Journal: Acta biochimica et biophysica Sinica

Article Title: Divergent roles of PKM2 in regulating PD-L1 and PD-L2 expression and their implications in human and mouse cancer models.

doi: 10.3724/abbs.2025019

Figure Lengend Snippet: Figure 1. PKM2 is inversely correlated with CD8+ T-cell infiltration in ESCC, as demonstrated by in silico analysis of data from the TCGA database Pearson correlation analysis was used to evaluate the relationship between PKM2 mRNA expression and the expression of immune checkpoint molecules, as well as that of tumor-associated lymphocyte markers. Tpm stands for transcripts per million.

Article Snippet: The membranes were incubated with rabbit polyclonal antibodies against PKM2 (15822-1-AP; Proteintech, Wuhan, China), PD-L1 (28076-1-AP), PD-L2 (18251-1-AP), GATA3 (66400-1-Ig), and βactin (66009-1-Ig).

Techniques: In Silico, Expressing

Journal: Acta biochimica et biophysica Sinica

Article Title: Divergent roles of PKM2 in regulating PD-L1 and PD-L2 expression and their implications in human and mouse cancer models.

doi: 10.3724/abbs.2025019

Figure Lengend Snippet: Figure 2. PKM2 regulates PD-L1 and PD-L2 expression in tumor cells (A) In HeLa cells, compared with controls, PKM2 knockout significantly reduced PD-L1 expression while increasing PD-L2 expression. A similar pattern was observed in the A549 lung cancer cell line, where PKM2 knockout notably increased PD-L2 expression and decreased PD-L1 expression. (B) Additionally, PKM2 down- regulates GATA3 expression in tumor cells. β-Actin served as the internal control. The images shown are representative samples, with band intensity variations quantified via ImageJ software.

Article Snippet: The membranes were incubated with rabbit polyclonal antibodies against PKM2 (15822-1-AP; Proteintech, Wuhan, China), PD-L1 (28076-1-AP), PD-L2 (18251-1-AP), GATA3 (66400-1-Ig), and βactin (66009-1-Ig).

Techniques: Expressing, Knock-Out, Control, Software

Journal: Acta biochimica et biophysica Sinica

Article Title: Divergent roles of PKM2 in regulating PD-L1 and PD-L2 expression and their implications in human and mouse cancer models.

doi: 10.3724/abbs.2025019

Figure Lengend Snippet: Figure 3. PKM2 knockout (KO) significantly suppressed tumor cell proliferation and metastasis in a HeLa cell xenograft mouse model (A) PKM2 knockout drastically inhibited the proliferation of HeLa cells in nude mice xenografted with both HeLa cells and PKM2-knockout HeLa cells (n = 5 per group, **P < 0.01). (B) Similarly, PKM2 knockout markedly reduced lung and liver metastases in nude mice after tail vein injection of PKM2- knockout HeLa cells (n = 5 per group). Representative hematoxylin‒eosin (H&E)-stained sections are shown. Scale bar: 25 μm. (C) Immunohistochemical analysis of PKM2, GATA3, PD-L1, PD-L2, and Ki-67 expression in tumor lesions from xenografted mice with HeLa and PKM2-knockout HeLa cells. Scale bar: 25 μm. Average optical density (AOD) scores were calculated for each section via ImageJ software. Statistical significance was determined via a two-tailed independent sample t test. *P < 0.05, **P < 0.01, ***P < 0.001 relative to the control. (D) Clonogenic assays were performed on A549 and HeLa cells with PKM2 knockout. Statistical analysis was conducted via a two-tailed independent sample t test. ***P < 0.001, ****P < 0.0001 compared with the control.

Article Snippet: The membranes were incubated with rabbit polyclonal antibodies against PKM2 (15822-1-AP; Proteintech, Wuhan, China), PD-L1 (28076-1-AP), PD-L2 (18251-1-AP), GATA3 (66400-1-Ig), and βactin (66009-1-Ig).

Techniques: Knock-Out, Injection, Staining, Immunohistochemical staining, Expressing, Software, Two Tailed Test, Control

Journal: Acta biochimica et biophysica Sinica

Article Title: Divergent roles of PKM2 in regulating PD-L1 and PD-L2 expression and their implications in human and mouse cancer models.

doi: 10.3724/abbs.2025019

Figure Lengend Snippet: Figure 5. GATA3 overexpression (OE) in PKM2-knockout A549 and HeLa cells restores effects similar to those observed with PKM2, enhancing PD-L1 and reducing PD-L2 expression (A) Overexpression of GATA3 in PKM2-knockout A549 and HeLa cells reinstates the original PKM2- induced regulation, where PD-L1 expression is increased and PD-L2 expression is decreased. (B) GATA3 overexpression also restored the migratory capacity of A549 and HeLa cells following PKM2 knockout. (C) Similarly, GATA3 overexpression re-established the invasive properties of A549 and HeLa cells after PKM2 knockout. Statistical analysis was performed via a two-tailed independent sample t test. *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001 compared with the control.

Article Snippet: The membranes were incubated with rabbit polyclonal antibodies against PKM2 (15822-1-AP; Proteintech, Wuhan, China), PD-L1 (28076-1-AP), PD-L2 (18251-1-AP), GATA3 (66400-1-Ig), and βactin (66009-1-Ig).

Techniques: Over Expression, Knock-Out, Expressing, Two Tailed Test, Control

Journal: Acta biochimica et biophysica Sinica

Article Title: Divergent roles of PKM2 in regulating PD-L1 and PD-L2 expression and their implications in human and mouse cancer models.

doi: 10.3724/abbs.2025019

Figure Lengend Snippet: Figure 6. PKM2 is inversely correlated with CD8+ T-cell infiltration in a xenograft model in C57BL/6N mice (A) Evaluation of lentiviral shRNA- mediated PKM2 knockdown efficiency at the mRNA level via qRT-PCR. (B) Confirmation of the knockdown efficiency via western blot analysis. (C) Detection of variations in PD-L1 and PD-L2 expression in AKR cells following PKM2 knockdown, as demonstrated by western blot analysis. (D) Development of a xenograft mouse model using the C57BL/6N strain. (E) Macroscopic examination of subcutaneous tumor growth in the PKM2-shRNA2 (n = 5) and control (n = 5) groups. (F) Quantitative analysis of tumor weight. (G) Representative immunohistochemistry images and corresponding quantifications showing differences in the expressions of GATA3, PKM2, PD-L1, PD-L2, CD206, and CD8+ T cells in tumor tissues from xenograft C57BL/6 N mice. Scale bar: 25 μm, magnification, 200×. Average optical density (AOD) scores were calculated for each section via ImageJ software. Differences between groups were analyzed via an independent sample t test. *P < 0.05, **P < 0.01, and ***P < 0.001 compared with the control group.

Article Snippet: The membranes were incubated with rabbit polyclonal antibodies against PKM2 (15822-1-AP; Proteintech, Wuhan, China), PD-L1 (28076-1-AP), PD-L2 (18251-1-AP), GATA3 (66400-1-Ig), and βactin (66009-1-Ig).

Techniques: shRNA, Knockdown, Quantitative RT-PCR, Western Blot, Expressing, Control, Immunohistochemistry, Software

Journal: Acta biochimica et biophysica Sinica

Article Title: Divergent roles of PKM2 in regulating PD-L1 and PD-L2 expression and their implications in human and mouse cancer models.

doi: 10.3724/abbs.2025019

Figure Lengend Snippet: Figure 7. Schematic diagram highlighting the species-dependent regulation of PKM2 In a human model, PKM2 enhances PD-L1 expression while suppressing PD-L2 expression, facilitating immune evasion through PD-L1 upregulation. Conversely, in a mouse model, PKM2 suppresses both PD-L1 and PD-L2 expression, leading to increased CD8+ T-cell infiltration upon PKM2 knockdown. KO, knockout; KD, knockdown; OE, overexpression. The blue arrow indicates downregulation; the red arrow indicates upregulation.

Article Snippet: The membranes were incubated with rabbit polyclonal antibodies against PKM2 (15822-1-AP; Proteintech, Wuhan, China), PD-L1 (28076-1-AP), PD-L2 (18251-1-AP), GATA3 (66400-1-Ig), and βactin (66009-1-Ig).

Techniques: Expressing, Knockdown, Knock-Out, Over Expression

Journal: Frontiers in Cell and Developmental Biology

Article Title: Genome-wide analysis reveals the MORC3-mediated repression of PD-L1 expression in head and neck cancer

doi: 10.3389/fcell.2024.1410130

Figure Lengend Snippet: MORC3 represses PD-L1 expression. (A) The expression levels of STAT1 and PD-L1 in CAL 27 cells with or without MORC3 knockdown were analyzed by qRT-PCR. (B) The expression levels of STAT1 and PD-L1 in CAL 27 cells with or without MORC3 overexpression were analyzed by qRT-PCR. *, p < 0.05. (C) The correlation between the expression levels of MORC3 and STAT1 and PD-L1 in single cells of OSCC were analyzed by reanalyzing the expression data of GSE10332 in the database of CancerSEA. (D) The expression levels of lncRNA LINC00880 and LINC02574 in CAL 27 cells with or without MORC3 knockdown were analyzed by qRT-PCR. (E) qRT-PCR analysis of LINC00880 expression in CAL 27 cells with or without MORC3 knockdown. (F) qRT-PCR analysis of the efficiency of LINC00880 knockdown by siRNA. Si0880 is a siRNA against LINC00880. (G) The expression level of PD-L1 in CAL 27 cells with or without LINC00880 knockdown was analyzed by qRT-PCR. *, p < 0.05. (H) The expression levels of LINC00880 in TCGA HNSCC were analyzed in OncoDB database.

Article Snippet: The membranes were blocked with 5% skimmed milk and incubated overnight at 4°C with following antibodies: mouse monoclonal anti-MORC3 antibody (Santa Cruz Biotechnology, United States), rabbit monoclonal anti- interferon induced protein with tetratricopeptide repeats 2 (IFIT2) antibody (abcam, United States), rabbit monoclonal anti-interferon induced transmembrane protein 3 (IFITM3) antibody (Cell Signaling Technology, United States), rabbit monoclonal anti-interferon induced protein with tetratricopeptide repeats 1 (IFIT1) antibody (Cell Signaling Technology, United States), rabbit monoclonal anti-cyclin D1 (CCND1) (ABclonal, China), rabbit monoclonal anti-cyclinD2 (CCND2) (ABclonal, China), rabbit polyclonal anti-JUN (ABclonal, China), rabbit polyclonal anti-interferon regulatory factor 7 (IRF7) (Proteintech, China), rabbit polyclonal anti-PD-L1 antibody (Proteintech, China), rabbit polyclonal anti-DExD/H-Box Helicase 60 (DDX60) antibody (Proteintech, China), and mouse anti-β-actin (Abmart, China).

Techniques: Expressing, Knockdown, Quantitative RT-PCR, Over Expression