147 research outputs found
Progress and challenges in large-scale expansion of human pluripotent stem cells
The constant supply of high cell numbers generated by defined, robust, and economically viable culture processes is indispensable for the envisioned application of human pluripotent stem cells (hPSCs) and their progenies for drug discovery and regenerative medicine. To achieve required cell numbers and to reduce process-related risks such as cell transformation, relative short batch-like production processes at industry- and clinically-relevant scale(s) must be developed and optimized. Here, we will review recent progress in the large-scale expansion of hPSCs with particular focus on suspension culture, which represents a universal strategy for controlled mass cell production. Another focus of the paper relates to bioreactor-based approaches, including technical aspects of bioreactor technologies and operation modes. Lastly, we will discuss current challenges of hPSC process engineering for enabling the transition from early stage process development to fully optimized hPSC production scale operation, a mandatory step for hPSCs’ industrial and clinical translation
Development of an animal-free methodology for mechanical performance assessment of engineered skin substitutes
Challenges of Periodontal Tissue Engineering: Increasing Biomimicry through 3D Printing and Controlled Dynamic Environment
In recent years, tissue engineering studies have proposed several approaches to regenerate periodontium based on the use of three-dimensional (3D) tissue scaffolds alone or in association with periodontal ligament stem cells (PDLSCs). The rapid evolution of bioprinting has sped up classic regenerative medicine, making the fabrication of multilayered scaffolds—which are essential in targeting the periodontal ligament (PDL)—conceivable. Physiological mechanical loading is fundamental to generate this complex anatomical structure ex vivo. Indeed, loading induces the correct orientation of the fibers forming the PDL and maintains tissue homeostasis, whereas overloading or a failure to adapt to mechanical load can be at least in part responsible for a wrong tissue regeneration using PDLSCs. This review provides a brief overview of the most recent achievements in periodontal tissue engineering, with a particular focus on the use of PDLSCs, which are the best choice for regenerating PDL as well as alveolar bone and cementum. Different scaffolds associated with various manufacturing methods and data derived from the application of different mechanical loading protocols have been analyzed, demonstrating that periodontal tissue engineering represents a proof of concept with high potential for innovative therapies in the near future
An electro-mechanical bioreactor providing physiological cardiac stimuli
In cardiac tissue engineering it has been widely demonstrated the fundamental role of physical stimuli in improving structural and functional properties of the engineered cardiac constructs. An electro-mechanical bioreactor has been designed and developed to provide physiological uniaxial stretching and electrical stimuli for inducing functional differentiation and promoting morphological and structural maturation of cultured cardiac constructs obtained from stem cell-seeded scaffolds. The bioreactor is composed of: a transparent and sterilizable culture chamber for housing four cell-seeded scaffolds and the culture medium (working volume = 70 ml); a mechanical stimulation system, with a dedicated grasping system, to provide cyclic stretching (strain up to 20%, cycling frequency up to 2 Hz); an electrical stimulation system to provide electrical monophasic square pulses (1-6 V/cm, 0.25-10 ms, 1-5 Hz); a recirculation system for the automated medium change; a control system for data acquisition and mechanical stimulation. Preliminary in-house tests confirmed the suitability and the performances of the bioreactor as regards fittingness of chamber isolation, grasping system, and physical stimulation systems. Cell culture tests are in progress for investigating the influence of stretching and electrical stimuli on development of engineered cardiac constructs. Due to its high versatility, this bioreactor is a multipurpose adaptable system for dynamic culture of cell-seeded scaffolds for tissue engineering research and application
Versatile electrical stimulator for providing cardiac-like electrical impulses in vitro
In the perspective of reliable methods alternative to in vivo animal testing for cardiac tissue engineering (CTE) research, the versatile electrical stimulator ELETTRA has been developed. ELETTRA delivers controlled and stable cardiac-like electrical impulses, and it can be coupled to already existing bioreactors for providing in vitro combined biomimetic culture conditions. Designed to be cost-effective and easy to use, this device could contribute to the reduction and replacement of in vivo animal experiments in CTE
Versatile electrical stimulator for cardiac tissue engineering—Investigation of charge-balanced monophasic and biphasic electrical stimulations
The application of biomimetic physical stimuli replicating the in vivo dynamic microenvironment is crucial for the in vitro development of functional cardiac tissues. In particular, pulsed electrical stimulation (ES) has been shown to improve the functional properties of in vitro cultured cardiomyocytes. However, commercially available electrical stimulators are expensive and cumbersome devices while customized solutions often allow limited parameter tunability, constraining the investigation of different ES protocols. The goal of this study was to develop a versatile compact electrical stimulator (ELETTRA) for biomimetic cardiac tissue engineering approaches, designed for delivering controlled parallelizable ES at a competitive cost. ELETTRA is based on an open-source micro-controller running custom software and is combinable with different cell/tissue culture set-ups, allowing simultaneously testing different ES patterns on multiple samples. In particular, customized culture chambers were appositely designed and manufactured for investigating the influence of monophasic and biphasic pulsed ES on cardiac cell monolayers. Finite element analysis was performed for characterizing the spatial distributions of the electrical field and the current density within the culture chamber. Performance tests confirmed the accuracy, compliance, and reliability of the ES parameters delivered by ELETTRA. Biological tests were performed on neonatal rat cardiac cells, electrically stimulated for 4 days, by comparing, for the first time, the monophasic waveform (electric field = 5 V/cm) to biphasic waveforms by matching either the absolute value of the electric field variation (biphasic ES at ±2.5 V/cm) or the total delivered charge (biphasic ES at ±5 V/cm). Findings suggested that monophasic ES at 5 V/cm and, particularly, charge-balanced biphasic ES at ±5 V/cm were effective in enhancing electrical functionality of stimulated cardiac cells and in promoting synchronous contraction
Bizonal cardiac engineered tissues with differential maturation features in a mid-throughput multimodal bioreactor
Functional three-dimensional (3D) engineered cardiac tissue (ECT) models are essential for effective drug screening and biological studies. Application of physiological cues mimicking those typical of the native myocardium is known to promote the cardiac maturation and functionality in vitro. Commercially available bioreactors can apply one physical force type at a time and often in a restricted loading range. To overcome these limitations, a millimetric-scalemicroscope-integrated bioreactor was developed to deliver multiple biophysical stimuli to ECTs. In this study, we showed that the single application of auxotonic loading (passive) generated a bizonal ECT with a unique cardiac maturation pattern. Throughout the statically cultured constructs and in the ECT region exposed to high passive loading, cardiomyocytes predominantly displayed a round morphology and poor contractility ability. The ECT region with a low passive mechanical stimulation instead showed both rat- and human-origin cardiac cell maturation and organization, as well as increased ECT functionality
An electro-mechanical bioreactor providing physiological cardiac stimuli
In cardiac tissue engineering it has been widely demonstrated the fundamental role of physical stimuli in improving structural and functional properties of the engineered cardiac constructs.
An electro-mechanical bioreactor has been designed and developed to provide physiological uniaxial stretching and electrical stimuli for inducing functional differentiation and promoting morphological and structural maturation of cultured cardiac constructs obtained from stem cell-seeded scaffolds.
The bioreactor is composed of: a transparent and sterilizable culture chamber for housing four cell-seeded scaffolds and the culture medium (working volume = 70 ml); a mechanical stimulation system, with a dedicated grasping system, to provide cyclic stretching (strain up to 20%, cycling frequency up to 2 Hz); an electrical stimulation system to provide electrical monophasic square pulses (1-6 V/cm, 0.25-10 ms, 1-5 Hz); a recirculation system for the automated medium change; a control system for data acquisition and mechanical stimulation. Preliminary in-house tests confirmed the suitability and the performances of the bioreactor as regards fittingness of chamber isolation, grasping system, and physical stimulation systems. Cell culture tests are in progress for investigating the influence of stretching and electrical stimuli on development of engineered cardiac constructs. Due to its high versatility, this bioreactor is a multipurpose adaptable system for dynamic culture of cell-seeded scaffolds for tissue engineering research and applications
Image-Based Three-Dimensional Analysis to Characterize the Texture of Porous Scaffolds
The aim of the present study is to characterize the microstructure of composite scaffolds for bone tissue regeneration containing different ratios of chitosan/gelatin blend and bioactive glasses. Starting from realistic 3D models of the scaffolds reconstructed from micro-CT images, the level of heterogeneity of scaffold architecture is evaluated performing a lacunarity analysis. The results demonstrate that the presence of the bioactive glass component affects not only macroscopic features such as porosity, but mainly scaffold microarchitecture giving rise to structural heterogeneity, which could have an impact on the local cell-scaffold interaction and scaffold performances. The adopted approach allows to investigate the scale-dependent pore distribution within the scaffold and the related structural heterogeneity features, providing a comprehensive characterization of the scaffold texture
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