Fig 1: Light microscopic images of K562 (left) and Hek293 (right): (A) alone, (B) HVEM untreated cells with CFSE CD4+ T cells at one tumor or normal to ten T cells, or (C) HVEM blocker treated cells with CFSE CD4+ T cells after K562 and Hek293 for 72 h of incubation. Data are representative of three separate experiments. Magnification: 100 μm.
Fig 2: Receiver operating characteristic curve for HVEM gene expression in patients with malignant ALL compared with non-malignant healthy controls: (A) in all samples, (B) in newly diagnosed ALL, (C) in remission ALL, and (D) in relapse/refractory. AUC, the area under the curve, values are 0.7762 (p = 0.0142), 0.8889 (p = 0.0133), 0.6543 (p = 0.1985), and 0.7472 (p = 0.0359), respectively, indicating that HVEM is a potential biomarker in ALL patients, especially in newly diagnosed patients and those in the relapse/refractory phase.
Fig 3: Light microscopy images and the expression of herpesvirus entry mediator (HVEM) surface protein on breast cancer (MCF-7), hepatocellular carcinomas (HepG2), chronic myelogenous leukemia (K562), and human embryonic kidney (Hek293) cells. The left column represents the microscope and a magnification of 100 pt using NIS-Elements F 4.00.00 software (Nikon Instruments, USA) under a light microscope (Nikon Eclipse, Fujisawa, Japan). Data collected from four separate experiments represents cell proliferation, morphology, and attachment nature. The right column shows histogram graphs representing the measurements of HVEM expression after staining with anti-HVEM-PE. The dotted-line histograms show the unstained control cells, whereas the line histograms represent the HVEM-stained cells. The numbers at the top of each histogram indicate the MFI of the unstained (left) and HVEM-PE (right). The number in the middle represents the percentage of HVEM-positive cells. The results were collected from two wells in three separate experiments. Magnification: 100 pt.
Fig 4: Proliferation of CFSE-labeled CD4+ T cells in response to (A) K562 tumor cells or (B) Hek293 healthy control cells without HVEM blocker treatment (left column), or after K562 and Hek293 treatment with HVEM blocker (middle column); and non-proliferated CFSE CD4+ T cells (right column) after 72 h of incubation at 37 °C. Data are representative of two separate experiments.
Fig 5: Mean ± SD of fluorescence intensity of CFSE-labeled CD4+ T cells before proliferation on day zero and after 72 h of incubation with either K562 tumor cells or with Hek293 normal cells that were left untreated or treated with 20 ng HVEM blocker. Cells were cultured at a ratio of one tumor or normal cell to ten CFSE CD4+ T cells. ** indicates strong significant differences between groups with p = 0.0033, whereas “ns” indicates no significant differences with p = 0.5918. The results were collected from three separate experiments.
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