The interplay between tissue growth and scaffold degradation in engineered tissue constructs

In vitro tissue engineering is emerging as a potential tool to meet the high demand for replacement tissue, caused by the increased incidence of tissue degeneration and damage. A key challenge in this field is ensuring that the mechanical properties of the engineered tissue are appropriate for the in vivo environment. Achieving this goal will require detailed understanding of the interplay between cell proliferation, extracellular matrix (ECM) deposition and scaffold degradation. In this paper, we use a mathematical model (based upon a multiphase continuum framework) to investigate the interplay between tissue growth and scaffold degradation during tissue construct evolution in vitro. Our model accommodates a cell population and culture medium, modelled as viscous fluids, together with a porous scaffold and ECM deposited by the cells, represented as rigid porous materials. We focus on tissue growth within a perfusion bioreactor system, and investigate how the predicted tissue composition is altered under the influence of (1) differential interactions between cells and the supporting scaffold and their associated ECM, (2) scaffold degradation, and (3) mechanotransduction-regulated cell proliferation and ECM deposition. Numerical simulation of the model equations reveals that scaffold heterogeneity typical of that obtained from $\mu $ CT scans of tissue engineering scaffolds can lead to significant variation in the flow-induced mechanical stimuli experienced by cells seeded in the scaffold. This leads to strong heterogeneity in the deposition of ECM. Furthermore, preferential adherence of cells to the ECM in favour of the artificial scaffold appears to have no significant influence on the eventual construct composition; adherence of cells to these supporting structures does, however, lead to cell and ECM distributions which mimic and exaggerate the heterogeneity of the underlying scaffold. Such phenomena have important ramifications for the mechanical integrity of engineered tissue constructs and their suitability for implantation in vivo.     

Influence of tissue- and cell-scale extracellular matrix distribution on the mechanical properties of tissue-engineered cartilage

The insufficient load-bearing capacity of today’s tissue- engineered (TE) cartilage limits its clinical application. Generally, cartilage TE studies aim to increase the extracellular matrix (ECM) content, as this is thought to determine the load-bearing properties of the cartilage. However, there are apparent inconsistencies in the literature regarding the correlation between ECM content and mechanical properties of TE constructs. In addition to the amount of ECM, the spatial inhomogeneities in ECM distribution at the tissue scale as well as at the cell scale may affect the mechanical properties of TE cartilage. The relative importance of such structural inhomogeneities on mechanical behavior of TE cartilage is unknown. The aim of the present study was, therefore, to theoretically elucidate the influence of these inhomogeneities on the mechanical behavior of chondrocyte-agarose TE constructs. A validated non-linear fiber-reinforced poro-elastic swelling cartilage model that can accommodate for effects of collagen reinforcement and swelling by proteoglycans was used. At the tissue scale, ECM was gradually varied from predominantly localized in the periphery of the TE construct toward an ECM-rich inner core. The effect of these inhomogeneities in relation to the total amount of ECM was also evaluated. At the cell scale, ECM was gradually varied from localized in the pericellular area, toward equally distributed throughout the interterritorial area. Results from the tissue-scale model indicated that localization of ECM in either the construct periphery or in the inner core may reduce construct stiffness compared with that of constructs with homogeneous ECM. Such effects are more significant at high ECM amounts. At the cell scale, localization of ECM around the cells significantly reduced the overall stiffness, even at low ECM amounts. The compressive stiffness gradually increased when ECM distribution became more homogeneous and the osmotic swelling pressure in the interterritorial area increased. We conclude that for the same amount of ECM content in TE cartilage constructs, superior mechanical properties can be achieved with more homogeneous ECM distribution at both tissue and cell scale. Inhomogeneities at the cell scale are more important than those at the tissue scale.     

Prediction of extracellular matrix stiffness in engineered heart valve tissues based on nonwoven scaffolds

The in vitro development of tissue engineered heart valves (TEHV) exhibiting appropriate structural and mechanical characteristics remains a significant challenge. An important step yet to be addressed is establishing the relationship between scaffold and extracellular matrix (ECM) mechanical properties. In the present study, a composite beam model accounting for nonwoven scaffold-ECM coupling and the transmural collagen concentration distribution was developed, and utilized to retrospectively estimate the ECM effective stiffness in TEHV specimens incubated under static and cyclic flexure conditions (Engelmayr Jr et~al. in Biomaterials 26(2):175-187 2005). The ECM effective stiffness was expressed as the product of the local collagen concentration and the collagen specific stiffness (i.e., stiffness/concentration), and was related to the overall TEHV effective stiffness via an empirically determined scaffold-ECM coupling parameter and measured transmural collagen concentration distributions. The scaffold-ECM coupling parameter was determined by flexural mechanical testing of polyacrylamide gels (i.e., ECM analogs) of variable stiffness and associated scaffold-polyacrylamide gel composites (i.e., engineered tissue analogs). The transmural collagen concentration distributions were quantified from fluorescence micrographs of picro-sirius red stained TEHV sections. As suggested by a previous structural model of the nonwoven scaffold (Engelmayr Jr and Sacks in J Biomech Eng 128(4):610-622, 2006), nonwoven scaffold-ECM composites did not follow a traditional rule of mixtures. The present study provided further evidence that the primary mode of reinforcement in nonwoven scaffold-ECM composites is an increase in the number fiber-fiber bonds with a concomitant increase in the effective stiffness of the spring-like fiber segments. Simulations of potential ECM deposition scenarios using the current model indicated that the present approach is sensitive to the specific time course of tissue deposition, and is thus very suitable for studies of ECM formation in engineered heart valve tissues.     

A mechanical composite spheres analysis of engineered cartilage dynamics

In the preparation of bioengineered reparative strategies for damaged or diseased tissues, the processes of biomaterial degradation and neotissue synthesis combine to affect the developing mechanical state of multiphase, composite engineered tissues. Here, cell-polymer constructs for engineered cartilage have been fabricated by seeding chondrocytes within three-dimensional scaffolds of biodegradable polymers. During culture, synthetic scaffolds degraded passively as the cells assembled an extracellular matrix (ECM) composed primarily of glycosaminoglycan and collagen. Biochemical and biomechanical assessment of the composite (cells, ECM, and polymer scaffold) were modeled at a unit-cell level to mathematically solve stress-strain relationships and thus construct elastic properties (n=4 samples per seven time points). This approach employed a composite spheres, micromechanical analysis to determine bulk moduli of: (1) the cellular-ECM inclusion within the supporting scaffold structure; and (2) the cellular inclusion within its ECM. Results indicate a dependence of constituent volume fractions with culture time (p<0.05). Overall mean bulk moduli were variably influenced by culture, as noted for the cell-ECM inclusion (K(c-m)=29.7 kPa, p=0.1439), the cellular inclusion (K(c)=5.5 kPa, p=0.0067), and its surrounding ECM (K(m)=373.9 kPa, p=0.0748), as well as the overall engineered construct (K=165.0 kPa, p=0.6899). This analytical technique provides a framework to describe the time-dependent contribution of cells, accumulating ECM, and a degrading scaffold affecting bioengineered construct mechanical properties.     

Modeling the dynamic composition of engineered cartilage

Mathematical models to describe extracellular matrix (ECM) deposition and scaffold degradation in cell-polymer constructs for the design of engineered cartilage were developed and validated. The ECM deposition model characterized a product-inhibition mechanism in the concentration of cartilage molecules, collagen and glycosaminoglycans (GAG). The scaffold degradation model used first-order kinetics to describe hydrolysis (not limited by diffusion) of biodegradable polyesters, polyglycolic acid and polylactic acid. Each model was fit to published accumulation and degradation data. As experimental validation, cell-polymer constructs (n=24) and unseeded scaffolds (n=24) were cultured in vitro. Biochemical assays for ECM content and measurements of scaffold mass were performed at 1, 2, 4, 6, 8, or 10 weeks (n=8 per time point). The models demonstrated a strong fit with published data and experimental results (R(2)=0.75 to 0.99) and predicted the temporal total construct mass with reasonable accuracy (30% RMS error). This approach can elucidate mechanisms governing accumulation/degradation and may be coupled with structure-function relationships to describe time-dependent changes in construct elastic properties.     

A validated model of GAG deposition,cell distribution,and growth of tissue engineered cartilage cultured in a rotating bioreactor

In this work a new phenomenological model of growth of cartilage tissue cultured in a rotating bioreactor is developed. It represents an advancement of a previously derived model of deposition of glycosaminoglycan (GAG) in engineered cartilage by (i) introduction of physiological mechanisms of proteoglycan accumulation in the extracellular matrix (ECM) as well as by correlating (ii) local cell densities and (iii) tissue growth to the ECM composition. In particular, previously established predictions and correlations of local oxygen concentrations and GAG synthesis rates are extended to distinguish cell secreted proteoglycan monomers free to diffuse in cell surroundings and outside from the engineered construct, from large aggrecan molecules, which are constrained within the ECM and practically immovable. The model includes kinetics of aggregation, that is, transformation of mobile GAG species into immobile aggregates as well as maintenance of the normal ECM composition after the physiological GAG concentration is reached by incorporation of a product inhibition term. The model also includes mechanisms of the temporal evolution of cell density distributions and tissue growth under in vitro conditions. After a short initial proliferation phase the total cell number in the construct remains constant, but the local cell distribution is leveled out by GAG accumulation and repulsion due to negative molecular charges. Furthermore, strong repulsive forces result in expansion of the local tissue elements observed macroscopically as tissue growth (i.e., construct enlargement). The model is validated by comparison with experimental data of (i) GAG distribution and leakage, (ii) spatial‐temporal distributions of cells, and (iii) tissue growth reported in previous works. Validation of the model predictive capability—against a selection of measured data that were not used to construct the model—suggests that the model successfully describes the interplay of several simultaneous processes carried out during in vitro cartilage tissue regeneration and indicates that this approach could also be attractive for application in other tissue engineering systems. Biotechnol. Bioeng. 2010. 105: 842–853. © 2009 Wiley Periodicals, Inc.     

Cell–scaffold interaction within engineered tissue

The structure of a tissue engineering scaffold plays an important role in modulating tissue growth. A novel gelatin–chitosan (Gel–Cs) scaffold with a unique structure produced by three-dimensional printing (3DP) technology combining with vacuum freeze-drying has been developed for tissue-engineering applications. The scaffold composed of overall construction, micro-pore, surface morphology, and effective mechanical property. Such a structure meets the essential design criteria of an ideal engineered scaffold. The favorable cell–matrix interaction supports the active biocompatibility of the structure. The structure is capable of supporting cell attachment and proliferation. Cells seeded into this structure tend to maintain phenotypic shape and secreted large amounts of extracellular matrix (ECM) and the cell growth decreased the mechanical properties of scaffold. This novel biodegradable scaffold has potential applications for tissue engineering based upon its unique structure, which acts to support cell growth.     

Dental pulp stem cell-derived extracellular matrix: autologous tool boosting bone regeneration

Background aimsTo facilitate artificial bone construct integration into a patient's body, scaffolds are enriched with different biologically active molecules. Among various scaffold decoration techniques, coating surfaces with cell-derived extracellular matrix (ECM) is a rapidly growing field of research. In this study, for the first time, this technology was applied using primary dental pulp stem cells (DPSCs) and tested for use in artificial bone tissue construction.MethodsRat DPSCs were grown on three-dimensional-printed porous polylactic acid scaffolds for 7 days. After the predetermined time, samples were decellularized, and the remaining ECM detailed proteomic analysis was performed. Further, DPSC-secreated ECM impact to mesenchymal stromal cells (MSC) behaviour as well as its role in osteoregeneration induction were analysed.ResultsIt was identified that DPSC-specific ECM protein network ornamenting surface-enhanced MSC attachment, migration and proliferation and even promoted spontaneous stem cell osteogenesis. This protein network also demonstrated angiogenic properties and did not stimulate MSCs to secrete molecules associated with scaffold rejection. With regard to bone defects, DPSC-derived ECM recruited endogenous stem cells, initiating the bone self-healing process. Thus, the DPSC-secreted ECM network was able to significantly enhance artificial bone construct integration and induce successful tissue regeneration.ConclusionsDPSC-derived ECM can be a perfect tool for decoration of various biomaterials in the context of bone tissue engineering.     

A triphasic constrained mixture model of engineered tissue formation under in vitro dynamic mechanical conditioning

While it has become axiomatic that mechanical signals promote in vitro engineered tissue formation, the underlying mechanisms remain largely unknown. Moreover, efforts to date to determine parameters for optimal extracellular matrix (ECM) development have been largely empirical. In the present work, we propose a two-pronged approach involving novel theoretical developments coupled with key experimental data to develop better mechanistic understanding of growth and development of dense connective tissue under mechanical stimuli. To describe cellular proliferation and ECM synthesis that occur at rates of days to weeks, we employ mixture theory to model the construct constituents as a nutrient-cell-ECM triphasic system, their transport, and their biochemical reactions. Dynamic conditioning protocols with frequencies around 1 Hz are described with multi-scale methods to couple the dissimilar time scales. Enhancement of nutrient transport due to pore fluid advection is upscaled into the growth model, and the spatially dependent ECM distribution describes the evolving poroelastic characteristics of the scaffold-engineered tissue construct. Simulation results compared favorably to the existing experimental data, and most importantly, distinguish between static and dynamic conditioning regimes. The theoretical framework for mechanically conditioned tissue engineering (TE) permits not only the formulation of novel and better-informed mechanistic hypothesis describing the phenomena underlying TE growth and development, but also the exploration/optimization of conditioning protocols in a rational manner.     

Enhancement of cell growth in tissue-engineering constructs under direct perfusion: Modeling and simulation

Perfusion bioreactors improve mass transfer in cell-scaffold constructs. We developed a mathematical model to simulate nutrient flow through cellular constructs. Interactions among cell proliferation, nutrient consumption, and culture medium circulation were investigated. The model incorporated modified Contois cell-growth kinetics that includes effects of nutrient saturation and limited cell growth. Nutrient uptake was depicted through the Michaelis-Menton kinetics. To describe the culture medium convection, the fluid flow outside the cell-scaffold construct was described by the Navier-Stokes equations, while the fluid dynamics within the construct was modeled by Brinkman's equation for porous media flow. Effects of the media perfusion were examined by including time-dependant porosity and permeability changes due to cell growth. The overall cell volume was considered to consist of cells and extracellular matrices (ECM) as a whole without treating ECM separately. Numerical simulations show when cells were cultured subjected to direct perfusion, they penetrated to a greater extent into the scaffold and resulted in a more uniform spatial distribution. The cell amount was increased by perfusion and ultimately approached an asymptotic value as the perfusion rates increased in terms of the dimensionless Peclet number that accounts for the ratio of nutrient perfusion to diffusion. In addition to enhancing the nutrient delivery, perfusion simultaneously imposes flow-mediated shear stress to the engineered cells. Shear stresses were found to increase with cell growth as the scaffold void space was occupied by the cell and ECM volumes. The macro average stresses increased from 0.2 mPa to 1 mPa at a perfusion rate of 20 microm/s with the overall cell volume fraction growing from 0.4 to 0.7, which made the overall permeability value decrease from 1.35 x 10(-2)cm(2) to 5.51 x 10(-4)cm(2). Relating the simulation results with perfusion experiments in literature, the average shear stresses were below the critical value that would induce the chondrocyte necrosis.     

Engineering cartilage tissues with the shape of human nasal alar by using chondrocyte macroaggregate--Experiment study in rabbit model

Despite of progresses in tissue engineering based on cell/scaffold strategy, uneven cell distribution as well as tissue formation in the scaffold, limited cell seeding efficiency and inflammatory reaction triggered by the degradation of scaffold remain problems to be resolved. In this study, we proposed a novel cell-macroaggregate cultivation system, and explored a feasible strategy to construct three-dimensional cartilage tissue with shape of human nasal alar by using cell macroaggregate. Isolated chondrocytes was cultured at high density to form a monolayer chondrocyte sheet as well as expanded for seeding on the sheet to produce mechanically operable cell macroaggregate. Chondrocyte macroaggregates were then fabricated into transplants with shape of nasal alar by using Internal support or External scaffold techniques; results of in vivo chondrogenesis were investigated in immunocompetent animal. Chondrocyte macroaggregates presented long survival time and good viability; constructs fabricated using both techniques can develop into tissues with characteristic structure of native cartilage, glycosaminoglycans as well as type II collagen were highly produced in the ECM of engineered cartilages. By placing hyaluronan ester film as Internal support, the predetermined shape of the chondrocyte macroaggregate can be well maintained. In contrast, due to the poor mechanical stability of grafts fabricated in External scaffold group, obvious deformation occurred in harvested specimens. The experiment proved the usefulness of chondrocyte macroaggregate in cartilage regeneration, and provided a new strategy to engineer cartilage with special shape by using cell macroaggregate/biodegradable support.     

Encapsulating chondrocytes in degrading PEG hydrogels with high modulus: engineering gel structural changes to facilitate cartilaginous tissue production

A major challenge when designing cell scaffolds for chondrocyte delivery in vivo is creating scaffolds with sufficient mechanical properties to restore initial function while simultaneously controlling temporal changes in the gel structure to facilitate tissue formation. To address this design challenge, degradable photocrosslinked hydrogels based on poly(ethylene glycol) were investigated. To alter the gel's initial mechanical properties, hydrogels were fabricated by varying the initial macromer concentration from 10% to 15% to 20%. A twofold increase in macromer concentration resulted in an eightfold increase in the initial compressive modulus from 60 to 500 kPa. Gel degradation was tailored by incorporating fast-degrading crosslinks that enable maximal extracellular matrix (ECM) diffusion with time and a minimal number of nondegrading (or slowly degrading) crosslinks to maintain scaffold integrity and prevent complete gel erosion during tissue formation. Chondrocytes encapsulated in these gels produced cartilaginous tissue rich in glycosaminoglycans and collagen as seen biochemically and histologically. Interestingly, mass loss appeared to more closely match tissue secretion in gels fabricated from a 15% macromer concentration. However, the spatial ECM distribution was grossly similar in all three gels. By tailoring gel degradation and controlling network evolution during degradation, gels with optimal properties can be fabricated to support initially physiologic compressive loads while simultaneously supporting the formation of a neotissue.     

l-glutamine is a key parameter in the immunosuppression phenomenon

The present work describes the influence of both vitamin C (VC) and mechanical stimulation on development of the extracellular matrix (ECM) and improvement in mechanical properties of a chondrocyte-agarose construct in a regenerating tissue disease model of hyaline cartilage. We used primary bovine chondrocytes and two types of VC, ascorbic acid (AsA) as an acidic form and ascorbic acid 2-phosphate (A2P) as a non-acidic form, and applied uniaxial compressive strain to the tissue model using a purpose-built bioreactor. When added to the medium in free-swelling culture conditions, A2P downregulated development of ECM and suppressed improvement of the tangent modulus more than AsA. By contrast, application of mechanical stimulation to the construct both increased the tangent modulus more than the free-swelling group containing A2P and enhanced the ECM network of inner tissue to levels nearly as high as the free-swelling group containing AsA. Thus, mechanical stimulation and strain appears to enhance the supply of nutrients and improve the synthesis of ECM via mechanotransduction pathways of chondrocytes. Therefore, we suggest that mechanical stimulation is necessary for homogenous development of ECM in a cell-associated construct with a view to implantation of a large-sized articular cartilage defect.     

Cell–scaffold mechanical interplay within engineered tissue

Effective tissue engineering requires appropriate selection of cells and scaffold, where the latter serves as a mechanical and biological support for cell growth and functionality. The optimal combination of cell source and scaffold properties can vary for each desired application. Such preconditions necessitate enhanced understanding of the interactions between cells and scaffold within engineered tissue. Several studies have examined the deforming effects cells induce in scaffolds via exertion of contractile forces. In contrast, other studies focus on the scaffold's biochemical and mechanical properties and their effects on cell behavior.This review summarizes the mechanical interplay between cells and scaffold within engineered tissue. We present evidence for contractile forces exerted by cells on three-dimensional (3D) scaffolds and discuss existing methods for their quantification. In addition, we address some theories related to the effects of scaffold stiffness and mechanical stimulation on cell behavior. Further understanding of the reciprocal effects between cells and scaffold will provide both enhanced knowledge regarding the expected properties of engineered tissue and more competent tissue regeneration techniques.     

Dynamic compressive loading of image-guided tissue engineered meniscal constructs

This study investigated the hypothesis that dynamic compression loading enhances tissue formation and increases mechanical properties of anatomically shaped tissue engineered menisci. Bovine meniscal fibrochondrocytes were seeded in 2%w/v alginate, crosslinked with CaSO(4), injected into μCT based molds, and post crosslinked with CaCl(2). Samples were loaded via a custom bioreactor with loading platens specifically designed to load anatomically shaped constructs in unconfined compression. Based on the results of finite element simulations, constructs were loaded under sinusoidal displacement to yield physiological strain levels. Constructs were loaded 3 times a week for 1 h followed by 1 h of rest and loaded again for 1 h. Constructs were dynamically loaded for up to 6 weeks. After 2 weeks of culture, loaded samples had 2-3.2 fold increases in the extracellular matrix (ECM) content and 1.8-2.5 fold increases in the compressive modulus compared with static controls. After 6 weeks of loading, glycosaminoglycan (GAG) content and compressive modulus both decreased compared with 2 week cultures by 2.3-2.7 and 1.5-1.7 fold, respectively, whereas collagen content increased by 1.8-2.2 fold. Prolonged loading of engineered constructs could have altered alginate scaffold degradation rate and/or initiated a catabolic cellular response, indicated by significantly decreased ECM retention at 6 weeks compared with 2 weeks. However, the data indicates that dynamic loading had a strikingly positive effect on ECM accumulation and mechanical properties in short term culture.     

A multifunctional 3D co-culture system for studies of mammary tissue morphogenesis and stem cell biology

Studies on the stem cell niche and the efficacy of cancer therapeutics require complex multicellular structures and interactions between different cell types and extracellular matrix (ECM) in three dimensional (3D) space. We have engineered a 3D in vitro model of mammary gland that encompasses a defined, porous collagen/hyaluronic acid (HA) scaffold forming a physiologically relevant foundation for epithelial and adipocyte co-culture. Polarized ductal and acinar structures form within this scaffold recapitulating normal tissue morphology in the absence of reconstituted basement membrane (rBM) hydrogel. Furthermore, organoid developmental outcome can be controlled by the ratio of collagen to HA, with a higher HA concentration favouring acinar morphological development. Importantly, this culture system recapitulates the stem cell niche as primary mammary stem cells form complex organoids, emphasising the utility of this approach for developmental and tumorigenic studies using genetically altered animals or human biopsy material, and for screening cancer therapeutics for personalised medicine.     

Engineered Smooth Muscle Tissues: Regulating Cell Phenotype with the Scaffold

Culturing cells on three-dimensional, biodegradable scaffolds may create tissues suitable either for reconstructive surgery applications or as novel in vitro model systems. In this study, we have tested the hypothesis that the phenotype of smooth muscle cells (SMCs) in three-dimensional, engineered tissues is regulated by the chemistry of the scaffold material. Specifically, we have directly compared cell growth and patterns of extracellular matrix (ECM) (e.g. , elastin and collagen) gene expression on two types of synthetic polymer scaffolds and type I collagen scaffolds. The growth rates of SMCs on the synthetic polymer scaffolds were significantly higher than on type I collagen sponges. The rate of elastin production by SMCs on polyglycolic acid (PGA) scaffolds was 3.5 +/- 1.1-fold higher than that on type I collagen sponges on Day 11 of culture. In contrast, the collagen production rate on type I collagen sponges was 3.3 +/- 1.1-fold higher than that on PGA scaffolds. This scaffold-dependent switching between elastin and collagen gene expression was confirmed by Northern blot analysis. The finding that the scaffold chemistry regulates the phenotype of SMCs independent of the scaffold physical form was confirmed by culturing SMCs on two-dimensional films of the scaffold materials. It is likely that cells adhere to these scaffolds via different ligands, as the major protein adsorbed from the serum onto synthetic polymers was vitronectin, whereas fibronectin and vitronectin were present at high density on type I collagen sponges. In summary, this study demonstrates that three-dimensional smooth muscle-like tissues can be created by culturing SMCs on three-dimensional scaffolds, and that the phenotype of the SMCs is strongly regulated by the scaffold chemistry. These engineered tissues provide novel, three-dimensional models to study cellular interaction with ECM in vitro.     

In silico multi‐scale model of transport and dynamic seeding in a bone tissue engineering perfusion bioreactor

Computer simulations can potentially be used to design, predict, and inform properties for tissue engineering perfusion bioreactors. In this work, we investigate the flow properties that result from a particular poly‐L ‐lactide porous scaffold and a particular choice of perfusion bioreactor vessel design used in bone tissue engineering. We also propose a model to investigate the dynamic seeding properties such as the homogeneity (or lack of) of the cellular distribution within the scaffold of the perfusion bioreactor: a pre‐requisite for the subsequent successful uniform growth of a viable bone tissue engineered construct. Flows inside geometrically complex scaffolds have been investigated previously and results shown at these pore scales. Here, it is our aim to show accurately that through the use of modern high performance computers that the bioreactor device scale that encloses a scaffold can affect the flows and stresses within the pores throughout the scaffold which has implications for bioreactor design, control, and use. Central to this work is that the boundary conditions are derived from micro computed tomography scans of both a device chamber and scaffold in order to avoid generalizations and uncertainties. Dynamic seeding methods have also been shown to provide certain advantages over static seeding methods. We propose here a novel coupled model for dynamic seeding accounting for flow, species mass transport and cell advection‐diffusion‐attachment tuned for bone tissue engineering. The model highlights the timescale differences between different species suggesting that traditional homogeneous porous flow models of transport must be applied with caution to perfusion bioreactors. Our in silico data illustrate the extent to which these experiments have the potential to contribute to future design and development of large‐scale bioreactors. Biotechnol. Bioeng. 2013; 110: 1221–1230. © 2012 Wiley Periodicals, Inc.     

组织工程皮肤生物反应器的研究与设计

目的设计一套生物反应器,能针对不同支架材料———细胞复合物进行构建组织工程皮肤。方法根据皮肤的自身生长特点和不同支架材料-细胞复合物的特性,模拟皮肤的生长环境和力学环境,通过生物反应器解决组织工程皮肤构建中支架的装夹和气液界面问题。结果生物反应器由控制系统和生物反应器主体两部分构成,能提供对多种皮肤细胞复合物的动态培养。结论皮肤生物反应器能够满足不同组织工程皮肤产品的需要。能够形成气液界面和模拟生物力学的刺激。