Ouabain promotes immune responses in WEHI‐3 cells to generate leukemia mice through enhancing phagocytosis and natural killer cell activities in vivo

Abstract

Ouabain, a cardiotonic steroid, was used for the treatment of heart failure and atrial fibrillation and induces cancer cell apoptosis in many human cancer cells including human leukemia cells.However, there are no reports to show the effects on immune responses in a leukemia mouse model. In this study, WEHI-3 cell generated leukemia mice were developed and treated by oral ouabain at 0, 0.75, 1.5, and 3 mg/kg for 15 days. Results indicated that ouabain did not affect body appearance, but decreased liver and spleen weights, B-and T-cell proliferation at all three doses treatment and increased CD19 cells at 3.0 mg/kg treatment, decreased CD3, CD11b, and Mac-3 cells levels compared with positive control. Furthermore, ouabain increased the macrophage phagocytosis from peripheral blood mononuclear cell and peritoneal cavity at all three doses treatment and increased NK cell activities. Ouabain restored GOT, GPT and LDH levels in WEHI-3 leukemia mice in vivo. K E Y W O R D S immune response, macrophage, natural killer cell, ouabain, phagocytosis Yung-Luen Shih, Hung-Sheng Shang, Hsu-Feng Lu, and Jing-Gung Chung have contributed equally to this study.

Cardamonin induces immune responses and enhances survival rate in WEHI‐3 cell–generated mouse leukemia in vivo

“…Animals were provided by National Laboratory Animal Center (Taipei, Taiwan), and experiments were performed based on animal ethical guidelines of the China Medical University. All animals were housed in the Animal Housing Facility Center of China Medical University (Taichung, Taiwan) under standard conditions (12 h light/dark cycles at 25°C) and standard rodent diet (Prolab RMH 2500; LabDiet, PMI Nutrition, Brentwood, Missouri) as previously described . This project was reviewed and approved by Institutional Animal Care and Use Committee of China Medical University (approval ID: 2018‐378).…”

“…Immune effects of cardamonin on mice in vivo were assayed using leukocyte population analysis and phagocytosis activity test as previously described . Briefly, forty mice are separated randomly into four groups (N = 10).…”

“…The animal studies were performed for 14 and 42 days for the evaluation of immune responses and survival rates, respectively. At the end of treatment, mice were weighed and then sacrificed by euthanasia with CO 2 as described . The flowchart of experimental design is summarized in Figure .…”

“…After 14‐day treatment, blood samples from individual mice were harvested for determining the leukocytes' cell markers as described previously . Leukocytes were individually incubated with fluorescein‐labeled antibodies, including phycoerythrin‐conjugated anti‐mouse CD3 (T cells) and CD19 (B cells) and fluorescein isothiocyanate (FITC)‐conjugated anti‐mouse CD11b (monocytes) and Mac‐3 (macrophages) antibodies, for 1 hour at 4°C and then quantified the cell marker population of leukocytes by flow cytometry as described …”

“…Phagocytosis of macrophages from each group was determined by flow cytometric assay as previous citation . Briefly, all macrophages were individually harvested from peripheral blood mononuclear cells (PBMCs) and peritoneum.…”