Orally Delivered miRNA Targeting TLR8 by Salmonella Suppresses Systemic Inflammation
- 주제(키워드) Salmonella , TLR8 , miRNA , allergy , inflammation
- 발행기관 고려대학교 대학원
- 지도교수 박용근
- 발행년도 2012
- 학위수여년월 2012. 2
- 학위구분 석사
- 학과 일반대학원 생명공학과
- 세부전공 의약생명공학전공
- 원문페이지 46 p
- 실제URI http://www.dcollection.net/handler/korea/000000032562
- 본문언어 영어
- 제출원본 000045697068
초록/요약
RNA interference has great potential as a medical therapy to treat disease. However, targeted induction of RNAi in vivo has met with significant challenges. In this study, plasmid of expressing miRNA was transformed into attenuated Salmonella. TLR8 activation leads to the secretion of proinflammatory cytokines and chemokines. Chemokines such as CCL17 express in chronic lesions [1]. In vitro infection studies with the recombinant Salmonella expressing TLR8 miRNA demonstrated that silencing of TLR8 expression in cultured RAW264.7 cell. In addition, TLR8 downregulation reduced expression of IL-1α, IL-1β, IL-18, CCL17 and CCL22. In vivo, the ST-miRTLR8 suppressed TLR8 in mice spleen. Moreover, in the mice skin, expression of TLR8 and IL-1α, IL-4, TNF-α were decreased by ST-miRTLR8. ST-miRTLR8 inhibited TNF-α, IL-4, IgE and induced IFN-γ in the total serum which with cutaneous disease in DNCB treated allergic mouse model. Furthermore, mice were significantly suppressed symptoms of cutaneous diseases such as scratching and epidermal thickness after oral administration with ST-miRTLR8. These results suggest that ST-miRTLR8 may be an effective genetic medicine for inflammatory skin disease.
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ABSTRACT 1
INTRODUCTION 2
MATERIALS AND METHODS 5
Construction of a plasmid vector capable of expressing TLR8 miRNA 5
Salmonella strain to express TLR8 miRNA and growth condition 5
Cell culture and TLR8 miRNA transfection 6
Invasion assay 7
Reverse transcription PCR 8
Allergic mouse model 9
Cytokine analysis and bacterial distribution in allergic mice 10
Enzyme-Linked Immunosorbent Assay 10
SDS-PAGE and western blot analysis 11
Behavioral observation 12
Histological observation of the skin 12
Flow cytometry 12
RESULTS 14
Construction of miRNA expression vectors 14
Cell infection by recombinant Salmonella 17
Effects of bacterially delivered miRNA on cytokine levels in cell 21
Modulation of TLR8, after treatment with ST-miRTLR8 in allergic mouse model 23
Reduced inflammatory effects of ST-miRTLR8 mice with cutaneous disease 26
DISCUSSION 32
REFERENCES 35
KOREAN ABSTRACT 39
