Nanobiohybrid-based artificial eye composed of eye organoid for drug screening
- 주제어 (키워드) Nanobiohybrid , Artificial eye , Retinal organoid , Thalamic organoid , Drug screening
- 발행기관 서강대학교 일반대학원
- 지도교수 최정우
- 발행년도 2024
- 학위수여년월 2024. 2
- 학위명 박사
- 학과 및 전공 일반대학원 화공생명공학과
- 실제URI http://www.dcollection.net/handler/sogang/000000076947
- UCI I804:11029-000000076947
- 본문언어 영어
- 저작권 서강대학교 논문은 저작권 보호를 받습니다.
초록
The research on artificial eyes has attracted a lot of attention in the biomedical fields due to its potential for a wide range of applications, including the development of eye-related disease model and drug evaluation. However, most of the current research focuses on reproducing the biological structure of the eye, and no studies have been conducted to regulate the electrical signals of an artificial eye to interact with other cells like a real human eye. In this study, a light-responsive nanobiohybrid artificial eye is developed for the regulation of electrical signal transfer to living cells for biomedical applications. To fabricate the nanobiohybrid artificial eye, biomaterials that generate electrical signals by light were introduced together with nanomaterials to improve their functionality. First, the bio-solar cell-based artificial eye composed of two types of light-harvesting biomolecules (bacteriorhodopsin and chlorophyllin) and Ni/TiO2 nanoparticles was developed for biomedical applications. Using the developed bio-solar cell, electrophysiological signals of the muscle cells at the neuromuscular junction were controlled through photocurrent transmission. In addition, retinal/thalamic assembloid composed of retinal organoids and Au nanomesh-encapsulated thalamic assembloid was developed for evaluation of anti- aging drug of the retina. The retinal/thalamic assembloid generated electrophysiological signals at the cellular level upon light irradiation. Furthermore, the anti-aging drug was evaluated using an aged model of retinal/thalamic assembloid, which was fabricated by blue light irradiation. Finally, the eye-blinking model was developed through the connection of retinal/thalamic assembloid, brain organoid, motor neuron spheroids, and muscle bundle on a chip. The results of the blinking motion in the presence or absence of retinal toxic chemical demonstrated the feasibility of assessing retinal toxicity using the developed eye-blinking model. In conclusion, the developed nanobiohybrid artificial eye can be applied to develop various kinds of eye disease models and therapeutics for eye diseases, as well as visual sensors for biohybrid robots.
more목차
1. Introduction 1
1.1 Overview 1
1.2 Objectives of this research 3
1.3 References 8
2. Nanobiohybrid solar cell-based artificial eye to regulate the electrical signal transmission to living cells for biomedical application 10
2.1 Introduction 10
2.2 Materials and methods 16
2.2.1 Materials 16
2.2.2 Synthesis of the Ni/TiO2 NPs 17
2.2.3 Fabrication of the artificial eye on the MEA 18
2.2.4 Electrochemical analysis of the artificial eye 19
2.2.5 Measurement of the photocurrent generated by the artificial eye 20
2.2.6 Cell culture on the artificial eye-modified MEA 20
2.2.7 Fixation of the cells for morphology investigation 21
2.2.8 Immunostaining of the NMJ 22
2.2.9 qPCR analysis of muscle cells and motor neurons 22
2.2.10 Electrophysiological signals measurement 23
2.2.11 Statistical Analysis 23
2.3 Results and Discussion 25
2.3.1 Development of the artificial eye 25
2.3.2 Photocurrent generation by the artificial eye 36
2.3.3 Formation of the NMJ on the artificial eye-modified MEA 51
2.3.4 Signal transmission to muscle cells at the NMJ using the artificial eye 63
2.4 Conclusion 73
2.5 References 76
3. Retinal/thalamic assembloid encapsulated by Au nanomesh to evaluate the anti-aging drug of retina 83
3.1 Introduction 83
3.2 Materials and methods 86
3.2.1 iPSC culture 86
3.2.2 Generation of retinal organoid 87
3.2.3 Generation of thalamic organoid 88
3.2.4 Fabrication of Au nanomesh 89
3.2.5 Generation of retinal/thalamic assembloid 89
3.2.6 Immunohistochemistry 90
3.2.7 qPCR analysis 90
3.2.8 Evaluation of electrophysiological activity 91
3.2.9 Reactive oxygen species (ROS) measurement 92
3.2.10 Light irradiation 92
3.3 Results and Discussion 94
3.3.1 Generation of retinal organoid and thalamic organoid 94
3.3.2 Fabrication of Au nanomesh introduced thalamic organoid 100
3.3.3 Fabrication of the retinal/thalamic assembloid 105
3.3.4 Investigation of the effect of blue light on the retinal/thalamic assembloid 109
3.3.5 Evaluation of the effect of lutein as an anti-aging drug using the retinal/thalamic assembloid 112
3.4 Conclusion 115
3.5 References 117
4. Eye-blinking model composed of retinal/thalamic assembloid, brain organoid, motor neuron spheroid, and muscle bundle for retinal toxicity assay 122
4.1 Introduction 122
4.2 Materials and methods 125
4.2.1 Generation of brain organoid 125
4.2.2 Generation of motor neuron spheroids 127
4.2.3 Generation of muscle bundle 128
4.2.4 Generation of retinla/thalamic assembloid 128
4.2.5 Tissue clearing and immunohistochemistry 130
4.2.6 qPCR analysis 132
4.2.7 Evaluation of electrophysiological activity 132
4.3 Results and Discussion 134
4.3.1 Generat ion and character izat ion of cerebral organoid, retinal/thalamic assembloid, and motor neuron spheroid 134
4.3.2 Verification of connection between organoids and motor neuron spheroids 138
4.3.3 Investigation of connection between muscle bundle and motor neuron spheroids 141
4.3.4 Fabrication of eye-blinking model 144
4.3.5 Performing retinal toxicity assay using eye-blinking model 146
4.4 Conclusion 148
4.5 References 149
5. Conclusion and Perspective 154
5.1 Conclusion 154
5.1.1 Nanobiohybrid Bio-Solar Cell-based Artifial Eye to Regulate the Electrical Signal Transmission to Living Cells for Biomedical Application 154
5.1.2 Retinal/thalamic assembloid encapsulated by Au nanomesh to evaluate the anti-aging drug of retina 158
5.1.3 Eye-blinking model composed of retinal/thalamic assembloid, brain organoid, motor neuron spheroid, and muscle bundle for retinal toxicity assay 159
5.2 Perspective of Further study 160
5.2.1 Introduction of nanomaterials into the eye-blinking model 160
5.2.2 Introduction of organoids with disease model 161
Vita 163
List of Publications 164