Journal: European Journal of Medical Research

Article Title: Olfactory mucosa-mesenchymal stem cells with overexpressed Nrf2 modulate angiogenesis and exert anti-inflammation effect in an in vitro traumatic brain injury model

doi: 10.1186/s40001-025-02344-6

Figure Lengend Snippet: Effects of OM-MSCs Nrf2 on the levels of relevant cytokines in PC12 cells. A Flow cytometry was used to characterize CD45, CD34, CD29, CD90, CD73, and CD105 of OM-MSCs. B RT-qPCR-based to assess transfection efficiency after overexpression of Nrf2 plasmid. OM-MSCs Nrf2 : NRF2 overexpression group. OM-MSCs (ctrl): Empty vector transfection group. OM-MSCs: Untransfected OM-MSCs. C Morphology of both OM-MSCs (ctrl) and OM-MSCs Nrf2 under the optical microscope. Scale bar: 100 μm. D The changes in cytokine concentrations in PC12 cells measured by ELISA, including IL4, IL10, IL13, VEGF-A, and TGF-β. Results of independent triplicates were expressed as mean ± standard deviation and the statistical difference between two groups was marked with asterisks (“ns” indicates no significant difference, * p < 0.05, and ** p < 0.01). OM-MSCs olfactory mucosa-mesenchymal stem cells, Nrf2 : Nuclear Factor Erythroid-Derived 2-Like 2, IL interleukin, VEGF-A vascular endothelial growth factor-A, TGF-β transforming growth factor beta

Article Snippet: Specifically, PC12 cells were cultured in Roswell Park Memorial Institute-1640 medium (#C3250, Solarbio® Life Sciences, China) supplemented with 15% horse serum (#S9050, Solarbio® Life Sciences, China) and 5% fetal bovine serum; BV-2 cells were cultured in high-glucose DMEM (#D5194, Solarbio® Life Sciences, China) supplemented with 10% fetal bovine serum; HUVECs were maintained in M199 medium (#11150067, Gibco, USA) including 15% fetal bovine serum, 1% penicillin–streptomycin, 30 μg/mL endothelial cell growth supplement (#211-GS, Sigma, Darmstadt, Germany), and 10 ng/mL epidermal growth factor (#E5036, Sigma, Germany).

Techniques: Flow Cytometry, Quantitative RT-PCR, Transfection, Over Expression, Plasmid Preparation, Microscopy, Enzyme-linked Immunosorbent Assay, Standard Deviation, Derivative Assay

Journal: European Journal of Medical Research

Article Title: Olfactory mucosa-mesenchymal stem cells with overexpressed Nrf2 modulate angiogenesis and exert anti-inflammation effect in an in vitro traumatic brain injury model

doi: 10.1186/s40001-025-02344-6

Figure Lengend Snippet: Effects of OM-MSCs Nrf2 on the survival and angiogenesis and proteins related to PI3K/AKT pathway and apoptosis in PC12 cells. A Effects of OM-MSCs (ctrl) and OM-MSCs Nrf2 on the survival of PC12 cells based on CCK-8 assay. B , C Angiogenesis assay in evaluating the angiogenesis of PC12 cells following the co-culture with OM-MSCs (ctrl) and OM-MSCs Nrf2 . scale bar: 200 μm D , E Ratio of p-PI3K to PI3K and p-AKT to AKT as well as BAX protein level in PC12 cells following the co-culture with OM-MSCs (ctrl) and OM-MSCs Nrf2 , and those cells treated with minocycline were applied as the positive control. Results of independent triplicates were expressed as mean ± standard deviation and the statistical difference between two groups was marked with asterisks ( * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001). OM-MSCs olfactory mucosa-mesenchymal stem cells, Nrf2 Nuclear Factor Erythroid-Derived 2-Like 2, OD optical density, p-PI3K phosphorylated-PI3K, PI3K Phosphoinositide 3-kinase, p-AKT phosphorylated-AKT, AKT protein kinase B, BAX BCL2 Associated X, GAPDH Glyceraldehyde-3-phosphate dehydrogenase

Article Snippet: Specifically, PC12 cells were cultured in Roswell Park Memorial Institute-1640 medium (#C3250, Solarbio® Life Sciences, China) supplemented with 15% horse serum (#S9050, Solarbio® Life Sciences, China) and 5% fetal bovine serum; BV-2 cells were cultured in high-glucose DMEM (#D5194, Solarbio® Life Sciences, China) supplemented with 10% fetal bovine serum; HUVECs were maintained in M199 medium (#11150067, Gibco, USA) including 15% fetal bovine serum, 1% penicillin–streptomycin, 30 μg/mL endothelial cell growth supplement (#211-GS, Sigma, Darmstadt, Germany), and 10 ng/mL epidermal growth factor (#E5036, Sigma, Germany).

Techniques: CCK-8 Assay, Angiogenesis Assay, Co-Culture Assay, Positive Control, Standard Deviation, Derivative Assay

Journal: European Journal of Medical Research

Article Title: Olfactory mucosa-mesenchymal stem cells with overexpressed Nrf2 modulate angiogenesis and exert anti-inflammation effect in an in vitro traumatic brain injury model

doi: 10.1186/s40001-025-02344-6

Figure Lengend Snippet: Relevant mechanisms related to OM-MSCs Nrf2 on the neuroinflammation mediators and glycolysis in PC12 cells. A Effects of OM-MSCs and OM-MSCs Nrf2 on the mRNA level of Nlrp3 , Il1b , Il18 , Ccl5 , P2ry12 , and Cx3cr1 in PC12 cells. B , C Effects of OM-MSCs (ctrl) and OM-MSCs Nrf2 on the ECAR ( B ) and OCR ( C ) of PC12 cells. Results of independent triplicates were expressed as mean ± standard deviation and the statistical difference between two groups was marked with asterisks ( * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001). OM-MSCs olfactory mucosa-mesenchymal stem cells, Nrf2 Nuclear Factor Erythroid-Derived 2-Like 2, Nlrp3 NOD-, LRR- and pyrin domain-containing protein 3; Il1b Interleukin 1 beta; Il18 Interleukin 18, Ccl5 C–C chemokine ligand 5, P2ry12 purinergic receptor P2Y12, Cx3cr1 C-X3-C motif chemokine receptor 1, ECAR extracellular acidification rate, oligo Oligomycin; OCR oxygen consumption rate, 2-DG 2-deoxy-D-glucose, FCCP Trifluoromethoxy carbonylcyanide phenylhydrazone

Article Snippet: Specifically, PC12 cells were cultured in Roswell Park Memorial Institute-1640 medium (#C3250, Solarbio® Life Sciences, China) supplemented with 15% horse serum (#S9050, Solarbio® Life Sciences, China) and 5% fetal bovine serum; BV-2 cells were cultured in high-glucose DMEM (#D5194, Solarbio® Life Sciences, China) supplemented with 10% fetal bovine serum; HUVECs were maintained in M199 medium (#11150067, Gibco, USA) including 15% fetal bovine serum, 1% penicillin–streptomycin, 30 μg/mL endothelial cell growth supplement (#211-GS, Sigma, Darmstadt, Germany), and 10 ng/mL epidermal growth factor (#E5036, Sigma, Germany).

Techniques: Standard Deviation, Derivative Assay

Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: Neutrophil-macrophage crosstalk via NETs–IL-17/VEGF/S100A9 axis promotes hepatocellular carcinoma progression

doi: 10.1186/s13046-025-03618-x

Figure Lengend Snippet: The levels of NETs increase in HCC patients, especially in advanced and metastatic stages. A A heatmap illustrating the differential expression patterns of NETs-related genes in normal and HCC samples based on TCGA database analysis. B Box plots depicting the expression differences of NETs-related genes between normal and HCC samples from the TCGA database. C , D Representative images of HE (Hematoxylin and eosin) and IF staining for CD66b (C) and CitH3 (D) in tissue sections from HCs and HCC patients. The mean IOD of CD66b or CitH3 was quantified (Image Pro Plus) from three random regions per section (HC, n = 5; HCC, n = 5) and shown as fold change relative to HCs. E Representative images of IF staining for CitH3 in peripheral blood neutrophils from HC and HCC patient F Western blot analysis of CitH3 expression in peripheral blood neutrophils from HCs ( n = 8) and HCC ( n = 8) patients, with band intensities normalized to β-actin and shown as fold change relative to controls G ELISA analysis of serum MPO-DNA levels in HCs and HCC patients. H ELISA analysis of serum MPO-DNA levels in HCC patients at different TNM stages (I–IV). I ELISA analysis of serum MPO-DNA levels in HCC patients with and without metastasis. J ELISA analysis of serum MPO-DNA levels in HCC patients with intrahepatic metastasis and extrahepatic metastasis. K Western blot analysis of CitH3 expression in peripheral blood neutrophils from HCC patients with ( n = 6) and without ( n = 6) extrahepatic metastasis. Band intensities were normalized to β-actin and quantified relative to the non-metastasis group White and black scale bars: 50 μm. Data are presented as median (IQR) (G-J) and other data are presented as mean ± SD. * p < 0.05, ** p < 0.01, *** p < 0.001. Student’s t test (C, D, F, K), Mann–Whitney U test (B, G, I, J), Kruskal–Wallis test (H)

Article Snippet: Neutrophils were isolated from the peripheral blood of healthy volunteers and HCC patients using the Human Peripheral Blood Neutrophil Isolation Kit (P9040, Solarbio, China), according to the manufacturer’s instructions.

Techniques: Quantitative Proteomics, Expressing, Staining, Western Blot, Enzyme-linked Immunosorbent Assay, MANN-WHITNEY

Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: Neutrophil-macrophage crosstalk via NETs–IL-17/VEGF/S100A9 axis promotes hepatocellular carcinoma progression

doi: 10.1186/s13046-025-03618-x

Figure Lengend Snippet: Activation of IL-17R/NF-κB signaling by NETs-attached IL-17 is responsible for NETs-induced M2 macrophage polarization. A Schematic illustration of the transcriptome sequencing protocol for THP1-M0 under three treatment conditions. The volcano plot shows significantly DEGs for Group B vs. A, Group C vs. A, and Group C vs. B. B KEGG pathway enrichment analysis of DEGs was conducted using the DAVID tool. C FCM analysis of the proportion of CD68 + CD206 + M2 derived from THP-1-M0 following 48-hour co-culture of HuH-7 cells with NETs and (or) AMG827. Quantification of the fold change in the proportion from three independent experiments is shown in the right panel. D Western blot analysis of IL-10 and VEGF expression in cell lysates following the same treatments as in C. Densitometric values were normalized to β-actin and compared to the control group, with quantification shown in the right panel E Western blot analysis of IL-17 expression in extracted and purified NETs from peripheral blood neutrophils of HCs ( n = 8) and HCC patients ( n = 8). F ELISA analysis of IL-17 levels carried by NETs standardized to 50 ng/µL in HCs ( n = 8) and patients with HCC ( n = 8). G Western blot analysis of IL-17 expression in peripheral blood neutrophils from HCs ( n = 8) and HCC ( n = 8) patients. Densitometric values were normalized to β-actin and shown as a fold change relative to HCs. H Representative IF images of IL-17 and NE staining in peripheral blood neutrophils with (lower panel) or without PMA stimulation (upper panel) for 4 h from HCs and HCC patients. I Western blot analysis was conducted to assess the expression levels of p-65 and p-p65(S536) in THP-1-M0 following 48 h co-culture of HuH-7 cells with NETs and (or) AMG827. Densitometric values were normalized to β-actin and shown as fold changes relative to control in the right panel. J FCM was used to analyze the proportion of CD68 + CD206 + M2 derived from THP-1-M0 following 48-hour co-culture of HuH-7 cells with NETs and (or) BAY11-7082. Quantification of the fold change in the proportion from three independent experiments is shown in the right panel. K IF analysis of CD206 + M2 macrophage from the above co-culture systems of J treatment for 48 h. L ELISA analysis of secretion levels of M2d-related cytokines VEGF, IL-10 and TGF-β, in the culture supernatant from the above co-culture systems of J treatment for 48 h White scale bars: 20 μm. Data are presented as mean ± SD. Ns, not significant. * p < 0.05, ** p < 0.01, *** p < 0.001. One-way ANOVA followed by the Newman-Keuls multiple comparison test (C, D, I, J, L); Student’s t test (F, G)

Article Snippet: Neutrophils were isolated from the peripheral blood of healthy volunteers and HCC patients using the Human Peripheral Blood Neutrophil Isolation Kit (P9040, Solarbio, China), according to the manufacturer’s instructions.

Techniques: Activation Assay, Sequencing, Derivative Assay, Co-Culture Assay, Western Blot, Expressing, Control, Purification, Enzyme-linked Immunosorbent Assay, Staining, Comparison

Journal: Nature Microbiology

Article Title: Evolution of enhanced innate immune suppression by SARS-CoV-2 Omicron subvariants

doi: 10.1038/s41564-023-01588-4

Figure Lengend Snippet: a – g , Calu-3 infection with 2,000 E copies per cell of Delta (yellow, Ο), BA.1 (blue, Ο), BA.2 (blue, Δ), BA.4 (purple, O) and BA.5 (purple, Δ), n = 3: mean viral E copies at 2 h.p.i. across three independent experiments ( a ); viral replication over time measured by RT–qPCR for intracellular E copies per microgram RNA ( b ); infection levels measured by nucleocapsid expression (% N+ by flow cytometry) ( c ); expression of IFNB , CXCL10 , IFIT1 , IFIT2 , RSAD2 , MX1 , MX2 and DDX58 in infected cells over time ( d ); IFNβ ( e ) and CXCL10 ( f ) secretion from infected Calu-3 cells measured by ELISA at 48 h.p.i.; rescue of viral replication by JAK1-inhibitor ruxolitinib in Calu-3 cells at 48 h.p.i., where relative infection levels are shown across three independent experiments determined by E copies per microgram RNA normalized to the median infection level of the untreated control ( g ). h – k , Primary bronchial HAEs were infected with the indicated variants at 1,500 E copies per cell: viral replication measured by intracellular E copies at 72 h.p.i. ( h ) and viral release into apical washes over time ( i ), with three biological replicates shown; expression of IFNB , CXCL10 , IFIT1 , IFIT2 , DDX58 and RSAD2 in HAEs at 72 h.p.i., with six biological replicates shown ( j ); intracellular viral E copies in HAEs in the presence or absence of 5 μM ruxolitinib at 72 h.p.i., with three biological replicates shown ( k ). For a , one-way analysis of variance (ANOVA) with Bonferroni post-test was used. n.s., not significant at P > 0.05 for all comparisons. For b – h and j , one-way ANOVA and Dunnett’s post-test were used. For i , two-way ANOVA with a Bonferroni post-test was used. For k , one-tailed unpaired Student’s t -test was used. Replicate measurements from one of three independent experiments. Fold change over mock is shown. Mean ± s.e.m. or individual datapoints are shown. h.p.i., hours post infection.

Article Snippet: IFNβ, IFNλ1/IFNλ3 and CXCL10 were measured using Human IFN-β Quantikine ELISA Kit, Human IL-29/IL-28B (IFNλ1/IFNλ3) DuoSet ELISA or Human CXCL10/IP-10 DuoSet ELISA reagents (Bio-Techne R&D Systems) according to the manufacturer’s instructions.

Techniques: Infection, Quantitative RT-PCR, Expressing, Flow Cytometry, Enzyme-linked Immunosorbent Assay, One-tailed Test

Journal: PLoS ONE

Article Title: High EMT Signature Score of Invasive Non-Small Cell Lung Cancer (NSCLC) Cells Correlates with NFκB Driven Colony-Stimulating Factor 2 (CSF2/GM-CSF) Secretion by Neighboring Stromal Fibroblasts

doi: 10.1371/journal.pone.0124283

Figure Lengend Snippet: Calu-1 and NCI-H1437 tumor cells were co-cultured with HDFs and two patient-derived NF/CAF pairs (NF1/CAF1, NF2/CAF2) for 24 h. Supernatants from mono- (green) and co-cultures (purple: Calu-1 and orange: NCI-H1437) were collected and tested for the indicated cytokines with at least three biological replicates, each representing two technical replicates (Cytokine Array), or three biological replicates, each representing three technical replicates (ELISA). (A) Cytokine Array: Relative pixel counts on the Y-axis. (B) ELISA: The cytokine level is depicted on the Y-axis [pg/ml]. Statistical analysis was performed by using Student’s t-test (for p-values see ).

Article Snippet: The following ELISA kits were purchased from R&D Systems for cross-validation of identified hits: IL-6 (D6050), IL-8 (D8000C), GCP-2 (DGC00), CXCL1 (DGR00), GM-CSF (DGM00) and VEGF (DVE00), respectively.

Techniques: Cell Culture, Derivative Assay, Enzyme-linked Immunosorbent Assay

Journal: PLoS ONE

Article Title: High EMT Signature Score of Invasive Non-Small Cell Lung Cancer (NSCLC) Cells Correlates with NFκB Driven Colony-Stimulating Factor 2 (CSF2/GM-CSF) Secretion by Neighboring Stromal Fibroblasts

doi: 10.1371/journal.pone.0124283

Figure Lengend Snippet: (A) Luciferase-based NFκB reporter activity in co-cultures of Calu-1 or NCI-H1437 with various FBs. 24 h after transfection of the respective cell type (depicted in the title of each graph) the NFκB-reporter plasmid-containing cell line was incubated with a non-transfected cell line as indicated on the X-axis for additional 24 h. The upper three graphs display studies with transfected FBs co-cultured with non-transfected tumor cell lines or with the non-transfected parental FB line, respectively. The lower two graphs exhibit the experiment with transfected tumor cell lines. Bars indicate fold changes of the Luciferase reporter signal mean values. For FBs lipofectamine transfection, for tumor cell lines electroporation transfection was applied. (B) NCI-H157 co-cultures were treated with trametinib and BI5700 for 24 h (4 μM each). The different FBs had been transfected prior with an NFκB-reporter plasmid. Luciferase units are depicted on the Y-axis. (C) BI5700 dose-dependent reduction of CSF2 transcription in HDFs. Relative CSF2 transcripts are depicted (RT-qPCR). Supernatant from a 24 h HDF mono- or Calu-1/HDF co-culture was transferred to an HDF mono-culture for another hour with or without BI5700 treatment. Incubation with RPMI 10% FCS medium served as a control. The level of CSF2 transcripts in the respective non BI5700-treated HDFs is indicated by [-∞]. Statistical analyses were performed on the mean values by unpaired comparisons using Student’s t-test (*p<0.05, **p<0.01, ***p<0.001, ****p<0.0001; n.s.: not significant). (D) ELISA for CSF2 (Y-axis [pg/ml]) of supernatants derived from mono- and co-cultures of invasive (Calu-1, NCI-H157) and non-invasive tumor cell lines (NCI-H1437) with HDFs. Supernatants were taken and replaced with fresh medium at the indicated time points (X-axis; time [h]). All experiments were performed in triplicates (n = 3).

Article Snippet: The following ELISA kits were purchased from R&D Systems for cross-validation of identified hits: IL-6 (D6050), IL-8 (D8000C), GCP-2 (DGC00), CXCL1 (DGR00), GM-CSF (DGM00) and VEGF (DVE00), respectively.

Techniques: Luciferase, Activity Assay, Transfection, Plasmid Preparation, Incubation, Cell Culture, Electroporation, Quantitative RT-PCR, Co-Culture Assay, Control, Enzyme-linked Immunosorbent Assay, Derivative Assay