Journal: Journal of Extracellular Biology

Article Title: Development of a Novel Extracellular Vesicle‐Based Biomarker Approach for Pediatric High‐Grade Glioma

doi: 10.1002/jex2.70117

Figure Lengend Snippet: PTPRZ1 and GLAST are reliable cell surface markers for radial glia (RG)‐like identity . (A) Single cell RNA‐seq analysis revealed 3 candidate genes highly specific to RG: GLAST, PTPRZ1 and ADGRV1 . (B) Multiple linear regression analysis showed the highest fitness with the combination of PTPRZ1 and GLAST (C) Double positive (DP) cells mainly fell into the RG and dividing cell cluster. (D) Elevated expression of other RG marker ( VIM, HES1, PAX6 and FABP7) in DP cells. Statistical analysis was performed using the Wilcoxon rank sum test (****: p < 0.0001).

Article Snippet: For nano‐flow cytometry analysis, 3 μL of SJ‐HGGX42‐derived EVs were incubated overnight at 4 degrees with gentle rotation with 1 μL of the antibody solution containing GLAST antibody conjugated to APC (1:800; Miltenyi Biotec, ACSA‐1, cat. no. 130‐123‐555, mouse monoclonal IgG2a) or PTP zeta antibody conjugated with FITC (1:1600; Bioss, cat. no. bs‐11327R‐FITC, rabbit polyclonal IgG).

Techniques: RNA Sequencing, Expressing, Marker

Journal: Journal of Extracellular Biology

Article Title: Development of a Novel Extracellular Vesicle‐Based Biomarker Approach for Pediatric High‐Grade Glioma

doi: 10.1002/jex2.70117

Figure Lengend Snippet: Expression of RG markers in pHGG patient‐derived cell lines . (A) RT‐qPCR of various HGG patient‐derived, control cells, and astrocytes showed the expression of RG‐like markers ( n = 3). (B) Representative flow cytometry density plots of HGG patient‐derived cell lines and astrocytes showing the percentage of double‐positive (PTPRZ1 + /GLAST + ) cells. The DP gate was set based on GLAST/PTPRZ1 expression on SH‐SY5Y (negative control) (C) Quantification of flow cytometry data, where each dot represents a biological replicate, and error bars indicate standard deviation ( n = 3–4). Statistical analysis was performed using the Kruskal‐Wallis test with multiple comparisons. *: p < 0.05, **: p < 0.01. (D) Live time‐lapse imaging from patient‐derived cell line SJ‐HGGX42 showing mitotic somal translocation (MST). Images acquired using 10X bright‐field microscopy over a 48‐h period with 2‐min intervals, under standard incubator conditions (37°C, 5% CO2).

Article Snippet: For nano‐flow cytometry analysis, 3 μL of SJ‐HGGX42‐derived EVs were incubated overnight at 4 degrees with gentle rotation with 1 μL of the antibody solution containing GLAST antibody conjugated to APC (1:800; Miltenyi Biotec, ACSA‐1, cat. no. 130‐123‐555, mouse monoclonal IgG2a) or PTP zeta antibody conjugated with FITC (1:1600; Bioss, cat. no. bs‐11327R‐FITC, rabbit polyclonal IgG).

Techniques: Expressing, Derivative Assay, Quantitative RT-PCR, Control, Flow Cytometry, Negative Control, Standard Deviation, Imaging, Translocation Assay, Microscopy

Journal: Journal of Extracellular Biology

Article Title: Development of a Novel Extracellular Vesicle‐Based Biomarker Approach for Pediatric High‐Grade Glioma

doi: 10.1002/jex2.70117

Figure Lengend Snippet: EVs derived from RG‐like glioma cells exhibited detectable levels of PTPRZ1 and GLAST markers . (A) Illustration of workflow for EV collection and characterization. (B) Nanoparticle tracking analysis (NTA) showing the size distribution of SJ‐HGGX42 EVs obtained by Nanosight NS300 (C) Transmission Electron Microscopy (TEM) image of EVs isolated from SJ‐HGGX42 cells by ExoQuick‐TC. (D) Western blot analysis of EV surface markers (CD63 and CD81) and negative control Calnexin. (E) Representative flow cytometry histograms of HGG patient‐derived EVs and astrocyte‐derived EVs, showing the APC signal (PTPRZ1 + /GLAST + EVs). The gate was set based on the control (secondary antibody only). (F) Quantification of flow cytometry data. Each dot represents a biological replicate and error bars indicate standard deviation ( n = 3–6). (G) Quantification of flow cytometry data of HGG patient‐derived EVs (SJ‐HGGX42) and astrocyte‐derived EVs isolated by ultracentrifugation, showing the APC signal (PTPRZ1 + /GLAST + EVs). Each dot represents a biological replicate and error bars indicate standard deviation ( n = 3). Statistical analysis was performed using one‐way ANOVA with multiple comparisons. **: p < 0.01, ****: p < 0.0001.

Article Snippet: For nano‐flow cytometry analysis, 3 μL of SJ‐HGGX42‐derived EVs were incubated overnight at 4 degrees with gentle rotation with 1 μL of the antibody solution containing GLAST antibody conjugated to APC (1:800; Miltenyi Biotec, ACSA‐1, cat. no. 130‐123‐555, mouse monoclonal IgG2a) or PTP zeta antibody conjugated with FITC (1:1600; Bioss, cat. no. bs‐11327R‐FITC, rabbit polyclonal IgG).

Techniques: Derivative Assay, Transmission Assay, Electron Microscopy, Isolation, Western Blot, Negative Control, Flow Cytometry, Control, Standard Deviation

Journal: iScience

Article Title: Cytohesin-4/ARF6 facilitates the progression of acute myeloid leukemia through activating PIK3R5/PI3K/AKT pathway

doi: 10.1016/j.isci.2025.112634

Figure Lengend Snippet: CYTH4 silencing delays the progression and extramedullary invasion of AML in a xenografts model (A) Representative images of the livers and spleens of the recipients following transplantation for five weeks. Quantifications of liver weight and spleen weight are shown on the right ( n = 3). Data are represented as mean ± SEM. Statistical significance was assessed by unpaired t test. (B) Human CD45 chimerism in the peripheral blood, bone marrow, liver, and spleen of recipients was assessed using flow cytometry following transplantation for five weeks ( n = 3). THP-1 cells (4.0 × 10 5 cells per mouse) were transplanted into NCG mice via tail vein injection. Following euthanizing, peripheral blood, femurs, livers, and spleens of the recipients were harvested for human CD45 chimerism analysis, with quantifications presented at the bottom. Data are represented as mean ± SEM. Statistical significance was assessed by unpaired t test. (C) Representative hematoxylin/eosin staining images for the femurs, livers, and spleens. The infiltrated leukemic cells are indicated by black arrows. (D) Kaplan-Meier survival curve for the mice transplanted with Sc shRNA/shCYTH4-transduced THP-1 cells ( n = 10 per group). (E) Flow cytometry analysis was conducted on the bone marrow and spleen of the xenograft recipient mice 16 h post tail vein injection for homing evaluation ( n = 3). Data are represented as mean ± SEM. Statistical significance was assessed by unpaired t test. ∗ p < 0.05, ∗∗ p < 0.01, and ∗∗∗ p < 0.001. Sc, scramble control shRNA; shCYTH4, CYTH4 shRNA.

Article Snippet: APC-conjugated mouse monoclonal anti-human CD45 , eBioscience , Cat no. 17-9459-42; RRID: AB_10718532.

Techniques: Transplantation Assay, Flow Cytometry, Injection, Staining, shRNA, Control