Engineered lumenized vascular networks and support matrix
Inventors
Glazier, James A. • Alileche, Abdelkrim • SHIRINIFARD, Abbas • Amarie, Dragos
Assignees
National Institutes of Health NIH • Indiana University Bloomington
Publication Number
US-10196596-B2
Publication Date
2019-02-05
Expiration Date
2031-03-15
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Abstract
Disclosed herein are capillary fabrication devices comprising living cells within a support medium. Culture of the cells produces viable lumenized capillary networks with natural or pre-determined geometries and ECM and basement membrane associated with the capillary networks. The capillary networks and the ECM and basement membrane detachable from the capillary networks are useful for tissue engineering applications.
Core Innovation
The invention relates to capillary fabrication devices comprising living cells within a support medium. Upon culturing, these devices produce viable lumenized capillary networks with natural or pre-determined geometries, along with associated extracellular matrix (ECM) and basement membrane. These capillary networks and ECM are detachable and useful for tissue engineering applications.
The problem being addressed is the limitation of existing in vitro angiogenesis techniques which are incapable of rapidly creating lumenized capillary networks with properties substantially similar to in vivo networks. Current tissue engineering methods are limited by difficulties in forming functional vascular networks, especially for thicker, metabolically demanding organs like heart and liver. Existing techniques cannot produce controlled-geometry microvessels capable of connecting to host vessels or preventing vascular regression.
Claims Coverage
The patent discloses one independent claim directed to a device for fabrication of engineered capillary networks. The main inventive features involve the composition and characteristics of the device's components, including the cells, cell-culture surface, and support medium.
Use of endothelial lineage cells with a non-treated polystyrene surface
The device specifically comprises endothelial lineage cells cultured on a non-treated polystyrene cell-culture surface, providing a unique environment for capillary network formation.
Low-density support-generating medium with specific gel-forming material dilution and thickness
The support-generating medium includes a gel-forming material substantially dissolved in liquid cell-culture medium in a ratio of 1:30 to 1:60, forming a support medium 20 to 100 microns thick that covers the endothelial cells and culture medium.
Inclusion of additional supportive cell types
The device may further comprise at least one additional cell type selected from pericytes, smooth muscle cells, fibroblasts, or any combination thereof to aid capillary network stability.
Modification of the cell-culture surface by hydrophobic patterning and polymer coatings
The non-treated polystyrene surface may include hydrophobic regions or coatings of temperature sensitive polymers, with possible modifications by etching, stamping, contact printing, or UV laser ablation to define cellular growth patterns.
Specific composition of cell-culture medium and gel-forming material
The liquid cell-culture medium may contain serum albumin, bicarbonate-base, HEPES buffer, and ECM proteins, while the gel-forming material can include proteins such as laminin, collagen IV, and others, with Matrigel™ as a preferred material, dissolved to yield specified ECM protein concentrations.
Overall, the claims focus on a capillary fabrication device characterized by endothelial lineage cells cultured on a non-treated polystyrene surface with a low-density, specifically diluted gel-forming support medium forming a thin support matrix, optionally incorporating additional supportive cell types and surface modifications to control capillary network formation and viability.
Stated Advantages
The capillary fabrication devices produce capillaries with morphology and tubule diameters closely matching in vivo capillaries, unlike standard devices producing abnormally elongated endothelial cells and non-functional tubes.
The devices create lumenized capillary networks that remain viable for up to four weeks, enabling longer term studies and applications unlike existing devices where networks degrade within 48 hours.
The support-generating medium permits normal cell motility and proliferation of non-endothelial lineage cells, making the devices less sensitive to initial cell density and allowing capillary formation from limited or autologous cell sources.
The use of non-treated polystyrene surfaces combined with the support-generating medium allows cell survival and differentiation into capillary networks, unlike cells plated on treated surfaces which form confluent undifferentiated layers.
Custom-patterned capillary fabrication devices enable formation of controlled-geometry capillary networks with predefined patterns to optimize blood perfusion and integration in engineered tissues.
The engineered ECM and basement membrane can be detached with minimal damage and are suitable for tissue repair and engineering, facilitating autologous or cross-species applications with reduced rejection risk.
Documented Applications
In vitro angiogenesis assays for screening angiogenesis promoters and inhibitors relevant to wound healing, age-related macular degeneration, diabetes, cancer, and other diseases.
Tissue engineering for creating vascular replacements capable of integration with host vessels, enabling formation of functional capillary networks for thicker, metabolically demanding organs like heart and liver.
Use of detached ECM and basement membrane as scaffolds to promote growth of the patient's own vasculature in tissue repair and engineering applications.
Cell-sheet engineering and bioprinting applications where layers of engineered capillary networks and ECM are combined with engineered tissue layers to enhance vascular network formation.
Implantation of viable capillary networks in vivo to aid tissue repair and regeneration, demonstrated by implantation onto chick chorioallantoic membrane.
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