💪Cell and Tissue Engineering Unit 9 – Blood Vessel Tissue Engineering

Key Concepts in Blood Vessel Engineering

• Involves creating functional blood vessels in the lab using cells, biomaterials, and engineering techniques
• Aims to address the shortage of viable blood vessels for transplantation and bypass surgeries
• Requires an understanding of the complex structure and function of native blood vessels
• Utilizes principles from cell biology, biomaterials science, and biomedical engineering
• Focuses on recreating the three main layers of blood vessels: tunica intima, tunica media, and tunica adventitia
  • Tunica intima consists of a single layer of endothelial cells that line the vessel lumen
  • Tunica media is composed of smooth muscle cells and elastic fibers that provide structural support and regulate vessel diameter
  • Tunica adventitia is the outermost layer made up of fibroblasts and connective tissue
• Involves selecting appropriate cell sources, biomaterials, and fabrication techniques to mimic native blood vessel properties
• Requires the use of bioreactor systems to provide mechanical and biochemical cues for cell growth and tissue maturation

Structure and Function of Blood Vessels

• Blood vessels are a critical component of the cardiovascular system that transport blood throughout the body
• Arteries carry oxygenated blood away from the heart to the tissues, while veins return deoxygenated blood to the heart
• Capillaries are the smallest blood vessels that facilitate the exchange of nutrients, oxygen, and waste products between blood and tissues
• The endothelial cells lining the vessel lumen play a crucial role in maintaining blood flow, preventing thrombosis, and regulating vascular tone
• Smooth muscle cells in the tunica media are responsible for vasoconstriction and vasodilation, which control blood pressure and flow
• The extracellular matrix (ECM) provides structural support and influences cell behavior through mechanical and biochemical signaling
  • ECM components include collagen, elastin, fibronectin, and proteoglycans
• Blood vessels exhibit unique mechanical properties, such as compliance and burst strength, which are essential for their function
  • Compliance refers to the ability of a vessel to expand and contract in response to changes in blood pressure
  • Burst strength is the maximum pressure a vessel can withstand before rupturing

Challenges in Blood Vessel Engineering

• Recreating the complex, hierarchical structure of native blood vessels with multiple cell types and ECM components
• Achieving appropriate mechanical properties, such as compliance and burst strength, to withstand physiological blood pressures and flow rates
• Ensuring proper cell alignment and organization, particularly for smooth muscle cells in the tunica media
• Promoting the formation of a confluent and functional endothelial cell layer to maintain blood flow and prevent thrombosis
• Encouraging the production and remodeling of ECM components by cells to mimic native tissue composition
• Preventing immune rejection and ensuring long-term patency of engineered blood vessels after implantation
• Scaling up the fabrication process to create blood vessels of clinically relevant sizes and lengths
• Developing cost-effective and efficient manufacturing methods for large-scale production of engineered blood vessels

Biomaterials for Vascular Scaffolds

• Biomaterials serve as temporary scaffolds to support cell growth and tissue formation in blood vessel engineering
• Ideal biomaterials should be biocompatible, biodegradable, and possess suitable mechanical properties
• Natural biomaterials, such as collagen, fibrin, and silk fibroin, are derived from biological sources and offer excellent biocompatibility
  • Collagen is the most abundant ECM protein in native blood vessels and promotes cell adhesion and proliferation
  • Fibrin, formed from the polymerization of fibrinogen, can be used to create hydrogels that mimic the blood clotting process
  • Silk fibroin, derived from silkworm cocoons, has high tensile strength and slow degradation rates
• Synthetic biomaterials, such as polyglycolic acid (PGA), polylactic acid (PLA), and poly(lactic-co-glycolic acid) (PLGA), offer tunable properties and reproducibility
  • PGA is a highly crystalline polymer that degrades rapidly and has been used to create porous vascular scaffolds
  • PLA is a more hydrophobic polymer with slower degradation rates and better mechanical properties than PGA
  • PLGA is a copolymer of PGA and PLA that allows for the adjustment of degradation rates and mechanical properties by varying the monomer ratios
• Decellularized tissue matrices, such as small intestinal submucosa (SIS) and umbilical cord vein (UCV), provide a natural ECM structure for cell growth
  • Decellularization involves removing cells from native tissues while preserving the ECM composition and structure
  • SIS is derived from the small intestine and has been used to create tubular scaffolds for blood vessel engineering
  • UCV is obtained from human umbilical cords and has a natural tubular structure with a confluent endothelial cell layer

Cell Sources and Considerations

• Selecting appropriate cell sources is crucial for the success of blood vessel engineering
• Autologous cells, derived from the patient, offer the advantage of avoiding immune rejection but may be limited in availability and functionality
  • Endothelial cells can be isolated from a patient's blood vessels (e.g., saphenous vein) or differentiated from autologous stem cells
  • Smooth muscle cells can be obtained from a patient's blood vessels or differentiated from autologous stem cells
• Allogeneic cells, derived from donors, provide a more readily available source but may require immunosuppression to prevent rejection
  • Umbilical cord-derived endothelial cells and smooth muscle cells have high proliferative capacity and can be expanded in vitro
  • Mesenchymal stem cells (MSCs) from bone marrow or adipose tissue can be differentiated into vascular cell types
• Induced pluripotent stem cells (iPSCs) can be generated from a patient's own cells and differentiated into vascular cell types, offering a potentially unlimited cell source
• Co-culture of endothelial cells and smooth muscle cells can promote cell-cell interactions and improve the overall function of engineered blood vessels
• Ensuring high cell viability, purity, and functionality during isolation, expansion, and seeding processes is essential for successful blood vessel engineering

Fabrication Techniques

• Various fabrication techniques have been developed to create 3D vascular constructs with desired geometries and properties
• Electrospinning involves using an electric field to draw polymer fibers from a solution, creating nanofibrous scaffolds that mimic the ECM structure
  • Electrospun scaffolds can be functionalized with bioactive molecules to promote cell adhesion and growth
  • Aligned electrospun fibers can guide cell orientation and improve mechanical properties
• Bioprinting uses 3D printing technology to precisely deposit cells and biomaterials in a layer-by-layer fashion, allowing for the creation of complex vascular networks
  • Extrusion-based bioprinting can be used to create tubular structures by depositing cell-laden hydrogels or bioinks
  • Laser-assisted bioprinting offers high resolution and can be used to create intricate vascular patterns
• Microfluidic systems can be employed to create perfusable vascular networks within hydrogels or other biomaterials
  • Sacrificial templates, such as carbohydrate glass or gelatin, can be printed and later removed to create hollow channels
  • Vasculogenesis-inspired approaches involve the self-assembly of endothelial cells into capillary-like networks within hydrogels
• Cell sheet engineering involves culturing cells on thermoresponsive surfaces that allow for the detachment of intact cell sheets, which can be stacked or rolled to form vascular constructs
• Decellularization and recellularization of native blood vessels can be used to create tissue-engineered vascular grafts with natural ECM structure and composition

Bioreactor Systems and Conditioning

• Bioreactor systems are used to provide controlled environments for the growth and maturation of engineered blood vessels
• Perfusion bioreactors allow for the continuous flow of culture medium through the vascular construct, mimicking physiological blood flow conditions
  • Pulsatile flow can be applied to simulate the cyclic stretching and relaxation of blood vessels in vivo
  • Shear stress generated by fluid flow can stimulate endothelial cell alignment and function
• Mechanical conditioning, such as cyclic stretching or pressure loading, can be applied to improve the mechanical properties and cell organization of engineered blood vessels
  • Cyclic stretching can promote the circumferential alignment of smooth muscle cells and the production of elastin fibers
  • Pressure loading can increase the burst strength and compliance of engineered blood vessels
• Biochemical conditioning involves the addition of growth factors, cytokines, or other signaling molecules to the culture medium to guide cell behavior and tissue development
  • Vascular endothelial growth factor (VEGF) can promote endothelial cell proliferation and migration
  • Transforming growth factor-beta (TGF-β) can stimulate smooth muscle cell differentiation and ECM production
• Monitoring and controlling environmental parameters, such as temperature, pH, and oxygen levels, is essential for maintaining cell viability and function within the bioreactor system
• Bioreactor systems can also be used for the dynamic seeding of cells onto scaffolds, promoting uniform cell distribution and attachment

Clinical Applications and Future Directions

• Engineered blood vessels have the potential to address the shortage of autologous and allogeneic grafts for various clinical applications
• Coronary artery bypass grafting (CABG) is a common procedure that requires the use of blood vessel grafts to bypass blocked coronary arteries
  • Tissue-engineered vascular grafts could provide an alternative to autologous saphenous vein or internal mammary artery grafts
  • Off-the-shelf availability of engineered grafts could reduce the need for invasive harvesting procedures and associated complications
• Peripheral artery disease (PAD) involves the narrowing or blockage of blood vessels in the legs, leading to reduced blood flow and tissue damage
  • Engineered blood vessels could be used as bypass grafts to restore blood flow to the affected limbs
  • Small-diameter vascular grafts are particularly challenging due to the high risk of thrombosis and intimal hyperplasia
• Hemodialysis access for patients with end-stage renal disease requires the creation of an arteriovenous fistula or graft for repeated needle punctures
  • Tissue-engineered vascular access grafts could provide a more durable and infection-resistant alternative to synthetic grafts
  • Endothelialization of the graft lumen could reduce the risk of thrombosis and improve long-term patency
• Future directions in blood vessel engineering include the development of more advanced biomaterials, such as smart or responsive materials that can adapt to the physiological environment
• Incorporating multiple cell types, such as pericytes or immune cells, could further improve the functionality and long-term stability of engineered blood vessels
• Developing efficient and scalable manufacturing processes, such as automated bioreactor systems or 3D bioprinting, will be essential for the clinical translation and commercialization of engineered blood vessels
• Conducting long-term preclinical and clinical studies to assess the safety, efficacy, and durability of tissue-engineered vascular grafts in various patient populations