human pd l1 b7 h1 fc chimera protein  (R&D Systems)

Journal: Cancer Research

Article Title: Nerves Stimulate Cross-talk Between Gastric Cancer and Group 3 Innate Lymphoid Cells to Enhance Immunosuppression

doi: 10.1158/0008-5472.CAN-25-3092

Figure Lengend Snippet: Elevated ABHD16A triggered by ACh was associated with poor prognosis in patients with gastric cancer. A, Representative IHC images of ABHD16A expression in human normal gastric tissues and gastric cancer tissues from the TMA with magnifications of 40×, 100×, and 200×. Scale bar for 200×, 100 μm. B–D, Relationship of ABHD16A IHC score to clinical stage ( B ), pT stage ( C ), and lymph node metastasis ( n = 80; D ). E, Cumulative survival curves of patients with gastric cancer with high or low ABHD16A expression based on the TMA. F, IHC images and score of ABHD16A in S100 + or S100 − gastric cancer tissues. Scale bar, 100 μm. G, Procedure for coculture of DRG neurons and gastric cancer cells. H, Western blotting was used to analyze ABHD16A levels in MFC cells cocultured with DRG neurons or DRG neuron–derived CM. GAPDH served as the loading control. I, Expression of HIF1A and ABHD16A in neurotransmitter [HA, dopamine (DA), 5-HT, norepinephrine (NE), ACh]-treated MFC cells and ACh-treated MGC-803 cells. J, ACh concentration was detected in DRG CM by ELISA. K, Tumor volume of orthotopic gastric cancer tumors with or without vagotomy. L, Representative mIF staining images of FOXP3, CD163, CD11b, CD8, PD-L1, and Pan-CK in gastric cancer tissues with high or low expression of ABHD16A. Scale bar, 100 μm. **, P < 0.01; ***, P < 0.001; ****, P < 0.0001.

Article Snippet: FITC anti-human CD3 (Thermo Fisher Scientific, cat. #11-0038-42, RRID: AB_2043831, 5 μL/1 × 10 6 cells), PE anti-human CD127 (BioLegend, cat. #351304, RRID: AB_10720185, 5 μL/1 × 10 6 cells), Brilliant Violet 421 anti-human CD294 (BioLegend, cat. #350112, RRID: AB_2562468, 5 μL/1 × 10 6 cells), APC anti-human CD117 (BioLegend, cat. #313206, RRID: AB_314985, 5 μL/1 × 10 6 cells), FITC anti-human CD19 (eBioscience, cat. #11-0199-42, RRID: AB_10669461, 5 μL/1 × 10 6 cells), FITC anti-human CD14 (eBioscience, cat. #11-0149-42, RRID: AB_10597597, 5 μL/1 × 10 6 cells), BV650 anti-human CD45 (eBioscience, cat. #416-0459-42, RRID: AB_2925684, 5 μL/1 × 10 6 cells), iFluor 647 anti-Ki67 (HUABIO, cat. #HA720163F, RRID: AB_3072100, 1 μL/1 × 10 6 cells), PE/Cyanine7 anti-human CD274 (Elabscience, cat. #E-AB-F1133H, 5 μL/1 × 10 6 cells), and PE anti-human CD273 (Elabscience, cat. #E-AB-F1175D, 5 μL/1 × 10 6 cells).

Techniques: Expressing, Western Blot, Derivative Assay, Control, Concentration Assay, Enzyme-linked Immunosorbent Assay, Staining

Journal: Cancer Research

Article Title: Nerves Stimulate Cross-talk Between Gastric Cancer and Group 3 Innate Lymphoid Cells to Enhance Immunosuppression

doi: 10.1158/0008-5472.CAN-25-3092

Figure Lengend Snippet: ABHD16A promotes ILC3 aggregation and IL22 release in the immune microenvironment. A, A total of 5 × 10 6 MFC-luc or Abhd16a -knockdown MFC-luc cells were injected into the epidermis of the greater curvature of the stomach in the 615 mice. Tumor growth was monitored weekly through bioluminescence imaging from the first day of MFC cell inoculation. Scale bar, 2.000e+4 – 5.000e + 5 p/s/cm 2 /sr. B and C, Representative images of the excised stomachs ( B ) and the orthotopic tumor volume ( C ) on day 28 after gastric cancer cells were implanted orthotopically ( n = 9 per group). D, H&E images of the stomach, peritoneum, liver, intestine, and lung on day 42 after MFC or Abhd16a -knockdown MFC cells were implanted into 615 mice. Scale bar, 400 μm. E–G, Representative images ( E ), volume ( F ), and weight ( G ) of control and Abhd16a -knockdown subcutaneous tumors ( n = 5 per group). H, IHC images of PD-L1 in control and Abhd16a -knockdown orthotopic and subcutaneous gastric cancer tissues. Scale bar, 500 μm. I, Detection of mouse cytokines in the TIF of orthotopic gastric cancer tissues by proteome profiler mouse XL cytokine array. J, Flow cytometry gating strategy and frequencies of ILC3s and Th17/Th22 cells out of CD45 + cells in control and Abhd16a -knockdown gastric cancer tissues in 615 mice. K, Flow cytometry gating strategy and frequencies of ILC3s out of ILCs in control and Abhd16a -knockdown gastric cancer tissues in Rag1 −/− mice. ***, P < 0.001.

Article Snippet: FITC anti-human CD3 (Thermo Fisher Scientific, cat. #11-0038-42, RRID: AB_2043831, 5 μL/1 × 10 6 cells), PE anti-human CD127 (BioLegend, cat. #351304, RRID: AB_10720185, 5 μL/1 × 10 6 cells), Brilliant Violet 421 anti-human CD294 (BioLegend, cat. #350112, RRID: AB_2562468, 5 μL/1 × 10 6 cells), APC anti-human CD117 (BioLegend, cat. #313206, RRID: AB_314985, 5 μL/1 × 10 6 cells), FITC anti-human CD19 (eBioscience, cat. #11-0199-42, RRID: AB_10669461, 5 μL/1 × 10 6 cells), FITC anti-human CD14 (eBioscience, cat. #11-0149-42, RRID: AB_10597597, 5 μL/1 × 10 6 cells), BV650 anti-human CD45 (eBioscience, cat. #416-0459-42, RRID: AB_2925684, 5 μL/1 × 10 6 cells), iFluor 647 anti-Ki67 (HUABIO, cat. #HA720163F, RRID: AB_3072100, 1 μL/1 × 10 6 cells), PE/Cyanine7 anti-human CD274 (Elabscience, cat. #E-AB-F1133H, 5 μL/1 × 10 6 cells), and PE anti-human CD273 (Elabscience, cat. #E-AB-F1175D, 5 μL/1 × 10 6 cells).

Techniques: Knockdown, Injection, Imaging, Control, Flow Cytometry

Journal: Cancer Research

Article Title: Nerves Stimulate Cross-talk Between Gastric Cancer and Group 3 Innate Lymphoid Cells to Enhance Immunosuppression

doi: 10.1158/0008-5472.CAN-25-3092

Figure Lengend Snippet: High expression of ABHD16A mediates the cumulative release of LysoPS into the TME. A, mIF images of the orthotopic gastric cancer (GC) tissues stained for PD-L1, RORC, and CD3. Yellow, PD-L1 + tumor cells; red, RORC + ILC3s; green, CD3 + cells. Scale bars, 100 μm (left) and 25 μm (right). B and C, Flow cytometry gating strategy and frequencies of ILC3s ( B ) and proportions of ILC3s in total CD45 + cells ( C ) isolated from control and Abhd16a -knockdown orthotopic and subcutaneous gastric cancer tissues ( n = 3 per group). D and E, Flow cytometry gating strategy and frequencies of ILC3s ( D ) and proportions of ILC3s in total CD45 + cells ( E ) derived from the peripheral blood of healthy controls (HC) and patients with gastric cancer ( n = 8 per group). F and G, LC-MS/MS analysis of 18:0 and 18:1 LysoPS in the control and Abhd16a -knockdown orthotopic ( F ) and subcutaneous ( G ) gastric cancer tissues ( n = 3 per group). H–J, ELISA was used to measure levels of LysoPS in gastric cancer cell supernatant ( H ), TIF ( I ), and in vitro tumor culture supernatant ( J ) of control and Abhd16a -knockdown orthotopic and subcutaneous gastric cancer tumors ( n = 3 per group). K, LysoPS levels in the serum of healthy controls and patients with gastric cancer detected by ELISA. *, P < 0.05; **, P < 0.01; ***, P < 0.001.

Article Snippet: FITC anti-human CD3 (Thermo Fisher Scientific, cat. #11-0038-42, RRID: AB_2043831, 5 μL/1 × 10 6 cells), PE anti-human CD127 (BioLegend, cat. #351304, RRID: AB_10720185, 5 μL/1 × 10 6 cells), Brilliant Violet 421 anti-human CD294 (BioLegend, cat. #350112, RRID: AB_2562468, 5 μL/1 × 10 6 cells), APC anti-human CD117 (BioLegend, cat. #313206, RRID: AB_314985, 5 μL/1 × 10 6 cells), FITC anti-human CD19 (eBioscience, cat. #11-0199-42, RRID: AB_10669461, 5 μL/1 × 10 6 cells), FITC anti-human CD14 (eBioscience, cat. #11-0149-42, RRID: AB_10597597, 5 μL/1 × 10 6 cells), BV650 anti-human CD45 (eBioscience, cat. #416-0459-42, RRID: AB_2925684, 5 μL/1 × 10 6 cells), iFluor 647 anti-Ki67 (HUABIO, cat. #HA720163F, RRID: AB_3072100, 1 μL/1 × 10 6 cells), PE/Cyanine7 anti-human CD274 (Elabscience, cat. #E-AB-F1133H, 5 μL/1 × 10 6 cells), and PE anti-human CD273 (Elabscience, cat. #E-AB-F1175D, 5 μL/1 × 10 6 cells).

Techniques: Expressing, Staining, Flow Cytometry, Isolation, Control, Knockdown, Derivative Assay, Liquid Chromatography with Mass Spectroscopy, Enzyme-linked Immunosorbent Assay, In Vitro

Journal: Cancer Research

Article Title: Nerves Stimulate Cross-talk Between Gastric Cancer and Group 3 Innate Lymphoid Cells to Enhance Immunosuppression

doi: 10.1158/0008-5472.CAN-25-3092

Figure Lengend Snippet: IL22 upregulates PD-L1 expression in gastric cancer cells through the UPR IRE1α–XBP1 axis. A, mIF images show the alterations of PD-L1 + tumor cells (purple), CD4 + (green), and CD8 + (red) T cells in orthotopic gastric cancer tumors from control and Abhd16a -knockdown mice following IL22 treatment. Scale bar, 50 μm. B, KEGG pathway enrichment analysis of RNA-seq data of gastric cancer tissues with or without IL22 treatment. C, RT-PCR was used to assess the mRNA expression of key downstream molecules of the UPR branches ( XBP1 , ATF4 , ATF6 ) in control and IL22RA1 -knockdown gastric cancer cells. D, Western blotting analysis of PD-L1 and XBP1s levels in control and IL22RA1 -knockdown MGC-803 cells treated with IL22 (100 μg/L). E, Western blotting detection of PD-L1 and XBP1s levels in XBP1- knockdown MGC-803 cells treated with IL22 and MGC-803 cells treated with IL22 or XBP1s inhibitor (STF083010, 30 μmol/L) in combination with IL22. F, The binding sequence of XBP1 on the CD274 promoter. G and H, ChIP ( G ) and luciferase reporter assay ( H ) showing the transcriptional regulation of CD274 by XBP1s under IL22 stimulation. I, Orthotopic gastric cancer mouse models ( n = 5 per group) were injected with anti-IL22 (200 μg per mouse), anti-CD90.2 antibody (150 μg per mouse), anti-CD90.2 antibody in combination with IL22 (500 ng per mouse), or anti-CD90.2 antibody in combination with XBP1s inhibitors (STF083010, 30 mg/kg) and IL22 for 2 weeks. IHC analysis was used to show IL22, XBP1s, and PD-L1 levels in gastric cancer tissues. Scale bar, 200 μm. J, Tumor volume of orthotopic gastric cancer models under treatments the same as in I . *, P < 0.05; **, P < 0.01; ***, P < 0.001; ns, nonsignificant.

Article Snippet: FITC anti-human CD3 (Thermo Fisher Scientific, cat. #11-0038-42, RRID: AB_2043831, 5 μL/1 × 10 6 cells), PE anti-human CD127 (BioLegend, cat. #351304, RRID: AB_10720185, 5 μL/1 × 10 6 cells), Brilliant Violet 421 anti-human CD294 (BioLegend, cat. #350112, RRID: AB_2562468, 5 μL/1 × 10 6 cells), APC anti-human CD117 (BioLegend, cat. #313206, RRID: AB_314985, 5 μL/1 × 10 6 cells), FITC anti-human CD19 (eBioscience, cat. #11-0199-42, RRID: AB_10669461, 5 μL/1 × 10 6 cells), FITC anti-human CD14 (eBioscience, cat. #11-0149-42, RRID: AB_10597597, 5 μL/1 × 10 6 cells), BV650 anti-human CD45 (eBioscience, cat. #416-0459-42, RRID: AB_2925684, 5 μL/1 × 10 6 cells), iFluor 647 anti-Ki67 (HUABIO, cat. #HA720163F, RRID: AB_3072100, 1 μL/1 × 10 6 cells), PE/Cyanine7 anti-human CD274 (Elabscience, cat. #E-AB-F1133H, 5 μL/1 × 10 6 cells), and PE anti-human CD273 (Elabscience, cat. #E-AB-F1175D, 5 μL/1 × 10 6 cells).

Techniques: Expressing, Control, Knockdown, RNA Sequencing, Reverse Transcription Polymerase Chain Reaction, Western Blot, Binding Assay, Sequencing, Luciferase, Reporter Assay, Injection

Journal: Cancer Research

Article Title: Nerves Stimulate Cross-talk Between Gastric Cancer and Group 3 Innate Lymphoid Cells to Enhance Immunosuppression

doi: 10.1158/0008-5472.CAN-25-3092

Figure Lengend Snippet: Combination therapy enhances the anti–PD-L1 immunotherapeutic effect in gastric cancer. A and B, After tumor formation, the orthotopic gastric cancer mice ( n = 5 per group) were treated with anti–PD-L1 (100 μg per mouse), GPR34 inhibitor (20 mg/kg), or XBP1s inhibitor (30 mg/kg) every 3 days or ACh inhibitor (2.5 mg/kg) daily. Combinations of anti–PD-L1 with each inhibitor followed the every 3-day dosing schedule for a total duration of 2 weeks via i.p. injection. Living images were used to monitor tumor progression at 5-day intervals from the time of drug administration ( A ); IHC and mIF were performed to detect PD-L1 and XBP1s levels and proportions of CD4 + (green) and CD8 + (red) T cells in gastric cancer tissues at the end of treatments ( B ). Scale bars, 1.000e+5 –∼ 5.000e + 5 p/s/cm 2 /sr for living images; 200 μm for IHC; 50 μm for immunofluorescence. C and D, Representative images ( C ) and tumor volume ( D ) of subcutaneous tumors. The administration protocol for the mice was consistent with the description provided in A and B . **, P < 0.01; ***, P < 0.001.

Article Snippet: FITC anti-human CD3 (Thermo Fisher Scientific, cat. #11-0038-42, RRID: AB_2043831, 5 μL/1 × 10 6 cells), PE anti-human CD127 (BioLegend, cat. #351304, RRID: AB_10720185, 5 μL/1 × 10 6 cells), Brilliant Violet 421 anti-human CD294 (BioLegend, cat. #350112, RRID: AB_2562468, 5 μL/1 × 10 6 cells), APC anti-human CD117 (BioLegend, cat. #313206, RRID: AB_314985, 5 μL/1 × 10 6 cells), FITC anti-human CD19 (eBioscience, cat. #11-0199-42, RRID: AB_10669461, 5 μL/1 × 10 6 cells), FITC anti-human CD14 (eBioscience, cat. #11-0149-42, RRID: AB_10597597, 5 μL/1 × 10 6 cells), BV650 anti-human CD45 (eBioscience, cat. #416-0459-42, RRID: AB_2925684, 5 μL/1 × 10 6 cells), iFluor 647 anti-Ki67 (HUABIO, cat. #HA720163F, RRID: AB_3072100, 1 μL/1 × 10 6 cells), PE/Cyanine7 anti-human CD274 (Elabscience, cat. #E-AB-F1133H, 5 μL/1 × 10 6 cells), and PE anti-human CD273 (Elabscience, cat. #E-AB-F1175D, 5 μL/1 × 10 6 cells).

Techniques: Injection, Immunofluorescence