The Regulatory Mechanisms of Immune Response by Natural Products on Macrophage and Mast cells

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Immune cells participate in the inflammatory response by recognizing and subsequently eliminating unwanted antigens. This response is induced by various inflammatory cytokines secreted by activated immune cells. However, overproduction of these cytokines can cause various inflammatory disorders. Recent studies reported that various natural products can regulate the activation of immune cells such as macrophages and mast cells. In particular, seaweed extract has been demonstrated to inhibit inflammatory cytokines produced by activated macrophages. Therefore, the aim of this study was to determine whether seaweeds inhibit the inflammatory cytokines secreted by activated macrophages.
In recent studies, seaweeds were shown to possess anti-inflammatory activity. However, the effect of Grateloupia lanceolata, which inhibits inflammatory cytokine secretion from activated macrophages, remains unknown. Thus, MTT assays were performed to examine the effect of an ethanolic extract of G.lanceolata (EGL) on macrophage cell viability. Moreover, real-time PCR and nitric oxide assays were also conducted to assess inflammatory cytokine expression and nitric oxide (NO) production following EGL treatment. To further elucidate the underlying mechanism of inflammatory cytokine production, protein levels of EGL-pretreated activated macrophages were examined by western blot analysis. Our results demonstrate that EGL was non-cytotoxic to macrophages and decreased inflammatory cytokine production, including IL-1, and NO. Moreover, EGL inhibited IL-1 and NO expression through the MAPK and NF-B signaling pathways. These findings provide evidence that EGL may be a candidate therapeutic agent for treating inflammatory diseases. Grateloupia lanceolata was first discovered in southern California. Typically, seaweeds reside in the sease of northeastern Asia and North America. The distribution of G. lanceolata has been limited to the eastern shores of Asia for a long time. Grateloupia lanceolata is a red algae consisting of cirtolline and taurine as major components. This seaweed has a short stalk appressorium with vertical width that can be separated into two or three branches. However, the biological functions of G. lanceolata remain poorly understood. Thus, investigation into the biological effect and mechanism of how this seaweed influences anti-inflammation and NO production at the molecular level would be invaluable. In this study, we showed that an extract of G. lanceolata inhibited inflammation during LPS-induced macrophage activation by controlling the expression of pro-inflammatory cytokines and genes via ERK phosphorylation.
Anaphylaxis is a rapidly occurring allergic reaction to any foreign substance, including venom from insects, foods, and medications, which may cause death. To prevent anaphylaxis, these triggers must be avoided. However, avoidance of many triggers is difficult. For this reason, development of immunotherapeutic adjuvants that suppress the allergic response is important for anaphylaxis control. Mast cells are one of the major inflammatory cells involved in the inflammatory response, which secrete several inflammatory cytokines such as tumor necrosis factor (TNF)-α, interleukin (IL)-6, IL-1β, and recruits other immune cells. Mast cells are also involved in many diseases such as sinusitis, rheumatoid arthritis, and asthma. Genistein, a phytoestrogen, has been reported to have antioxidative and anti-inflammatory activities. However, the effects of genistein on the anti-inflammatory response of mast cells remain unknown. In this study, we investigated the anti-inflammatory effects of genistein on mast cells. We found that genistein significantly decreased mRNA levels of IL-6 and IL-1β as well as IL-6 production in phorbol 12-myristate 13-acetate (PMA)/A23187-induced mast cells. Moreover, we found that genistein inhibited phosphorylation of ERK 1/2 in PMA/A23187-induced mast cell activation. However, phosphorylation of p38 was not altered. In addition, our findings suggest that genistein inhibited the inflammatory status of mast cells through inhibition of the ERK pathway.
Alternative Title
대식세포와 비만세포에서 천연물에 의한 면역반응 조절 기전 연구
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Dong Hwan Kim
일반대학원 생명과학
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Table Of Contents
국문초록 4
I. Introduction 6
II. Materials and Methods 8
II-1. Cell culture 8
II-2. Purification of total extraction of G. lanceolata 8
II-3. Cell viability 9
II-4. Nitric oxide assay 9
II-5. Real time PCR and reverse transcription (RT)-PCR 9
II-6. Western blot analysis 10
II-7. Statistical analysis 11
III. Results 12
III-1. Preparation of G. lanceolata extract 12
III-2. EGL inhibits NO production in LPS-stimulated macrophage 15
III-3. EGL suppresses the expression levels of pro-inflammatory genes 19
III-4. EGL inhibits LPS-induced ERK and JNK phosphorylation but not p38 MAP kinase 21
lV. Discussion 25
V. References 27
I. Introduction 31
II. Materials and Methods 34
II-1. Cell culture and genistein treatment 34
II-2. Cell viability 34
II-3. Polymerase chain reaction and reverse transcription (RT)-PCR 34
II-4. Western blot analysis 35
II-5. Enzyme-linked immunosorbent assay 36
II-6. Statistical anaylsis 37
III. Results 38
III-1. Genistein had no effect on cytotoxicity in a human mast cell line (HMC-1) 38
III-2. Genistein suppresses expression levels of pro-inflammatory cytokines in PMA/A23187-induced HMC-1 40
III-3. Genistein significantly inhibits IL-6 production in PMA/A23187-induced HMC-1 42
III-4. Genistein regulates phosphorylation of ERK 44
IV. Discussion 47
V. References 50
Vl. Acknowledgements 55
김동환. (2013). The Regulatory Mechanisms of Immune Response by Natural Products on Macrophage and Mast cells.
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