Survey
* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
Atherosclerosis wikipedia , lookup
Lymphopoiesis wikipedia , lookup
Polyclonal B cell response wikipedia , lookup
Immune system wikipedia , lookup
Adaptive immune system wikipedia , lookup
Cancer immunotherapy wikipedia , lookup
Adoptive cell transfer wikipedia , lookup
Psychoneuroimmunology wikipedia , lookup
Rate of Phagocytosis in Macrophages Stimulated by Astragalus membranaceus Curtis Dobrowolski Department of Biology and Marine Science Jacksonville University Introduction Macrophages are the first line of defense in the human immune system Phagocytose Important antigen presenting cells, required for adaptive immune response Introduction Advantages of increased phagocytosis Removal of pathogen More T cell interaction, faster adaptive immune response More memory More Antibody production More cytokines released increased inflammation and response from other immune cells www.phototakeusa.com Introduction Astragalus membranaceus Shrub native to east Asia, primarily China A. membranaceus is a common medicinal herb used in eastern medicine One of the 50 fundamental Chinese medicinal herbs Huáng qí- root tea Used primarily when symptoms of an infection first arise Shown to increase antibody production in mice (1) Increased cytokine production (2) http://nccam.nih.gov/health/astragalus/ 50 Fundamental Chinese Medicinal Herbs 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 Agastache rugosa Alangium chininese Anemone chininese Anisodus tanguticus Aridisia j aponica Aster tataricus Astragalus membranaceus Camillia sinesis- Traditional Black Tea Cannabis sativa- Marijuana Cathamus tinctorius Cinnamomum cassia- Cinnamon Cissampelous pareira Coptis chinensis Corydalis ambigua Croton tiglium Daphne genkwa Datura metel Datura tatula Dendrobium nobile -Orchid Dichroa febrifuga Ephedra sinica Eucommia ulmoides Euphorbia pekinensis Flueggea suffruticosa Forsythia suspensa 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 Gentiana loureiroi Gleditsia sinensis Glycyrrhiza uralensis Hydnocarpus anthelmintica Ilex purpurea Leonurus japonicus Ligusticum wallichii Lobella chinensis Phellodendron amurense Platycladus orientalis Pseudolarix amabilis Psilopeganum sinense Pueraria lobata Rauwolfia serpentina Rehmannia glutenosa Rheum officinale Rhododendron tsinghaiense Saussurea costus Schisandrac hinensis Scutellaria baicalensis Stemona tuberosa Stephania tetrandra Styphnolobium japonicum Trichosanthes kirilowii Wiksotroemia indica Hypothesis In vitro phagocytosis in macrophages increases when subjected to A. membranaceus Methods IC-21 macrophages were cultured in tissue culturing flasks Macrophages grown RPMI 1640 media supplemented with 10% fetal bovine serum and 1% pen/strep antibiotic Incubated at 37°C with 5% CO2 until cells were fully confluent www.millan.com Methods Macrophages were lifted with Ca2+ depleted PBS Solution PBS= phosphate buffered saline PBS/macrophage solution was stained with trypan blue vital dye and counted using a hemocytometer Using (Q1+Q2+Q3+Q4)*104=cells/ml equation concentrations were determined www.ruf.rice.edu Methods Using the count from the hemocytometer we placed 300,000 cells in each micro-chamber Equal amount of RPMI 1640 media were added to each microchamber Incubated for minimum of 3 hours www.usascientific.com Methods Media/PBS solution was removed Macrophages were incubated for a minimum of 24 hours to allow the medias to have its effects on the macrophages Media alone 0.3% EtOH Control 1% astragalus Control EtOH Control 1%A.M. 1%A.M. Control EtOH Control 1%A.M. 1%A.M. Methods Media was removed 1μ diameter fluorescently tagged latex beads were layered on top of macrophages, at a concentration of 20 beads per cell Media alone, EtOH and astragalus media were then added to the respective wells Methods Media and beads were removed and wells were washed 3 times To remove beads that have not been phagocytosed Wells were removed and slides were stained with protocol hema 3 stain Increasing phagocytosis More cells phagocytosing Subset of cells phagocytosing more Methods Data collected Total number of cells (75-175 cells counted per experimental well) Number of cells that phagocytosed any beads Number of cells that phagocytosed >1 bead Rate of phagocytosis calculated Results 3.00% Rate of Phagocytosis of Any Number of Beads Media Alone 2.50% N=9 EtOH Control 1% A.M. Media for 60 Min Rate of Phagocytosis 2.00% 1% A.M. Media for 90 Min 1.50% 1.00% 0.50% 0.00% P Value 0.396169 P Value 0.000538 P Value 0.000463 Results 0.60% Rate of Phagocytosis of More than One Bead Media Alone Rate of Phagocytosis of More than One Bead 0.50% N=9 EtOH Control 1% A.M. Media for 60 Min 0.40% 1% A.M. Media for 90 Min 0.30% 0.20% 0.10% 0.00% -0.10% P value 0.3867711 P value 0.445056 P value 0.0167912 Conclusions The rate of in vitro phagocytosis in macrophages increases when subjected to A. membranaceus Could be beneficial in helping immuno-compromised patients HIV/AIDS patients Genetic disorders More research is need to fully understand A. membranaceus’ immuno-stimulating properties and benefits Acknowledgements I would like to thank Dr. Karen Jackson PhD, for all of her help in culturing macrophages in a less than ideal environment Also I would like to thank Sally and Judy in the Science and Math division office, for signing me out a lab key nearly everyday of the year References Zhao, K S, Mancini, C, et al., “Enhancement of the immune response in mice by Astragalus membranaceus extracts.”, Immunopharmacology, 1990 Nov-Dec; 20(2): 225-233 2. Bedir E, Pugh N, et al., “Immunostimulatory effects of cycloartane-type triterpene glycosides from astragalus species.”, Biol Pharm Bull, 2000 Jul; 23(7): 834-7 1.