Journal: Clinical and Experimental Medicine

Article Title: The mechanism of S100A8/A9 heterodimer promoting atherosclerotic thrombosis

doi: 10.1007/s10238-025-02016-z

Figure Lengend Snippet: S100A8/A9 heterodimer promotes platelet adhesion and aggregation via EGFR/PI3K/AKT and EGFR/p38 MAPK axes ( A ) Representative images of the morphological changes observed by TEM in washed human platelets treated with vehicle (DMSO), 10 µg/mL S100A8/A9 heterodimer, AG1478 or LY294002. ( B ) Representative images of the platelet adhesion test of washed human platelets treated with vehicle (DMSO), 1 µg/mL S100A8/A9 heterodimer, AG1478 or LY294002. The statistics are shown in the histogram below. ( C ) Platelet aggregation induced by ADP, AA or EPI in PRP treated with vehicle (DMSO), 1 µg/mL S100A8/A9 heterodimer, AG1478 or LY294002. The statistics are shown in the histogram below. ( D ) Expression of p-p38 and p38 in the platelets harvested from the reaction cells of the platelet aggregometer. The statistics are shown in the histogram below. ( E ) Expression of p-PI3K, PI3K, p-AKT and AKT in the platelets harvested from the reaction cells of the platelet aggregometer. The statistics are shown in the histogram below. n = 3 independent experiments. Significant results are presented as * p <0.05, ** P <0.01, or *** P <0.001.

Article Snippet: The roles of S100A8/A9 heterodimer and EGFR signaling pathway in platelet shape, adhesion and aggregation were investigated using recombinant S100A8/A9 heterodimer (Sino Biological, Beijing, China, Cat: CT002-H0822B) or specific inhibitors EGFR inhibitor (AG1478, 10 μM, MedChem Express, Monnouth Junction, NJ, USA, Cat: HY-13524) and PI3K inhibitor (LY294002, 10 μM, MedChem Express, Monnouth Junction, NJ, USA, Cat: HY-10108).

Techniques: Expressing

Journal: Clinical and Experimental Medicine

Article Title: The mechanism of S100A8/A9 heterodimer promoting atherosclerotic thrombosis

doi: 10.1007/s10238-025-02016-z

Figure Lengend Snippet: S100A8/A9 heterodimer promotes platelet αⅡbβ3 activation and TXB 2 secretion via EGFR signaling pathway ( A ) Activity of CD62P in washed human platelets treated with Vehicle (DMSO), ADP, ADP + S100A8/A9, ADP + S100A8/A9 + AG1478 or ADP + S100A8/A9 + LY294002. The statistics are shown in the histogram below. ( B ) Representative phalloidin-stained images of washed human platelets treated with vehicle (DMSO), 1 µg/mL S100A8/A9 heterodimer, AG1478 or LY294002 spreading on immobilized fibrinogen for every 20 min. The statistics are shown in the histogram below. ( C ) Clot retraction of human platelets treated with vehicle (DMSO), 1 µg/mL S100A8/A9 heterodimer, AG1478 or LY294002. The statistics are shown in the histogram below. ( D ) TXB 2 secretion detected with ELISA in human platelets treated with Vehicle (DMSO), ADP, ADP + S100A8/A9, AG1478 or LY294002. n = 3 independent experiments. Significant results are presented as * P <0.05, ** P <0.01, or *** P <0.001. Non-significant results are presented as ns.

Article Snippet: The roles of S100A8/A9 heterodimer and EGFR signaling pathway in platelet shape, adhesion and aggregation were investigated using recombinant S100A8/A9 heterodimer (Sino Biological, Beijing, China, Cat: CT002-H0822B) or specific inhibitors EGFR inhibitor (AG1478, 10 μM, MedChem Express, Monnouth Junction, NJ, USA, Cat: HY-13524) and PI3K inhibitor (LY294002, 10 μM, MedChem Express, Monnouth Junction, NJ, USA, Cat: HY-10108).

Techniques: Activation Assay, Activity Assay, Staining, Enzyme-linked Immunosorbent Assay

Journal: Clinical and Experimental Medicine

Article Title: The mechanism of S100A8/A9 heterodimer promoting atherosclerotic thrombosis

doi: 10.1007/s10238-025-02016-z

Figure Lengend Snippet: The role of S100A8/A9 heterodimer produced by endothelial cells stimulated by NETs in thrombosis is shown. Neutrophil extracellular traps (NETs) induce pyroptosis and increase S100A8/A9 heterodimer expression in HUVECs via ROS/NF-κB/Caspase-1 axis and S100A8/A9 heterodimer activates platelets via the EGFR/PI3K/AKT and EGFR/p38 MAPK signaling pathways, driving thrombus formation.

Article Snippet: The roles of S100A8/A9 heterodimer and EGFR signaling pathway in platelet shape, adhesion and aggregation were investigated using recombinant S100A8/A9 heterodimer (Sino Biological, Beijing, China, Cat: CT002-H0822B) or specific inhibitors EGFR inhibitor (AG1478, 10 μM, MedChem Express, Monnouth Junction, NJ, USA, Cat: HY-13524) and PI3K inhibitor (LY294002, 10 μM, MedChem Express, Monnouth Junction, NJ, USA, Cat: HY-10108).

Techniques: Produced, Expressing, Protein-Protein interactions

Journal: Virulence

Article Title: Streptococcus mutans -derived extracellular vesicles promote herpes simplex virus infection in oral epithelia

doi: 10.1080/21505594.2025.2602261

Figure Lengend Snippet: Effect of the EGFR – ERK pathway on the Sm-EV-dependent increase of HSV-1 production in HOK cells. (A) Cells were pretreated with EGFR antagonist (AG1478) for 3 h and then treated with Sm EVs for 24 h. After infection with HSV-1 (MOI 0.1) for another 48 h, the cells were stained with fluorescent gD antibody. Fluorescence intensity was quantified using zen microscopy software. (B) The protein level of HSV-1 gD was analyzed using western blot analysis under the same conditions. EGFR, erk, and NF-κB phosphorylation levels were analyzed using western blot analysis under the same conditions. (C) Cells were pretreated with erk antagonist (PD98059) for 3 h and then treated with Sm EVs for 24 h. After infection with HSV-1 (MOI 0.1) for another 48 h, western blot analysis was performed to measure gD protein levels and erk phosphorylation. β-actin was used as an internal loading control. (D) Cells were pretreated with AG1478 for 3 h and then treated with Sm EVs for 24 h. After infection with HSV-1 (MOI 0.1) for another 48 h, qPCR was performed to measure IFN mRNA level. (E) Cytokine ELISA was performed to measure the levels of secreted IFN proteins under the same conditions. Results are presented as the mean ± standard deviation of three independent experiments. * p < 0.05, ** p < 0.01, and *** p < 0.001.

Article Snippet: The EGFR inhibitor AG1478 and the ERK inhibitor PD98059 were provided by MedChemExpress (NJ, USA).

Techniques: Infection, Staining, Fluorescence, Microscopy, Software, Western Blot, Phospho-proteomics, Control, Enzyme-linked Immunosorbent Assay, Standard Deviation

Journal: Virulence

Article Title: Streptococcus mutans -derived extracellular vesicles promote herpes simplex virus infection in oral epithelia

doi: 10.1080/21505594.2025.2602261

Figure Lengend Snippet: Quantification of HSV-1 production by plaque assay following Sm EV treatment and EGFR – ERK inhibition. (A) HOK cells were pretreated with Sm EVs (1 × 10 3 particles/cell) for 24 h, followed by HSV-1 infection (MOI 0.1) under the indicated conditions: non-treated (NT), AG1478 (10 μM), or PD98059 (20 μM). After 48 h, culture supernatants were collected, and HSV-1 titers were determined by plaque assay. (B) Representative plaque images from culture supernatants. Results are presented as the mean ± standard deviation of three independent experiments. * p < 0.05, ** p < 0.01, and *** p < 0.001.

Article Snippet: The EGFR inhibitor AG1478 and the ERK inhibitor PD98059 were provided by MedChemExpress (NJ, USA).

Techniques: Plaque Assay, Inhibition, Infection, Standard Deviation

Journal: Sexual Medicine

Article Title: Hinokiflavone alleviates high-fat diet-induced erectile dysfunction via the EGFR/PI3K/Akt/eNOS signaling pathway

doi: 10.1093/sexmed/qfaf059

Figure Lengend Snippet: HINO alleviates PA-induced endothelial dysfunction by activating the EGFR and PI3K/Akt/eNOS Signaling pathways. (A-D) Representative immunoblotting images (A) for p-PI3K, PI3K, p-Akt, Akt, p-eNOS, and eNOS after treatment with EGFR inhibitor AG1478 (10 μM, 24 h) in RCCECs. Semi-quantification of p-PI3K/PI3K (B), p-Akt/Akt (C), and p-eNOS/eNOS (D) was conducted with data presented as mean ± SD (n = 3). (E-G) Representative immunoblotting images (E) for p-Akt, Akt, p-eNOS, and eNOS after treatment with PI3K inhibitor LY294002 (10 μM, 24 h) in RCCECs. Semi-quantification of p-Akt/Akt (F) and p-eNOS/eNOS was conducted with data presented as mean ± SD (n = 3). (H-I) Representative immunoblotting images for p-eNOS and eNOS after treatment with Akt inhibitor MK-2206 (10 μM, 24 h) in RCCECs. Semi-quantification of p-eNOS/eNOS (I) was conducted with data presented as mean ± SD (n = 3). (J-K) Representative images (J) and semi-quantification (K) of nitric oxide release using DAF-FM from RCCECs treated with inhibitor AG1478, LY294002, MK-2206, and L-NAME. Scale bars, 100 μm. Data are presented as mean ± SD (n = 3).

Article Snippet: Additionally, an inhibitor working solution was formulated by dissolving powdered AG1478 (Cat# HY-13524, MCE, Shanghai, China), LY294002 (Cat# HY-10108, MCE, Shanghai, China), MK-2206 dihydrochloride (Cat# HY-10358, MCE, Shanghai, China), and L-NAME hydrochloride (Cat# HY-18729A, MCE, Shanghai, China) in DMSO.

Techniques: Protein-Protein interactions, Western Blot

Journal: bioRxiv

Article Title: CD163⁺/Dab2⁺ Macrophages Alleviate Cardiac Hypertrophy via Nrg2/ErbB4-Mediated Mitochondrial Reprogramming

doi: 10.1101/2025.05.22.655661

Figure Lengend Snippet: A, Representative flow plots of F4/80, CD11b, CD163, Dab2 and Nrg2 expression in polarized BMDMs. B, ELISA analysis of Nrg2 levels in CD163 - , CD163 + /Dab2 - , and CD163 + /Dab2 + macrophages (n=3). * P <0.05; ** P <0.01. C, Representative immunofluorescence images of phalloidin staining in NRVMs cultured with different medium and treated with vehicle or Ang II for 48 hours. Scale bar=100 μm. D, Relative cell surface area of NRVMs cultured with different medium following vehicle or Ang II treatment (n=6). * P <0.05; *** P <0.001; ns, not significant. E, RT-qPCR analysis of the mRNA levels of Anp and Myh7 in NRVMs cultured with different medium and treated with vehicle or Ang II for 48 hours (n=5). * P <0.05; ** P <0.01; *** P <0.001; ns, not significant. F, Representative immunofluorescence images of phalloidin staining in NRVMs treated with recombinant Nrg2 and inhibitor AG1478 in response to vehicle or Ang II treatment for 48 hours, respectively. Scale bar=100 μm. G, Relative cell surface area of NRVMs treated with recombinant Nrg2 and/or inhibitor AG1478 in response to vehicle or Ang II treatment for 48 hours, respectively (n=5). *** P <0.001; ns, not significant. H, RT-qPCR analysis of the mRNA levels of Anp, Bnp and Myh7 in NRVMs treated with recombinant Nrg2 and and inhibitor AG1478 in response to vehicle or Ang II treatment for 48 hours, respectively (n=3). * P <0.05; ** P <0.01; *** P <0.001. I, Protein expression of Anp and Myh7 in NRVMs treated with recombinant Nrg2 and inhibitor AG1478 in response to vehicle or Ang II treatment for 48 hours, respectively

Article Snippet: For the activation and blockade of the Nrg2 and ErbB4 pathway, 1 week after TAC surgery, mice were randomized and continuously treated with either vehicle or recombinant Nrg2 (produced by our lab) at a dosage of 500 μg/kg/d and inhibitor AG1478 (HY-13524, MCE) at a dosage of 10 mg/kg/d for 7 days.

Techniques: Expressing, Enzyme-linked Immunosorbent Assay, Immunofluorescence, Staining, Cell Culture, Quantitative RT-PCR, Recombinant

Journal: bioRxiv

Article Title: CD163⁺/Dab2⁺ Macrophages Alleviate Cardiac Hypertrophy via Nrg2/ErbB4-Mediated Mitochondrial Reprogramming

doi: 10.1101/2025.05.22.655661

Figure Lengend Snippet: A, Schematic diagram illustrating the experimental design involving the injection of recombinant Nrg2 and inhibitor AG1478 into a mouse model of cardiac hypertrophy. Wildtype mice at 6 weeks of age were subjected to transverse aortic constriction (TAC) or Sham surgery. After 1 week, recombinant Nrg2 or Nrg2 and inhibitor AG1478 were continuously administered into the mice for 7 days. These mice groups were denoted as Sham, TAC, TAC+Nrg2, and TAC+Nrg2+AG1478. B, Representative images of hearts at 8 weeks post-surgery from mice subjected to Sham or TAC surgery and injected with recombinant Nrg2 or Nrg2 and inhibitor AG1478. C, Heart weight/body weight (HW/BW) ratios at 8 weeks post-surgery for mice subjected to Sham or TAC surgery and injected with recombinant Nrg2 or Nrg2 and inhibitor AG1478 (Sham, n=8; TAC, n=7; TAC+Nrg2, n=8; TAC+Nrg2+AG1478, n=8). ** P <0.01; *** P <0.001; ns, not significant. D and E, M-mode echocardiography images (D) and quantified parameters (E) of the left ventricular (LV) chamber at 8 weeks post-surgery in mice subjected to Sham or TAC surgery and injected with recombinant Nrg2 or Nrg2 and inhibitor AG1478 (Sham, n=8; TAC, n=7; TAC+Nrg2, n=8; TAC+Nrg2+AG1478, n=8). ** P <0.01; *** P <0.001; ns, not significant. EF, ejection fraction; FS, fractional shortening; CO, cardiac output. F, Hematoxylin-eosin (H&E) (scale bar=1 mm) and Wheat germ agglutinin (WGA) staining (scale bar=50 μm) showing morphology and cell boundaries of heart sections 8 weeks after TAC with indicated treatment regimens. G, Quantification of the cardiomyocyte cross-sectional areas of the indicated groups (n=6 mice per group). *** P <0.001; ns, not significant. H, The mRNA levels of hypertrophic marker genes Anp, Bnp, Myh7 and Acta1 in hearts of the indicated groups (n=6 mice per group). * P <0.05; *** P <0.001; ns, not significant. I, The mRNA levels of fibrosis marker genes Col1a1, Col3a1, Ctgf and MMP9 in hearts of the indicated groups (n=6 mice per group). * P <0.05; ** P <0.01; *** P <0.001; ns, not significant. J, Masson’s trichrome staining showing fibrosis in heart sections 8 weeks after TAC with indicated treatment regimens. Scale bar=50 μm. K, Quantification of cardiac fibrosis in tissue shown in panel J (n=6 mice per group). * P <0.05; *** P <0.001.

Article Snippet: For the activation and blockade of the Nrg2 and ErbB4 pathway, 1 week after TAC surgery, mice were randomized and continuously treated with either vehicle or recombinant Nrg2 (produced by our lab) at a dosage of 500 μg/kg/d and inhibitor AG1478 (HY-13524, MCE) at a dosage of 10 mg/kg/d for 7 days.

Techniques: Injection, Recombinant, Staining, Marker