인체유래 중간엽 줄기세포의 기관지천식 알레르기 염증반응 조절효과

학위논문 (박사) -- 서울대학교 대학원 : 의과대학 협동과정임상약리학전공, 2021. 2. 조상헌.Background: Asthma is a chronic inflammatory disorder of the airways characterized by variable airflow limitation and airway hyperresponsiveness. The therapeutic effects of human mesenchymal stem cells derived from umbilical cord blood (UCB-MSCs) or umbilical cord (UC-MSCs) on asthma are not clearly understood. This study aims to evaluate the effect of UCB-MSCs and UC-MSCs on allergic airway inflammation in experimental animal models. Methods: Six-week-old female BALB/c mice were used in the following two groups according to the timing of MSC administration: the preventive MSCs group (UCB-MSCs intravenous injection before the ovalbumin (OVA) challenge) and the inhibitory MSCs group (UC-MSCs intravenous injection during the OVA challenge). Each group was evaluated for airway hyperresponsiveness, histological changes, cytokines, allergen-specific antibodies, and population of innate and adaptive immune cells. Furthermore, in vitro experiments revealed the effect of UC-MSCs treatment on macrophages from bronchoalveolar lavage (BAL) fluid, which were stimulated with interleukin (IL)-4/IL-13. Transwell assays were performed to determine the paracrine effect of UC-MSCs promoting macrophage polarization. Results: Intravenous administration of UCB-MSCs significantly reduced airway hyperresponsiveness to methacholine and eosinophil counts in BAL cells. Treatment with UCB-MSCs resulted in a significant reduction in airway inflammation, mucus production, and goblet cell hyperplasia. There was a significant decrease in serum OVA-specific immunoglobulin E (IgE) and IgG1 levels, along with T helper 2 (Th2) cytokine production (IL-4, IL-5, and IL-13) in the lung and spleen tissues, whereas an increased percentage of regulatory T (Treg) cells was observed after intravenous administration of UCB-MSCs. Intravenous UC-MSCs treatment also significantly decreased the numbers of type 2 and type 3 innate lymphoid cells, dendritic cells, and macrophages in the lungs of asthmatic mice. Alveolar macrophages (CD11c+CD11b-) and resident alveolar macrophages (SiglecF+CD11c+CD11b-) were prominently increased after UC-MSCs treatment. However, the number of interstitial macrophages (CD11b+) and their subtypes, including M1 (CD206-CD86+) and M2 (CD206+CD86-) macrophages, was significantly decreased after intravenous UC-MSCs treatment. Of the two subsets, the number of M2 macrophages was more effectively reduced. IL-4- and IL-13-treated M2 expression (CD206) was reduced after UC-MSCs treatment in vitro. Similar results were observed through paracrine action. Conclusion: The results of this study suggest that UCB-MSCs and UC-MSCs exert cross-species anti-inflammatory activity, which might be mediated by a Th1/Th2 shift, Treg cell induction, and macrophage switching. UCB-MSCs and UC-MSCs not only affect adaptive immunity but also innate immunity. Preclinical findings may provide important clues to support further UCB-MSCs and UC-MSCs research for the development of a new asthma treatment strategy.서론: 천식은 기도 폐쇄와 기도 과민성을 특징으로 하는 만성 염증성 기도 질환으로, 천식에서 제대혈 및 제대 유래 중간엽 줄기세포의 치료 효과는 아직까지 명확하게 알려져 있지 않다. 이 연구는 천식 실험동물모델에서 인체 유래 (제대 혈 및 제대) 중간엽 줄기세포의 알레르기 염증반응 조절 효과를 선천면역 및 적응면역에 관련된 세포들의 반응을 통해 확인해 보기 위해 수행되었다. 방법: 천식 동물 모델인 BALB/c 마우스의 복강으로 난알부민을 투여하여 감작시킨 이후, 난알부민 기도투여로 알레르기성 천식을 유도하기 전 또는 후에 꼬리 정맥으로 제대혈 또는 제대유래 중간엽 줄기세포를 주입한다. 이후 마우스의 혈청, 기관지폐포세척액, 폐조직, 비장세포에서 줄기세포 투여에 따른 염증세포 분획, 조직학적 변화, 혈청 특이 알레르겐 항체, 염증 사이토카인 분비, 호산구, T세포와 조절 T 세포, 선천성 림프구 세포, 수지상세포와 대식 세포를 생체 내 실험(in vivo)을 통하여 확인하였다. 또한 제대유래 중간엽 줄기세포의 주변 분비 작용(paracrine effect)을 확인하고자 M2 대식세포주를 이용하여 생체외(in vitro)실험을 진행하였다. 결과: 제대혈유래 중간엽 줄기세포는 기도과민성을 감소시키고 조직에서 호산구 침윤, 점액과 술잔세포의 과형성을 유의하게 감소시켰다. 기관지폐포세척액에서 호산구를 비롯한 염증세포들을 감소시키고 혈청에서 난알부민 특이 IgE와 IgG1 항체를 의미 있게 줄였다. 또한 제대혈유래 중간엽 줄기세포는 기관지폐포세척액과 폐/비장 조직에서 Th2 사이토카인(IL-4, 5, 13)을 의미 있게 감소시키며 조절 사이토카인(IL-10, TGF-β)을 의미 있게 증가시켰다. 아울러 폐 림프절 및 비장 조직에서 조절 T세포의 증가도 관찰되었다. 제대유래 중간엽 줄기세포는 폐조직에서 type 2, 3 선천성 림프구 세포, 수지상세포와 대식세포의 수를 의미 있게 감소시켰다. 대식세포 중 폐포 대식세포는 제대유래 중간엽 줄기세포 치료 후 의미 있게 증가한 반면, 간질성 대식세포는 감소하였으며 간질성 대식세포군 들 중 하위 유형인 M1, M2, M2a, M2c도 간질성 대식세포와 비슷한 양상을 보였다. 생체외 실험을 통해 제대유래 중간엽 줄기세포의 주변 분비 작용도 확인하였다. 결론: 인체 유래 중간엽 줄기세포는 Th1/Th2 세포 면역 편향, 조절 T 세포 등 적응면역계, 그리고 선천성 림프구 세포와 대식세포 등 선천면역계의 변화를 통해 천식 알레르기 염증반응을 조절하는 역할이 있음을 확인할 수 있었다. 이를 통해 선천면역계 및 적응면역계에 모두 관여하는 제대 및 제대혈 유래 중간엽 줄기세포가 천식 치료제로 효과적임을 확인할 수 있었다.1. Introduction 1 2. Methods 2 Literature review of stem cells in animal asthma models 2 Preparation of hMSCs 3 Murine asthma model with or without MSCs treatment 5 Measurement of airway hyperresponsiveness 8 Analysis of bronchoalveolar lavage fluid 8 Histopathological analysis 9 Analysis of cytokine expression 9 Analysis of serum OVA-specific antibody 11 Flow cytometric analysis 11 Real time quantitative PCR to assess macrophage polarization 13 3. Statistical analysis 15 4. Results 15 4.1 Literature review of stem cells as anti-asthmatic therapeutics in animal asthma models 15 4.2 Experiment I assessing the effect of UCB-MSCs on the adaptive immune responses in asthma 19 AHR to methacholine 19 BAL and histologic findings 20 Measurement of OVA-specific immunoglobulin 23 Levels of cytokines measured by RT-qPCR and ELISA 25 Comparison of regulatory T cells in lymph nodes 28 4.3 Experiment II assessing the effect of UC-MSCs on the innate immune responses in asthma 30 AHR to methacholine 30 BAL and histologic findings 31 Changes in lymphoid lineages by UC-MSCs treatment 33 Changes in dendritic cells and macrophages by UC-MSCs treatment 39 Impact of UC-MSCs treatment on macrophage expression 46 5. Discussion 47 6. References 58 7. Table 74 8. Abstract 79Docto

불환지폐의 가치결정원리에 대한 비판적 고찰

학위논문(석사)--서울대학교 대학원 :경제학부 경제학전공,2001.Maste

(A)s tudy on the value and price of software : focused on the information goods value controversy

학위논문(박사) --서울대학교 대학원 :경제학부,2008.2.Docto

Usefulness of Interactive e-learning education program for asthma guideline

학위논문 (석사)-- 서울대학교 대학원 : 의학과(면역학전공), 2015. 8. 민경업.Introduction: Effective educational tools and implementation strategies are important for the dissemination of guideline. We developed the interactive computer-based education program of asthma guideline named as Virtual Learning Center for Asthma Management and evaluated its usefulness in terms of the improvement of awareness and satisfaction. Methods: 170 physicians were enrolled from six tertiary hospitals. They utilized the interactive education program for 2 weeks for the learning of asthma management guideline. We compared awareness of asthma guideline before and after utilization of the program and investigated the user satisfaction with questionnaire survey. Results: Mean age of study subjects was 28.2±3.10 and 78(49.4%) of subjects were male. The total score of awareness of asthma guideline was significantly improved from 80.3±6.39 to 85.1±6.90 (p<0.001). All categories of awareness including knowledge, attitude and practice were improved significantly after learning with the program (p<0.001). Satisfactions of users were high in the aspects of usefulness, convenience, interest, improvement of understanding of guideline and elevation of confidence. The most useful section of the program was virtual cyber management of asthma patients. Conclusion: Interactive education program is a useful and effective tool for dissemination of asthma guideline.I. 서론 1 II. 연구 재료와 방법 2 1. 연구 대상 2 2. Virtual Learning Center for Asthma Management (VLCAM)프로그램 2 3. 설문 항목 3 4. 통계 4 III. 연구 결과 5 1. 조사 대상 분석 5 2. VLCAM 프로그램 사용 전후 천식 진료지침의 인지도 변화 5 3. VLCAM 프로그램의 유용성 평가 6 IV. 고찰 7 V. 참고 문헌 12 VI. Tables and figures 15 VII. 별첨: 천식 인지도 평가 및 프로그램 만족도 조사 문항 내용 23 VIII. 영문 초록 37Maste