Journal: Microbiology Spectrum

Article Title: Toxoplasma gondii impairs CX3CL1/fractalkine shedding from mouse cortical neurons, leading to microglia activation

doi: 10.1128/spectrum.01074-25

Figure Lengend Snippet: Characterization of neuron-derived angiogenic factors in latently infected cultures. Neuronal conditioned media (nCM) was collected at 7 dpi from two independent cultures and applied to angiogenic proteome array membranes. One representative array of uninfected culture is depicted in ( A ). Target proteins are spotted in duplicates, marked with blue squares, and their identities are revealed by the numbers written next to each square, as follows: CX3CL1 (1), IGFBP-2 (2), IGFBP-3 (3), CCL-2 (4), CCL-3 (5), NOV (6), PDGF-AA (7), CXCL12 (8), and VEGF (9). Densitometric analyses of two control and two infected cultures are shown in ( B ) for each detected analyte. Each dot in graphs represent the signal intensity in pixels from two spots average from each membrane, for each analyte.

Article Snippet: Secretion of angiogenesis-related protein levels was detected using a Proteome Profiler Mouse Angiogenesis Antibody Array kit (R&D Systems) according to the manufacturer’s instructions.

Techniques: Derivative Assay, Infection, Control, Membrane

Journal: Microbiology Spectrum

Article Title: Toxoplasma gondii impairs CX3CL1/fractalkine shedding from mouse cortical neurons, leading to microglia activation

doi: 10.1128/spectrum.01074-25

Figure Lengend Snippet: Angiogenic factors identified in neuronal cultures. Based on the mini proteome assays, three main regulators of endothelial cell biology were detected: VEGF, ADAMTS1 and endothelin-1. Neuronal cultures were analyzed by RT-qPCR at 7 dpi for Vegfa , Adamts1 , Tsp1 , Lrp1 , Edn1, and Tgfb1 expression ( A ). Data are shown as the relative expression, normalized by the controls (represented by the red dashed line). Mouse brain cortices were analyzed after 10 and 40 days of T. gondii infection ( B–M ) by RT-qPCR or western blotting. Vegfa ( B ), VEGFR1 ( Flt1 , C ), VEGFR2 ( Flk1 , D ) transcripts remained unaltered in infected brains, whereas VEGFR2 protein levels were increased at 40 dpi ( E and F ). Adamts1 ( G ), Tsp1 ( I ), Tgfb1 ( J ), and Edn1 ( K ) mRNA levels were increased at 40 dpi in infected cortices, and Lrp1 ( I ) had no significant changes. No changes were found in endothelin-1 protein levels ( L and M ). ( F and M ) show representative blots for VEGFR2 and Endothelin-1, respectively. For VEGFR, signal from the strongest Ponceau-stained band signal was used as loading control, and for endothelin-1, α-tubulin was used. Each dot in ( A ) corresponds to independent neuronal cultures; in ( B–L ), dots represent independent mice from at least two rounds of infections. * P < 0.05, ** P < 0.01. Unpaired Student’s T test in ( B ) and two-way ANOVA with Bonferroni post-test in ( B–L ).

Article Snippet: Secretion of angiogenesis-related protein levels was detected using a Proteome Profiler Mouse Angiogenesis Antibody Array kit (R&D Systems) according to the manufacturer’s instructions.

Techniques: Quantitative RT-PCR, Expressing, Infection, Western Blot, Staining, Control

Journal: Microbiology Spectrum

Article Title: Toxoplasma gondii impairs CX3CL1/fractalkine shedding from mouse cortical neurons, leading to microglia activation

doi: 10.1128/spectrum.01074-25

Figure Lengend Snippet: Endothelium-modulating growth factors released by T. gondii -infected neurons. Mini-proteome assays using nCM revealed differential secretion of matrikines IGFBP-2, IGFBP-3, and NOV, as well as of PDGF-AA. Neuronal cultures were infected with T. gondii ME49 strain tachyzoites, and mRNA was analyzed at 7 dpi for Igfp2 , Igfbp3 , Ccn3 (NOV), Ccn1 (Cyr61), Pdgfb, and Pdgfra by RT-qPCR ( A ). Data are shown as the relative expression, normalized by the controls (represented by the red dashed line). IGFBP3, Cyr61, and PDGFRA transcripts were significantly increased in infected cultures. Mouse brain cortices were analyzed after 10 and 40 days of T. gondii infection ( B–H ) by RT-qPCR for the same target genes, as well as for IGFBP3 receptor, TMEM219 ( D ). IGFBP3 was significantly increased in mouse cortices at 40 dpi. Each dot in ( A ) corresponds to independent neuronal cultures; in ( B–H ), dots represent independent mice from at least two rounds of infections. * P < 0.05, ** P < 0.01. Unpaired Student’s T test in ( A ) and two-way ANOVA with Bonferroni post-test in ( B–H ).

Article Snippet: Secretion of angiogenesis-related protein levels was detected using a Proteome Profiler Mouse Angiogenesis Antibody Array kit (R&D Systems) according to the manufacturer’s instructions.

Techniques: Infection, Quantitative RT-PCR, Expressing

Journal: Microbiology Spectrum

Article Title: Toxoplasma gondii impairs CX3CL1/fractalkine shedding from mouse cortical neurons, leading to microglia activation

doi: 10.1128/spectrum.01074-25

Figure Lengend Snippet: CX3CL1/Fractalkine pathway is disrupted by T. gondii latent infection in neurons. Fractalkine was consistently reduced in infected nCMs as shown by mini-proteome assays. Cx3cl1 gene secretion was drastically reduced in infected cultures, as shown by ELISA ( A ) although no changes to gene expression were observed ( B ). Cathepsin S ( Ctss ), Adam10, and Adam17 , known to be involved in fractalkine shedding, had their expression levels assessed by RT-qPCR ( C–E ). Fractalkine immunoreactivity and localization were analyzed by confocal microscopy ( F ). Neurons were stained with anti-β-III-tubulin (in red) and CX3CL1 (in green). Insets in F show CX3CL1 (in green) and DAPI (in blue) signals in higher magnification from the areas delimited by the dashed line squares. CX3CL1/fractalkine fluorescent signal was increased in T. gondii -infected dishes as compared to controls ( G ). CX3CL1 was decreased in brain cortices of infected mouse at 10 and 40 dpi, as shown by RT-qPCR ( H ) and western blotting ( I and J ), whereas Cx3cr1 and Ctss transcripts were increased at 40 dpi ( K and L ). ADAM10 ( M ) and ADAM17 ( N ) had no change to mRNA levels, but ADAM17 protein levels were increased at 40 dpi ( O and P ). ( J and P ) show representative blots for CX3CL1 and ADAM17, respectively. * P < 0.05, ** P < 0.01, *** P < 0.001. Unpaired Student’s t -test in ( A–E and G ); two-way ANOVA with Bonferroni post-test in ( H–O ). Each dot in graphs in ( A–E ) corresponds to independent neuronal culture preparation; in ( G ), to microscopic fields (with 20× objective) from two independent cultures; in ( H–O ), to individual mouse cortices, from at least two rounds of infection. Scale bars: 50 µm.

Article Snippet: Secretion of angiogenesis-related protein levels was detected using a Proteome Profiler Mouse Angiogenesis Antibody Array kit (R&D Systems) according to the manufacturer’s instructions.

Techniques: Infection, Enzyme-linked Immunosorbent Assay, Gene Expression, Expressing, Quantitative RT-PCR, Confocal Microscopy, Staining, Western Blot

Journal: Advanced Science

Article Title: ASH2L‐K312‐Lac Stimulates Angiogenesis in Tumors to Expedite the Malignant Progression of Hepatocellular Carcinoma

doi: 10.1002/advs.202509477

Figure Lengend Snippet: ASH2L lactylation affects angiogenesis in HCC tumor cells via VEGFA. a) The developed Huh7‐ASH2L‐WT and Huh7‐ASH2L‐K312R cells were subjected to transcriptome sequencing (bulk mRNA‐seq). b) Bulk mRNA‐seq data analysis showed that ASH2L‐K312‐lac effectively facilitated the upregulation of VEGFA expression to promote angiogenesis in HCC. c) Analysis using a Proteome Profiler Human Angiogenesis Antibody Array Detection Kit revealed reduced VEGFA secretion in Huh7‐ASH2L‐K312R cells. d) Quantitative polymerase chain reaction (qPCR) and enzyme‐linked immunosorbent assay (ELISA) results revealed reduced VEGFA expression and secretion in Huh‐7 cells with the K312R mutation. e) Tube formation assays revealed that the angiogenic potential of HUVECs to form capillary‐like structures in the supernatant of Huh7‐ASH2L‐K312R cells was significantly lower than that of Huh7‐ASH2L‐WT group. f) The supernatants of Huh7‐ASH2L‐WT and Huh7‐ASH2L‐K312R cells were separately combined with Matrigel for use in Matrigel plug assays in mice. g) Images of the retrieved Matrigel plugs. Compared with the Matrigel plug containing supernatant from the control group, those containing supernatant from the ASH2L‐K312R group showed more vascular infiltration (left), supporting the IHC results (right).

Article Snippet: A Proteome Profiler Human Angiogenesis Antibody Array Kit (ARY007, R&D Systems) was used to detect changes in cytokine levels in the cell supernatants.

Techniques: Sequencing, Expressing, Ab Array, Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay, Mutagenesis, Control

Journal: Matrix biology : journal of the International Society for Matrix Biology

Article Title: Role of syndecan-4 in angiogenesis and vasculogenic mimicry in triple negative breast cancer cells.

doi: 10.1016/j.matbio.2025.02.002

Figure Lengend Snippet: Fig. 2. The angiogenic profile of TNBC cells is altered by SDC4 knockdown. The profile of angiogenesis-regulating molecules secreted by control and SDC4 siRNA transfected cells, both cultivated alone and in a 3D co-culture system with HUVECs, was evaluated using the Proteome Profiler TM Human Angiogenesis Antibody Array (A-D). Conditioned media from the secretome of control or SDC4 siRNA-treated TNBC cells, co-cultured in a 3D with HUVECs were collected for analysis. (E-H) Immunoreactive signals on array membranes with colored boxes highlighting common and cell type-specific dysregulated factors.

Article Snippet: Proteome profiler human angiogenesis array The expression levels of 55 angiogenesis-related proteins in the supernatant of 3D co-cultures and TNBC cells was analyzed using the Proteome ProfilerTM Human Angiogenesis Antibody array (R&D Systems, cat. ARY007, Minneapolis, MN, USA), following the manufacturer’s instructions.

Techniques: Knockdown, Control, Transfection, Co-Culture Assay, Ab Array, Cell Culture