Recellularization of decellularized human adipose-tissue-derived extracellular matrix sheets with other human cell types

Decellularized human extracellular matrices (ECMs) are an extremely appealing biomaterial for tissue engineering and regenerative medicine. In this study, we decellularized human adipose tissue, fabricated a thin ECM sheet and explored the potential of this human adipose-derived ECM sheet as a substrate to support the formation of tissues other than adipose tissue. Acellular ECM sheets were fabricated from human adipose tissue through successive physical and chemical treatments: homogenization, centrifugation, casting, freeze-drying and sodium dodecyl sulfate treatment. The ECM sheets exhibited good mechanical properties, despite their porous structure. They degraded quickly in the presence of collagenase and the degradation rate increased with the collagenase concentration in phosphate-buffered saline. Five different human cell types, covering a broad range of cells and applications (normal human dermal fibroblasts, human aortic smooth muscle cells, human chondrocytes, human umbilical vein endothelial cells and human adipose-derived stem cells), were seeded onto the ECM sheets. All the human cell types spread well, proliferated and were successfully integrated into the decellularized ECM sheet. Overall, the results suggest that recellularized ECM sheets are a promising substitute for defective or damaged human tissues.     

Decellularized omentum as novel biologic scaffold for reconstructive surgery and regenerative medicine

Homologous tissues, such as adipose tissue, may be an interesting source of acellular scaffolds, maintaining a complex physiological three-dimensional (3D) structure, to be recellularized with autologous cells. The aim of the present work is to evaluate the possibility of obtaining homologous acellular scaffolds from decellularization of the omentum, which is known to have a complex vascular network. Adult rat and human omenta were treated with an adapted decellularization protocol involving mechanical rupture (freeze-thaw cycles), enzymatic digestion (trypsin, lipase, deoxyribonuclease, ribonuclease) and lipid extraction (2-propanol). Histological staining confirmed the effectiveness of decellularization, resulting in cell-free scaffolds with no residual cells in the matrix. The complex 3D networks of collagen (azan-Mallory), elastic fibers (Van Gieson), reticular fibers and glycosaminoglycans (PAS) were maintained, whereas Oil Red and Sudan stains showed the loss of lipids in the decellularized tissue. The vascular structures in the tissue were still visible, with preservation of collagen and elastic wall components and loss of endothelial (anti-CD31 and -CD34 immunohistochemistry) and smooth muscle (anti-alpha smooth muscle actin) cells. Fat-rich and well vascularized omental tissue may be decellularized to obtain complex 3D scaffolds preserving tissue architecture potentially suitable for recellularization. Further analyses are necessary to verify the possibility of recolonization of the scaffold by adipose-derived stem cells in vitro and then in vivo after re implantation, as already known for homologus implants in regenerative processes.Key words: omentum, scaffold, decellularization, adipose tissue engineering, regenerative medicine, microvascularization     

Optimized decellularization protocol including α-Gal epitope reduction for fabrication of an acellular porcine annulus fibrosus scaffold

Recent advances in tissue engineering have led to potential new strategies, especially decellularization protocols from natural tissues, for the repair, replacement, and regeneration of intervertebral discs. This study aimed to validate our previously reported method for the decellularization of annulus fibrosus (AF) tissue and to quantify potentially antigenic α-Gal epitopes in the decellularized tissue. Porcine AF tissue was decellularized using different freeze–thaw temperatures, chemical detergents, and incubation times in order to determine the optimal method for cell removal. The integrity of the decellularized material was determined using biochemical and mechanical tests. The α-Gal epitope was quantified before and after decellularization. Decellularization with freeze–thaw in liquid nitrogen, an ionic detergent (0.1% SDS), and a 24 h incubation period yielded the greatest retention of GAG and collagen relative to DNA reduction when tested as single variables. Combined, these optimal decellularization conditions preserved more GAG while removing the same amount of DNA as the conditions used in our previous study. Components and biomechanical properties of the AF matrix were retained. The decellularized AF scaffold exhibited suitable immune-compatibility, as evidenced by successful in vivo remodeling and a decrease in the α-Gal epitope. Our study defined the optimal conditions for decellularization of porcine AF tissues while preserving the biological composition and mechanical properties of the scaffold. Under these conditions, immunocompatibility was evidenced by successful in vivo remodeling and reduction of the α-Gal epitope in the decellularized material. Decellularized AF scaffolds are potential candidates for clinical applications in spinal surgery.     

Region-specific constitutive modeling of the plantar soft tissue

Recent research has shown that hyperelastic properties of the plantar soft tissue consisting of adipose tissue and fibrous septa change from region to region. However, relatively little research has been conducted to develop analytical or computational models to describe the region-specific behavior of the plantar soft tissue. The objective of the research is to develop a region-specific constitutive model of the plantar soft tissue. Plantar soft tissue specimens were dissected from six regions [subcalcaneal (CA), sublateral (LA), subnavicular (Nav), 1st, 3rd, and 5th submetatarsal (M1, M3, M5)] from cadaveric foot samples, and a picrosirius red staining technique was used to visualize the collagen fibers in fibrous septa. The volume fractions of adipose tissue and fibrous septa and the volume fractions of the principal orientations of the fibrous septa were calculated with the intensity gradient method. Region-specific constitutive models were then developed in finite element analysis considering the microstructure of the plantar soft tissue. The hyperelastic region specific material properties of the plantar soft tissue were validated with experimental unconfined compression tests and indentation tests from the literature. The results show that the models give reasonable predictions of the stiffness of the soft tissue within a standard deviation of the tests. The region-specific constitutive models help to explain how changes in the constituents are related to mechanical behavior of the soft tissue on a region specific basis.     

Method for characterizing viscoelasticity of human gluteal tissue

Characterizing compressive transient large deformation properties of biological tissue is becoming increasingly important in impact biomechanics and rehabilitation engineering, which includes devices interfacing with the human body and virtual surgical guidance simulation. Individual mechanical in vivo behaviour, specifically of human gluteal adipose and passive skeletal muscle tissue compressed with finite strain, has, however, been sparsely characterised. Employing a combined experimental and numerical approach, a method is presented to investigate the time-dependent properties of in vivo gluteal adipose and passive skeletal muscle tissue. Specifically, displacement-controlled ramp-and-hold indentation relaxation tests were performed and documented with magnetic resonance imaging. A time domain quasi-linear viscoelasticity (QLV) formulation with Prony series valid for finite strains was used in conjunction with a hyperelastic model formulation for soft tissue constitutive model parameter identification and calibration of the relaxation test data. A finite element model of the indentation region was employed. Strong non-linear elastic but linear viscoelastic tissue material behaviour at finite strains was apparent for both adipose and passive skeletal muscle mechanical properties with orthogonal skin and transversal muscle fibre loading. Using a force-equilibrium assumption, the employed material model was well suited to fit the experimental data and derive viscoelastic model parameters by inverse finite element parameter estimation. An individual characterisation of in vivo gluteal adipose and muscle tissue could thus be established. Initial shear moduli were calculated from the long-term parameters for human gluteal skin/fat: G(∞,S/F)=1850 Pa and for cross-fibre gluteal muscle tissue: G(∞,M)=881 Pa. Instantaneous shear moduli were found at the employed ramp speed: G(0,S/F)=1920 Pa and G(0,M)=1032 Pa.     

Preparation and evaluation of decellularized porcine carotid arteries cross-linked by genipin: the preliminary results

Decellularized arteries have been considered as promising scaffolds for small-diameter vascular substitutes. However, weakened mechanical properties, immunological rejection and rapid degradation after transplantation still exist after decellularization. Previous studies indicated that genipin cross-linking can solve these problems. Therefore, genipin was selected as the cross-linking agent for the pre-treatment of decellularized arteries in our study. Histological analysis, scanning electron microscopy, mechanical properties analysis and subcutaneous embedding experiment were adopted to investigate the efficiency of decellularization and the effect of genipin cross-linking on improving mechanical, structural and biological properties of decellularized arteries. Decellularization protocols based on three trypsin concentrations were used to prepare decellularized arteries, after decellularization, arteries were cross-linked with genipin. Results showed that 0.5% trypsin was the most efficient concentration to remove cellular components and preserve ECM. However, mechanical properties of 0.5% trypsin decellularized arteries weakened significantly, while genipin cross-linking improved mechanical properties of decellularized arteries to the same level as fresh arteries. After 4 weeks subcutaneous embedding, cross-linked arteries caused the mildest inflammatory response. In conclusion, genipin could be employed as an ideal cross-linking agent to strengthen mechanical properties, enhance the resistance to degradation and reduce the antigenicity of decellularized arteries for small-diameter blood vessel tissue engineering applications.     

Scale-dependent mechanical properties of native and decellularized liver tissue

Decellularization, a technique used in liver regenerative medicine, is the removal of all the cellular components from a tissue or organ, leaving behind an intact structure of extracellular matrix. The biomechanical properties of this novel scaffold material are currently unknown and are important due to the mechanosensitivity of liver cells. Characterizing this material is important for bioengineering liver tissue from this decellularized scaffold as well as creating new 3-dimensional mimetic structures of liver extracellular matrix. This study set out to characterize the biomechanical properties of perfused liver tissue in its native and decellularized states on both a macro- and nano-scale. Poroviscoelastic finite element models were then used to extract the fluid and solid mechanical properties from the experimental data. Tissue-level spherical indentation-relaxation tests were performed on 5 native livers and 8 decellularized livers at two indentation rates and at multiple perfusion rates. Cellular-level spherical nanoindentation was performed on 2 native livers and 1 decellularized liver. Tissue-level results found native liver tissue to possess a long-term Young’s modulus of 10.5 kPa and decellularized tissue a modulus of 1.18 kPa. Cellular-level testing found native tissue to have a long-term Young’s modulus of 4.40 kPa and decellularized tissue to have a modulus of 0.91 kPa. These results are important for regenerative medicine and tissue engineering where cellular response is dependent on the mechanical properties of the engineered scaffold.     

Lung tissue engineering and preservation of alveolar microstructure using a novel casting method

Abstract

We used a rat model to decellularize and seed alveolar cells on a three-dimensional lung scaffold to preserve alveolar microarchitecture. We verified the preservation of terminal respiratory structure by casting and by scanning electron microscopy (SEM) of the casts after decellularization. Whole lungs were obtained from 12 healthy Sprague-Dawley rats, cannulated through the trachea under sterile conditions, and decellularized using a detergent-based method. Casting of both natural and decellularized lungs was performed to verify preservation of the inner microstructure of scaffolds for further cell seeding. Alveolar cell seeding was performed using green fluorescent protein (GFP) lung cells and non-GFP lung cells, and a peristaltic pump. We assessed cell seeding using histological and immunohistochemical staining, and enzymatic evaluation. All cellular components were removed completely from the scaffolds, and histological staining and SEM of casts were used to verify the preservation of tissue structure. Tensile tests verified conservation of biomechanical properties. The hydroxyproline content of decellularized lungs was similar to native lung. Histological and immunohistochemical evaluations showed effective cell seeding on decellularized matrices. Enzymatic measurement of trypsin and alpha 1 antitrypsin suggested the potential functional properties of the regenerated lungs. Casts produced by our method have satisfactory geometrical properties for further cell seeding of lung scaffolds. Preservation of micro-architecture and terminal alveoli that was confirmed by SEM of lung casts increases the probability of an effective cell seeding process.     

Culture on Tissue‐Specific Coatings Derived from α‐Amylase‐Digested Decellularized Adipose Tissue Enhances the Proliferation and Adipogenic Differentiation of Human Adipose‐Derived Stromal Cells

While extracellular matrix (ECM)‐derived coatings have the potential to direct the response of cell populations in culture, there is a need to investigate the effects of ECM sourcing and processing on substrate bioactivity. To develop improved cell culture models for studying adipogenesis, the current study examines the proliferation and adipogenic differentiation of human adipose‐derived stem/stromal cells (ASCs) on a range of ECM‐derived coatings. Human decellularized adipose tissue (DAT) and commercially available bovine tendon collagen (COL) are digested with α‐amylase or pepsin to prepare the coatings. Physical characterization demonstrates that α‐amylase digestion generates softer, thicker, and more stable coatings, with a fibrous tissue‐like ultrastructure that is lost in the pepsin‐digested thin films. ASCs cultured on the α‐amylase‐digested ECM have a more spindle‐shaped morphology, and proliferation is significantly enhanced on the α‐amylase‐digested DAT coatings. Further, the α‐amylase‐digested DAT provides a more pro‐adipogenic microenvironment, based on higher levels of adipogenic gene expression, glycerol‐3‐phosphate dehydrogenase (GPDH) enzyme activity, and perilipin staining. Overall, this study supports α‐amylase digestion as a new approach for generating bioactive ECM‐derived coatings, and demonstrates tissue‐specific bioactivity using adipose‐derived ECM to enhance ASC proliferation and adipogenic differentiation.     

Engineering of vascularized adipose constructs

Adipose tissue engineering offers a promising alternative to the current surgical techniques for the treatment of soft tissue defects. It is a challenge to find the appropriate scaffold that not only represents a suitable environment for cells but also allows fabrication of customized tissue constructs, particularly in breast surgery. We investigated two different scaffolds for their potential use in adipose tissue regeneration. Sponge-like polyurethane scaffolds were prepared by mold casting with methylal as foaming agent, whereas polycaprolactone scaffolds with highly regular stacked-fiber architecture were fabricated with fused deposition modeling. Both scaffold types were seeded with human adipose tissue-derived precursor cells, cultured and implanted in nude mice using a femoral arteriovenous flow-through vessel loop for angiogenesis. In vitro, cells attached to both scaffolds and differentiated into adipocytes. In vivo, angiogenesis and adipose tissue formation were observed throughout both constructs after 2 and 4?weeks, with angiogenesis being comparable in seeded and unseeded constructs. Fibrous tissue formation and adipogenesis were more pronounced on polyurethane foam scaffolds than on polycaprolactone prototyped scaffolds. In conclusion, both scaffold designs can be effectively used for adipose tissue engineering.     

Morphofunctional characterization of decellularized vena cava as tissue engineering scaffolds

Clinical experience for peripheral arterial disease treatment shows poor results when synthetic grafts are used to approach infrapopliteal arterial segments. However, tissue engineering may be an option to yield surrogate biocompatible neovessels. Thus, biological decellularized scaffolds could provide natural tissue architecture to use in tissue engineering, when the absence of ideal autologous veins reduces surgical options. The goal of this study was to evaluate different chemical induced decellularization protocols of the inferior vena cava of rabbits. They were decellularized with Triton X100 (TX100), sodium dodecyl sulfate (SDS) or sodium deoxycholate (DS). Afterwards, we assessed the remaining extracellular matrix (ECM) integrity, residual toxicity and the biomechanical resistance of the scaffolds. Our results showed that TX100 was not effective to remove the cells, while protocols using SDS 1% for 2 h and DS 2% for 1 h, efficiently removed the cells and were better characterized. These scaffolds preserved the original organization of ECM. In addition, the residual toxicity assessment did not reveal statistically significant changes while decellularized scaffolds retained the equivalent biomechanical properties when compared with the control. Our results concluded that protocols using SDS and DS were effective at obtaining decellularized scaffolds, which may be useful for blood vessel tissue engineering.     

Natural Scaffolds for Renal Differentiation of Human Embryonic Stem Cells for Kidney Tissue Engineering

Despite the enthusiasm for bioengineering of functional renal tissues for transplantation, many obstacles remain before the potential of this technology can be realized in a clinical setting. Viable tissue engineering strategies for the kidney require identification of the necessary cell populations, efficient scaffolds, and the 3D culture conditions to develop and support the unique architecture and physiological function of this vital organ. Our studies have previously demonstrated that decellularized sections of rhesus monkey kidneys of all age groups provide a natural extracellular matrix (ECM) with sufficient structural properties with spatial and organizational influences on human embryonic stem cell (hESC) migration and differentiation. To further explore the use of decellularized natural kidney scaffolds for renal tissue engineering, pluripotent hESC were seeded in whole- or on sections of kidney ECM and cell migration and phenotype compared with the established differentiation assays for hESC. Results of qPCR and immunohistochemical analyses demonstrated upregulation of renal lineage markers when hESC were cultured in decellularized scaffolds without cytokine or growth factor stimulation, suggesting a role for the ECM in directing renal lineage differentiation. hESC were also differentiated with growth factors and compared when seeded on renal ECM or a new biologically inert polysaccharide scaffold for further maturation. Renal lineage markers were progressively upregulated over time on both scaffolds and hESC were shown to express signature genes of renal progenitor, proximal tubule, endothelial, and collecting duct populations. These findings suggest that natural scaffolds enhance expression of renal lineage markers particularly when compared to embryoid body culture. The results of these studies show the capabilities of a novel polysaccharide scaffold to aid in defining a protocol for renal progenitor differentiation from hESC, and advance the promise of tissue engineering as a source of functional kidney tissue.     

The Acinar Cage: Basement Membranes Determine Molecule Exchange and Mechanical Stability of Human Breast Cell Acini

The biophysical properties of the basement membrane that surrounds human breast glands are poorly understood, but are thought to be decisive for normal organ function and malignancy. Here, we characterize the breast gland basement membrane with a focus on molecule permeation and mechanical stability, both crucial for organ function. We used well-established and nature-mimicking MCF10A acini as 3D cell model for human breast glands, with ether low- or highly-developed basement membrane scaffolds. Semi-quantitative dextran tracer (3 to 40 kDa) experiments allowed us to investigate the basement membrane scaffold as a molecule diffusion barrier in human breast acini in vitro. We demonstrated that molecule permeation correlated positively with macromolecule size and intriguingly also with basement membrane development state, revealing a pore size of at least 9 nm. Notably, an intact collagen IV mesh proved to be essential for this permeation function. Furthermore, we performed ultra-sensitive atomic force microscopy to quantify the response of native breast acini and of decellularized basement membrane shells against mechanical indentation. We found a clear correlation between increasing acinar force resistance and basement membrane formation stage. Most important native acini with highly-developed basement membranes as well as cell-free basement membrane shells could both withstand physiologically relevant loads (≤ 20 nN) without loss of structural integrity. In contrast, low-developed basement membranes were significantly softer and more fragile. In conclusion, our study emphasizes the key role of the basement membrane as conductor of acinar molecule influx and mechanical stability of human breast glands, which are fundamental for normal organ function.     

Mechanical properties of anterior malleolar ligament from experimental measurement and material modeling analysis

In this paper, mechanical properties of the anterior malleolar ligament (AML) of human middle ear were studied through the uniaxial tensile, stress relaxation and failure tests. The digital image correlation (DIC) method was used to assess the boundary effect in experiments and calculate the strain on specimens. The constitutive behavior of the AML was described by a transversely isotropic hyperelastic model which consists of a first-order Ogden model augmented by a I (4)-type reinforcing term. The material parameters of the model were estimated and the viscoelasticity of the AML was illustrated by hysteresis phenomena and stress relaxation function. The mechanical strength of the AML was obtained through the failure test and the mean ultimate stress and stretch ratio were measured as 1.05 MPa and 1.51, respectively. Finally, a linear Young's modulus-stress relationship of the AML was derived based on constitutive equation of the AML within a stress range of 0-0.5 MPa.     

Modeling material-degradation-induced elastic property of tissue engineering scaffolds

The mechanical properties of tissue engineering scaffolds play a critical role in the success of repairing damaged tissues/organs. Determining the mechanical properties has proven to be a challenging task as these properties are not constant but depend upon time as the scaffold degrades. In this study, the modeling of the time-dependent mechanical properties of a scaffold is performed based on the concept of finite element model updating. This modeling approach contains three steps: (1) development of a finite element model for the effective mechanical properties of the scaffold, (2) parametrizing the finite element model by selecting parameters associated with the scaffold microstructure and/or material properties, which vary with scaffold degradation, and (3) identifying selected parameters as functions of time based on measurements from the tests on the scaffold mechanical properties as they degrade. To validate the developed model, scaffolds were made from the biocompatible polymer polycaprolactone (PCL) mixed with hydroxylapatite (HA) nanoparticles and their mechanical properties were examined in terms of the Young modulus. Based on the bulk degradation exhibited by the PCL/HA scaffold, the molecular weight was selected for model updating. With the identified molecular weight, the finite element model developed was effective for predicting the time-dependent mechanical properties of PCL/HA scaffolds during degradation.     

Digital image correlation and finite element modelling as a method to determine mechanical properties of human soft tissue in vivo

The mechanical properties of human soft tissue are crucial for impact biomechanics, rehabilitation engineering, and surgical simulation. Validation of these constitutive models using human data remains challenging and often requires the use of non-invasive imaging and inverse finite element (FE) analysis. Post-processing data from imaging methods such as tagged magnetic resonance imaging (MRI) can be challenging. Digital image correlation (DIC), however, is a relatively straightforward imaging method. DIC has been used in the past to study the planar and superficial properties of soft tissue and excised soft tissue layers. However, DIC has not been used to non-invasive study of the bulk properties of human soft tissue in vivo. Thus, the goal of this study was to assess the use of DIC in combination with FE modelling to determine the bulk material properties of human soft tissue. Indentation experiments were performed on a silicone gel soft tissue phantom. A two camera DIC setup was then used to record the 3D surface deformation. The experiment was then simulated using a FE model. The gel was modelled as Neo-Hookean hyperelastic, and the material parameters were determined by minimising the error between the experimental and FE data. The iterative FE analysis determined material parameters (μ=1.80 kPa, K=2999 kPa) that were in close agreement with parameters derived independently from regression to uniaxial compression tests (μ=1.71 kPa, K=2857 kPa). Furthermore the FE model was capable of reproducing the experimental indentor force as well as the surface deformation found (R²=0.81). It was therefore concluded that a two camera DIC configuration combined with FE modelling can be used to determine the bulk mechanical properties of materials that can be represented using hyperelastic Neo-Hookean constitutive laws.     

Validation of endogenous control genes in human adipose tissue: relevance to obesity and obesity-associated type 2 diabetes mellitus.

The aim of the present study was to test the influence of obesity and the presence of type 2 diabetes mellitus (T2DM) on the expression of ten housekeeping genes and of the 18S rRNA in a group of human adipose tissue samples from the omental and subcutaneous depot. Adipose tissue biopsies were obtained by laparoscopic surgery from lean and obese patients. After the extraction, mRNA levels in adipose tissue samples were quantified by real-time PCR using the commercial HUMAN ENDOGENOUS CONTROL PLATES. From the genes analyzed, 18S rRNA exhibited the most stable expression levels in both depots regardless of the pathophysiological conditions of obesity and obesity-associated T2DM. Contrarily, GAPD was the gene with the highest variation in its expression levels, being upregulated (8.0-fold) in the obese group and downregulated (3.5-fold) in obesity-associated T2DM. Our results show that 18S rRNA may be the most suitable gene for normalization in expression studies performed in human adipose tissue samples obtained from patients suffering from obesity and/or obesity-associated T2DM, whereas GAPD is less appropriate for comparison purposes under these circumstances.     

Human breast cancer decellularized scaffolds promote epithelial-to-mesenchymal transitions and stemness of breast cancer cells in vitro

Breast cancer, with unsatisfactory survival rates, is the leading cause of cancer-related death in women worldwide. Recent advances in the genetic basis of breast cancer have benefitted the development of gene-based medicines and therapies. Tissue engineering technologies, including tissue decellularizations and reconstructions, are potential therapeutic alternatives for cancer research and tissue regeneration. In our study, human breast cancer biopsies were decellularized by a detergent technique, with sodium lauryl ether sulfate (SLES) solution, for the first time. And the decellularization process was optimized to maximally maintain tissue microarchitectures and extracellular matrix (ECM) components with minimal DNA compounds preserved. Histology analysis and DNA quantification results confirmed the decellularization effect with maximal genetic compounds removal. Quantification, immunofluorescence, and histology analyses demonstrated better preservation of ECM components in 0.5% SLES-treated scaffolds. Scaffolds seeded with MCF-7 cells demonstrated the process of cell recellularization in vitro, with increased cell migration, proliferation, and epithelial-to-mesenchymal transition (EMT) process. When treated with 5-fluorouracil, the expressions of stem cell markers, including Oct4, Sox2, and CD49F, were maximally maintained in the recellularized scaffold with decreased apoptosis rates compared with monolayer cells. These results showed that the decellularized breast scaffold model with SLES treatments would help to simulate the pathogenesis of breast cancer in vitro. And we hope that this model could further accelerate the development of effective therapies for breast cancer and benefit drug screenings.     

Characterization of Mechanical and Biological Properties of 3-D Scaffolds Reinforced with Zinc Oxide for Bone Tissue Engineering

A scaffold for bone tissue engineering should have highly interconnected porous structure, appropriate mechanical and biological properties. In this work, we fabricated well-interconnected porous β-tricalcium phosphate (β-TCP) scaffolds via selective laser sintering (SLS). We found that the mechanical and biological properties of the scaffolds were improved by doping of zinc oxide (ZnO). Our data showed that the fracture toughness increased from 1.09 to 1.40 MPam^1/2, and the compressive strength increased from 3.01 to 17.89 MPa when the content of ZnO increased from 0 to 2.5 wt%. It is hypothesized that the increase of ZnO would lead to a reduction in grain size and an increase in density of the strut. However, the fracture toughness and compressive strength decreased with further increasing of ZnO content, which may be due to the sharp increase in grain size. The biocompatibility of the scaffolds was investigated by analyzing the adhesion and the morphology of human osteoblast-like MG-63 cells cultured on the surfaces of the scaffolds. The scaffolds exhibited better and better ability to support cell attachment and proliferation when the content of ZnO increased from 0 to 2.5 wt%. Moreover, a bone like apatite layer formed on the surfaces of the scaffolds after incubation in simulated body fluid (SBF), indicating an ability of osteoinduction and osteoconduction. In summary, interconnected porous β-TCP scaffolds doped with ZnO were successfully fabricated and revealed good mechanical and biological properties, which may be used for bone repair and replacement potentially.