mkn 45  (ATCC)

Journal: bioRxiv

Article Title: Patchouli alcohol suppresses gastric cancer growth and immune evasion via inhibition of the NF-κB/PD-L1 axis

doi: 10.64898/2026.03.17.712304

Figure Lengend Snippet: Inhibitory effects of PA on the viability of gastric cancer cells and the expression of PD-L1 and NF-κB. (A) mRNA expression levels of NF-κB and PD-L1 in human gastric mucosal cells and gastric cancer cell lines. (B) Viability of the gastric cancer cell lines HGC-27 and MKN-45 treated with increasing concentrations of PA (0 mM, 0.1 mM, 0.25 mM, 0.5 mM, 1 mM, 10 mM, 20 mM and 50 mM) for 24 hours, 48 hours, and 72 hours. (C) Effect of PA treatment on the mRNA expression of NF-κB and PD-L1 in HGC-27 and MKN-45 cells. (D) Effect of PA treatment on the protein expression of NF-κB and PD-L1 in HGC-27 and MKN-45 cells. * P < 0.05, ** P < 0.01, *** P < 0.001; n=3 independent experiments.

Article Snippet: SureScript TM First-Strand cDNA Synthesis Kit (Cat: QP056T; GeneCopeia, USA), 2×SYBR Green qPCR Master Mix (None ROX) (Cat: G3320-05; Servicebio, China), TriQuick Reagent (Cat: R1100, Solarbio, China), MKN-45 (CL-0292, Procell, China), HGC-27 (CL-0107, Procell, China), GES-1 (CL-0563, Procell, China), Cell Counting Kit-8 (Cat: G4103, Servicebio, China), Patchouli alcohol (Cat: MUST-24053118, Purity: 99.63%, Must Bio-Technology, China), Multiskan FC Microplate Photometer (DNM-9602, Perlang, China), inverted microscope (OPTOP OD630K, Sunny Hengpin, China), chemiluminescence imaging system (ChemiScope6100, Clin, China), microplate reader (RMR-2208LZN; Ranhui, China), Invitrogen Power Blotter (Model PB0010; Thermo Fisher Scientific, China), anti-NF-kB p65 antibody (ab16502; abcam, UK), NF-κB p65 polyclonal antibody (Cat: 10745-1-AP, Proteintech, China), recombinant anti-PD-L1 antibody [EPR19759] (ab213524, Abcam, UK), PD-L1/CD274 (C-terminal) polyclonal antibody (Cat: 28076-1-AP; Proteintech, China), primary and secondary antibody diluent for immunostaining (Cat: 36206ES76; YEASEN, China), goat anti-rabbit IgG H&L (Alexa Fluor ® 488) (ab150077; abcam, UK), fluorescence microscope (BH2-RFCA; OLYMPUS, Japan), Annexin V-FITC/PI apoptosis kit (Cat: AP101-100-kit, MultiSciences, China), BD FACSCalibur TM Flow Cytometer (E97501093, BD Biosciences, USA), Human Tumor Necrosis Factor Alpha (TNF-α) ELISA Kit (JL10208, Jianglai Biological, China), Human Interferon (IFN-γ) detection kit (JL12152, Jianglai Biological, China), TUNEL Apoptosis Detection Kit-FITC (50T) (MK1027, BOSTER, China), Dual Luciferase Reporter Gene Assay Kit (Cat: RG009, Beyotime, China), Plasmid Mini Preparation Kit (BioDev-Tech, China), pmirGLO (Honeke, China), restriction endonuclease (Thermo, USA), and GloMax ® 96 Microplate Luminometer (Promega, USA).

Techniques: Expressing

Journal: bioRxiv

Article Title: Patchouli alcohol suppresses gastric cancer growth and immune evasion via inhibition of the NF-κB/PD-L1 axis

doi: 10.64898/2026.03.17.712304

Figure Lengend Snippet: PA pretreatment sensitizes gastric cancer cells to PBMC-mediated death through the activation of CD8⁺ T cells. (A) Apoptosis of HGC-27 and MKN-45 cells analyzed by flow cytometry. (B) Analysis of the cytotoxic activity of PBMCs against gastric cancer cells by LDH release assays and concentrations of IFN-γ, TNF-α, and granzyme B in the coculture supernatant measured by ELISA. (C, D) Immunophenotypic analysis of the cocultured PBMCs. Flow cytometry was used to quantify the proportions of CD3⁺CD8⁺ T cells (C) and IFN-γ–producing CD8⁺ (IFN-γ⁺CD8⁺) T cells (D) . * P < 0.05, ** P < 0.01, *** P < 0.001, vs. the control group; n=3 independent experiments.

Article Snippet: SureScript TM First-Strand cDNA Synthesis Kit (Cat: QP056T; GeneCopeia, USA), 2×SYBR Green qPCR Master Mix (None ROX) (Cat: G3320-05; Servicebio, China), TriQuick Reagent (Cat: R1100, Solarbio, China), MKN-45 (CL-0292, Procell, China), HGC-27 (CL-0107, Procell, China), GES-1 (CL-0563, Procell, China), Cell Counting Kit-8 (Cat: G4103, Servicebio, China), Patchouli alcohol (Cat: MUST-24053118, Purity: 99.63%, Must Bio-Technology, China), Multiskan FC Microplate Photometer (DNM-9602, Perlang, China), inverted microscope (OPTOP OD630K, Sunny Hengpin, China), chemiluminescence imaging system (ChemiScope6100, Clin, China), microplate reader (RMR-2208LZN; Ranhui, China), Invitrogen Power Blotter (Model PB0010; Thermo Fisher Scientific, China), anti-NF-kB p65 antibody (ab16502; abcam, UK), NF-κB p65 polyclonal antibody (Cat: 10745-1-AP, Proteintech, China), recombinant anti-PD-L1 antibody [EPR19759] (ab213524, Abcam, UK), PD-L1/CD274 (C-terminal) polyclonal antibody (Cat: 28076-1-AP; Proteintech, China), primary and secondary antibody diluent for immunostaining (Cat: 36206ES76; YEASEN, China), goat anti-rabbit IgG H&L (Alexa Fluor ® 488) (ab150077; abcam, UK), fluorescence microscope (BH2-RFCA; OLYMPUS, Japan), Annexin V-FITC/PI apoptosis kit (Cat: AP101-100-kit, MultiSciences, China), BD FACSCalibur TM Flow Cytometer (E97501093, BD Biosciences, USA), Human Tumor Necrosis Factor Alpha (TNF-α) ELISA Kit (JL10208, Jianglai Biological, China), Human Interferon (IFN-γ) detection kit (JL12152, Jianglai Biological, China), TUNEL Apoptosis Detection Kit-FITC (50T) (MK1027, BOSTER, China), Dual Luciferase Reporter Gene Assay Kit (Cat: RG009, Beyotime, China), Plasmid Mini Preparation Kit (BioDev-Tech, China), pmirGLO (Honeke, China), restriction endonuclease (Thermo, USA), and GloMax ® 96 Microplate Luminometer (Promega, USA).

Techniques: Activation Assay, Flow Cytometry, Activity Assay, Enzyme-linked Immunosorbent Assay, Control

Journal: bioRxiv

Article Title: Patchouli alcohol suppresses gastric cancer growth and immune evasion via inhibition of the NF-κB/PD-L1 axis

doi: 10.64898/2026.03.17.712304

Figure Lengend Snippet: Patchouli alcohol suppresses NF-κB-induced PD-L1 upregulation in gastric cancer cells. (A) western blotting analysis of AKT, NF-κB, and PD-L1 protein expression in HGC-27 and MKN-45 cells treated as indicated. (B) RT‒qPCR analysis of NF-κB mRNA expression in HGC-27 and MKN-45 cells under various treatments. (C, D) Immunofluorescence staining was used to measure the levels of NF-κB in HGC-27 (C) and MKN-45 (D) cells; OE-NF-κB: overexpression of NF-κB; shR-NF-κB: shRNA targeting NF-κB. Scale bar, 100 μm. * P < 0.05, ** P < 0.01, *** P < 0.001, vs. the control group; # P < 0.05, ## P < 0.01, ### P < 0.001, vs. the OE-NF-κB+0.5 mM PA group; & P < 0.05, &&& P < 0.001, vs. the OE-NF-κB group; n=3 independent experiments.

Article Snippet: SureScript TM First-Strand cDNA Synthesis Kit (Cat: QP056T; GeneCopeia, USA), 2×SYBR Green qPCR Master Mix (None ROX) (Cat: G3320-05; Servicebio, China), TriQuick Reagent (Cat: R1100, Solarbio, China), MKN-45 (CL-0292, Procell, China), HGC-27 (CL-0107, Procell, China), GES-1 (CL-0563, Procell, China), Cell Counting Kit-8 (Cat: G4103, Servicebio, China), Patchouli alcohol (Cat: MUST-24053118, Purity: 99.63%, Must Bio-Technology, China), Multiskan FC Microplate Photometer (DNM-9602, Perlang, China), inverted microscope (OPTOP OD630K, Sunny Hengpin, China), chemiluminescence imaging system (ChemiScope6100, Clin, China), microplate reader (RMR-2208LZN; Ranhui, China), Invitrogen Power Blotter (Model PB0010; Thermo Fisher Scientific, China), anti-NF-kB p65 antibody (ab16502; abcam, UK), NF-κB p65 polyclonal antibody (Cat: 10745-1-AP, Proteintech, China), recombinant anti-PD-L1 antibody [EPR19759] (ab213524, Abcam, UK), PD-L1/CD274 (C-terminal) polyclonal antibody (Cat: 28076-1-AP; Proteintech, China), primary and secondary antibody diluent for immunostaining (Cat: 36206ES76; YEASEN, China), goat anti-rabbit IgG H&L (Alexa Fluor ® 488) (ab150077; abcam, UK), fluorescence microscope (BH2-RFCA; OLYMPUS, Japan), Annexin V-FITC/PI apoptosis kit (Cat: AP101-100-kit, MultiSciences, China), BD FACSCalibur TM Flow Cytometer (E97501093, BD Biosciences, USA), Human Tumor Necrosis Factor Alpha (TNF-α) ELISA Kit (JL10208, Jianglai Biological, China), Human Interferon (IFN-γ) detection kit (JL12152, Jianglai Biological, China), TUNEL Apoptosis Detection Kit-FITC (50T) (MK1027, BOSTER, China), Dual Luciferase Reporter Gene Assay Kit (Cat: RG009, Beyotime, China), Plasmid Mini Preparation Kit (BioDev-Tech, China), pmirGLO (Honeke, China), restriction endonuclease (Thermo, USA), and GloMax ® 96 Microplate Luminometer (Promega, USA).

Techniques: Western Blot, Expressing, Immunofluorescence, Staining, Over Expression, shRNA, Control

Journal: bioRxiv

Article Title: Patchouli alcohol suppresses gastric cancer growth and immune evasion via inhibition of the NF-κB/PD-L1 axis

doi: 10.64898/2026.03.17.712304

Figure Lengend Snippet: NF-κB directly regulates PD-L1 promoter activity in gastric cancer cells. (A, B) Validation of NF-κB overexpression or knockdown in MKN-45 cells and HGC-27 cells. (C) Luciferase reporter assays in HGC-27 cells. (D) Luciferase reporter assays in MKN-45 cells. OE-NF-κB: overexpression of NF-κB; OE-NC: overexpression negative control; shR-NF-κB: shRNA targeting NF-κB; shR-NC: shRNA negative control. *** P < 0.001, * P < 0.05, vs. the OE-NC group; # P < 0.05, ## P < 0.01, vs. the shR-NC group; n=3 independent experiments.

Article Snippet: SureScript TM First-Strand cDNA Synthesis Kit (Cat: QP056T; GeneCopeia, USA), 2×SYBR Green qPCR Master Mix (None ROX) (Cat: G3320-05; Servicebio, China), TriQuick Reagent (Cat: R1100, Solarbio, China), MKN-45 (CL-0292, Procell, China), HGC-27 (CL-0107, Procell, China), GES-1 (CL-0563, Procell, China), Cell Counting Kit-8 (Cat: G4103, Servicebio, China), Patchouli alcohol (Cat: MUST-24053118, Purity: 99.63%, Must Bio-Technology, China), Multiskan FC Microplate Photometer (DNM-9602, Perlang, China), inverted microscope (OPTOP OD630K, Sunny Hengpin, China), chemiluminescence imaging system (ChemiScope6100, Clin, China), microplate reader (RMR-2208LZN; Ranhui, China), Invitrogen Power Blotter (Model PB0010; Thermo Fisher Scientific, China), anti-NF-kB p65 antibody (ab16502; abcam, UK), NF-κB p65 polyclonal antibody (Cat: 10745-1-AP, Proteintech, China), recombinant anti-PD-L1 antibody [EPR19759] (ab213524, Abcam, UK), PD-L1/CD274 (C-terminal) polyclonal antibody (Cat: 28076-1-AP; Proteintech, China), primary and secondary antibody diluent for immunostaining (Cat: 36206ES76; YEASEN, China), goat anti-rabbit IgG H&L (Alexa Fluor ® 488) (ab150077; abcam, UK), fluorescence microscope (BH2-RFCA; OLYMPUS, Japan), Annexin V-FITC/PI apoptosis kit (Cat: AP101-100-kit, MultiSciences, China), BD FACSCalibur TM Flow Cytometer (E97501093, BD Biosciences, USA), Human Tumor Necrosis Factor Alpha (TNF-α) ELISA Kit (JL10208, Jianglai Biological, China), Human Interferon (IFN-γ) detection kit (JL12152, Jianglai Biological, China), TUNEL Apoptosis Detection Kit-FITC (50T) (MK1027, BOSTER, China), Dual Luciferase Reporter Gene Assay Kit (Cat: RG009, Beyotime, China), Plasmid Mini Preparation Kit (BioDev-Tech, China), pmirGLO (Honeke, China), restriction endonuclease (Thermo, USA), and GloMax ® 96 Microplate Luminometer (Promega, USA).

Techniques: Activity Assay, Biomarker Discovery, Over Expression, Knockdown, Luciferase, Negative Control, shRNA