mitochondrial and (C) total reactive oxygen species (ROS) levels. (D) Cell viability, defined as Annexin V - /7-AAD - cells. (E) Fatty acid oxidation (FAO), measured via oxygen consumption rate and compared to the FAO inhibitor etomoxir. (F) CD25 surface expression and (G) intracellular Ki67 expression were assessed after 72 hours of TMZ treatment. (H) Cell viability (Annexin V - /7-AAD - ) after 72 hours. (I) Mitochondrial membrane potential measured with JC-1 dye after 72 hours. (J) Mitochondrial and (K) total ROS levels after 72 hours of TMZ. Data represents n = 6 donors. Paired t-tests were used for statistical analysis. CO: Control; TMZ: Trimetazidine; φ: mitochondrial membrane potential.

Article Snippet: Mice were enrolled when fasting glucose reached 150-180 mg/dL and received 62.5 mg/L Trimetazidine dihydrochloride (MedChemExpress, NJ, USA) in drinking water (approx.

Techniques: Activation Assay, Membrane, Expressing, Control

Journal: International Journal of Biological Sciences

Article Title: Metabolic Adaptation of CD8⁺ T Cells Limits the Efficacy of Fatty Acid Oxidation Inhibition in Type 1 Diabetes

doi: 10.7150/ijbs.125649

Figure Lengend Snippet: Trimetazidine delays diabetes onset and reduces islet T cell infiltration in dysglycemic NOD mice. (A) Schematic overview of the experiment, starting with 68 female NOD/ShiLtJ mice. Of these, 55 mice reached a fasting blood glucose of 150 mg/dL but no more than 180 mg/dL and were treated with TMZ in drinking water. 20 randomly allocated mice sacrificed after 1 week, while the remaining 35 were followed until diabetes onset. In total, 28 of these mice developed diabetes, defined as fasting blood glucose ≥200 mg/dL sustained for one week, measured twice per week. (B) Proportion of colony animals enrolled in the study over time. n= 80. (C) Representative images showing β-cell (insulin) and α-cell (glucagon) distribution in dysglycemic mice. (D) Quantification of β-cell (insulin) to α-cell (glucagon) ratios in pancreatic islets after 1 week of treatment. Mean total islet area analyzed per mouse: 137,460 µm². n= 9-10. (E) Representative images of CD4 + and CD8 + T cell infiltration in pancreatic islets (white outlines). (F-G) Quantification of CD4 + and CD8 + T cell infiltration into pancreatic islets. Mean total islet area analyzed per mouse: 234,377 µm². n= 10. (H) Representative images of FoxP3 + regulatory T cells. (I) Quantification of FoxP3 + regulatory T cells in pancreatic islets. Mean total islet area analyzed per mouse: 137,460 µm². n= 9-10. (J) Representative images CD68 + macrophages. (K) Quantification of CD68 + macrophage in pancreatic islets. Mean total islet area analyzed per mouse: 125,565 µm². n= 10. (L) Kaplan-Meier survival curve showing delayed diabetes onset in TMZ-treated mice following intervention. P-value calculated with the Gehan-Breslow-Wilcoxon test. n= 14. (M) Fasting blood glucose levels during the 3-week follow-up, with area under the curve (AUC) comparison. Two-way ANOVA mixed-effects model with Geisser-Greenhouse correction; p-value reflects the time x treatment interaction. (N-Q) Quantification of (N) CD4 + (N), CD8 + (O), regulatory T cells (P), and macrophages (Q) infiltration in diabetic mice treated with TMZ. Mean total islet area analyzed per mouse: (N-O) 74,548 µm². n= 13-14; (P) 65,402 µm². n= 14; (Q) 95,811 µm². n= 14. Unpaired t-test or Mann-Whitney test were used for statistical analysis. CO = Control, TMZ = Trimetazidine.

Article Snippet: Mice were enrolled when fasting glucose reached 150-180 mg/dL and received 62.5 mg/L Trimetazidine dihydrochloride (MedChemExpress, NJ, USA) in drinking water (approx.

Techniques: Comparison, MANN-WHITNEY, Control

Journal: International Journal of Biological Sciences

Article Title: Metabolic Adaptation of CD8⁺ T Cells Limits the Efficacy of Fatty Acid Oxidation Inhibition in Type 1 Diabetes

doi: 10.7150/ijbs.125649

Figure Lengend Snippet: Trimetazidine reduces mitochondrial and inflammatory stress-associated cell populations in dysglycemic pancreatic tissue. (A) Uniform Manifold Approximation and Projection (UMAP) of scRNA-seq data from pre-diabetic mice, showing six major pancreatic cell clusters: acinar cells, mitochondrially active stromal cells, stressed cells, ductal cells, endothelial cells, and immune cells. (B) Clustergrammer heatmap illustrating cluster-specific gene expression profiles. Genes enriched in mitochondrially active stromal cells and stressed cells are highlighted. (C) Reactome pathway enrichment analysis (via Enrichr) of genes significantly upregulated in mitochondrially active stromal cells. (D) Reactome pathway enrichment analysis based on the heatmap-defined gene cluster and significantly upregulated genes in stressed cells. (E) UMAPs showing pancreatic cells from control and TMZ-treated pre-diabetic mice. (F) Statistical analysis of cluster-specific cell frequency changes in TMZ-treated animals.

Article Snippet: Mice were enrolled when fasting glucose reached 150-180 mg/dL and received 62.5 mg/L Trimetazidine dihydrochloride (MedChemExpress, NJ, USA) in drinking water (approx.

Techniques: Gene Expression, Control

Journal: International Journal of Biological Sciences

Article Title: Metabolic Adaptation of CD8⁺ T Cells Limits the Efficacy of Fatty Acid Oxidation Inhibition in Type 1 Diabetes

doi: 10.7150/ijbs.125649

Figure Lengend Snippet: Prophylactic TMZ treatment does not delay diabetes onset or alter islet immune infiltration in NOD mice. (A) Experimental timeline: Female NOD/ShiLtJ mice received TMZ via drinking water from 5 weeks of age. A cohort of 13-14 mice was sacrificed at 15 weeks for analysis prior to diabetes onset, while 20 mice were monitored until either diabetes onset (defined as fasting blood glucose ≥200 mg/dL for one week) or 30 weeks of age, whichever comes first. (B) Quantification of β-cell (insulin) to α-cell (glucagon) ratios in pancreatic islets from pre-diabetes onset mice. Mean total islet area analyzed per mouse: 175,385 µm². n= 13-14. (C) Representative images showing insulin + β-cells and glucagon + α-cells in islets from pre-diabetes onset mice. (D) Quantification of CD4 + and (E) CD8 + T cell infiltration in islets from pre-diabetes onset mice. Mean total islet area analyzed per mouse: 112,745 µm². n= 11-13. (F) Representative images of CD4 + and CD8 + T cell infiltration in islets from pre-diabetes onset mice. (G) Quantification of FoxP3 + regulatory T cells in islets from pre-diabetes onset mice. Mean total islet area analyzed per mouse: 175,385 µm². n= 13-14. (H) Representative images of FoxP3 + regulatory T cell infiltration in islets from pre-diabetes onset mice. (I) Quantification of CD68 + macrophage in islets from pre-diabetes onset mice. Mean total islet area analyzed per mouse: 202,532 µm². n= 13-14. (J) Representative images of CD68 + macrophage infiltration in islets from pre-diabetes onset mice. (K) Kaplan-Meier analysis of diabetes onset in TMZ-treated and control mice. P-value calculated with the Gehan-Breslow-Wilcoxon test. n= 20. (L) Final diabetes incidence in both groups. n=20. Statistical comparison was performed using Fisher's exact test. (M) Representative images of insulin and glucagon staining in diabetic pancreases. (N) Quantification of β-cell (insulin) to α-cell (glucagon) ratios in diabetic islets. Mean total islet area analyzed per mouse: 88,582 µm². n=16-17. (O) Quantification of CD4 + T cell infiltration in diabetic islets. Mean total islet area analyzed per mouse: 77,100 µm². n= 14-16. (P) Quantification of CD8 + T cell infiltration in diabetic islets. Mean total islet area analyzed per mouse: 77,100 µm². n= 17. (Q) Quantification of FoxP3 + T cell infiltration in diabetic islets. Mean total islet area analyzed per mouse: 88,582 µm². n= 16-18. (R) Quantification of CD68 + macrophages in diabetic islets. Mean total islet area analyzed per mouse: 130,986 µm². n= 17. CO: Control; TMZ: Trimetazidine. Unpaired t-test or Mann-Whitney test were used for statistical analysis.

Article Snippet: Mice were enrolled when fasting glucose reached 150-180 mg/dL and received 62.5 mg/L Trimetazidine dihydrochloride (MedChemExpress, NJ, USA) in drinking water (approx.

Techniques: Control, Comparison, Staining, MANN-WHITNEY

Journal: International Journal of Biological Sciences

Article Title: Metabolic Adaptation of CD8⁺ T Cells Limits the Efficacy of Fatty Acid Oxidation Inhibition in Type 1 Diabetes

doi: 10.7150/ijbs.125649

Figure Lengend Snippet: Trimetazidine modulates CD4 + and CD8 + T cell differentiation in vitro . Naïve human CD4 + and CD8 + T cells were cultured under stimulatory conditions with or without TMZ for 17 days. Surface expression of CCR7 and CD45RA was used to define T cell subsets by flow cytometry: naïve (T N ; CCR7 + CD45RA + ), central memory (T CM ; CCR7 + CD45RA - ), and effector memory (T EM ; CCR7 - CD45RA - ). (A) Percentage of CD4 + T N cells across days 7-17 and time-course analysis from day 0 to 17. (B) CD4 + T CM cells across days 7-17 and time-course analysis. (C) CD4 + T EM cells across days 7-17 and time-course analysis. (D) CD8 + T N cells across days 7-17 and time-course analysis. (E) CD8 + T CM cells across days 7-17 and time-course analysis. (F) CD8 + T EM cells across days 7-17 and time-course analysis. Data represent n = 6 donors. Paired t-test was used for statistical analysis. * indicates p < 0.05. CO: Control; TMZ: Trimetazidine.

Article Snippet: Mice were enrolled when fasting glucose reached 150-180 mg/dL and received 62.5 mg/L Trimetazidine dihydrochloride (MedChemExpress, NJ, USA) in drinking water (approx.

Techniques: Cell Differentiation, In Vitro, Cell Culture, Expressing, Flow Cytometry, Control

Journal: International Journal of Biological Sciences

Article Title: Metabolic Adaptation of CD8⁺ T Cells Limits the Efficacy of Fatty Acid Oxidation Inhibition in Type 1 Diabetes

doi: 10.7150/ijbs.125649

Figure Lengend Snippet: Metabolic compensation of CD8 + T cells under Trimetazidine treatment. Naïve human CD8 + T cells were cultured under stimulatory conditions with or without TMZ for 17 days. Surface expression of CCR7 and CD45RA was used to define T cell subsets by flow cytometry: naïve (T N ; CCR7 + CD45RA + ), central memory (T CM ; CCR7 + CD45RA - ), effector memory (T EM ; CCR7 - CD45RA - ), and T EMRA (CCR7 - CD45RA + ). (A-D) Representative heatmap-style dot plot from intracellular flow cytometry showing expression of CPT1A (carnitine-palmitoyl-transferase-1A) (A), GLUT1 (glucose transporter 1) (B), PRDX2 (peroxiredoxin-2) (C), and IDH2 (NADP + -dependent isocitrate dehydrogenase 2) (D) in control CD8 + T cell subsets (T N , T CM , T EM ) at day 10. Dot color shows mean fluorescence intensity, thus indicates protein abundance. (E-H) Heatmaps of z-scored expression levels for CPT1A (E), GLUT1 (F), PRDX2 (G), and IDH2 (H) across CD8 + T cell subsets and all time points (day 0, 3, 7, 10, 14, 17) under control and TMZ conditions. (I-L) Quantification of CPT1A expression over time in CD8 + T N (I), T CM (J), T EM (K), and T EMRA (L) subsets. (M) Heatmap-style dot plot showing CPT1A expression across all subsets at day 17, highlighting elevated expression in all TMZ-treated populations. Data represent n = 5-6 donors. Paired t-test was used for statistical analysis. * indicates p < 0.05. CO: Control; TMZ: Trimetazidine.

Article Snippet: Mice were enrolled when fasting glucose reached 150-180 mg/dL and received 62.5 mg/L Trimetazidine dihydrochloride (MedChemExpress, NJ, USA) in drinking water (approx.

Techniques: Cell Culture, Expressing, Flow Cytometry, Control, Fluorescence, Quantitative Proteomics

Journal: International Dental Journal

Article Title: Ethno-Dentistry of Medicinal Plants Used in North Waziristan, Pakistan

doi: 10.1016/j.identj.2023.10.001

Figure Lengend Snippet: List of ethnodentistry medicinal plants with scientific name, local name, family name, voucher No., growth form, part used, UVs, URs, FC, RFC, and FL.

Article Snippet: Vitex negundo L. SR-13171 , Arwanad , Verbenaceae , Shrub , Leaves, Twigs , Paste, toothbrush , Toothache, gum infections, bad breath, cleaning teeth , 35 , 0.27 , 0.69 , 24 , 74.29.

Techniques: Cannabis, Infection