Journal: Cell communication and signaling : CCS

Article Title: HMGB3 promotes the malignant phenotypes and stemness of epithelial ovarian cancer through the MAPK/ERK signaling pathway.

doi: 10.1186/s12964-023-01172-7

Figure Lengend Snippet: Fig. 1 HMGB3 promotes ovarian cancer proliferation. A pLKO.1, HMGB3 shRNA-1 (shHMGB3-1), HMGB3 shRNA-2 (shHMGB3-2), pCMV, and pCMV HMGB3 plasmids were stably transfected into A2780 and SKOV3 cells. HMGB3 protein levels were determined by western blot. B Cells were seeded into 96-well plates and cultured for 1, 2, 3, 4, and 5 days, and an MTT assay was performed to assess cell viability. C A clonogenic assay was used to assess the colony formation efficiency of A2780 and SKOV3 cells with HMGB3 knocked down or overexpressed. D Quantification of the number of clones in C. E Proliferation of A2780 and SKOV3 cells with HMGB3 knocked down or overexpressed detected by EdU assay. Nuclei were stained using DAPI. Scale bar, 10 μm. F Quantification of the ratio of EdU positive cells in (E). Data are presented as the mean ± SEM, *p < 0.05, **p < 0.01, n = 3

Article Snippet: Antibodies against HMGB3 (27465-1-AP), ETS-1 (12118-1-AP), MEK1/2 (11049-1-AP), CCND1 (60186- 1-Ig), c-Myc (10828-1-AP), SOX2 (11064-1-AP), and ALDH1A1 (15910-1-AP) were purchased from Proteintech (Wuhan, China).

Techniques: shRNA, Stable Transfection, Transfection, Western Blot, Cell Culture, MTT Assay, Clonogenic Assay, Clone Assay, EdU Assay, Staining

Journal: Cell communication and signaling : CCS

Article Title: HMGB3 promotes the malignant phenotypes and stemness of epithelial ovarian cancer through the MAPK/ERK signaling pathway.

doi: 10.1186/s12964-023-01172-7

Figure Lengend Snippet: Fig. 3 RNA sequencing analysis of signaling pathways involved in HMGB3 function. A2780 cells were transfected with HMGB3 siRNA (siHMGB3) or negative control siRNA (Ctr) for 48 h. High-throughput RNA sequencing analysis was used to compare mRNA expression profiles of the siHMGB3 and Ctr groups. A Volcano plot showing differentially expressed genes (DEGs) between the siHMGB3 and Ctr groups. In total, 91 genes were up-regulated (Up) and 679 genes were down-regulated (Down); other genes expression levels were not significantly altered (no-DEGs) (B) Kyoto Encyclopedia of Genes and Genomes enrichment analysis of genes down-regulated in the siHMGB3 group relative to the Ctr group. C Heatmap showing genes involved in stem cell pluripotency and the MAPK signaling pathway down-regulated in the siHMGB3 group relative to the Ctr group. D A2780 cells were transfected with HMGB3 siRNA (siHMGB3) or negative control siRNA (Ctr) for 48 h, and qRT-PCR used to verify down-regulation of representative genes in the siHMGB3 group relative to the Ctr group. Data are presented as the mean ± SEM, *p < 0.05, **p < 0.01, n = 3

Article Snippet: Antibodies against HMGB3 (27465-1-AP), ETS-1 (12118-1-AP), MEK1/2 (11049-1-AP), CCND1 (60186- 1-Ig), c-Myc (10828-1-AP), SOX2 (11064-1-AP), and ALDH1A1 (15910-1-AP) were purchased from Proteintech (Wuhan, China).

Techniques: RNA Sequencing, Protein-Protein interactions, Transfection, Negative Control, High Throughput Screening Assay, Expressing, Quantitative RT-PCR

Journal: Cell communication and signaling : CCS

Article Title: HMGB3 promotes the malignant phenotypes and stemness of epithelial ovarian cancer through the MAPK/ERK signaling pathway.

doi: 10.1186/s12964-023-01172-7

Figure Lengend Snippet: Fig. 4 HMGB3 activates the MAPK/ERK signaling pathway in ovarian cancer cells. A p-MEK1/2, MEK1/2, p-ERK1/2, ERK1/2, ETS-1, CCND1, c-Myc, HMGB3, and β-actin protein levels of in A2780 and SKOV3 cells with HMGB3 knocked down or overexpressed detected by western blot. B Quantification of the protein levels in (A). C Protein levels in ovarian cancer cells transfected with pLKO.1, HMGB3 shRNA-1 (shHMGB3-1) and/or pCMV HMGB3 detected by western blot. (D) Quantification of the protein levels in (C). Data are presented as the mean ± SEM, *p < 0.05, **p < 0.01, n = 3

Article Snippet: Antibodies against HMGB3 (27465-1-AP), ETS-1 (12118-1-AP), MEK1/2 (11049-1-AP), CCND1 (60186- 1-Ig), c-Myc (10828-1-AP), SOX2 (11064-1-AP), and ALDH1A1 (15910-1-AP) were purchased from Proteintech (Wuhan, China).

Techniques: Western Blot, Transfection, shRNA

Journal: Cell communication and signaling : CCS

Article Title: HMGB3 promotes the malignant phenotypes and stemness of epithelial ovarian cancer through the MAPK/ERK signaling pathway.

doi: 10.1186/s12964-023-01172-7

Figure Lengend Snippet: Fig. 6 HMGB3 promotes the malignant phenotypes of ovarian cancer via the MAPK/ERK signaling pathway. A A2780 and SKOV3 cells with HMGB3 overexpression were seeded into 6 cm dishes and then treated with or without AZD6244 (5 µM)/PD0325901 (10 µM) for 24 h. p-ERK1/2, ERK1/2, HMGB3, and β-actin protein levels detected by western blot. B A2780 and SKOV3 cells with HMGB3 overexpression were seeded into 96-well plates and then treated with or without AZD6244 (5 µM)/PD0325901 (10 µM) for 72 h. Cell viability was detected by MTT assay. C A2780 and SKOV3 cells with HMGB3 overexpression were treated with or without AZD6244 (1 µM)/PD0325901 (2 µM) for 1–2 weeks. Colony formation efficiency was assessed by clonogenic assay. D Quantification of the number of clones in (C). E A2780 and SKOV3 cells with HMGB3 overexpression were seeded into transwell plates and then treated with or without AZD6244 (5 µM)/PD0325901 (10 µM) for 12–24 h. Cell migration and invasion were evaluated by transwell assay. Scale bar, 50 μm. F Quantification of the number of cells in (E). Data are presented as the mean ± SEM, **p < 0.01, n = 3

Article Snippet: Antibodies against HMGB3 (27465-1-AP), ETS-1 (12118-1-AP), MEK1/2 (11049-1-AP), CCND1 (60186- 1-Ig), c-Myc (10828-1-AP), SOX2 (11064-1-AP), and ALDH1A1 (15910-1-AP) were purchased from Proteintech (Wuhan, China).

Techniques: Over Expression, Western Blot, MTT Assay, Clonogenic Assay, Clone Assay, Migration, Transwell Assay

Journal: Cell communication and signaling : CCS

Article Title: HMGB3 promotes the malignant phenotypes and stemness of epithelial ovarian cancer through the MAPK/ERK signaling pathway.

doi: 10.1186/s12964-023-01172-7

Figure Lengend Snippet: Fig. 7 HMGB3 promotes ovarian cancer cell proliferation in vivo through the MAPK/ERK signaling pathway. A2780 cells (5 × 106) transfected with pLKO.1, HMGB3 shRNA-1 (shHMGB3-1), or HMGB3 shRNA-2 (shHMGB3-2) were subcutaneously injected into nude mice. Mice were divided into three groups: pLKO.1 (Ctr), shHMGB3-1, and shHMGB3-2 (n = 6 per group). Two weeks post-injection, mice were euthanized and the xenograft tumors were removed. A Tumors from each group are shown. B Tumor volumes of each group. C Body weight of each group. D p-MEK1/2, MEK1/2, p-ERK1/2, ERK1/2, HMGB3, and β-actin protein levels in tumor tissues detected by western blot. E Quantification of the protein levels in (D). A2780 cells (5 × 106) transfected with pCMV or pCMV HMGB3 were subcutaneously injected into nude mice. Mice were divided into three groups: pCMV (Ctr), pCMV HMGB3, and pCMV HMGB3 + AZD6244 (n = 6 per group). One group of mice received an intraperitoneal injection of AZD6244 (25 mg/ kg) once a day. Two weeks post-injection, mice were euthanized and xenograft tumors were removed. F Tumors from each group are shown. G Tumor volumes of each group. H Body weight of each group. I p-MEK1/2, MEK1/2, p-ERK1/2, ERK1/2, HMGB3, and β-actin protein levels in tumor tissues detected by western blot. J Quantification of the protein levels in (I). Data are presented as mean ± SEM, #p > 0.05, **p < 0.01, n = 6

Article Snippet: Antibodies against HMGB3 (27465-1-AP), ETS-1 (12118-1-AP), MEK1/2 (11049-1-AP), CCND1 (60186- 1-Ig), c-Myc (10828-1-AP), SOX2 (11064-1-AP), and ALDH1A1 (15910-1-AP) were purchased from Proteintech (Wuhan, China).

Techniques: In Vivo, Transfection, shRNA, Injection, Western Blot

Journal: Cell communication and signaling : CCS

Article Title: HMGB3 promotes the malignant phenotypes and stemness of epithelial ovarian cancer through the MAPK/ERK signaling pathway.

doi: 10.1186/s12964-023-01172-7

Figure Lengend Snippet: Fig. 8 Schematic summary of the study findings. HMGB3 overexpression activates the MAPK/ERK signaling pathway, thereby promoting ovarian cancer proliferation, metastasis, and stemness. Inhibition of the MAPK/ERK signaling pathway using specific inhibitors counteracts the effects of HMGB3

Article Snippet: Antibodies against HMGB3 (27465-1-AP), ETS-1 (12118-1-AP), MEK1/2 (11049-1-AP), CCND1 (60186- 1-Ig), c-Myc (10828-1-AP), SOX2 (11064-1-AP), and ALDH1A1 (15910-1-AP) were purchased from Proteintech (Wuhan, China).

Techniques: Over Expression, Inhibition

Journal: Nature communications

Article Title: Decoding the genomic landscape of chromatin-associated biomolecular condensates.

doi: 10.1038/s41467-024-51426-2

Figure Lengend Snippet: Fig. 4 | Experimental validation of potential biomolecular condensates. a Immunofluorescence images of mESC showing that SUPT6H (green) colocalizes with CTR9 (red) and SUPT5H (grey) in puncta. DNA was stained with DAPI (blue). This experimental result was consistent across two independent cell-seeding, fixation, and co-IF staining experiments (each contains three slides). Scale bar: 10 μm. b Line scans of the images of a cell co-stained for SUPT6H, CTR9 and SUPT5H, at the position depicted by the white line. The direction is from the green tick to the purple tick, and the two arrows refer to two representative puncta. FRAP experiments for SUPT6H (c), CTR9 (d) and SUPT5H (e). Left, representative images of the FRAP experiment. The white arrow refers to the punctum undergoing bleaching. Right, quantification of FRAP data for puncta of SUPT6H (n = 6), CTR9 (n = 5) and SUPT5H (n = 5). Puncta were photobleached at t = 0 s, and data were plot

Article Snippet: Following this, the ConA beads bound cells were collected using a magnet and resuspended in 100 μl of antibody buffer containing either 2 μl of DDX21 (Proteintech, 10528-1- AP, lot # 00088037), 4 μl of CTR9 (Bethyl Laboratories, A301-395A, lot # 4), 4 μl of SUPT6H (Novus Biologicals, NB100-2582, lot # 2 A), 1.5 μl of SS18 (Cell Signaling Technology, 21792 (D6I4Z), lot # 1), 1.5 μl of EP300 (Santa Cruz, sc-48343 (F-4), lot # A1323), or 0.5 μl of ELL3 (generously gifted by Prof. Chengqi Lin, Southeast University, China) primary Nature Communications | (2024) 15:6952 13 antibody respectively.

Techniques: Biomarker Discovery, Staining

Journal: Journal of nanobiotechnology

Article Title: m6A-modified exosome-derived circHIF1α binding to KH domain of IGF2BP3 mediates DNA damage and arrests G1/S transition phase to resists bacterial infection in bacteremia.

doi: 10.1186/s12951-024-02932-4

Figure Lengend Snippet: Fig. 3 Exosome and exosome-derived circHIF1α inhibits PK-15 cells proliferation. A–B The relative expression of circHIF1α in the precipitation or super natant of PIEC/PK-15 cells after PIEC cells treated with G. parasuis. C The relative expression of circHIF1α in PK-15 cells treated with exosomes from over expressed circHIF1α PIEC cells. D–G The proliferation of PK-15 cells treated with exosomes from PIEC cells after circHIF1α overexpression or knockdown was evaluated by CCK-8 (D and F) and EdU assay (E and G). Scale bar, 75 μm. H–I. The proliferation of PK-15 cells transfected with circHIF1α siRNA or overexpression vector was evaluated by CCK-8 (H) and EdU assay (I). Scale bar, 100 μm. J Mass spectrometric data of interaction between circHIF1α and ptotein showed the important correlated biological process. K The pathway result of mass spectrometry after RNA pulldown assay

Article Snippet: In the immunofluorescence assay, PK-15 cells were transiently transfected with Cy3labelled circHIF1α and proceeded to FISH assay, and then incubated with anti-IGF2BP3 antibody (Proteintech, Chicago, USA) to observe the colocalization of circHIF1α and IGF2BP3.

Techniques: Derivative Assay, Expressing, Over Expression, Knockdown, CCK-8 Assay, EdU Assay, Transfection, Plasmid Preparation, Mass Spectrometry

Journal: Journal of nanobiotechnology

Article Title: m6A-modified exosome-derived circHIF1α binding to KH domain of IGF2BP3 mediates DNA damage and arrests G1/S transition phase to resists bacterial infection in bacteremia.

doi: 10.1186/s12951-024-02932-4

Figure Lengend Snippet: Fig. 4 Exosome and exosome-derived circHIF1α promotes PK-15 cells DNA damage, and mediates the G1/S phase. A–B The DNA damage level of PK-15 cells treated with exosomes from PIEC cells after circHIF1α overexpression or knockdown was detected by γ-H2AX immunofluorescence. Scale bar, 75 μm. C Cell cycle analysis was executed by flow cytometry when PK-15 cells were treated with exosomes from PIEC cells after circHIF1α knockdown. D Western blot showing the levels of DNA damage-related proteins, including γ-H2AX, NPM1, p53, and p-p53 in PK-15 cells transfected with ov-circHIF1α or ov-pLC5. E Cell cycle analysis was executed by flow cytometry after circHIF1α overexpression for 24 h. F The expression level of cell cycle-related proteins was detected in PK-15 cells after circHIF1α overexpression for 24 h by Western blot. Data are represented as mean ± SD. NS, not signifcant, *p < 0.05, **p < 0.01, ***p < 0.001

Article Snippet: In the immunofluorescence assay, PK-15 cells were transiently transfected with Cy3labelled circHIF1α and proceeded to FISH assay, and then incubated with anti-IGF2BP3 antibody (Proteintech, Chicago, USA) to observe the colocalization of circHIF1α and IGF2BP3.

Techniques: Derivative Assay, Over Expression, Knockdown, Immunofluorescence, Cell Cycle Assay, Flow Cytometry, Western Blot, Transfection, Expressing

Journal: Journal of nanobiotechnology

Article Title: m6A-modified exosome-derived circHIF1α binding to KH domain of IGF2BP3 mediates DNA damage and arrests G1/S transition phase to resists bacterial infection in bacteremia.

doi: 10.1186/s12951-024-02932-4

Figure Lengend Snippet: Fig. 5 CircHIF1α reduces bacterial adhesion and invasion for PK-15 cell. A–C The quantity of PK-15 cells infected with G. parasuis (A)/S. aureus (B)/SS2 (C) was identified by the viable counting method. D–F PK-15 cells infected with G. parasuis(D) /S. aureus (E)/SS2 (F) was identified by IF. Scale bar, 25 μm. G–I The quantity of PK-15 cells infected with G. parasuis (G) /S. aureus (H) /SS2 (I) was identified after PK-15 cells were treated with 250nM NSC 80,467 by bacteria adhesion and invasion assays. J–L. The quantity of PK-15 cells infected with G. parasuis (J), S. aureus (K), and SS2 (L) was identified after PK-15 cells were treated with 2mM Thymidine by bacteria adhesion and invasion assays. Data are represented as mean ± SD. NS, not signifcant, *p < 0.05, **p < 0.01, ***p < 0.001

Article Snippet: In the immunofluorescence assay, PK-15 cells were transiently transfected with Cy3labelled circHIF1α and proceeded to FISH assay, and then incubated with anti-IGF2BP3 antibody (Proteintech, Chicago, USA) to observe the colocalization of circHIF1α and IGF2BP3.

Techniques: Infection, Bacteria

Journal: Journal of nanobiotechnology

Article Title: m6A-modified exosome-derived circHIF1α binding to KH domain of IGF2BP3 mediates DNA damage and arrests G1/S transition phase to resists bacterial infection in bacteremia.

doi: 10.1186/s12951-024-02932-4

Figure Lengend Snippet: Fig. 8 Exosomal circHIF1α resists bacterial infection in vivo. A Left, Bioluminescent image showed localization of G. parasuis-exosome (20 mg/kg) or Mock-exosome (20 mg/kg) in mice after injecting through the tail vein at different times. Right, the fluorescence intensity of exosomes in vivo. n = 6 mice/ group. B The fluorescence intensity of G. parasuis-exosome (20 mg/kg) or Mock-exosome (20 mg/kg) in different organs of mice. n = 6 mice/group. C–E The effect of exosomes and circHIF1α on the survival of mice after G. parasuis (C)/SS2 (D)/S. aureus (E) infection. n = 6 mice/group. F–H The effect of exo somes and circHIF1α on G. parasuis (F)/SS2 (G)/S. aureus (H) count of different organs after mice were infected with G. parasuis. I H&E staining of various organs after mice were infected with G. parasuis followed treating with exosome or circHIF1α. Scale bar, 50 μm. n = 6 mice/group. Data are represented as mean ± SD. *p < 0.05, **p < 0.01

Article Snippet: In the immunofluorescence assay, PK-15 cells were transiently transfected with Cy3labelled circHIF1α and proceeded to FISH assay, and then incubated with anti-IGF2BP3 antibody (Proteintech, Chicago, USA) to observe the colocalization of circHIF1α and IGF2BP3.

Techniques: Infection, In Vivo, Fluorescence, Staining

Journal: Journal of nanobiotechnology

Article Title: m6A-modified exosome-derived circHIF1α binding to KH domain of IGF2BP3 mediates DNA damage and arrests G1/S transition phase to resists bacterial infection in bacteremia.

doi: 10.1186/s12951-024-02932-4

Figure Lengend Snippet: Fig. 9 Proposed model for the potential function of exosomal circHIF1α in bacterial infection progression

Article Snippet: In the immunofluorescence assay, PK-15 cells were transiently transfected with Cy3labelled circHIF1α and proceeded to FISH assay, and then incubated with anti-IGF2BP3 antibody (Proteintech, Chicago, USA) to observe the colocalization of circHIF1α and IGF2BP3.

Techniques: Infection