Fig 2: Effect of SF and HD EOs on TNF-α, IL-2, and IL-6 production by LPS-activated RAW 264.7. The results are stated as the means ± SD of three replicates. a, b, c & d, significant (p < 0.05) compared to untreated cells, LPS-only treated cells, quercetin, or SF

Fig 3: CRTC1 in LLC cells inhibits T-cell proliferation and cytotoxicity via Notch1/Akt. (A) Western blot analysis of CRTC1, Notch1, and p-Akt expression in LLC cells transfected with plasmids knocking down CRTC1 (si-CRTC1) and overexpressing Notch1 (oe-Notch1) or their negative controls (si-NC or oe-NC), followed by co-culture with activated CTLL-2 cells (1:10 ratio, 24 h). (B) CCK-8 assay assessing LLC cell viability. (C-D) Flow cytometry analysis of apoptosis in LLC cells. (E) CCK-8 assay measuring T-cell viability. (F) Flow cytometry analysis of apoptosis in T cells. (G) Western blot analysis of PD-L1, CXCL10, and CXCL11 expression in co-cultures. (H) ELISA detecting CXCL10 and CXCL11 secretion in supernatants. (I) ELISA measuring IFN-γ and IL-2 levels in supernatants. (J) LDH release assay evaluating T-cell cytotoxicity. n = 3. The p-values in panels A-K were calculated using ANOVA with Tukey’s post hoc test. *p < 0.05 vs. Control. #p < 0.05 vs. T cells + si-NC. &p < 0.05 vs. T cells + si-CRTC1 + oe-NC.

Fig 4: CRTC1 knockdown synergizes with PD-L1 blockade to suppress tumor growth in vivo. (A) Western blot analysis of CRTC1, Notch1, and p-Akt expression in Lewis xenograft tumors from C57BL/6J mice treated with Atezolizumab combined with plasmids knocking down CRTC1 (si-CRTC1) and overexpressing Notch1 (oe-Notch1) or their negative controls (si-NC or oe-NC). (B) Tumor size in mice bearing Lewis xenografts treated with Atezolizumab, CRTC1 knockdown, and Notch1 overexpression. (C) Tumor volume and weight of Lewis xenografts under combined treatment. (D) Representative H&E staining images of Lewis xenografts post-treatment. Scale bar = 100 μm (top) and 25 μm (bottom). (E) IHC analysis of PD-L1 expression in Lewis xenograft tumors. Scale bar = 100 μm (top) and 25 μm (bottom). (F) Western blot analysis of CXCL10 and CXCL11 expression in tumors. (G) Flow cytometry analysis of CD3+ T-cell infiltration in tumors. (H) ELISA measuring serum IFN-γ and IL-2 levels. n = 5. The p-values in panels (A, C, E–H) were calculated using ANOVA with Tukey’s post hoc test. *p < 0.05 vs. Model. #p < 0.05 vs. Atezolizumab + si-NC. &p < 0.05 vs. Atezolizumab + si-CRTC1 + oe-NC.

Fig 5: CRTC1 in LLC cells suppresses T-cell proliferation and cytotoxicity. (A) Western blot analysis of CRTC1 expression in LLC cells transfected with plasmid overexpressing (oe-CRTC1) or its negative control (oe-NC) and co-cultured with activated CTLL-2 cells at a 1:10 ratio for 24 h (B) CCK-8 assay measuring viability of LLC cells. (C) Flow cytometry analysis of apoptosis in LLC cells. (D) CCK-8 assay assessing T-cell viability. (E-F) Flow cytometry analysis of apoptosis in T cells. (G) Western blot analysis of PD-L1, CXCL10, and CXCL11 expression in co-cultures. (H) ELISA detecting CXCL10 and CXCL11 secretion in supernatants. (I) ELISA measuring IFN-γ and IL-2 levels in supernatants. (J) LDH release assay evaluating T-cell cytotoxicity via supernatant LDH concentration. n = 3. The p-values in panels (A–J) were calculated using ANOVA with Tukey’s post hoc test. *p < 0.05 vs. Control. #p < 0.05 vs. T cells + oe-NC.