Journal: Stem Cell Research & Therapy

Article Title: Mesenchymal stem cells combined with liraglutide relieve acute lung injury through apoptotic signaling restrained by PKA/β-catenin

doi: 10.1186/s13287-020-01689-5

Figure Lengend Snippet: Liraglutide reduces apoptosis of hCMSCs via PKA/β-catenin pathway. a Western blot and b RT-qPCR verify the knockdown effects of three Si-GLP-1R in hCMSCs. c , d Western blot was used to detect of β-catenin and p-β-catenin expression under the stimulation of LPS by adding 20 μM H89 or 100 nM Si-GLP-1R and liraglutide. e The expression of apoptotic proteins Bax, Bcl-2, cleaved caspase-9, and cleaved caspase-3 was detected by western blot with PKA inhibitor H89 and liraglutide. f The expression of GLP-1R and apoptotic proteins Bax, Bcl-2, cleaved caspase-9, and cleaved caspase-3 were detected by western blot with Si-GLP-1R and liraglutide. Error bars represent mean ± SD from three independent experiments. Compared with Si-con group, *** P < 0.001

Article Snippet: To verify the PKA/β-catenin pathway, MSCs were exposed to 10 nM liraglutide or 100 nM SiRNA or 20 μM PKA inhibitor H89 (MCE, Cat. No. 130964-39-5, USA) on the premise of exposure to 30 μg/mL LPS.

Techniques: Western Blot, Quantitative RT-PCR, Knockdown, Expressing

Journal: Cell reports

Article Title: Retrograde adenosine/A 2A receptor signaling facilitates excitatory synaptic transmission and seizures

doi: 10.1016/j.celrep.2024.114382

Figure Lengend Snippet: (A) Left, Diagram illustrating the recording configuration. MC and MPP EPSCs were recorded from the same GC and evoked with stimulation electrodes placed in the inner and middle molecular layer, respectively. Right, Current clamp recording showing GC theta-burst firing (GC TBF). LTP induction protocol (GC TBF) was composed of 10 bursts at 5 Hz of 5 action potentials at 50 Hz, repeated 4 times every 5 s. (B) Left, Representative traces before (1) and after (2) GC TBF delivery. Right, Time-course plot showing that GC TBF induced LTP at MC-GC but not at MPP-GC synapses. (C) GC TBF-induced LTP was associated with significant reduction in PPR and CV (n = 13 cells). ** p < 0.01, *** p < 0.001. (D) LTP was abolished when TrkB was conditionally knocked out from from GCs (Post TrkB cKO, TrkBfl/fl mice injected in the dorsal blade with AAV5.CaMKII.Cre.GFP). LTP was unaffected in control animals (Control, TrkBfl/fl mice injected in the dorsal blade with AAV5.CaMKII.eGFP). (E) LTP was normally induced when loading PKI6-22 (2.5 μM) in GCs via the recording pipette but completely blocked when the cell-permeable PKA inhibitor PKI14-22 myristoylated (1 μM) was bath applied. (F) Summary bar graph showing the magnitude of GC TBF-induced LTP in the presence of DGC-IV (1 μM), when TrkB was conditionally knocked out from MCs (Pre TrkB cKO), when loading the PKI6-22 (2.5 μM) in GCs, and in the presence of D-APV (50 μM). LTP was abolished in the presence of the TrkB antagonist ANA-12 (15 μM), when Botox (0.5 μM) was loaded postsynaptically, in postsynaptic BDNF and TrkB cKO mice, and during bath application of the PKA inhibitors H89 (10 μM) or myristoylated PKI14-22 μM). Time-course summary plots are shown in Figure S1. ** p < 0.01, *** p < 0.001. (G) Scheme illustrating the emerging model for the mechanism underlying GC TBF-LTP. GC TBF triggers postsynaptic BDNF release and subsequent TrkB activation in GCs (1). Presynaptic PKA is then engaged downstream of postsynaptic BDNF/TrkB signaling (2), suggesting the requirement of a retrograde signal. Lastly, presynaptic PKA activation resulted in a long-lasting increase in glutamate release (3). Numbers in parentheses indicate the number of cells. Data are presented as mean ± SEM.

Article Snippet: H89 dihydrochloride , Tocris Bioscience , Cat#2910.

Techniques: Injection, Control, Transferring, Activation Assay

Journal: Cell reports

Article Title: Retrograde adenosine/A 2A receptor signaling facilitates excitatory synaptic transmission and seizures

doi: 10.1016/j.celrep.2024.114382

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: H89 dihydrochloride , Tocris Bioscience , Cat#2910.

Techniques: Virus, Plasmid Preparation, Recombinant, Software

Journal: Frontiers in Pharmacology

Article Title: Dopamine Homeostasis Imbalance and Dopamine Receptors-Mediated AC/cAMP/PKA Pathway Activation are Involved in Aconitine-Induced Neurological Impairment in Zebrafish and SH-SY5Y Cells

doi: 10.3389/fphar.2022.837810

Figure Lengend Snippet: Effects of SCH23390, sumanirole and H-89 on intracellular Ca 2+ changes induced by aconitine in zebrafish larvaes. (A) Effects of SCH23390 on intracellular Ca 2+ changes induced by aconitine. (B) Sumanirole suppressed aconitine-induced the increase of intracellular Ca 2+ . (C) Pretreatment with SCH23390 and sumanirole inhibited the rise of intracellular Ca 2+ aconitine-stimulated. (D) H-89 inhibited the increase of intracellular Ca 2+ induced by aconitine. All photographs were taken under a confocal microscope (100×). The values are expressed as mean ± SD, n = 8. * p < 0.05, ** p < 0.01, *** p < 0.001 vs. control groups. # p < 0.05, ## p < 0.01, ### p < 0.001 vs. aconitine-treated groups.

Article Snippet: D1R antagonist SCH23390, D2R agonist sumanirole, and PKA antagonist H-89 were provided by MCE (United States).

Techniques: Microscopy, Control

Journal: Signal Transduction and Targeted Therapy

Article Title: Protein C receptor maintains cancer stem cell properties via activating lipid synthesis in nasopharyngeal carcinoma

doi: 10.1038/s41392-021-00866-z

Figure Lengend Snippet: Signaling cascade induced by activated PROCR in NPC. a Ca 2+ flux dynamics labeled by fluo-4 in PROCR overexpression or knockout cells. Living cells were observed right after the dye supplementation. The corresponding right panels show the statistical analysis of fluorescence intensity. b , c Western blotting analysis of lipid metabolism related genes expression and stem cell markers expression in vector or PROCR overexpressing cells with different drugs treatment. 8-Bromo-cAMP, PKA agonist; H 89 2HCl, PKA antagonist; PDTC, NFκB inhibitor. d ELISA detection of the cellular cAMP content in PROCR overexpressing cells. e Sphere forming assay of the sorted CD45 - EPCAM + PROCR + cells treated with different drugs. f , g The cell content of triglycerides and cholesterol in PROCR overexpressing cells treated with different drugs. h ChIP-PCR detection of the direct transcriptional regulation of NFκB on FASN or PTGS2 expression. i Subcutaneous xenograft tumor volume of PROCR overexpressing cells treated with different drugs; n = 5 for each group. All mice received APC activation. j Percentages of GFP + cells from xenograft tumor nodules in the lungs of mice treated with different drugs. The drugs were administered every week; ns Not significant, * P < 0.05, *** P < 0.001

Article Snippet: For cell culture, the drugs treatment dosages were as follows: APC (100 nM, 55-3-08 A, Pepmic, Suzhou, China), PKA agonist (8-Bromo-cAMP, 10 μM, S7857, Selleck, Houston, TX, USA), PKA antagonist (H 89 2HCl, 10 μM, S1582, Selleck), NFκB inhibitor PDTC (Pyrrolidinedithiocarbamate ammonium, 10 μM, S3633, Selleck), Ca 2+ -ATPase inhibitor (Thapsigargin, 1 μM, ab120286, Abcam), FASN inhibitor (C75, 50 μM, S8915, Selleck), PTGS1 and PTGS2 inhibitor (Ketoprofen, 50 μM, S1645, Selleck), cAMP agonist (Forskolin, 10 μM, S2449, Selleck) and calcium chelator (BAPTA-AM, 10 μM, HY-100545, MedChemExpress).

Techniques: Labeling, Over Expression, Knock-Out, Fluorescence, Western Blot, Expressing, Plasmid Preparation, Enzyme-linked Immunosorbent Assay, Activation Assay

Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: Histone deacetylase HDAC7 restricts CD8 + T cell tumor infiltration and limits immunotherapy sensitivity in bladder cancer: reversal by pinocembrin

doi: 10.1186/s13046-025-03585-3

Figure Lengend Snippet: Retrospective clinical analysis indicates that HDAC7 is associated with the immunotherapy response in BCa. A Retrospective analysis of the IMvigor210 cohort identified HDAC1 and HDAC7 as BCa immunotherapy response-associated HDACs. B Flowchart illustrating the screening and follow-up of BCa patients who received adjuvant immunotherapy at our center, with analysis of the correlation between HDAC1 or HDAC7 expression and immunotherapy response. C Correlation analysis of HDAC1 or HDAC7 expression with the prognosis of BCa patients receiving adjuvant immunotherapy ( n = 20). D Querying the TISIDB database revealed a negative correlation between HDAC7 expression and immunotherapy response in BCa. E Western blot analysis of HDAC7 expression in BCa tissues and adjacent non-tumor tissues. F IHC of HDAC7 expression in BCa tissues and adjacent non-tumor tissues. G qRT-PCR analysis of HDAC7 expression in 40 paired BCa tissues and adjacent non-tumor tissues (* P < 0.05). H Correlation between HDAC7 expression and overall prognosis in 40 BCa patients. I qRT-PCR analysis of HDAC7 expression in 7 BCa cell lines and one normal urothelial cell line (* P < 0.05, ** P < 0.01, *** P < 0.001). Data are mean ± SD, n = 3

Article Snippet: HDAC7 recombinant protein was obtained from SIGNALCHEM (cat. H89-31G), and Pinocembrin was obtained from the MedChemExpress (CAS: 205370-59−8).

Techniques: Adjuvant, Expressing, Western Blot, Quantitative RT-PCR

Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: Histone deacetylase HDAC7 restricts CD8 + T cell tumor infiltration and limits immunotherapy sensitivity in bladder cancer: reversal by pinocembrin

doi: 10.1186/s13046-025-03585-3

Figure Lengend Snippet: HDAC7 inhibits CD8 + T cell chemotaxis and anti-tumor activity in vitro. A Analysis of IMvigor210 cohort data shows negative correlation between HDAC7 expression and CD8 + T cell chemotaxis. B Flow cytometry confirms the efficiency of CD8 + T cell sorting using CD3 and CD8 antibodies. C Microscopy of CD8 + T cell morphology before and after activation. D Schematic of the co-culture model. E In vitro co-culture shows that HDAC7 inhibits CD8 + T cell chemotaxis (** P < 0.01, *** P < 0.001). F HDAC7 knockdown in T24 cells increases IFN-γ and granzyme B production by CD8 + T cells (* P < 0.05, ** P < 0.01). (G) HDAC7 overexpression in UMUC3 cells inhibits IFN-γ and granzyme B production by CD8 + T cells (** P < 0.01). H CD8 + T cell cytotoxicity is enhanced when co-cultured with HDAC7 knockdown T24 cells and reduced when co-cultured with HDAC7-overexpressing UMUC3 cells. Data are mean ± SD, n = 3

Article Snippet: HDAC7 recombinant protein was obtained from SIGNALCHEM (cat. H89-31G), and Pinocembrin was obtained from the MedChemExpress (CAS: 205370-59−8).

Techniques: Chemotaxis Assay, Activity Assay, In Vitro, Expressing, Flow Cytometry, FACS, Microscopy, Activation Assay, Co-Culture Assay, Knockdown, Over Expression, Cell Culture

Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: Histone deacetylase HDAC7 restricts CD8 + T cell tumor infiltration and limits immunotherapy sensitivity in bladder cancer: reversal by pinocembrin

doi: 10.1186/s13046-025-03585-3

Figure Lengend Snippet: HDAC7 inhibits CD8 + T cell chemotaxis and immunotherapy sensitivity in BCa in vivo. A Schematic of the HuNOG mouse model and experimental design for studying HDAC7’s effect on CD8 + T cell chemotaxis and immunotherapy response. B Flow cytometry analysis of human CD45 positivity in HuNOG mice. C Representative tumor images from HuNOG mice injected with HDAC7 knockdown or control T24 cells and treated with PD-1 or IgG antibodies ( n = 3). D Tumor volume in HuNOG mice measured every 3 days (** P < 0.01, *** P < 0.001). E Tumor weight assessment in HuNOG mice (** P < 0.01). F IHC analysis of CD3 and CD8 in tumors from HuNOG mice, showing increased CD8 + T cell infiltration with HDAC7 knockdown. G – H. Flow cytometry of CD8, IFN-γ, and granzyme B in tumor tissues, confirming that HDAC7 knockdown enhances CD8 + T cell infiltration and anti-tumor activity. Data are mean ± SD, n = 3

Article Snippet: HDAC7 recombinant protein was obtained from SIGNALCHEM (cat. H89-31G), and Pinocembrin was obtained from the MedChemExpress (CAS: 205370-59−8).

Techniques: Chemotaxis Assay, In Vivo, Flow Cytometry, Injection, Knockdown, Control, Activity Assay

Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: Histone deacetylase HDAC7 restricts CD8 + T cell tumor infiltration and limits immunotherapy sensitivity in bladder cancer: reversal by pinocembrin

doi: 10.1186/s13046-025-03585-3

Figure Lengend Snippet: HDAC7 inhibits CCL5 expression and impedes CD8 + T cell infiltration in BCa. A Heat map of mRNA sequencing in three pairs of HDAC7 knockdown and control T24 cells. B Volcano plot of mRNA sequencing showing differentially expressed genes, including CCL5. C GSEA indicates enrichment of differentially expressed genes in the T cell chemotaxis pathway. D Analysis of the IMvigor210 cohort shows a negative correlation between HDAC7 and CCL5 expression. E – F Correlation between CCL5 expression and immune cell infiltration across cancers. G Pearson correlation between HDAC7 and CCL5 expression in 40 BCa tissues. H – I qRT-PCR analysis in T24 and UMUC3 cells demonstrates that HDAC7 knockdown upregulates, whereas overexpression downregulates, CCL5 expression (* P < 0.05, ** P < 0.01, *** P < 0.001). J – K ELISA shows that HDAC7 inhibits CCL5 protein expression (* P < 0.05, ** P < 0.01). Data are mean ± SD, n = 3

Article Snippet: HDAC7 recombinant protein was obtained from SIGNALCHEM (cat. H89-31G), and Pinocembrin was obtained from the MedChemExpress (CAS: 205370-59−8).

Techniques: Expressing, Sequencing, Knockdown, Control, Chemotaxis Assay, Quantitative RT-PCR, Over Expression, Enzyme-linked Immunosorbent Assay

Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: Histone deacetylase HDAC7 restricts CD8 + T cell tumor infiltration and limits immunotherapy sensitivity in bladder cancer: reversal by pinocembrin

doi: 10.1186/s13046-025-03585-3

Figure Lengend Snippet: HDAC7 targets SRSF7 via non-histone deacetylation to regulate CCL5 expression. A-B ChIP-qPCR with H3K9ac and H3K27ac antibodies examines HDAC7’s effect on acetylation in the CCL5 promoter region. C CO-IP and mass spectrometry identify HDAC7 binding to SRSF7. D Integration of mass spectrometry and RBPsuite predictions identifies SRSF7 as the mediator of HDAC7-regulated CCL5 expression. E CO-IP confirms HDAC7–SRSF7 interaction. F – G HDOCK analysis reveals HDAC7 binding to SRSF7 at Lys24, a potential deacetylation site. H – I Western blot analysis of the effects of HDAC7 on SRSF7 acetylation (Lys24) and protein expression in T24 and UMUC3 cells. J – K CO-IP analysis of how SRSF7 Lys24 mutation affects HDAC7–SRSF7 binding in T24 cells. Data are mean ± SD, n = 3

Article Snippet: HDAC7 recombinant protein was obtained from SIGNALCHEM (cat. H89-31G), and Pinocembrin was obtained from the MedChemExpress (CAS: 205370-59−8).

Techniques: Expressing, ChIP-qPCR, Co-Immunoprecipitation Assay, Mass Spectrometry, Binding Assay, Western Blot, Mutagenesis

Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: Histone deacetylase HDAC7 restricts CD8 + T cell tumor infiltration and limits immunotherapy sensitivity in bladder cancer: reversal by pinocembrin

doi: 10.1186/s13046-025-03585-3

Figure Lengend Snippet: HDAC7 deacetylates SRSF7, promoting BTRC-induced ubiquitination and degradation of SRSF7. A PhosphoSitePlus analysis predicts acetylation and ubiquitination of SRSF7 at Lys24. B – C Ubibrowser and HDOCK predict BTRC binding to SRSF7 at Lys24. D CO-IP confirms BTRC–SRSF7 interaction. E CO-IP shows that HDAC7 knockdown promotes SRSF7 expression and reduces BTRC-SRSF7 binding in T24 cells. CO-IP shows that HDAC7 overexpression inhibits SRSF7 expression and increases BTRC-SRSF7 binding in UMUC3 cells. F CO-IP reveals that HDAC7 knockdown inhibits SRSF7 ubiquitination in T24 cells. G In T24 cells, HDAC7 overexpression increases SRSF7 ubiquitination, which is reduced by concurrent BTRC knockdown. H – I Western blot analysis shows that HDAC7 overexpression inhibits SRSF7 expression, which is restored by BTRC knockdown in T24 and UMUC3 cells. CO-IP analysis of SRSF7 Lys24 mutation on BTRC–SRSF7 binding in T24 cells. Data are mean ± SD, n = 3

Article Snippet: HDAC7 recombinant protein was obtained from SIGNALCHEM (cat. H89-31G), and Pinocembrin was obtained from the MedChemExpress (CAS: 205370-59−8).

Techniques: Ubiquitin Proteomics, Binding Assay, Co-Immunoprecipitation Assay, Knockdown, Expressing, Over Expression, Western Blot, Mutagenesis

Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: Histone deacetylase HDAC7 restricts CD8 + T cell tumor infiltration and limits immunotherapy sensitivity in bladder cancer: reversal by pinocembrin

doi: 10.1186/s13046-025-03585-3

Figure Lengend Snippet: SRSF7 inhibits exon skipping of CCL5 pre-mRNA, thereby promoting mRNA maturation. A Analysis of alternative splicing (AS-Novel) in transcriptome sequencing data showed that exon skipping (ES) events accounted for 67.55% of all events. B AS-Novel analysis identified exon 2 (E2) skipping in CCL5. C ESE finder 3.0 predicted that CCL5 E2 contains exonic splicing enhancer (ESE) regions. D RIP assay confirmed the binding between SRSF7 and CCL5 pre-mRNA (*** P < 0.001). E Primers spanning E2 and targeting E2-skipping were designed; PCR amplification and agarose gel electrophoresis were used to assess changes in CCL5 alternative splicing following SRSF7 knockdown in T24 cells. F – G qRT-PCR showed that SRSF7 knockdown inhibited CCL5 expression in T24 and UMUC3 cells (* P < 0.05, *** P < 0.001). H – I ELISA showed reduced CCL5 protein production following SRSF7 knockdown in T24 and UMUC3 cells (** P < 0.01, *** P < 0.001). J – K qRT-PCR and ELISA-based rescue experiments demonstrated that HDAC7 knockdown increases CCL5 expression, which is repressed when SRSF7 is simultaneously knocked down in T24 cells (* P < 0.05, ** P < 0.01). Data are mean ± SD, n = 3

Article Snippet: HDAC7 recombinant protein was obtained from SIGNALCHEM (cat. H89-31G), and Pinocembrin was obtained from the MedChemExpress (CAS: 205370-59−8).

Techniques: Alternative Splicing, Sequencing, Binding Assay, Amplification, Agarose Gel Electrophoresis, Knockdown, Quantitative RT-PCR, Expressing, Enzyme-linked Immunosorbent Assay

Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: Histone deacetylase HDAC7 restricts CD8 + T cell tumor infiltration and limits immunotherapy sensitivity in bladder cancer: reversal by pinocembrin

doi: 10.1186/s13046-025-03585-3

Figure Lengend Snippet: Virtual screening identifies Pinocembrin as a specific HDAC7 inhibitor. A Workflow of virtual screening for HDAC7 inhibitors. B Chemical structure of Pinocembrin and its predicted binding score with HDAC7. C Schematic diagrams showing Pinocembrin binding to HDAC7 in cartoon and surface representations. D Three-dimensional (3D) and two-dimensional (2D) views of Pinocembrin binding to HDAC7. In the 3D diagram, the C backbone of HDAC7 is green, N atoms blue, O atoms magenta, H atoms white, and Pinocembrin sky blue. Purple dashed lines indicate hydrogen bond lengths, with longer lines representing weaker interactions. E–F. MST experiments evaluate the binding affinity of Pinocembrin to HDAC7. G–H. T24 and MB49 cells were treated with increasing concentrations of Pinocembrin (10, 50, 100, 200 µM); HDAC activity was assessed in cells with different HDAC7 expression levels using an HDAC Activity Assay Kit (* P < 0.05, ** P < 0.01, *** P < 0.001). Data are mean ± SD, n = 3

Article Snippet: HDAC7 recombinant protein was obtained from SIGNALCHEM (cat. H89-31G), and Pinocembrin was obtained from the MedChemExpress (CAS: 205370-59−8).

Techniques: Binding Assay, Activity Assay, Expressing, HDAC Activity Assay

Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: Histone deacetylase HDAC7 restricts CD8 + T cell tumor infiltration and limits immunotherapy sensitivity in bladder cancer: reversal by pinocembrin

doi: 10.1186/s13046-025-03585-3

Figure Lengend Snippet: Schematic model of the Pinocembrin/HDAC7/BTRC/SRSF7/CCL5 regulatory and functional network

Article Snippet: HDAC7 recombinant protein was obtained from SIGNALCHEM (cat. H89-31G), and Pinocembrin was obtained from the MedChemExpress (CAS: 205370-59−8).

Techniques: Functional Assay