Recapitulating Blood-Brain-Barrier Driven Traumatic Brain Injury Cascades In Vitro via a Multi- Deformation Responsive 3D Platform = 다중 기계변형 반응형 3 차원 플랫폼을 이용한 뇌혈관장벽 기반 외상성 뇌손상 연쇄반응의 체외 재구성

목차 (Table of Contents)

• ABSTRACT ........................................................................................................................... i
• 국문 초록 ............................................................................................................................... v
• TABLE OF CONTENTS ..................................................................................................... xi
• LIST OF TABLES ............................................................................................................. xiv
• LIST OF FIGURES ............................................................................................................. xv
• NOMENCLATURE ......................................................................................................... xviii
• CHAPTER 1. Introduction: Conceptual Framework for BBB-Centric TBI Modeling ........ 1
• 1.1 Clinical and Mechanistic Overview of Traumatic Brain Injury .............................. 1
• 1.1.1 Epidemiology and Socioeconomic Burden ....................................... 1
• 1.1.2 Classification and Modes of Injury ................................................... 3
• 1.1.3 Acute and Chronic Pathophysiological Cascades ............................. 4
• 1.1.4 Rationale for a Vascular-Focused Perspective .................................. 6
• 1.2 The Blood-Brain Barrier as a Primary Target in TBI .............................................. 8
• 1.2.1 Structural and Functional Overview of the Human BBB .................. 8
• 1.2.2 Biomechanical Vulnerability of the BBB ........................................ 10
• 1.2.3 Inflammation Amplification Following BBB Disruption ............... 12
• 1.2.4 Astrocyte Reactivity in the Post-TBI Microenvironment ............... 14
• 1.2.5 Inflammatory Cytokine Signatures: LCN2, IL-1β, ICAM-1 ........... 16
• 1.3 In Vivo and In Vitro Modeling of TBI: Approaches and Gaps ............................. 18
• 1.3.1. Animal Models ............................................................................... 18
• 1.3.2. In Vitro Neurocentric Models ........................................................ 21
• 1.3.3. Need for Vascular-Interated Models .............................................. 23
• 1.4 Platform Design Principles for 3D Strain-induced Deformation Responsive BBB
• Modeling ............................................................................................................................. 27
• 1.4.1. Recapitulating the Human BBB In Vitro ....................................... 27
• 1.4.2. Precise and Multimodal Mechanical Injury application ................. 28
• 1.4.3. Capturing Barrier-Specific Injury Cascades ................................... 32
• 1.5 Research Objective and Platform Modeling Focus ............................................... 33
• CHAPTER 2. Materials & Methods ................................................................................... 36
• 2.1 Material and Reagants ......................................................................................... 36
• 2.2 Methods ............................................................................................................... 41
• 2.2.1 Collagen 1 Preparation and 3D ECM Compartment Assembly ............. 41
• 2.2.2 Endothelial Seeding and 3D BBB Tri-culture ........................................ 42
• 2.2.3 Post-injury Media Collection and Cytokine Quantification ................... 43
• 2.2.4 Immunofluorescence Staining and Confocal Imaging ........................... 44
• 2.2.5 Permeability and Barrier Integrity Assay ............................................... 46
• 2.2.6 Reactive Oxygen Species Detection ....................................................... 47
• 2.3 Data Analysis and Statistics ................................................................................ 48
• CHAPTER 3. Engineering of a Multi-Injury TBI platform with an integrated hyperelastic
• PDMS BBB chip ................................................................................................................. 50
• 3.1 System Overview ................................................................................................... 50
• 3.1.1. Platform Architecture and Integration of Multi-Modal Injury Modules 50
• 3.1.2. Mechanical, Optical, and Biological Design Constraints ...................... 53
• 3.2 Fabrication of the hyperelastic PDMS-based BBB chip ........................................ 56
• 3.2.1. Structural Configuration with Integrated Perfusion and Penetration
• Access .............................................................................................................. 56
• 3.2.2. Chip Assembly and Surface Functionalization ...................................... 58
• 3.3 Implementation and validation of tensile stretch injury actuation ......................... 61
• 3.3.1. Displacement-Controlled Clamp and Linear Drive System .................. 61
• 3.3.2. Strain Range Calibration and Elastic Recovery ..................................... 62
• 3.3.3. Validation of Repeatable Stretch-Induced Deformation ....................... 72
• 3.4 Implementation and validation of circumferential tangential strain deformation
• injury actuation .................................................................................................................... 72
• 3.4.1. Camshaft-Based Actuation System and Deformation Calculation ........ 72
• 3.4.2. Validation of Repeatable Circumferential Tangential Strain-Induced
• Deformation ..................................................................................................... 78
• CHAPTER 4. Formation and Evaluation of a 3D BBB Model Within the Hyperelastic PDMS
• Chip ..................................................................................................................................... 81
• 4.1. Design and Spatial Configuration of the 3D BBB Microenvironment ................. 81
• 4.1.1. Structural Layout of Brain Parenchyma and Endothelial Lumen .......... 81
• 4.1.2. Viability and Structural integrity of Tri-Culture Components .............. 83
• 4.2. Morphological and Functional Evaluation of the 3D BBB Microenvironment .... 86
• 4.2.1. Morphological Assessment and Junctional Protein Expression ............ 86
• 4.2.2. Permeability Evaluation Using FITC-Dextran Diffusion Assay ........... 87
• CHAPTER 5. BBB-Centric Analysis of Post-TBI Cascades in vitro ................................. 91
• 5.1. Deformation Type Specific Evaluation Strategy .................................................. 91
• 5.1.1. Deformation Type-Specific Application and Experimental Timeline ... 91
• 5.1.2. Metrics for Barrier Breakdown and Inflammatory Cascade Analysis ... 92
• 5.2. Structural Breakdown and Cellular Disruption Post-Injury .................................. 93
• 5.2.1. Microvascular Deformation and BBB Disruption ................................. 93
• 5.2.2. Induction of Astrocyte Reactivity Following TBI ................................. 99
• 5.3. Neuroinflammatory Cascade Triggered by BBB Disruption .............................. 105
• 5.3.1. Endothelial and Astrocyte-Derived Cytokine Secretion (IL-1β, LCN2)
• ....................................................................................................................... 105
• 5.3.2. Comparative Analysis of Astrocyte Activation by TNF-α Versus
• Mechanical Injury .......................................................................................... 108
• 5.4. Therapeutic Modulation and Neurovascular Expansion of the In Vitro TBI Platform
• ........................................................................................................................................... 110
• 5.4.1 Therapeutic Modulation of Astrocytic Inflammation via Peptide
• Intervention .................................................................................................... 110
• 5.4.2. Neurotoxicity Evaluation via BBB-conditioned iPSC-derived neuron
• culture ............................................................................................................ 113
• CHAPTER 6. Conclusion & Discussion ........................................................................... 118
• REFERENCES .................................................................................................................. 125