Biomanufacturing and Rapid Forming Technology Key Laboratory of Beijing

Beijing, China

Biomanufacturing and Rapid Forming Technology Key Laboratory of Beijing

Beijing, China
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Zhang T.,Tsinghua University | Zhang T.,Biomanufacturing and Rapid Forming Technology Key Laboratory of Beijing | Zhang T.,Innovation International | Zhang H.,Tsinghua University | And 10 more authors.
Biofabrication | Year: 2017

Integrative osteochondral repair is a useful strategy for cartilage-defect repair. To mimic the microenvironment, it is necessary that scaffolds effectively mimic the extracellular matrix of natural cartilage and subchondral bone. In this study, biomimetic osteochondral scaffolds containing an oriented cartilage layer, a compact layer, and a three-dimensional (3D)-printed core-sheath structured-bone layer were developed. The oriented cartilage layer was designed to mimic the structural and material characteristics of native cartilage tissue and was fabricated with cartilage matrix-chitosan materials, using thermal-induced phase-separation technology. The 3D-printed core-sheath structured-bone layer was fabricated with poly(L-lactide-co-glycolide)/β-tricalcium phosphate-collagen materials by low-temperature deposition technology, using a specially designed core-sheath nozzle, and was designed to mimic the mechanical characteristics of subchondral bone and improve scaffold hydrophilicity. The compact layer was designed to mimic the calcified-layer structure of natural cartilage to ensure the presence of different suitable microenvironments for the regeneration of bone and cartilage. A dissolving-bonding process was developed to effectively combine the three parts together, after which the bone and cartilage scaffolds exhibited good mechanical properties and hydrophilicity. Additionally, goat autologous bone mesenchymal stem cells (BMSCs) were isolated and then seeded into the bone and cartilage layers, respectively, and following a 1 week culture in vitro, the BMSC-scaffold constructs were implanted into a goat articular-defect model. Our results indicated that the scaffolds exhibited good biocompatibility, and 24 weeks after implantation, the femoral condyle surface was relatively flat and consisted of a large quantity of hyaloid cartilage. Furthermore, histological staining revealed regenerated trabecular bone formed in the subchondral bone-defect area. These results provided a new method to fabricate biomimetic osteochondral scaffolds and demonstrated their effectiveness for future clinical applications in cartilage-defect repair. © 2017 IOP Publishing Ltd.

Mao X.,Tsinghua University | Mao X.,Biomanufacturing and Rapid Forming Technology Key Laboratory of Beijing | Zhang L.,Tsinghua University | Zhang L.,Biomanufacturing and Rapid Forming Technology Key Laboratory of Beijing | And 4 more authors.
Physics of Fluids | Year: 2017

Droplet formation via the oscillations of a tapered capillary tube is experimentally and numerically investigated using incompressible, low-viscosity Newtonian liquids. As in many other common methods of droplet generation, this technique features a transient flow that is directed out of a nozzle. However, due to the interactions of the oscillations, the tube, and the fluids, the flow rate upstream of the nozzle cannot be directly obtained. In this study, the motion of the tube is measured under the activation of a specific waveform, and the flow inside the tube and drop formation are further numerically studied using a non-inertial reference system in which the tube is stationary. The mechanism of ejection is quantitatively explained by analyzing the temporal variation in the velocity and pressure distributions inside the tube. The dynamics of drop formation, the drop velocity, and the drop radius are studied as functions of the dimensionless groups that govern the problem, including the Ohnesorge number Oh, the Weber number We, the gravitational Bond number G, and various length scale ratios. The results show that droplets are generated due to the inertia of the liquid and velocity amplification in the tapered section. By influencing the balance between the viscous effect and inertial effect of the liquid along the entire tube, the length scale ratios affect the evolution of the transient flow at the nozzle and eventually influence the drop radius and velocity. For liquids with viscosities close to that of pure water, the critical Reynolds number, at which a drop can be generated, linearly depends on the Z number (the reciprocal of Oh) at the nozzle. © 2017 Author(s).

Zhao L.,Tsinghua University | Zhao L.,Biomanufacturing and Rapid Forming Technology Key Laboratory of Beijing | Zhao L.,Key Laboratory for Advanced Materials Processing Technology | Yan K.C.,The College of New Jersey | And 10 more authors.
Journal of Manufacturing Science and Engineering, Transactions of the ASME | Year: 2015

Drop-on-demand (DOD) microdroplet formation and deposition play an important role in additive manufacturing, particularly in printing of three-dimensional (3D) in vitro biological models for pharmacological and pathological studies, for tissue engineering and regenerative medicine applications, and for building of cell-integrated microfluidic devices. In development of a DOD based microdroplet deposition process for 3D cell printing, the droplet formation, controlled on-demand deposition and at the single-cell level, and most importantly, maintaining the viability and functionality of the cells during and after the printing are all remaining to be challenged. This report presents our recent study on developing a novel DOD based microdroplet deposition process for 3D printing by utilization of an alternating viscous and inertial force jetting (AVIFJ) mechanism. The results include an analysis of droplet formation mechanism, the system configuration, and experimental study of the effects of process parameters on microdroplet formation. Sodium alginate solutions are used for microdroplet formation and deposition. Key process parameters include actuation signal waveforms, nozzle dimensional features, and solution viscosity. Sizes of formed microdroplets are examined by measuring the droplet diameter and velocity. Results show that by utilizing a nozzle at a 45 μm diameter, the size of the formed microdroplets is in the range of 52-72 μm in diameter and 0.4-2.0 m/s in jetting speed, respectively. Reproducibility of the system is also examined and the results show that the deviation of the formed microdroplet diameter and the droplet deposition accuracy is within 6% and 6.2 μm range, respectively. Experimental results demonstrate a high controllability and precision for the developed DOD microdroplet deposition system with a potential for precise cell printing. Copyright © 2015 by ASME.

Zhang T.,Tsinghua University | Zhang T.,Key Laboratory for Advanced Materials Processing Technology | Zhang T.,Biomanufacturing and Rapid Forming Technology Key Laboratory of Beijing | Yan K.C.,The College of New Jersey | And 7 more authors.
Biofabrication | Year: 2013

Recent development in bioprinting technology enables the fabrication of complex, precisely controlled cell-encapsulated tissue constructs. Bioprinted tissue constructs have potential in both therapeutic applications and nontherapeutic applications such as drug discovery and screening, disease modelling and basic biological studies such as in vitro tissue modelling. The mechanical properties of bioprinted in vitro tissue models play an important role in mimicking in vivo the mechanochemical microenvironment. In this study, we have constructed three-dimensional in vitro soft tissue models with varying structure and porosity based on the 3D cell-assembly technique. Gelatin/alginate hybrid materials were used as the matrix material and cells were embedded. The mechanical properties of these models were assessed via compression tests at various culture times, and applicability of three material constitutive models was examined for fitting the experimental data. An assessment of cell bioactivity in these models was also carried out. The results show that the mechanical properties can be improved through structure design, and the compression modulus and strength decrease with respect to time during the first week of culture. In addition, the experimental data fit well with the Ogden model and experiential function. These results provide a foundation to further study the mechanical properties, structural and combined effects in the design and the fabrication of in vitro soft tissue models. © 2013 IOP Publishing Ltd.

Zhao Y.,Tsinghua University | Zhao Y.,Biomanufacturing and Rapid Forming Technology Key Laboratory of Beijing | Yao R.,Tsinghua University | Yao R.,Biomanufacturing and Rapid Forming Technology Key Laboratory of Beijing | And 12 more authors.
Biofabrication | Year: 2014

Advances in three-dimensional (3D) printing have enabled the direct assembly of cells and extracellular matrix materials to form in vitro cellular models for 3D biology, the study of disease pathogenesis and new drug discovery. In this study, we report a method of 3D printing for Hela cells and gelatin/alginate/fibrinogen hydrogels to construct in vitro cervical tumor models. Cell proliferation, matrix metalloproteinase (MMP) protein expression and chemoresistance were measured in the printed 3D cervical tumor models and compared with conventional 2D planar culture models. Over 90% cell viability was observed using the defined printing process. Comparisons of 3D and 2D results revealed that Hela cells showed a higher proliferation rate in the printed 3D environment and tended to form cellular spheroids, but formed monolayer cell sheets in 2D culture. Hela cells in 3D printed models also showed higher MMP protein expression and higher chemoresistance than those in 2D culture. These new biological characteristics from the printed 3D tumor models in vitro as well as the novel 3D cell printing technology may help the evolution of 3D cancer study. © 2014 IOP Publishing Ltd.

Ouyang L.,Tsinghua University | Ouyang L.,Biomanufacturing and Rapid Forming Technology Key Laboratory of Beijing | Yao R.,Tsinghua University | Yao R.,Biomanufacturing and Rapid Forming Technology Key Laboratory of Beijing | And 5 more authors.
Biofabrication | Year: 2015

3D printing has evolved into a versatile technology for fabricating tissue-engineered constructs with spatially controlled cells and biomaterial distribution to allow biomimicking of in vivo tissues. In this paper, we reported a novel study of 3D printing of cell lines derived from human embryonic kidney tissue into a macroporous tissue-like construct. Nozzle temperature, chamber temperature and the composition of the matrix material were studied to achieve high cell viability (>90%) after 3D printing and construct formation. Long-term construct stability with a clear grid structure up to 30 days was observed. Cells continued to grow as cellular spheroids with strong cell-cell interactions. Two transfected cell lines of HEK 293FT were also 3D printed and showed normal biological functions, i.e. protein synthesis and gene activation in responding to small molecule stimulus. With further refinement, this 3D cell printing technology may lead to a practical fabrication of functional embryonic tissues in vitro. © 2015 IOP Publishing Ltd.

Zhao Y.,Tsinghua University | Zhao Y.,Biomanufacturing and Rapid Forming Technology Key Laboratory of Beijing | Li Y.,Tsinghua University | Li Y.,Biomanufacturing and Rapid Forming Technology Key Laboratory of Beijing | And 8 more authors.
Biofabrication | Year: 2015

Three-dimensional (3D) cell printing technology has provided a versatile methodology to fabricate cell-laden tissue-like constructs and in vitro tissue/pathological models for tissue engineering, drug testing and screening applications. However, it still remains a challenge to print bioinks with high viscoelasticity to achieve long-term stable structure and maintain high cell survival rate after printing at the same time. In this study, we systematically investigated the influence of 3D cell printing parameters, i.e. composition and concentration of bioink, holding temperature and holding time, on the printability and cell survival rate in microextrusion-based 3D cell printing technology. Rheological measurements were utilized to characterize the viscoelasticity of gelatin-based bioinks. Results demonstrated that the bioink viscoelasticity was increased when increasing the bioink concentration, increasing holding time and decreasing holding temperature below gelation temperature. The decline of cell survival rate after 3D cell printing process was observed when increasing the viscoelasticity of the gelatin-based bioinks. However, different process parameter combinations would result in the similar rheological characteristics and thus showed similar cell survival rate after 3D bioprinting process. On the other hand, bioink viscoelasticity should also reach a certain point to ensure good printability and shape fidelity. At last, we proposed a protocol for 3D bioprinting of temperature-sensitive gelatin-based hydrogel bioinks with both high cell survival rate and good printability. This research would be useful for biofabrication researchers to adjust the 3D bioprinting process parameters quickly and as a referable template for designing new bioinks. © 2015 IOP Publishing Ltd.

Ouyang L.,Tsinghua University | Ouyang L.,Biomanufacturing and Rapid Forming Technology Key Laboratory of Beijing | Yao R.,Tsinghua University | Yao R.,Biomanufacturing and Rapid Forming Technology Key Laboratory of Beijing | And 7 more authors.
Biofabrication | Year: 2015

With the ability to manipulate cells temporarily and spatially into three-dimensional (3D) tissue-like construct, 3D bioprinting technology was used in many studies to facilitate the recreation of complex cell niche and/or to better understand the regulation of stem cell proliferation and differentiation by cellular microenvironment factors. Embryonic stem cells (ESCs) have the capacity to differentiate into any specialized cell type of the animal body, generally via the formation of embryoid body (EB), which mimics the early stages of embryogenesis. In this study, extrusion-based 3D bioprinting technology was utilized for biofabricating ESCs into 3D cell-laden construct. The influence of 3D printing parameters on ESC viability, proliferation, maintenance of pluripotency and the rule of EB formation was systematically studied in this work. Results demonstrated that ESCs were successfully printed with hydrogel into 3D macroporous construct. Upon process optimization, about 90% ESCs remained alive after the process of bioprinting and cell-laden construct formation. ESCs continued proliferating into spheroid EBs in the hydrogel construct, while retaining the protein expression and gene expression of pluripotent markers, like octamer binding transcription factor 4, stage specific embryonic antigen 1 and Nanog. In this novel technology, EBs were formed through cell proliferation instead of aggregation, and the quantity of EBs was tuned by the initial cell density in the 3D bioprinting process. This study introduces the 3D bioprinting of ESCs into a 3D cell-laden hydrogel construct for the first time and showed the production of uniform, pluripotent, high-throughput and size-controllable EBs, which indicated strong potential in ESC large scale expansion, stem cell regulation and fabrication of tissue-like structure and drug screening studies. © 2015 IOP Publishing Ltd.

Liu Y.,Drexel University | Hamid Q.,Drexel University | Snyder J.,Massachusetts Institute of Technology | Wang C.,Drexel University | And 3 more authors.
Rapid Prototyping Journal | Year: 2016

Purpose - This paper aims to present a solid freeform fabrication-based in situ three-dimensional (3D) printing method. This method enables simultaneous cross-linking alginate at ambient environmental conditions (temperature and pressure) for 3D-laden construct fabrication. The fabrication feasibility and potentials in biomedical applications were evaluated. Design/methodology/approach - Fabrication feasibility was evaluated as the investigation of fabrication parameters on strut formability (the capability to fabricate a cylindrical strut in the same diameter as dispensing tip) and structural stability (the capability to hold the fabricated 3D-laden construct against mechanical disturbance). Potentials in biomedical application was evaluated as the investigation on structural integrity (the capability to preserve the fabricated 3D-laden construct in cell culture condition). Findings - Strut formability can be achieved when the flow rate of alginate suspension and nozzle travel speed are set according to the dispensing tip size, and extruded alginate was cross-linked sufficiently. A range of cross-linking-related fabrication parameters was determined for sufficient cross-link. The structural stability and structural integrity were found to be controlled by alginate composition. An optimized setting of the alginate composition and the fabrication parameters was determined for the fabrication of a desired stable scaffold with structural integrity for 14 days. Originality/value - This paper reports that in situ 3D printing is an efficient method for 3D-laden construct fabrication and its potentials in biomedical application. © Emerald Group Publishing Limited.

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