Journal: Journal of Pharmaceutical Analysis

Article Title: Oxymatrine, a novel TLR2 agonist, promotes megakaryopoiesis and thrombopoiesis through the STING/NF-κB pathway

doi: 10.1016/j.jpha.2024.101054

Figure Lengend Snippet: Oxymatrine induces megakaryocyte (MK) differentiation by activating stimulator of interferon genes (STING)/nuclear factor-kappaB (NF-κB) pathway. (A) Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis for the potential pathway by which oxymatrine promotes MK differentiation. (B–E) Western blot (WB) analysis of the expression of pathway: extracellular regulated kinase 1/2 (ERK1/2)/STING/inhibitory subunit of nuclear factor kappa B alpha (IκBα)/NF-κB by which oxymatrine promotes MK differentiation. Representative immunoblot images and biochemical quantification of the ERK1/2 (B), STING (C), IκBα (D), and NF-κB (E) pathway after the treatment with oxymatrine (10, 20, and 40 μM) in Meg-01 cells for five days ( n = 3 per group). (F–I) Representative immunoblot images and biochemical quantification of the transcription factors (TFs) associated with MK differentiation: runt-related transcription factor 1 (RUNX1) (F), nuclear factor erythroid 2 (NF-E2) (G), early growth response protein 1 (EGR1) (H), and cellular oncogene fos (FOS) (I) ( n = 3 per group). (J) Representative immunofluorescence images of toll-like receptor 2 (TLR2) fluorescently triple-stained with p-STING and p-NF-κB, respectively, on bone marrow (BM) cells after 12 days of treatment with oxymatrine in a mouse model of radiation thrombocytopenia. Blue for the 4′,6-diamidino-2-phenylindole (DAPI), purple for the TLR2, red for the p-STING, and green for the p-NF-κB. (K, L) Representative images of immunofluorescence of the nuclear translocation of NF-E2 (K) and RUNX1 (L) in Meg-01 cells after the treatment of oxymatrine for five days. Excitation wavelengths: 470 nm for NF-E2 and RUNX1 and 405 nm for DAPI. Data represent the mean ± standard deviation (SD) of three independent experiments. ∗ P < 0.05, ∗∗ P < 0.01, and ∗∗∗ P < 0.001 vs. the control group. EGFR: epidermal growth factor receptor; IL-17: interleukin 17; VEGF: vascular endothelial growth factor ; TNF: tumor necrosis factor; JAK-STAT: janus kinase-signal transducer and activator of transcription; PI3K-Akt: phospholnositide-3 kinase-threonine kinase B; MAPK: mitogen-activated protein kinase; FoxO: forkhead box protein O; GAPDH: glyceraldehyde-3-phosphate dehydrogenase.

Article Snippet: After the addition of 5% bovine serum albumin (BSA), the cells were incubated at 4 °C overnight with anti-RUNX1 and anti-NF-E2 antibodies (Proteintech Group, Inc., Wuhan, China).

Techniques: Western Blot, Expressing, Immunofluorescence, Staining, Translocation Assay, Standard Deviation, Control

Journal: Journal of Pharmaceutical Analysis

Article Title: Oxymatrine, a novel TLR2 agonist, promotes megakaryopoiesis and thrombopoiesis through the STING/NF-κB pathway

doi: 10.1016/j.jpha.2024.101054

Figure Lengend Snippet: The schematic illustration of oxymatrine regulating megakaryocyte (MK) differentiation and thrombopoiesis. Oxymatrine targets toll-like receptor 2 (TLR2) and activates the downstream extracellular regulated kinase 1/2 (ERK1/2)/stimulator of interferon genes (STING)/inhibitory subunit of nuclear factor kappa B alpha (IκBα)/nuclear factor-kappaB (NF-κB) signaling pathway, which leads to the high expression of transcription factors (TFs): early growth response 1 (EGR1), nuclear factor erythroid 2 (NF-E2), cellular oncogene fos (FOS), and runt-related transcription factor 1 (RUNX1). As a result, the activated transcription factors promote the expression of megakaryopoiesis and thrombopoiesis related genes, which contribute to the MK maturation and platelet production and finally recover the platelet levels in adiation-induced thrombocytopenia mice. RIT: radiation-induced thrombocytopenia; HSC: hematopoietic stem cell; ERK: extracellular regulated kinase; Ub: ubiquitin.

Article Snippet: After the addition of 5% bovine serum albumin (BSA), the cells were incubated at 4 °C overnight with anti-RUNX1 and anti-NF-E2 antibodies (Proteintech Group, Inc., Wuhan, China).

Techniques: Expressing

Journal: Journal of Pharmaceutical Analysis

Article Title: Oxymatrine, a novel TLR2 agonist, promotes megakaryopoiesis and thrombopoiesis through the STING/NF-κB pathway

doi: 10.1016/j.jpha.2024.101054

Figure Lengend Snippet: Oxymatrine induces megakaryocyte (MK) differentiation by activating stimulator of interferon genes (STING)/nuclear factor-kappaB (NF-κB) pathway. (A) Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis for the potential pathway by which oxymatrine promotes MK differentiation. (B–E) Western blot (WB) analysis of the expression of pathway: extracellular regulated kinase 1/2 (ERK1/2)/STING/inhibitory subunit of nuclear factor kappa B alpha (IκBα)/NF-κB by which oxymatrine promotes MK differentiation. Representative immunoblot images and biochemical quantification of the ERK1/2 (B), STING (C), IκBα (D), and NF-κB (E) pathway after the treatment with oxymatrine (10, 20, and 40 μM) in Meg-01 cells for five days ( n = 3 per group). (F–I) Representative immunoblot images and biochemical quantification of the transcription factors (TFs) associated with MK differentiation: runt-related transcription factor 1 (RUNX1) (F), nuclear factor erythroid 2 (NF-E2) (G), early growth response protein 1 (EGR1) (H), and cellular oncogene fos (FOS) (I) ( n = 3 per group). (J) Representative immunofluorescence images of toll-like receptor 2 (TLR2) fluorescently triple-stained with p-STING and p-NF-κB, respectively, on bone marrow (BM) cells after 12 days of treatment with oxymatrine in a mouse model of radiation thrombocytopenia. Blue for the 4′,6-diamidino-2-phenylindole (DAPI), purple for the TLR2, red for the p-STING, and green for the p-NF-κB. (K, L) Representative images of immunofluorescence of the nuclear translocation of NF-E2 (K) and RUNX1 (L) in Meg-01 cells after the treatment of oxymatrine for five days. Excitation wavelengths: 470 nm for NF-E2 and RUNX1 and 405 nm for DAPI. Data represent the mean ± standard deviation (SD) of three independent experiments. ∗ P < 0.05, ∗∗ P < 0.01, and ∗∗∗ P < 0.001 vs. the control group. EGFR: epidermal growth factor receptor; IL-17: interleukin 17; VEGF: vascular endothelial growth factor ; TNF: tumor necrosis factor; JAK-STAT: janus kinase-signal transducer and activator of transcription; PI3K-Akt: phospholnositide-3 kinase-threonine kinase B; MAPK: mitogen-activated protein kinase; FoxO: forkhead box protein O; GAPDH: glyceraldehyde-3-phosphate dehydrogenase.

Article Snippet: The PVDF membranes were subjected to antibody probing using the following antibodies: TLR2 (TU362792S; Abmart (Shanghai) Co., Ltd., Shanghai, China), p-STING (TA7416S; Abmart (Shanghai) Co., Ltd.), STING (TD12090S; Abmart (Shanghai) Co., Ltd.), p-inhibitory subunit of nuclear factor kappa B alpha (p-IκBα) (2589; Cell Signaling Technology), IκBα (4814; Cell Signaling Technology), p-NF-κB (3033; Cell Signaling Technology), NF-κB (10745-1-AP; Proteintech Group, Inc., Chicago, IL, USA), EGR1 (22008-1-AP; Proteintech Group, Inc.), FOS (ab11959; Abcam plc, Cambridge, UK), NF-E2 (11089-1-AP; Proteintech Group, Inc.), RUNX1 (25315-1-AP; Proteintech Group, Inc.), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) (60004-1-lg; Proteintech Group, Inc.).

Techniques: Western Blot, Expressing, Immunofluorescence, Staining, Translocation Assay, Standard Deviation, Control

Journal: Journal of Pharmaceutical Analysis

Article Title: Oxymatrine, a novel TLR2 agonist, promotes megakaryopoiesis and thrombopoiesis through the STING/NF-κB pathway

doi: 10.1016/j.jpha.2024.101054

Figure Lengend Snippet: The schematic illustration of oxymatrine regulating megakaryocyte (MK) differentiation and thrombopoiesis. Oxymatrine targets toll-like receptor 2 (TLR2) and activates the downstream extracellular regulated kinase 1/2 (ERK1/2)/stimulator of interferon genes (STING)/inhibitory subunit of nuclear factor kappa B alpha (IκBα)/nuclear factor-kappaB (NF-κB) signaling pathway, which leads to the high expression of transcription factors (TFs): early growth response 1 (EGR1), nuclear factor erythroid 2 (NF-E2), cellular oncogene fos (FOS), and runt-related transcription factor 1 (RUNX1). As a result, the activated transcription factors promote the expression of megakaryopoiesis and thrombopoiesis related genes, which contribute to the MK maturation and platelet production and finally recover the platelet levels in adiation-induced thrombocytopenia mice. RIT: radiation-induced thrombocytopenia; HSC: hematopoietic stem cell; ERK: extracellular regulated kinase; Ub: ubiquitin.

Article Snippet: The PVDF membranes were subjected to antibody probing using the following antibodies: TLR2 (TU362792S; Abmart (Shanghai) Co., Ltd., Shanghai, China), p-STING (TA7416S; Abmart (Shanghai) Co., Ltd.), STING (TD12090S; Abmart (Shanghai) Co., Ltd.), p-inhibitory subunit of nuclear factor kappa B alpha (p-IκBα) (2589; Cell Signaling Technology), IκBα (4814; Cell Signaling Technology), p-NF-κB (3033; Cell Signaling Technology), NF-κB (10745-1-AP; Proteintech Group, Inc., Chicago, IL, USA), EGR1 (22008-1-AP; Proteintech Group, Inc.), FOS (ab11959; Abcam plc, Cambridge, UK), NF-E2 (11089-1-AP; Proteintech Group, Inc.), RUNX1 (25315-1-AP; Proteintech Group, Inc.), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) (60004-1-lg; Proteintech Group, Inc.).

Techniques: Expressing

Journal: Journal of Pharmaceutical Analysis

Article Title: Oxymatrine, a novel TLR2 agonist, promotes megakaryopoiesis and thrombopoiesis through the STING/NF-κB pathway

doi: 10.1016/j.jpha.2024.101054

Figure Lengend Snippet: Oxymatrine induces megakaryocyte (MK) differentiation by activating stimulator of interferon genes (STING)/nuclear factor-kappaB (NF-κB) pathway. (A) Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis for the potential pathway by which oxymatrine promotes MK differentiation. (B–E) Western blot (WB) analysis of the expression of pathway: extracellular regulated kinase 1/2 (ERK1/2)/STING/inhibitory subunit of nuclear factor kappa B alpha (IκBα)/NF-κB by which oxymatrine promotes MK differentiation. Representative immunoblot images and biochemical quantification of the ERK1/2 (B), STING (C), IκBα (D), and NF-κB (E) pathway after the treatment with oxymatrine (10, 20, and 40 μM) in Meg-01 cells for five days ( n = 3 per group). (F–I) Representative immunoblot images and biochemical quantification of the transcription factors (TFs) associated with MK differentiation: runt-related transcription factor 1 (RUNX1) (F), nuclear factor erythroid 2 (NF-E2) (G), early growth response protein 1 (EGR1) (H), and cellular oncogene fos (FOS) (I) ( n = 3 per group). (J) Representative immunofluorescence images of toll-like receptor 2 (TLR2) fluorescently triple-stained with p-STING and p-NF-κB, respectively, on bone marrow (BM) cells after 12 days of treatment with oxymatrine in a mouse model of radiation thrombocytopenia. Blue for the 4′,6-diamidino-2-phenylindole (DAPI), purple for the TLR2, red for the p-STING, and green for the p-NF-κB. (K, L) Representative images of immunofluorescence of the nuclear translocation of NF-E2 (K) and RUNX1 (L) in Meg-01 cells after the treatment of oxymatrine for five days. Excitation wavelengths: 470 nm for NF-E2 and RUNX1 and 405 nm for DAPI. Data represent the mean ± standard deviation (SD) of three independent experiments. ∗ P < 0.05, ∗∗ P < 0.01, and ∗∗∗ P < 0.001 vs. the control group. EGFR: epidermal growth factor receptor; IL-17: interleukin 17; VEGF: vascular endothelial growth factor ; TNF: tumor necrosis factor; JAK-STAT: janus kinase-signal transducer and activator of transcription; PI3K-Akt: phospholnositide-3 kinase-threonine kinase B; MAPK: mitogen-activated protein kinase; FoxO: forkhead box protein O; GAPDH: glyceraldehyde-3-phosphate dehydrogenase.

Article Snippet: After the addition of 5% bovine serum albumin (BSA), the cells were incubated at 4 °C overnight with anti-RUNX1 and anti-NF-E2 antibodies (Proteintech Group, Inc., Wuhan, China).

Techniques: Western Blot, Expressing, Immunofluorescence, Staining, Translocation Assay, Standard Deviation, Control

Journal: Journal of Pharmaceutical Analysis

Article Title: Oxymatrine, a novel TLR2 agonist, promotes megakaryopoiesis and thrombopoiesis through the STING/NF-κB pathway

doi: 10.1016/j.jpha.2024.101054

Figure Lengend Snippet: The schematic illustration of oxymatrine regulating megakaryocyte (MK) differentiation and thrombopoiesis. Oxymatrine targets toll-like receptor 2 (TLR2) and activates the downstream extracellular regulated kinase 1/2 (ERK1/2)/stimulator of interferon genes (STING)/inhibitory subunit of nuclear factor kappa B alpha (IκBα)/nuclear factor-kappaB (NF-κB) signaling pathway, which leads to the high expression of transcription factors (TFs): early growth response 1 (EGR1), nuclear factor erythroid 2 (NF-E2), cellular oncogene fos (FOS), and runt-related transcription factor 1 (RUNX1). As a result, the activated transcription factors promote the expression of megakaryopoiesis and thrombopoiesis related genes, which contribute to the MK maturation and platelet production and finally recover the platelet levels in adiation-induced thrombocytopenia mice. RIT: radiation-induced thrombocytopenia; HSC: hematopoietic stem cell; ERK: extracellular regulated kinase; Ub: ubiquitin.

Article Snippet: After the addition of 5% bovine serum albumin (BSA), the cells were incubated at 4 °C overnight with anti-RUNX1 and anti-NF-E2 antibodies (Proteintech Group, Inc., Wuhan, China).

Techniques: Expressing