Journal: Nature Communications

Article Title: High-fat diet disrupts a septal control on feeding to promote obesity in male mice

doi: 10.1038/s41467-025-68010-x

Figure Lengend Snippet: a Representative image of AAV-DIO-Gad2 expression in the LS. Scale bar, 500 μm. This pattern of expression and projection was consistently observed across 3 independent biological replicates. b Left: western blot analysis of Gad2 expression in the EGFP (Control) and Gad2 (Gad2 overexpression) groups (β-actin was an internal reference). Right: quantification of Gad2 signal intensity for the EGFP and Gad2 groups ( p < 0.05). c Quantification of GABA expression levels for the EGFP and Gad2 groups ( p < 0.01). d Schematic of the experimental design. e – g Liquid food intake in EGFP vs. Gad2-overexpressing mice. Left: licking behavior; middle: cumulative licks; right: total intake of standard liquid food ( e ), sucrose solution ( f ) and Ensure ( g ) (standard liquid food: p = 0.94; sucrose solution: p < 0.001; Ensure: p < 0.001). h Viral strategy and timeline for overexpression Gad2 in LS GABA neurons. i Quantification of energy intake (left) and body weight (right) for the Chow+EGFP, Chow+Gad2, HFD + EGFP, and HFD+Gad2 groups [Energy intake (virus: F 1,28 = 18.1, p < 0.001; diet: F 1,28 = 29.8; p < 0.001; time: F 3.08,86.25 = 33.3, p < 0.001). Body weight (virus: F 1,28 = 10.1, p < 0.01; diet: F 1,28 = 10.5; p < 0.01; time: F 1.848,51.75 = 219.9, p < 0.001)]. j Quantification of Gad2 expression level (virus: F 1,27 = 6.2, p < 0.05; diet: F 1,27 = 1895; p < 0.001; interaction: F 1,27 = 7.9, p < 0.01). Statistics: ( b , c , e–g ) Mann-Whitney U-test, ( i ) three-way ANOVA followed by post hoc test using two-stage step-up method of Benjamini, Krieger and Yekutieli, ( j ) two-way ANOVA followed by post hoc test using two-stage step-up method of Benjamini, Krieger and Yekutieli, with detailed statistics provided in Supplementary Data . Sample sizes are indicated in the figures. ns, no significant difference, * p < 0.05, ** p < 0.01, *** p < 0.001. Data are presented as mean ± SEM. All tests are two-sided. Source data are provided as a Source Data file.

Article Snippet: The primary antibodies used were rabbit anti-Gad65 (1:3000, Proteintech, Cat# 20746-1-AP) and mouse anit-β-actin (1:3000, Proteintech, Cat# 66009), The secondary antibodies used were goat anti-rabbit (1:2000, ThermoFisher, Cat# 32460) or goat anti-mouse (1:2000, ThermoFisher, Cat# 62-6520).

Techniques: Expressing, Western Blot, Control, Over Expression, Virus, MANN-WHITNEY

Journal: Oncogene

Article Title: GABAergic signaling contributes to tumor cell invasion and poor overall survival in colorectal cancer

doi: 10.1038/s41388-025-03546-2

Figure Lengend Snippet: A GAD1 expression of KRAS wild-type (WT) CRC cell lines plotted against their metastatic potential in mice (DepMap). B GAD1 expression of KRAS mutant CRC cell lines plotted against their metastatic potential in mice (DepMap). C Schematic of OOC model of tumor cell lines invading from the top channel into the bottom endothelial channel. D On-chip tumor invasion of GAD1 knockdown HCT116 tumor cells, normalized to control shRNA. Data was analyzed using an unpaired t-test. N = 6 chips; *** P = 0.0005. E On-chip tumor invasion of HCT116 tumor cells treated with GAD1 inhibitor 3-MPA (5 μM), normalized to vehicle control. Data was analyzed using an unpaired t-test. N = 6 chips; *** P = 0.0006. F On-chip tumor invasion of HCT116 tumor cells with increasing concentrations of exogenous GABA. Data was analyzed using a one-way ANOVA with Šídák’s multiple comparison test. N = 6–8 chips; **** P < 0.0001. G Intracellular [ 13 C 4 ]GABA or unlabeled GABA was measured via mass spectrometry-based metabolomics in the HCT116 tumor-chips after the addition of exogenous GABA for six days. N = 3 chips. The intensities of the internal standard 13 C 11 Tryptophan (Trp) were plotted as a control. H On-chip GABA staining in the top channel (1) representing non-invaded HCT116 tumor cells, and in the bottom channel (2) capturing invaded HCT116 tumor cells. Scale bars represent 200 μm in the top channel image and 100 μm in the bottom channel images. Individual data points are shown and mean ± SEM are represented. All schematics were made in or are from BioRender.

Article Snippet: GAD1 human shRNA lentiviral particles (Santa Cruz Biotechnology, Inc., #sc- sc-35435-V) and control shRNA lentiviral particles (Santa Cruz Biotechnology, Inc., #sc-108080) were used to produce HCT116 H2B-GFP GAD1 knock down cells at a multiplicity of infection (MOI) of 2 and 2 mg/mL of puromycin (Santa Cruz Biotechnology, #sc-108071) was used to select for knocked down cells.

Techniques: Expressing, Mutagenesis, Knockdown, Control, shRNA, Comparison, Mass Spectrometry, Staining

Journal: Oncogene

Article Title: GABAergic signaling contributes to tumor cell invasion and poor overall survival in colorectal cancer

doi: 10.1038/s41388-025-03546-2

Figure Lengend Snippet: Forest plots show the association of GAD1 tumor expression as a continuous variable with PFS ( A ) and OS ( B ) across treatment arms; Forest plots show the association of ABAT tumor expression with PFS ( C ) and OS ( D ) across treatment arms. Kaplan-Meier curves show PFS ( E ) and OS ( F ) stratified by GAD1 tumor expression tertiles (T1 low, T2 middle, and T3 high) in the overall population of the CALGB/SWOG 80405 trial. The statistical comparison is performed for T3 vs T1. Kaplan-Meier curves show PFS ( G ) and OS ( H ) stratified by ABAT tumor expression tertiles in the overall population of the CALGB/SWOG 80405 trial. The statistical comparison is performed for T3 vs T1. P values are based on log-rank test for PFS and OS in the univariate categorical analysis, and likelihood ratio test in the univariate continuous and multivariable Cox proportional hazards regression model. bev bevacizumab, cet cetuximab, CI confidence interval, HR hazard ratio, OS overall survival, PFS progression-free survival.

Article Snippet: GAD1 human shRNA lentiviral particles (Santa Cruz Biotechnology, Inc., #sc- sc-35435-V) and control shRNA lentiviral particles (Santa Cruz Biotechnology, Inc., #sc-108080) were used to produce HCT116 H2B-GFP GAD1 knock down cells at a multiplicity of infection (MOI) of 2 and 2 mg/mL of puromycin (Santa Cruz Biotechnology, #sc-108071) was used to select for knocked down cells.

Techniques: Expressing, Comparison

Journal: Oncogene

Article Title: GABAergic signaling contributes to tumor cell invasion and poor overall survival in colorectal cancer

doi: 10.1038/s41388-025-03546-2

Figure Lengend Snippet: A GAD1 mRNA expression from TCGA in primary colon cancer tumors. N = 196 patients with KRAS , NRAS , or BRAF mutant tumors; N = 201 patients with KRAS , NRAS , or BRAF wildtype tumors. Individual data points are shown and the median with interquartile range is represented. Data was analyzed with an unpaired t-test; **** P < 0.0001. B GAD1 mRNA expression from the Gene Expression Omnibus (GEO), accession GSE39582 , in primary colon cancer tumors. N = 220 patients with KRAS or BRAF mutant tumors; N = 276 patients with KRAS or BRAF wildtype tumors. Individual data points are shown and the median with interquartile range is represented. Data was analyzed with an unpaired t-test; **** P < 0.0001. C Kaplan-Meier curve with univariate analysis of the survival of patients with KRAS , NRAS , or BRAF mutated CRC tumors based on high versus low expression of GAD1 (defined as above or below the median GAD1 mRNA expression z-score of 0.3). Data was extracted from the TCGA. N = 254 patients. Data was analyzed using a log-rank (Mantel-Cox test) and a hazard ratio (HR) with 95% confidence interval (CI) is reported. D Kaplan-Meier curve with univariate analysis of the survival of patients with KRAS or BRAF mutated CRC tumors based on high versus low expression of GAD1 (defined as above or below the median GAD1 mRNA expression of 3.4). Data was extracted from the GEO, accession GSE39582 . N = 217 patients. Data was analyzed using a log-rank (Mantel-Cox test) and a hazard ratio (HR) with 95% confidence interval (CI) is reported. All schematics were made in or are from BioRender.

Article Snippet: GAD1 human shRNA lentiviral particles (Santa Cruz Biotechnology, Inc., #sc- sc-35435-V) and control shRNA lentiviral particles (Santa Cruz Biotechnology, Inc., #sc-108080) were used to produce HCT116 H2B-GFP GAD1 knock down cells at a multiplicity of infection (MOI) of 2 and 2 mg/mL of puromycin (Santa Cruz Biotechnology, #sc-108071) was used to select for knocked down cells.

Techniques: Expressing, Mutagenesis, Gene Expression