천식 환자에서 Thromboxane A2 receptor 유전자 발현에 미치는 MicroRNA-149의 조절 작용
Regulatory Function of MicroRNA-149 on Thromboxane A2 Receptor Gene Expression in Asthmatics
- 주제(키워드) Asthma , miR-149 , TBXA2R
- 발행기관 아주대학교
- 지도교수 박해심
- 발행년도 2016
- 학위수여년월 2016. 8
- 학위명 석사
- 학과 및 전공 일반대학원 의생명과학과
- 실제URI http://www.dcollection.net/handler/ajou/000000022723
- 본문언어 영어
- 저작권 아주대학교 논문은 저작권에 의해 보호받습니다.
초록/요약
Background and objectives: The regulation of inflammatory responses in human diseases is mediated by the coordinated control of inflammatory gene expression at the transcriptional level as well as at the post-transcriptional level. One mechanism of post-transcriptional regulation is through control by microRNAs, which bind to the 3′ untranslated region (3′ UTR) of target genes and regulate mRNA stability or translation. The aim of this genetic association study was to identify miR-149 polymorphisms and evaluate their relationship with asthma with a view to further understanding the pathogenesis of asthma. Subjects and methods: One hundred and eighty-four patients with asthma and 168 healthy normal controls (NC) were enrolled in this study. Single nucleotide polymorphisms (SNPs) in miR-149 (rs2292832) were genotyped using a TaqMan® allelic discrimination assay. Plasma miR-149 expression was evaluated using quantitative RT-PCR with a TaqMan® MicroRNA Reverse Transcription Kit and TaqMan® MicroRNA Assays. The expression of the thromboxane A2 receptor (TBXA2R) gene in peripheral blood mononuclear cells (PBMCs) was measured by quantitative RT-PCR. Human Embryonic Kidney 293T (HEK293T) cells were transfected with an EGFP-tagged TBXA2R 3′UTR plasmid construct and a miR-149 mimic or a miR-149 inhibitor using lipofectamine. EGFP expression was evaluated by measuring fluorescence intensity. Results: Asthma patients had a significantly lower frequency of the miR-149 C allele compared to that of the NC group (P = 0.014). The expression levels of plasma miR-149 in asthma patients were significantly lower than that of the NC group (P = 0.002). Plasma miR-149 expression levels showed a negative correlation with both total IgE (r = -0.320; P = 0.027) and peripheral eosinophil count (r = -0.311, P = 0.030). The expression of the TBXA2R gene in PBMCs was significantly higher in asthma patients compared to that in the NC group (P = 0.024). Transfection with the miR-149 mimic significantly suppressed TBXA2R expression (P = 0.04), whereas transfection with the miR-149 inhibitor significantly increased TBXA2R expression (P = 0.02). Conclusion: These findings suggest that the miR-149 C-allele (rs2292832) is associated with the phenotype of asthma and that miR-149 negatively regulates TBXA2R expression. Therefore, miR-149 may contribute to the pathogenesis of asthma.
more목차
Ⅰ. INTRODUCTION
Ⅱ. MATERIALS AND METHODS
1. Study subjects
2. Genotyping of miR-149 single nucleotide polymorphisms
3. Quantitative measurement of miR-149 expression in plasma
4. Quantitative measurement of TBXA2R expression in PBMCs
5. Plasmid construction for 3’UTR of TBXA2R
6. Transfection of miR-149 into HEK293T cell
7. Statistical analysis
Ⅲ. RESULTS
1. Clinical characteristics of study subjects
2. Genetic association of miR-149 gene polymorphism with asthma
3. Decreased expression of plasma miR-149 in asthmatics
4. Enhanced expression of TBXA2R in asthmatics
5. Negative regulation of TBXA2R expression by miR-149
Ⅳ. DISCUSSION
Ⅴ. CONCLUSION
REFERENCES
국문요약
