Search Results (8)

Bioreactors used for the maturation of cell-seeded tissue-engineered scaffolds should essentially mimic the dynamic in vivo environments experienced by the native tissues they intend to substitute. In addition to perfusion of growth medium to facilitate continuous mass transfer, application of appropriate mechanical stimulation is important to enhance cellular responses in scaffolds for tissues such as tendons, skin, and cardiac muscle that experience dynamic loading. This study focuses on the development of a multi-modal custom bioreactor capable of applying cyclic tensile stimulation and perfusion within physiologically relevant ranges while minimizing shear stress detrimental to cells seeded on scaffolds. To validate the bioreactor design and operation, we assessed the effects of tensile stimulation (0.1 Hz, 2000 cycles/day) and perfusion (media flow rate = 0.15 mL/min) over 21 days on the biofunctional performance of 3D-bioplotted polycaprolactone (PCL) auxetic scaffolds with a representative design (missing-rib pattern) characterized by negative Poisson’s ratio similar to the aforementioned soft tissues. The scaffold had a tensile yield strain of 9.14%, yield strength of 0.25 MPa, elastic modulus of 2.85 MPa, and ultimate tensile strength (UTS) of 1.32 MPa. The application of perfusion and tensile stimulation (0–5% cyclic strain) for 21 days did not adversely affect the yield strength and elastic modulus of the scaffold but affected its UTS (22.5% decrease) compared to the control cultured without perfusion or stimulation. Notably, it resulted in significantly improved fibroblast cellular responses (DNA = 29 µg/g sample and collagen = 371.78 µg/g sample) compared to the control (7.52 µg/g sample and 163.51 µg/g sample, respectively). These results validate the bioreactor system operation and the ability of multi-modal stimulation to control biofunctional responses of auxetic scaffolds, which will serve as the basis for future studies that will optimize auxetic scaffold design and dynamic culture parameters for NPR tissue-specific applications. Full article

In the context of bone fractures, the influence of the mechanical environment on the healing outcome is widely accepted, while its influence at the cellular level is still poorly understood. This study explores the influence of mechanical load on naïve mesenchymal stem cell (MSC) differentiation, focusing on hypertrophic chondrocyte differentiation. Unlike primary bone healing, which involves the direct differentiation of MSCs into bone-forming cells, endochondral ossification uses an intermediate cartilage template that remodels into bone. A high-throughput uniaxial bioreactor system (StrainBot) was used to apply varying percentages of strain on naïve MSCs encapsulated in GelMa hydrogels. This research shows that cyclic uniaxial compression alone directs naïve MSCs towards a hypertrophic chondrocyte phenotype. This was demonstrated by increased cell volumes and reduced glycosaminoglycan (GAG) production, along with an elevated expression of hypertrophic markers such as MMP13 and Type X collagen. In contrast, Type II collagen, typically associated with resting chondrocytes, was poorly detected under mechanical loading alone conditions. The addition of chondrogenic factor TGFβ1 in the culture medium altered these outcomes. TGFβ1 induced chondrogenic differentiation, as indicated by higher GAG/DNA production and Type II collagen expression, overshadowing the effect of mechanical loading. This suggests that, under mechanical strain, hypertrophic differentiation is hindered by TGFβ1, while chondrogenesis is promoted. Biochemical analyses further confirmed these findings. Mechanical deformation alone led to a larger cell size and a more rounded cell morphology characteristic of hypertrophic chondrocytes, while lower GAG and proteoglycan production was observed. Immunohistology staining corroborated the gene expression data, showing increased Type X collagen with mechanical strain. Overall, this study indicates that mechanical loading alone drives naïve MSCs towards a hypertrophic chondrocyte differentiation path. These insights underscore the critical role of mechanical forces in MSC differentiation and have significant implications for bone healing, regenerative medicine strategies and rehabilitation protocols. Full article

The present work described a bio-functionalized 3D fibrous construct, as an interactive teno-inductive graft model to study tenogenic potential events of human mesenchymal stem cells collected from Wharton’s Jelly (hWJ-MSCs). The 3D-biomimetic and bioresorbable scaffold was functionalized with nanocarriers for the local controlled delivery of a teno-inductive factor, i.e., the human Growth Differentiation factor 5 (hGDF-5). Significant results in terms of gene expression were obtained. Namely, the up-regulation of Scleraxis (350-fold, p ≤ 0.05), type I Collagen (8-fold), Decorin (2.5-fold), and Tenascin-C (1.3-fold) was detected at day 14; on the other hand, when hGDF-5 was supplemented in the external medium only (in absence of nanocarriers), a limited effect on gene expression was evident. Teno-inductive environment also induced pro-inflammatory, (IL-6 (1.6-fold), TNF (45-fold, p ≤ 0.001), and IL-12A (1.4-fold)), and anti-inflammatory (IL-10 (120-fold) and TGF-β1 (1.8-fold)) cytokine expression upregulation at day 14. The presented 3D construct opens perspectives for the study of drug controlled delivery devices to promote teno-regenerative events. Full article

Formation of a barrier capable of protecting tissue from external damage, chemical factors, and pathogens is one of the main functions of the epidermis. Furthermore, upon development and during aging, mechanoprotective epidermal functions change dramatically. However, comparative studies between embryonic and adult skin in comparison to skin equivalents are still scarce which is especially due to the lack of appropriate measurement systems with sufficient accuracy and long-term tissue compatibility. Our studies fill this gap by developing a combined bioreactor and tensile testing machine for biomechanical analysis of living epithelia. Based on this tissue stretcher, our data clearly show that viscoelastic and plastic deformation behavior of embryonic and adult skin differ significantly. Tissue responses to static strain compared to cyclic strain also show a clear dependence on differentiation stage. Multilayered unkeratinized epidermis equivalents, on the other hand, respond very similar to mechanical stretch as adult tissue. This mechanical similarity is even more evident after a single cycle of mechanical preconditioning. Our studies therefore suggest that skin equivalents are well suited model systems to analyze cellular interactions of epidermal cells in natural tissues. Full article

The central nervous system (CNS) does not recover from traumatic axonal injury, but the peripheral nervous system (PNS) does. We hypothesize that this fundamental difference in regenerative capacity may be based upon the absence of stimulatory mechanical forces in the CNS due to the protective rigidity of the vertebral column and skull. We developed a bioreactor to apply low-strain cyclic axonal stretch to adult rat dorsal root ganglia (DRG) connected to either the peripheral or central nerves in an explant model for inducing axonal growth. In response, larger diameter DRG neurons, mechanoreceptors and proprioceptors showed enhanced neurite outgrowth as well as increased Activating Transcription Factor 3 (ATF3). Full article

We developed a (three-dimensional) 3D scaffold, we named HY-FIB, incorporating a force-transmission band of braided hyaluronate embedded in a cell localizing fibrin hydrogel and poly-lactic-co-glycolic acid (PLGA) nanocarriers as transient components for growth factor controlled delivery. The tenogenic supporting capacity of HY-FIB on human-Bone Marrow Mesenchymal Stem Cells (hBM-MSCs) was explored under static conditions and under bioreactor-induced cyclic strain conditions. HY-FIB elasticity enabled to deliver a mean shear stress of 0.09 Pa for 4 h/day. Tendon and cytokine marker expression by hBM-MSCs were studied. Results: hBM-MSCs embedded in HY-FIB and subjected to mechanical stimulation, resulted in a typical tenogenic phenotype, as indicated by type 1 Collagen fiber immunofluorescence. RT-qPCR showed an increase of type 1 Collagen, scleraxis, and decorin gene expression (3-fold, 1600-fold, and 3-fold, respectively, at day 11) in dynamic conditions. Cells also showed pro-inflammatory (IL-6, TNF, IL-12A, IL-1β) and anti-inflammatory (IL-10, TGF-β1) cytokine gene expressions, with a significant increase of anti-inflammatory cytokines in dynamic conditions (IL-10 and TGF-β1 300-fold and 4-fold, respectively, at day 11). Mechanical signaling, conveyed by HY-FIB to hBM-MSCs, promoted tenogenic gene markers expression and a pro-repair cytokine balance. The results provide strong evidence in support of the HY-FIB system and its interaction with cells and its potential for use as a predictive in vitro model. Full article

Polyhydroxyalkanoates (PHA) are microbial biopolyesters utilized as “green plastics”. Their production under controlled conditions resorts to bioreactors operated in different modes. Because PHA biosynthesis constitutes a multiphase process, both feeding strategy and bioreactor operation mode need smart adaptation. Traditional PHA production setups based on batch, repeated batch, fed-batch or cyclic fed-batch processes are often limited in productivity, or display insufficient controllability of polyester composition. For highly diluted substrate streams like is the case of (agro) industrial waste streams, fed-batch enhanced by cell recycling has recently been reported as a viable tool to increase volumetric productivity. As an emerging trend, continuous fermentation processes in single-, two- and multi-stage setups are reported, which bring the kinetics of both microbial growth and PHA accumulation into agreement with process engineering and allow tailoring PHA’s molecular structure. Moreover, we currently witness an increasing number of CO₂-based PHA production processes using cyanobacteria; these light-driven processes resort to photobioreactors similar to those used for microalgae cultivation and can be operated both discontinuously and continuously. This development is parallel to the emerging use of methane and syngas as abundantly available gaseous substrates, which also calls for bioreactor systems with optimized gas transfer. The review sheds light on the challenges of diverse PHA production processes in different bioreactor types and operational regimes using miscellaneous microbial production strains such as extremophilic Archaea, chemoheterotrophic eubacteria and phototrophic cyanobacteria. Particular emphasis is dedicated to the limitations and promises of different bioreactor–strain combinations and to efforts devoted to upscaling these processes to industrially relevant scales. Full article

The objective of this study was to design and validate a unique bioreactor design for applying spatially selective, linear, cyclic strain to degradable and non-degradable polymeric fabric scaffolds. This system uses a novel three-clamp design to apply cyclic strain via a computer controlled linear actuator to a specified zone of a scaffold while isolating the remainder of the scaffold from strain. Image analysis of polyethylene terephthalate (PET) woven scaffolds subjected to a 3% mechanical stretch demonstrated that the stretched portion of the scaffold experienced 2.97% ± 0.13% strain (mean ± standard deviation) while the unstretched portion experienced 0.02% ± 0.18% strain. NIH-3T3 fibroblast cells were cultured on the PET scaffolds and half of each scaffold was stretched 5% at 0.5 Hz for one hour per day for 14 days in the bioreactor. Cells were checked for viability and proliferation at the end of the 14 day period and levels of glycosaminoglycan (GAG) and collagen (hydroxyproline) were measured as indicators of extracellular matrix production. Scaffolds in the bioreactor showed a seven-fold increase in cell number over scaffolds cultured statically in tissue culture plastic petri dishes (control). Bioreactor scaffolds showed a lower concentration of GAG deposition per cell as compared to the control scaffolds largely due to the great increase in cell number. A 75% increase in hydroxyproline concentration per cell was seen in the bioreactor stretched scaffolds as compared to the control scaffolds. Surprisingly, little differences were experienced between the stretched and unstretched portions of the scaffolds for this study. This was largely attributed to the conditioned and shared media effect. Results indicate that the bioreactor system is capable of applying spatially-selective, linear, cyclic strain to cells growing on polymeric fabric scaffolds and evaluating the cellular and matrix responses to the applied strains. Full article