초록 ▼

본 연구팀은 혈관 생성(vasculogenesis) 방식을 이용하여 사람의 동맥경화 미세환경을 모사하고 취약 플라그(vulnerable plaque)의 파열 매커니즘을 규명하는 미세유체소자의 개발을 목표로 하여 연구를 진행하였다.

1. 동맥경화 미세환경 조성이 가능한 미세유체소자의 개발
- 정수압차이나 펌프를 이용하여 혈관 네트워크 내 유동을 정교하게 조절
- 젤 채널에 혈관네트워크의 비계역할을 하는 하이드로겔(hydrogel)을 주입하여 3차원 배양환경 조성
- 혈관세포의 분화를 촉진하는 생화학적 성장

본 연구팀은 혈관 생성(vasculogenesis) 방식을 이용하여 사람의 동맥경화 미세환경을 모사하고 취약 플라그(vulnerable plaque)의 파열 매커니즘을 규명하는 미세유체소자의 개발을 목표로 하여 연구를 진행하였다.

1. 동맥경화 미세환경 조성이 가능한 미세유체소자의 개발
- 정수압차이나 펌프를 이용하여 혈관 네트워크 내 유동을 정교하게 조절
- 젤 채널에 혈관네트워크의 비계역할을 하는 하이드로겔(hydrogel)을 주입하여 3차원 배양환경 조성
- 혈관세포의 분화를 촉진하는 생화학적 성장인자의 포함 가능

2. 소자 내에서의 혈관형성법 최적화
- 사람 혈관세포를 이용하여 관류 가능한 삼차원 혈관 네트워크 형성

3. 동맥경화에 적합한 혈관모델 형성과 미세환경 구축
- 동맥경화 초기 과정 중 대식세포의 사멸체로 인한 혈관벽의 병리적 기능장애 모사
- 단백질 분해효소 억제제를 배양환경에 포함하여 하이드로겔의 지속성 향상

(출처 : 보고서 초록)

Abstract ▼

Ⅱ. Research Background
Many previous studies have shown that the rupture of a vulnerable plaque has a great impact on the occurrence and progression of arteriosclerosis, but the basal mechanism has not been elucidated in detail. Although many animal models and in vitro two-dimensional static expe

Ⅱ. Research Background
Many previous studies have shown that the rupture of a vulnerable plaque has a great impact on the occurrence and progression of arteriosclerosis, but the basal mechanism has not been elucidated in detail. Although many animal models and in vitro two-dimensional static experiments have been attempted, considerable problems have been pointed out in mimicking arteriosclerotic microenvironment of a human. Therefore, by using microfluidic devices, we tried to mimic the microenvironments of arteriosclerosis and investigated the underlying mechanism for rupture of a plaque.

Ⅲ. Research Description
We would develop a technique that reflects the dynamic characteristics of blood vessels in the atherosclerotic model and induces rupture of the plaque by applying vasculogenesis on a microfluidic device.

Ⅳ. Research Achievement
1. Development of a Microfluidic Device
The developed microfluidic device has the following characteristics. The gel channels were formed between two media channels using trapezoidal posts. A three-dimensional culture environment was established by using hydrogel as a scaffold for vasculogenesis.
It is also possible to form a concentration gradient of chemical factors across the hydrogel. Additionally, by connecting a pump to the media channel, the flow rate in the device can be precisely controlled. Furthermore, since the PDMS is transparent material, it is easy to get three-dimensional images and the cost is relatively low, so that a large number of devices can be obtained.

2. Fabrication of Microfluidic Devices and Optimization of Vasculogenesis
(1) Fabrication process of designed microfluidic device
Using the designed master mold as a master mold for soft lithography, which is one of the microfabrication methods, PDMS devices with the channel pattern was obtained and channels were formed by bonding with a cover glass.
(2) Optimization of vasculogenesis
Human vascular endothelial cells were mixed with fibrin gel and cultured in the microfluidic device with growth factors. Five days later, perfusable microvascular networks were generated.

3. Development of in vitro Microvascular Model Suitable for Atherosclerotic Microenvironment
In order to solve weak stability and short duration of existing in vitro vascular models, we sought the following strategies.
(1) Investigation of the dysfunction of the endothelial wall during the early stages of atherosclerosis
The pathological dysfunction of the endothelial wall due to the death of macrophages was mimicked. The apoptosis of macrophages was induced with UV lamps and then the apoptotic cells were injected into the microfluidic device where the endothelial wall was formed and the permeability was determined by measuring fluorescent intensity of VE-cadherin.
(2) Improvement of hydrogel persistence
In order to prevent the degradation of hydrogels in the vasculogenesis, 6-aminocaproic acid (EACA) which is one of the protease inhibitors was included in the culture environment. EACA was found to increase the persistence of the hydrogel by interfering with the action of the plasmin released from the endothelial cells.

Ⅴ. Needs for Further Research
1. Improvement of Hydrogel Persistence
It has been shown that EACA is involved in a complex mechanism between ANG-1, which is known to help stabilize the vessel wall, but the underlying mechanism has not yet been elucidated. In addition, it is necessary to investigate the complex role of EACA in vasculogenesis and to optimize the use of EACA in vasculogenesis.

2. Improvement of Stability of Vascular Networks by Co-culturing with Mesenchymal Stem Cells
Mesenchymal stem cells are known to differentiated into muscle cells that surround vascular endothelial cells during vasculogenesis and secrete cytokines that promote vasculogenesis. Therefore, we will focus on co-culturing of vascular endothelial cells with mesenchymal stem cells in a microfluidic device to improve the stability of the vascular networks.

3. Inducing Plaque Formation and Rupture of Plaques in the Atherosclerotic Microenvironment
UV-induced apoptotic macrophages, LDL cholesterol, and foam cells will be injected into the microfluidic device in which the vascular network is formed to induce the dysfunction of vascular wall or plaque formation. Quantitative confirmation of the role of each factor will help to optimize the method of inducing plaque formation and plaque rupture in the atherosclerotic microenvironment.

(출처 : SUMMARY)

목차 Contents

• 표지 ... 1제 출 문 ... 2보고서 초록 ... 3요 약 문 ... 4SUMMARY ... 6CONTENTS ... 9목차 ... 10제 1 장 연구개발과제의 개요 ... 11제 2 장 국내외 기술개발 현황 ... 14제 3 장 연구개발수행 내용 및 결과 ... 15 1절 동맥경화 미세환경 조성이 가능한 미세유체소자 개발 ... 15 2절 미세유체소자의 제작과 소자 내에서의 혈관형성법 최적화 ... 16 1. 설계한 미세유체소자의 제작 과정 ... 16 2. 혈관형성법 최적화 ... 16 3절 동맥경화 미세환경에 적합한 혈관 모델 개발 ... 17 1. 동맥경화 초기 과정 중 혈관벽의 병리적 기능장애 모사 ... 17 2. 혈관 네트워크의 안정성과 지속성 향상 ... 18제 4 장 목표달성도 및 관련분야에의 기여도 ... 22 1절 연구개발목표 달성도 ... 22 2절 관련분야 기술발전에의 기여도 ... 23제 5 장 연구개발결과의 활용계획 ... 24 1절 향후 추가연구의 필요성 ... 24 1. EACA를 활용한 하이드로겔의 지속성 향상 ... 24 2. 중간엽줄기세포와 공동배양으로 혈관 네트워크의 안정성 향상 ... 24 3. 동맥경화 환경에서 혈관벽 내 플라그 형성과 플라그 파열 유도 ... 25 2절 타 연구에의 응용 ... 25제 6 장 연구개발과정에서 수집한 해외과학기술정보 ... 26제 7 장 참고문헌 ... 27끝페이지 ... 31