Decellularized extracellular matrix biomaterials for regenerative therapies: Advances, challenges and clinical prospects.

Tissue engineering and regenerative medicine have shown potential in the repair and regeneration of tissues and organs via the use of engineered biomaterials and scaffolds. However, current constructs face limitations in replicating the intricate native microenvironment and achieving optimal regenerative capacity and functional recovery. To address these challenges, the utilization of decellularized tissues and cell-derived extracellular matrix (ECM) has emerged as a promising approach. These biocompatible and bioactive biomaterials can be engineered into porous scaffolds and grafts that mimic the structural and compositional aspects of the native tissue or organ microenvironment, both in vitro and in vivo. Bioactive dECM materials provide a unique tissue-specific microenvironment that can regulate and guide cellular processes, thereby enhancing regenerative therapies. In this review, we explore the emerging frontiers of decellularized tissue-derived and cell-derived biomaterials and bio-inks in the field of tissue engineering and regenerative medicine. We discuss the need for further improvements in decellularization methods and techniques to retain structural, biological, and physicochemical characteristics of the dECM products in a way to mimic native tissues and organs. This article underscores the potential of dECM biomaterials to stimulate in situ tissue repair through chemotactic effects for the development of growth factor and cell-free tissue engineering strategies. The article also identifies the challenges and opportunities in developing sterilization and preservation methods applicable for decellularized biomaterials and grafts and their translation into clinical products.

Decellularized Tissue-Induced Cellular Recruitment for Tissue Engineering and Regenerative Medicine.

Biomaterials that recapitulate the native in vivo microenvironment are promising to facilitate tissue repair and regeneration when used in combination with relevant growth factors (GFs), chemokines, cytokines, and other small molecules and cell sources. However, limitations with the use of exogenous factors and ex vivo cell expansion has prompted cell-/GF-free tissue engineering strategies. Additionally, conventional chemotaxis assays for studying cell migration behavior provide limited information, lack long-term stability, and fail to recapitulate physiologically relevant conditions. In this study, articular cartilage tissue-based biomaterials were developed via a rapid tissue decellularization protocol. The decellularized tissue was further processed into a hydrogel through solubilization and self-assembly. Chemotactic activity of the tissue-derived gel was investigated using sophisticated cellular migration assays. These tissue-derived extracellular matrix (ECM) biomaterials retain biochemical cues of native tissue and stimulate the chemotactic migration of hBMSCs in 2D and 3D cell migration models using a real-time chemotaxis assay. This strategy, in a way, developed a new paradigm in tissue engineering where cartilage tissue repair and regeneration can be approached with decellularized cartilage tissue in the place of an engineered matrix. This strategy can be further expanded for other tissue-based ECMs to develop cell-/GF-free tissue engineering and regenerative medicine strategies for recruiting endogenous cell populations to facilitate tissue repair and regeneration.

Nanofiber matrix formulations for the delivery of Exendin-4 for tendon regeneration: In vitro and in vivo assessment.

Tendon and ligament injuries are the most common musculoskeletal injuries, which not only impact the quality of life but result in a massive economic burden. Surgical interventions for tendon/ligament injuries utilize biological and/or engineered grafts to reconstruct damaged tissue, but these have limitations. Engineered matrices confer superior physicochemical properties over biological grafts but lack desirable bioactivity to promote tissue healing. While incorporating drugs can enhance bioactivity, large matrix surface areas and hydrophobicity can lead to uncontrolled burst release and/or incomplete release due to binding. To overcome these limitations, we evaluated the delivery of a peptide growth factor (exendin-4; Ex-4) using an enhanced nanofiber matrix in a tendon injury model. To overcome drug surface binding due to matrix hydrophobicity of poly(caprolactone) (PCL)-which would be expected to enhance cell-material interactions-we blended PCL and cellulose acetate (CA) and electrospun nanofiber matrices with fiber diameters ranging from 600 to 1000 nm. To avoid burst release and protect the drug, we encapsulated Ex-4 in the open lumen of halloysite nanotubes (HNTs), sealed the HNT tube endings with a polymer blend, and mixed Ex-4-loaded HNTs into the polymer mixture before electrospinning. This reduced burst release from ∼75% to ∼40%, but did not alter matrix morphology, fiber diameter, or tensile properties. We evaluated the bioactivity of the Ex-4 nanofiber formulation by culturing human mesenchymal stem cells (hMSCs) on matrix surfaces for 21 days and measuring tenogenic differentiation, compared with nanofiber matrices in basal media alone. Strikingly, we observed that Ex-4 nanofiber matrices accelerated the hMSC proliferation rate and elevated levels of sulfated glycosaminoglycan, tendon-related genes (Scx, Mkx, and Tnmd), and ECM-related genes (Col-I, Col-III, and Dcn), compared to control. We then assessed the safety and efficacy of Ex-4 nanofiber matrices in a full-thickness rat Achilles tendon defect with histology, marker expression, functional walking track analysis, and mechanical testing. Our analysis confirmed that Ex-4 nanofiber matrices enhanced tendon healing and reduced fibrocartilage formation versus nanofiber matrices alone. These findings implicate Ex-4 as a potentially valuable tool for tendon tissue engineering.

Smart Orthopedic Biomaterials and Implants.

Musculoskeletal injuries including bone defects continue to present a significant challenge in orthopedic surgery due to suboptimal healing. Bone reconstruction strategies focused on the use of biological grafts and bone graft substitutes in the form of biomaterials-based 3D structures in fracture repair. Recent advances in biomaterials science and engineering have resulted in the creation of intricate 3D bone-mimicking structures that are mechanically stable, biodegradable, and bioactive to support bone regeneration. Current efforts are focused on improving the biomaterial and implant physicochemical properties to promote interactions with the host tissue and osteogenesis. The "smart" biomaterials and their 3D structures are designed to actively interact with stem/progenitor cells and the extracellular matrix (ECM) to influence the local environment towards osteogenesis and de novo tissue formation. This article will summarize such smart biomaterials and the methodologies to apply either internal or external stimuli to control the tissue healing microenvironment. A particular emphasis is also made on the use of smart biomaterials and strategies to create functional bioactive implants for bone defect repair and regeneration.

Amorphous silica fiber matrix biomaterials: An analysis of material synthesis and characterization for tissue engineering.

Silica biomaterials including Bioglass offer great biocompatibility and bioactivity but fail to provide pore and degradation features needed for tissue engineering. Herein we report on the synthesis and characterization of novel amorphous silica fiber matrices to overcome these limitations. Amorphous silica fibers were fused by sintering to produce porous matrices. The effects of sacrificial polymer additives such as polyvinyl alcohol (PVA) and cellulose fibers (CF) on the sintering process were also studied. The resulting matrices formed between sintering temperatures of 1,350-1,550 °C retained their fiber structures. The matrices presented pores in the range of 50-200 µm while higher sintering temperatures resulted in increased pore diameter. PVA addition to silica significantly reduced the pore diameter and porosity compared with silica matrices with or without the addition of CF. The PVA additive morphologically appeared to fuse the silica fibers to a greater extent and resulted in significantly higher compressive modulus and strength than the rest of the matrices synthesized. These matrices lost roughly 30% of their original mass in an in vitro degradation study over 40 weeks. All matrices absorbed 500 wt% of water and did not change in their overall morphology, size, or shape with hydration. These fiber matrices supported human mesenchymal stem cell adhesion, proliferation, and mineralized matrix production. Amorphous silica fiber biomaterials/matrices reported here are biodegradable and porous and closely resemble the native extracellular matrix structure and water absorption capacity. Extending the methodology reported here to alter matrix properties may lead to a variety of tissue engineering, implant, and drug delivery applications.

Insulin-Functionalized Bioactive Fiber Matrices with Bone Marrow-Derived Stem Cells in Rat Achilles Tendon Regeneration.

Approximately half of annual musculoskeletal injuries in the US involve tendon tears. The naturally hypocellular and hypovascular tendon environment makes tendons injury-prone and heal slowly. Tendon tissue engineering strategies often use biomimetic scaffolds combined with bioactive factors and/or cells to enhance healing. FDA-approved growth factors to promote tendon healing are lacking, which highlights the need for safe and effective bioactive factors. Our previous work evaluated insulin as a bioactive factor and identified an optimal dose to promote in vitro mesenchymal stem cell survival, division, and tenogenesis. The present work evaluates the ability of insulin-functionalized electrospun nanofiber matrices with or without mesenchymal stem cells to enhance tendon repair in a rat Achilles injury model. Electrospun nanofiber matrices were functionalized with insulin, cultured with or without mesenchymal stem cells, and sutured to transected Achilles tendons in rats. We analyzed rat tendons 4 and 8 weeks after surgery for the tendon morphology, collagen production, and mechanical properties. Bioactive insulin-functionalized fiber matrices with mesenchymal stem cells resulted in significantly increased collagen I and III at 4 and 8 weeks postsurgery. Additionally, these matrices supported highly aligned collagen fibrils in the regenerated tendon tissue at 8 weeks. However, treatment- and control-regenerated tissues had similar tensile properties at 8 weeks, which were less than that of the native Achilles tendon. Our preliminary results establish the benefits of insulin-functionalized fiber matrices in promoting higher levels of collagen synthesis and alignment needed for functional recovery of tendon repair.

Engineering biomaterials to 3D-print scaffolds for bone regeneration: practical and theoretical consideration.

There are more than 2 million bone grafting procedures performed annually in the US alone. Despite significant efforts, the repair of large segmental bone defects is a substantial clinical challenge which requires bone substitute materials or a bone graft. The available biomaterials lack the adequate mechanical strength to withstand the static and dynamic loads while maintaining sufficient porosity to facilitate cell in-growth and vascularization during bone tissue regeneration. A wide range of advanced biomaterials are being currently designed to mimic the physical as well as the chemical composition of a bone by forming polymer blends, polymer-ceramic and polymer-degradable metal composites. Transforming these novel biomaterials into porous and load-bearing structures via three-dimensional printing (3DP) has emerged as a popular manufacturing technique to develop engineered bone grafts. 3DP has been adopted as a versatile tool to design and develop bone grafts that satisfy porosity and mechanical requirements while having the ability to form grafts of varied shapes and sizes to meet the physiological requirements. In addition to providing surfaces for cell attachment and eventual bone formation, these bone grafts also have to provide physical support during the repair process. Hence, the mechanical competence of the 3D-printed scaffold plays a key role in the success of the implant. In this review, we present various recent strategies that have been utilized to design and develop robust biomaterials that can be deployed for 3D-printing bone substitutes. The article also reviews some of the practical, theoretical and biological considerations adopted in the 3D-structure design and development for bone tissue engineering.

Bio-inspired zonal-structured matrices for bone-cartilage interface engineering.

Design and development of scaffold structures for osteochondral (OC) interface regeneration is a significant engineering challenge. Recent efforts are aimed at recapitulating the unique compositional and hierarchical structure of an OC interface. Conventional scaffold fabrication techniques often have limited design control and reproducibility, and the development of OC scaffolds with zonal hierarchy and structural integrity between zones is especially challenging. In this study, a series of multi-zonal and gradient structures were designed and fabricated using three-dimensional bioprinting. We developed OC scaffolds with bi-phasic and tri-phasic configurations to support the zonal structure of OC tissue, and gradient scaffold configurations to enable smooth transitions between the zones to more closely mimic a bone-cartilage interface. A biodegradable polymer, polylactic acid, was used for the fabrication of zonal/gradient scaffolds to provide mechanical strength and support OC function. The formation of the multi-zonal and gradient scaffolds was confirmed through scanning electron microscopy imaging and micro-computed tomography scanning. Precisely controlled hierarchy with tunable porosity along the scaffold length established the formation of the bio-inspired scaffolds with different zones/gradient structure. In addition, we also developed a novel bioprinting method to selectively introduce cells into desired scaffold zones of the zonal/gradient scaffolds via concurrent printing of a cell-laden hydrogel within the porous template. Live/dead staining of the cell-laden hydrogel introduced in the cartilage zone showed uniform cell distribution with high cell viability. Overall, our study developed bio-inspired scaffold structures with structural hierarchy and mechanical integrity for bone-cartilage interface engineering.

Correction: Growing a backbone - functional biomaterials and structures for intervertebral disc (IVD) repair and regeneration: challenges, innovations, and future directions.

Correction for 'Growing a backbone - functional biomaterials and structures for intervertebral disc (IVD) repair and regeneration: challenges, innovations, and future directions' by Matthew D. Harmon et al., Biomater. Sci., 2020, 8, 1216-1239, DOI: .

Biomaterial-directed cell behavior for tissue engineering.

Successful tissue regeneration strategies focus on the use of novel biomaterials, structures, and a variety of cues to control cell behavior and promote regeneration. Studies discovered how biomaterial/ structure cues in the form of biomaterial chemistry, material stiffness, surface topography, pore, and degradation properties play an important role in controlling cellular events in the contest of in vitro and in vivo tissue regeneration. Advanced biomaterials structures and strategies are developed to focus on the delivery of bioactive factors, such as proteins, peptides, and even small molecules to influence cell behavior and regeneration. The present article is an effort to summarize important findings and further discuss biomaterial strategies to influence and control cell behavior directly via physical and chemical cues. This article also touches on various modern methods in biomaterials processing to include bioactive factors as signaling cues to program cell behavior for tissue engineering and regenerative medicine.

Gradient scaffold with spatial growth factor profile for osteochondral interface engineering.

Osteochondral (OC) matrix design poses a significant engineering challenge due to the complexity involved with bone-cartilage interfaces. To better facilitate the regeneration of OC tissue, we developed and evaluated a biodegradable matrix with uniquely arranged bone and cartilage supporting phases: a poly(lactic-co-glycolic) acid (PLGA) template structure with a porosity gradient along its longitudinal axis uniquely integrated with hyaluronic acid hydrogel. Micro-CT scanning and imaging confirmed the formation of an inverse gradient matrix. Hydroxyapatite was added to the PLGA template which was then plasma-treated to increase hydrophilicity and growth factor affinity. An osteogenic growth factor (bone morphogenetic protein 2; BMP-2) was loaded onto the template scaffold via adsorption, while a chondrogenic growth factor (transforming growth factor beta 1; TGF-ß1) was incorporated into the hydrogel phase. Confocal microscopy of the growth factor loaded matrix confirmed the spatial distribution of the two growth factors, with chondrogenic factor confined to the cartilaginous portion and osteogenic factor present throughout the scaffold. We observed spatial differentiation of human mesenchymal stem cells (hMSCs) into cartilage and bone cells in the scaffoldsin vitro: cartilaginous regions were marked by increased glycosaminoglycan production, and osteogenesis was seen throughout the graft by alizarin red staining. In a dose-dependent study of BMP-2, hMSC pellet cultures with TGF-ß1 and BMP-2 showed synergistic effects on chondrogenesis. These results indicate that development of an inverse gradient matrix can spatially distribute two different growth factors to facilitate chondrogenesis and osteogenesis along different portions of a scaffold, which are key steps needed for formation of an OC interface.

Evaluation of Autologously Derived Biomaterials and Stem Cells for Bone Tissue Engineering.

Despite progress, clinical translation of tissue engineering (TE) products/technologies is limited. A significant effort is underway to develop biomaterials and cells through a minimally modified process for clinical translation of TE products. Recently, bone marrow aspirate (BMA) was identified as an autologous source of cells for TE applications and is currently being tested in clinical therapies, but the isolation methods need improvement to avoid potential for contamination and increase progenitor cell yield. To address these issues, we reproducibly processed human peripheral blood (PB) and BMA to develop autologously derived biomaterials and cells. We demonstrated PB-derived biomaterial/gel cross-linking and fibrin gel formation with varied gelation times as well as biocompatibility through support of human bone marrow-derived stem cell survival and growth in vitro. Next, we established a plastic culture-free process that concentrates and increases the yield of CD146+/CD271+ early mesenchymal progenitor cells in BMA (concentrated BMA [cBMA]). cBMA exhibited increased colony formation and multipotency (including chondrogenic differentiation) in vitro compared with standard BMA. PB-derived gels encapsulated with cBMA also demonstrated increased cell proliferation and enhanced mineralization when assessed for bone TE in vitro. This strategy can potentially be developed for use in any tissue regeneration application; however, bone regeneration was used as a test bed for this study.

Bioactive polymeric materials and electrical stimulation strategies for musculoskeletal tissue repair and regeneration.

Electrical stimulation (ES) is predominantly used as a physical therapy modality to promote tissue healing and functional recovery. Research efforts in both laboratory and clinical settings have shown the beneficial effects of this technique for the repair and regeneration of damaged tissues, which include muscle, bone, skin, nerve, tendons, and ligaments. The collective findings of these studies suggest ES enhances cell proliferation, extracellular matrix (ECM) production, secretion of several cytokines, and vasculature development leading to better tissue regeneration in multiple tissues. However, there is still a gap in the clinical relevance for ES to better repair tissue interfaces, as ES applied clinically is ineffective on deeper tissue. The use of a conducting material can transmit the stimulation applied from skin electrodes to the desired tissue and lead to an increased function on the repair of that tissue. Ionically conductive (IC) polymeric scaffolds in conjunction with ES may provide solutions to utilize this approach effectively. Injectable IC formulations and their scaffolds may provide solutions for applying ES into difficult to reach tissue types to enable tissue repair and regeneration. A better understanding of ES-mediated cell differentiation and associated molecular mechanisms including the immune response will allow standardization of procedures applicable for the next generation of regenerative medicine. ES, along with the use of IC scaffolds is more than sufficient for use as a treatment option for single tissue healing and may fulfill a role in interfacing multiple tissue types during the repair process.

Growing a backbone - functional biomaterials and structures for intervertebral disc (IVD) repair and regeneration: challenges, innovations, and future directions.

Back pain and associated maladies can account for an immense amount of healthcare cost and loss of productivity in the workplace. In particular, spine related injuries in the US affect upwards of 5.7 million people each year. The degenerative disc disease treatment almost always arises due to a clinical presentation of pain and/or discomfort. Preferred conservative treatment modalities include the use of non-steroidal anti-inflammatory medications, physical therapy, massage, acupuncture, chiropractic work, and dietary supplements like glucosamine and chondroitin. Artificial disc replacement, also known as total disc replacement, is a treatment alternative to spinal fusion. The goal of artificial disc prostheses is to replicate the normal biomechanics of the spine segment, thereby preventing further damage to neighboring sections. Artificial functional disc replacement through permanent metal and polymer-based components continues to evolve, but is far from recapitulating native disc structure and function, and suffers from the risk of unsuccessful tissue integration and device failure. Tissue engineering and regenerative medicine strategies combine novel material structures, bioactive factors and stem cells alone or in combination to repair and regenerate the IVD. These efforts are at very early stages and a more in-depth understanding of IVD metabolism and cellular environment will also lead to a clearer understanding of the native environment which the tissue engineering scaffold should mimic. The current review focusses on the strategies for a successful regenerative scaffold for IVD regeneration and the need for defining new materials, environments, and factors that are so finely tuned in the healthy human intervertebral disc in hopes of treating such a prevalent degenerative process.

Evaluation of an Engineered Hybrid Matrix for Bone Regeneration via Endochondral Ossification.

Despite its regenerative ability, long and segmental bone defect repair remains a significant orthopedic challenge. Conventional tissue engineering efforts induce bone formation through intramembranous ossification (IO) which limits vascular formation and leads to poor bone regeneration. To overcome this challenge, a novel hybrid matrix comprised of a load-bearing polymer template and a gel phase is designed and assessed for bone regeneration. Our previous studies developed a synthetic ECM, hyaluronan (HA)-fibrin (FB), that is able to mimic cartilage-mediated bone formation in vitro. In this study, the well-characterized HA-FB hydrogel is combined with a biodegradable polymer template to form a hybrid matrix. In vitro evaluation of the matrix showed cartilage template formation, cell recruitment and recruited cell osteogenesis, essential stages in endochondral ossification. A transgenic reporter-mouse critical-defect model was used to evaluate the bone healing potential of the hybrid matrix in vivo. The results demonstrated host cell recruitment into the hybrid matrix that led to new bone formation and subsequent remodeling of the mineralization. Overall, the study developed and evaluated a novel load-bearing graft system for bone regeneration via endochondral ossification.

Synthesis and characterization of photocrosslinkable hydrogels from bovine skin gelatin.

Hydrogels that mimic native tissues chemically and structurally have been increasingly sought for a wide variety of tissue engineering applications. Gelatin can be naturally derived from different sources and functionalized to fabricate hydrogels that exhibit high cytocompatibility and favorable biodegradable properties. The amino groups on the gelatin backbone can be substituted by adding varying proportions of methacrylic anhydride (MAA) to create biomimetic hydrogels which can be used as tissue engineering scaffolds. Gelatin from different sources yields hydrogels with distinctive physical, chemical, and biological properties. In this work, gelatin from bovine skin was used to fabricate hydrogels with varying degrees of crosslinking content using 1, 4, 7, and 10 mL MAA. The material properties of these hydrogels were characterized. The cytocompatibility of the gelatin-based hydrogels was studied using L6 rat myoblasts. The hydrogels from bovine skin gelatin exhibit mechanical properties that are conducive for applications which require substrates to propagate cell growth, migration, and proliferation rapidly. These hydrogels exhibit exceptional tunability behavior which makes them useful and applicable to culture different cell types.

Self-neutralizing PLGA/magnesium composites as novel biomaterials for tissue engineering.

Controlling acidic degradation of biodegradable polyesters remains a major clinical challenge. This work presents a simple and effective strategy of developing polyester composites with biodegradable magnesium metal or alloys. PLGA samples with compositions of 1, 3, 5, and 10 wt% magnesium were produced using a simple solvent-casting method, which resulted in composite films with near uniform Mg metal/alloy particle dispersion. Degradation study of the composite films showed that all compositions higher than 1 wt% magnesium were able to extend the duration of degradation, and buffer acidic pH resulting from PLGA degradation. PLGA composite with 5 wt% of magnesium showed near-neutral degradation pattern under sink conditions. Magnesium addition also showed improved mechanical characteristics in terms of the tensile modulus. In vitro experiments conducted by seeding PLGA composites with MC3T3-E1 pre-osteoblasts demonstrated increased ALP expression and cellular mineralization. The established new biodegradable polymer-metal system provides a useful biomaterial platform with a wide range of applications in biomedical device development and scaffold-based tissue engineering.

Hybrid extracellular matrix design for cartilage-mediated bone regeneration.

Recapitulating long bone repair through endochondral ossification (EO) is increasingly becoming a more popular approach. A successful EO Process depends greatly on the establishment of a healthy hypertrophic-cartilage template (HCT). The aim of this work is to design a hydrogel system, which closely mimics the extracellular matrix of HCT. We examined the combinatorial effect of two commonly used hydrogels for bone and cartilage regeneration strategies, hyaluronan (HA) and fibrin (FB), to induce HCT formation. Hydrogel combinations were evaluated using a clinically relevant cell source, human bone marrow mesenchymal stem cells (hBMSCs). The results establish that with increasing HA (50-90%) the chondrogenic and its subsequent hypertrophy trend improved, with 70:30 HA:FB combination showing the highest and most uniform expression of chondrogenic and hypertrophic stage specific markers. This combination also showed superior support for cell micro-aggregation and differentiation. Thus, 70:30 HA-FB matrix demonstrated a healthy formation of chondrogenic and hypertrophic stages with rich stage-specific ECM components. This study demonstrates that with the appropriate hydrogel design it is possible to develop effective tissue engineering therapies for bone defect repair and regeneration through endochondral ossification by establishing a healthy HCT. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 106B: 300-309, 2018.

* Harnessing External Cues: Development and Evaluation of an In Vitro Culture System for Osteochondral Tissue Engineering.

Over the last decade, engineered structures have been developed for osteochondral (OC) tissue regeneration. While the optimal structure design is yet to be determined, these scaffolds require in vitro evaluation before clinical use. However, the means by which complex scaffolds, such as OC scaffolds, can be tested are limited. Taking advantage of a mesenchymal stem cell's (MSC's) ability to respond to its surrounding we harness external cues, such as the cell's mechanical environment and delivered factors, to create an in vitro culture system for OC tissue engineering with a single cell source on a gradient yet integrated scaffold system. To do this, the effect of hydrogel stiffness on the expression of human MSCs (hMSCs) chondrogenic differentiation was studied using histological analysis. Additionally, hMSCs were also cultured in different combinations of chondrogenic and osteogenic media to develop a co-differentiation media suitable for OC lineage differentiation. A uniquely graded (density-gradient matrix) OC scaffold with a distal cartilage hydrogel phase specifically tailored to support chondrogenic differentiation was cultured using a newly developed "simulated in vivo culture method." The scaffold's culture in co-differentiation media models hMSC infiltration into the scaffold and subsequent differentiation into the distal cartilage and proximal bone layers. Cartilage and bone marker staining along with specific matrix depositions reveal the effect of external cues on the hMSC differentiation. As a result of these studies a model system was developed to study and culture OC scaffolds in vitro.

Patient-Derived and Intraoperatively Formed Biomaterial for Tissue Engineering.

In this chapter, we introduce a completely intraoperative procedure for obtaining a patient-derived biomaterial in cell therapy and tissue engineering applications. An automated device for processing human peripheral-blood ensures a reproducible method for retrieving the patient's cellular-rich as well as cellular-poor plasma. By substituting calcium for animal-derived thrombin, we engineer a completely autologous hydrogel that eliminates the risk of disease transmission and lowers FDA regulation hurdles. Through this chapter, we will discuss a bedside protocol developed to prepare a patient-derived hydrogel. This method can be effectively used to develop a completely intraoperative tissue engineering strategy (CITES) that can be easily translated into the clinic for surgical use.