Journal: Cancer Immunology, Immunotherapy : CII

Article Title: Immunological synapse formation as a key mechanism in T cell-dependent bispecific antibody-mediated immune activation and cytotoxicity

doi: 10.1007/s00262-025-04036-w

Figure Lengend Snippet: Cytotoxic activity and inflammatory cytokine release in co-culture systems. a–c After co-culturing WT or KO, T cells, and TDB, the color reaction with WST-8 reagent in an hour was measured. The co-culture times were 24 h, 48 h, and 72 h of co-culture, respectively. Red: WT + T cell + TDB, blue: KO + T cell + TDB. Error bars represent the mean ± SD ( n = 6), ** p < 0.005, *** p < 0.0005, n.s. not significant. d Measurement of IFN-γ release by ELISA after 24 h of co-culture time. Co-culture supernatants were obtained and used for assay using the ELISA Kit. Red: WT + T cell + TDB, blue: KO + T cell + TDB, gray: negative control (T cell + TDB). Red letters are the results of the WT and N/C statistical tests, and blue letters are the results of the KO and N/C statistical tests. Error bars represent the mean ± SD ( n = 3), *** p < 0.0005, n.s. not significant. e IFN-γ release at TDB: 10 ng/mL for each co-culture time. Error bars represent the mean ± SD ( n ≧ 3), *** p < 0.0005, n.s. not significant. f Cancer cell death when the IFN-γ release measured in e is added to the standard. From left to right, IFN-γ release at 6 h, 24 h, 48 h, and 72 h was added. Error bars represent the mean ± SD ( n ≧ 3), *** p < 0.0005, n.s. not significant. g-i TNF-α version of the same experiment as d – f

Article Snippet: T cells (E:T = 5:1, 1.0 × 10 6 cells/50 μL/well) were added and co-cultured. hEx3 was added at a final concentration of 0.1–100 ng/mL in 100 μL of medium and cultured for 24 h. The culture supernatant was then collected and assayed using human IFN-gamma ELISA Kit, human TNF-alpha ELISA Kit, human CXCL10 ELISA Kit (Proteintech, Rosemont, IL), and human HMGB1 ELISA Kit (Chondrex, Woodinville, WA) according to the protocols provided.

Techniques: Activity Assay, Co-Culture Assay, Enzyme-linked Immunosorbent Assay, Negative Control

Journal: Cell reports. Medicine

Article Title: Targeting neoadjuvant chemotherapy-induced metabolic reprogramming in pancreatic cancer promotes anti-tumor immunity and chemo-response.

doi: 10.1016/j.xcrm.2023.101234

Figure Lengend Snippet: Figure 6. Targeting CD36 synergistically promoted AG-mediated killing of PDAC in preclinical models (A) Visual presentation of subcutaneous xenograft murine PDAC tumor models (C57 mice) for each group. (B) Measurement of tumor volumes showed CD36 blockage synergistically promoted AG-mediated killing of PDAC in subcutaneous xenograft murine PDAC tumor models. (C) Measurement of tumor weights showed CD36 blockage synergistically promoted AG-mediated killing of PDAC in subcutaneous xenograft murine PDAC tumor models (n = 5). (D) Representative IHC staining showed Ki67 expression in subcutaneous xenografts treated with different regimens. (E) t-Distributed stochastic neighbor embedding (TSNE) analyses showed the clustering for CD36+ CD8+ T cells and GZMB+ CD8+ T cells. (F) Flow cytometry revealed that more CD8+ T cells infiltrated PDAC with NAC, while the percentage of CD36+ CD8+ T cells also increased (n = 5) (mean with standard deviation). (G) ELISA results showed the combination of AG and CD36 blockade significantly improved IFN-g and tumor necrosis factor a (TNF-a) levels intratumorally (n = 5). (H) Representative image of orthotopic murine models of PDAC. (I) Kaplan-Meier curve revealed the combination of CD36 blockade and AG significantly prolonged the survival interval of mice that received orthotopic PDAC cell transplantation (n = 10). Circle or square referred to a happened event (death or censored). Censored event means the mice is still alive at the time point that we ended follow-up. (J) CD36 blockade synergistically with AG regimens optimally narrowed the PDAC tumor size in a humanized PDX model (n = 10). (K) Representative IHC staining image of CD36-high and -low PDAC. (L) Kaplan-Meier curve showed increased CD36 expression predicted worse prognosis of PDAC patients with adjuvant AG chemotherapy. The statistical sig- nificance shown in this figure was detected using t test.

Article Snippet: The ELISA kits used in the present study were as follows: Human IFN-gamma ELISA Kit (absin, abs510007); Human IL-2 ELISA Kit (Boster, EK0397); Mouse TNF alpha ELISA Kit (absin, abs520010) and Mouse IFN- gamma ELISA Kit (absin, abs520007).

Techniques: Immunohistochemistry, Expressing, Flow Cytometry, Standard Deviation, Enzyme-linked Immunosorbent Assay, Transplantation Assay, Adjuvant

Journal: Analytical Cellular Pathology (Amsterdam)

Article Title: Exosomal PDL1 Suppresses the Anticancer Activity of CD8 + T Cells in Hepatocellular Carcinoma

doi: 10.1155/2024/1608582

Figure Lengend Snippet: Regulatory effect of exosomal PDL1 on proliferation of CD8 + T and hepatic cancer cells. (A–D): qRT-PCR (A and C) and western blot (B and D) for PDL1 level in Huh7 cells with the overexpression or knockdown of PDL1. (E) Western blot for exosomal PDL1 level in supernatants of PDL1-silenced Huh7 cells. (F and G): TEM (F) and NTA (G) for isolated exosomes derived from the supernatants of PDL1-silenced Huh7 cells. (H) Proliferation of Huh7 cells with the overexpression or silencing of PDL1, as assayed by CCK8. (I) Proliferation of CD8 + T cells with the treatments of exosomes derived from the supernatants of Huh7 cells, as assayed by CCK8. PDL1 was overexpressed or silenced in Huh7 cells. (J) ELISA for TNF α and IFN- γ levels in supernatants of human CD8 + T cells after treatments with exosomes. Data were analyzed by Student's two-sided t -test and are represented as mean ± SD. ∗∗ p <0.01, ∗∗∗ p <0.001.

Article Snippet: ELISA plates (96-well) (Biolegend) that coated with Human PDL1 antibody (DY156, R&D SYSTEMS, Minneapolis, MN, USA), Human TNF α ELISA Kit (KIT10602, SinoBiology, Beijing, China), Mouse TNF α Matched ELISA Antibody Pair Set (SEKA50349, SinoBiology), Human IFN gamma ELISA Kit (KIT11725A, SinoBiology), and Mouse IFN gamma ELISA Kit (KIT50709, SinoBiology) were used for detecting protein levels of PDL1, TFN α , and IFN- γ in samples according to manufactures' instructions.

Techniques: Quantitative RT-PCR, Western Blot, Over Expression, Knockdown, Isolation, Derivative Assay, Enzyme-linked Immunosorbent Assay

Journal: Analytical Cellular Pathology (Amsterdam)

Article Title: Exosomal PDL1 Suppresses the Anticancer Activity of CD8 + T Cells in Hepatocellular Carcinoma

doi: 10.1155/2024/1608582

Figure Lengend Snippet: Isolation of exosomes derived from the supernatants of cultured Hepa1–6 cells with different levels of PDL1. (A–D): qRT-PCR (A and C) and western blot (B and D) for PDL1 level in Hepa1–6 cells with the overexpression or knockdown of PDL1. (E) Western blot for protein level of PDL1, TSG101, and GAPDH in exosomes derived from the supernatants of PDL1-silenced Hepa1–6 cells. (F and G) TEM and NTA analysis for the isolated exosomes derived from the supernatants of Pdl1-silenced Hepa1–6 cells. (H) Proliferation of Hepa1–6 cells with the overexpression or silencing of Pdl1, as assayed by CCK8. (I) Proliferation of CD8 + T cells after treatments of exosomes derived from the supernatants of Hepa1–6 cells, as assayed by CCK8. Pdl1 was overexpressed or silenced in Hepa1–6 cells. (J) ELISA for TNF α and IFN- γ levels in supernatants of mice CD8 + T cells after treatments with exosomes. Data were analyzed by Student's two-sided t -test and are represented as mean ± SD. ∗∗∗ p <0.001.

Article Snippet: ELISA plates (96-well) (Biolegend) that coated with Human PDL1 antibody (DY156, R&D SYSTEMS, Minneapolis, MN, USA), Human TNF α ELISA Kit (KIT10602, SinoBiology, Beijing, China), Mouse TNF α Matched ELISA Antibody Pair Set (SEKA50349, SinoBiology), Human IFN gamma ELISA Kit (KIT11725A, SinoBiology), and Mouse IFN gamma ELISA Kit (KIT50709, SinoBiology) were used for detecting protein levels of PDL1, TFN α , and IFN- γ in samples according to manufactures' instructions.

Techniques: Isolation, Derivative Assay, Cell Culture, Quantitative RT-PCR, Western Blot, Over Expression, Knockdown, Enzyme-linked Immunosorbent Assay

Journal: Cell Reports Medicine

Article Title: Immunomodulatory effects and improved outcomes with cisplatin- versus carboplatin-based chemotherapy plus atezolizumab in urothelial cancer

doi: 10.1016/j.xcrm.2024.101393

Figure Lengend Snippet:

Article Snippet: Human IFNg ELISA kit , BD Biosciences , Cat.# 555142.

Techniques: Blocking Assay, Recombinant, Cell Isolation, Activation Assay, Enzyme-linked Immunosorbent Assay, Staining, Software