Speaker
Description
Introduction
One of the major challenges in tissue engineering of three-dimensional (3D) functional tissues is achieving vascularization, which is critical for developing large, viable, and physiologically relevant in vitro models. Constructs exceeding 40 µm in thickness require vascular networks to sustain cell survival1, and blood vessels play a key role in supporting tissue homeostasis through dynamic cell–vessel interactions. In engineered skin models, the lack of an endothelial barrier can significantly impact drug and cosmetic testing outcomes. In native skin, vascularization primarily occurs through angiogenesis — the sprouting of new vessels from pre-existing ones2. To address this, silk-based biomaterials have emerged as promising candidates due to their outstanding biocompatibility, mechanical properties, and ability to support cellular processes critical for tissue integration3.
Methods
We vascularized an already optimized 3D skin model5 using a pure silk fibroin membrane (KLISBio) integrated into a custom holder system. Endothelial cells (ECs) and mesenchymal stromal cells (hMSCs) were co-cultured on the silk membrane, supporting vascular network formation within a biomimetic dermal matrix of methacrylated gelatin (GelMA) containing fibroblasts (HFF-1). Constructs were cultured under pro-angiogenic stimulation with vascular endothelial growth factor (VEGF). Characterization included LIVE/DEAD viability assays, immunofluorescence imaging (whole-mount and cryosectioned samples), and droplet digital PCR (ddPCR) analysis of angiogenesis-related gene expression.
Results
Preliminary results confirmed excellent cell viability and integration across the constructs, as evidenced by LIVE/DEAD staining and immunofluorescence analysis. Endothelial sprouting behavior and early matrix remodeling were observed, indicating active vascular network formation. Comparative ddPCR analysis showed upregulation of key angiogenic markers in the silk-based constructs compared to commercial membrane controls. Immunofluorescence imaging highlighted enhanced endothelial organization and vessel-like structures within the silk platform, supporting its superior ability to promote vascular morphogenesis and functional dermal tissue development.
Discussion
These findings demonstrate the potential of silk fibroin membranes to create robust, vascularized 3D skin models with enhanced physiological relevance. Compared to conventional commercial membranes, the silk platform provides a more supportive environment for endothelial migration, organization, and angiogenesis, likely due to its favorable mechanical and biological properties. This work paves the way for developing next-generation skin models for regenerative medicine, disease modeling, and high-fidelity drug screening applications.
References
1. Hwang, D. G., Choi, Y. & Jang, J. 3D Bioprinting-Based Vascularized Tissue Models Mimicking Tissue-Specific Architecture and Pathophysiology for in vitro Studies. Front Bioeng Biotechnol 9, 685507 (2021).
2. Gao, C. et al. Strategies for vascularized skin models in vitro. Biomater. Sci. 10, 4724–4739 (2022).
3. Madappura, A. P. & Madduri, S. A comprehensive review of silk-fibroin hydrogels for cell and drug delivery applications in tissue engineering and regenerative medicine. Computational and Structural Biotechnology Journal 21, 4868–4886 (2023).
4. Villata, S. et al. Broadly Accessible 3D In Vitro Skin Model as a Comprehensive Platform for Antibacterial Therapy Screening. ACS Appl. Mater. Interfaces (2024) doi:10.1021/acsami.4c16397.
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