Effects of diallyl trisulfide on induction of apoptotic death in murine leukemia WEHI‐3 cells in vitro and alterations of the immune responses in normal and leukemic mice in vivo

Abstract

Diallyl trisulfide (DATS), a chemopreventive dietary constituent and extracted from garlic, has been shown to against cultured many types of human cancer cell liens but the fate of apoptosis in murine leukemia cells in vitro and immune responses in leukemic mice remain elusive. Herein, we clarified the actions of DATS on growth inhibition of murine leukemia WEHI-3 cells in vitro and used WEHI-3 cells to generate leukemic mice in vivo, following to investigate the effects of DATS in animal model. In in vitro study, DATS induced apoptosis of WEHI-3 cells through the G0/G1 phase arrest and induction of caspase-3 activation. In in vivo study DATS decreased the weight of spleen of leukemia mice but did not affect the spleen weight of normal mice. DATS promoted the immune responses such as promotions of the macrophage phagocytosis and NK cell activities in WEHI-3 leukemic and normal mice. However, DATS only promotes NK cell activities in normal mice. DATS increases the surface markers of CD11b and Mac-3 in leukemia mice but only promoted CD3 in normal mice. In conclusion, the present study indicates that DATS induces cell death through induction of apoptosis in mice leukemia WHEI-3 cells. DATS also promotes immune responses in leukemia and normal mice in vivo.

Chitosan promotes immune responses, ameliorating total mature white blood cell numbers, but increases glutamic oxaloacetic transaminase and glutamic pyruvic transaminase, and ameliorates lactate dehydrogenase levels in leukemia mice in vivo

“…The WEHI-3 murine acute myelomonocytic leukemia cell line was purchased from the Food Industry Research and Development Institute (Hsinchu, Taiwan, Republic of China). Cells were cultured in 75 cm 2 tissue culture flasks containing RPMI 1640 medium supplemented with 10% FBS, 2 mM L-glutamine and antibiotics (100 U/ml penicillin and 100 µg/ml streptomycin) and placed in a humidified atmosphere of 5% CO 2 at 37°C ( 21 ). Cells were allowed to re-equilibrate for 24 h prior to use.…”

“…Mice were maintained in stainless steel mesh-bottomed cages with specified pathogen-free conditions in the animal center of China Medical University (Taichung, Taiwan, Republic of China) as in a previous study ( 22 ). All animals procedures were carried out following the institutional guidelines for animal welfare of China Medical University and were firstly approved by the Institutional Animal Care and Use Committee of China Medical University as described previously ( 21 ).…”

“…Chitosan dissolved in acetic acid was administered by oral gavage individually to each mouse in groups IV and V every 2 days for 15 days. All mice from each group were individual weighed during the oral treatment and after treatment, and all mice were weighed and sacrificed by euthanasia with CO 2 as described previously ( 21 ).…”

“…After treatment, all mice were weighed, blood were collected by cardiocentesis under general anesthesia using a disposable syringe with needle (20-gauge), and liver and spleens were dissected and weighed ( 23 ). Collected blood (1 ml/mouse) was lysed with 1X Pharm Lyse™ lysing buffer (BD Biosciences) to destroy red blood cells and leukocytes, as described previously ( 21 ). Following centrifugation at 1,500 × g for 15 min at 4°C, white blood cells were collected and stained with phycoerythrin (PE)-labeled anti-mouse CD3 (BD553062; BD Biosciences, San Jose, CA, USA), PE-labeled anti-mouse CD19 (BD553786; BD Biosciences), fluorescein isothiocyanate (FITC)-labeled anti-mouse CD11b (BD553310; BD Biosciences) and FITC-labeled anti-mouse Mac-3 (BD553324; BD Biosciences) antibodies at 1:12 dilution of each for 30 min at 4°C.…”

“…Following centrifugation at 1,500 × g for 15 min at 4°C, white blood cells were collected and stained with phycoerythrin (PE)-labeled anti-mouse CD3 (BD553062; BD Biosciences, San Jose, CA, USA), PE-labeled anti-mouse CD19 (BD553786; BD Biosciences), fluorescein isothiocyanate (FITC)-labeled anti-mouse CD11b (BD553310; BD Biosciences) and FITC-labeled anti-mouse Mac-3 (BD553324; BD Biosciences) antibodies at 1:12 dilution of each for 30 min at 4°C. All samples were analyzed by flow cytometry (BD Biosciences) and quantified using CellQuest software version 5.2.1 (BD Biosciences), as previously described ( 21 ).…”