scholarly journals Medium Perfusion Flow Improves Osteogenic Commitment of Human Stromal Cells

2019 ◽  
Vol 2019 ◽  
pp. 1-10 ◽  
Author(s):  
Alice Pasini ◽  
Joseph Lovecchio ◽  
Giulia Ferretti ◽  
Emanuele Giordano
Keyword(s):  
Tissue Engineering ◽  
Cell Culture ◽  
Stromal Cells ◽  
Waste Product ◽  
Type I ◽  
Cell Viability Assay ◽  
Perfusion Flow ◽  
Dynamic Cell ◽  
Flow Stimulation

Dynamic culture protocols have recently emerged as part of (bone) tissue engineering strategies due to their ability to represent a more physiological cell environment in vitro. Here, we described how a perfusion flow induced by a simple bioreactor system improves proliferation and osteogenic commitment of human bone marrow stromal cells. L88/5 cells were cultured in poly(methyl methacrylate) custom-milled communicating well plates, in the presence of an osteogenic cocktail containing 1α,25-dihydroxyvitamin D3, L-ascorbic acid 2-phosphate, andβ-glycerophosphate. The dynamic cell culture was maintained under perfusion flow stimulation at 1 mL/min for up to 4 days and compared with a static control condition. A cell viability assay showed that the proliferation associated with the dynamic cell culture was 20% higher vs. the static condition. A significantly higher upregulation of the osteogenic markers runt-related transcription factor 2 (RUNX2), collagen type I (COL1A1), osteocalcin (BGLAP), alkaline phosphatase (ALPL), and osteopontin (SPP1) was detected when the perfusion flow stimulation was administered to the cells treated with the osteogenic cocktail. An in silico analysis showed that in the dynamic cell culture condition (i) the shear stress in the proximity of the cell layer approximates 10-3 Pa, (ii) the nutrient and the waste product concentration is more homogeneously distributed than in the static counterpart, and (iii) perfusion flow was associated with higher nutrient consumption. In summary, increased cell proliferation and enhanced early phenotype commitment indicate that dynamic cell culture conditions, delivered via bioreactor systems, produce an enhanced in vitro environment for both basic and translational research in tissue engineering and regenerative medicine.

Laser microstructured scaffolds for tissue engineering under dynamic cell culture conditions

2020 ◽  
Author(s):  
Ελευθερία Μπαμπαλιάρη
Keyword(s):  
Tissue Engineering ◽  
Cell Culture ◽  
Culture Conditions ◽  
Dynamic Cell ◽  
Cell Culture Conditions

Παρόλο που το περιφερικό νευρικό σύστημα εμφανίζει υψηλότερο ρυθμό αναγέννησης από εκείνο του κεντρικού νευρικού συστήματος μέσω αυθόρμητης αναγέννησης μετά από έναν τραυματισμό, η καθοδηγούμενη αξονική νευρική αναγέννηση και η λειτουργική αποκατάσταση είναι αρκετά σπάνια. Συνεπώς, η ανάπτυξη επιτυχημένων μεθόδων για την καθοδήγηση της νευρικής ανάπτυξης, «in vitro», είναι υψίστης σημασίας. Έχει αναφερθεί λεπτομερώς ότι η τοπογραφία του υποστρώματος επηρεάζει την ανάπτυξη, τον προσανατολισμό και τη διαφοροποίηση των νευρικών κυττάρων. Ωστόσο, η συνδυασμένη δράση της διατμητικής τάσης και της τοπογραφίας του υποστρώματος στην νευρική ανάπτυξη έχει ελάχιστα μελετηθεί, παρόλο που οι διατμητικές τάσεις είναι ευρέως γνωστό ότι διαδραματίζουν καθοριστικό ρόλο στην οργάνωση, ανάπτυξη και λειτουργία των ιστών. Σε αυτή τη μελέτη, ένα σύστημα μικροροών ακριβούς ελεγχόμενης ροής με συγκεκριμένους ειδικά σχεδιασμένους θαλάμους, που ενσωματώνουν μικροδομημένα υποστρώματα λέιζερ, αναπτύχθηκε για να μελετηθεί η συνδυασμένη δράση της διατμητικής τάσης και της τοπογραφίας υποστρώματος στην ανάπτυξη, στον προσανατολισμό, στην επιμήκυνση και στη διαφοροποίηση νευρικών κυττάρων. Πολυμερικά μικροδομημένα υποστρώματα, με ελεγχόμενη γεωμετρία και κανονικότητα μοτίβου, κατασκευάστηκαν με χρήση υπερβραχέων παλμών λέιζερ. Πραγματοποιήθηκε συγκριτική μελέτη μεταξύ στατικών και δυναμικών κυτταρικών καλλιεργειών για να αξιολογηθεί η συνεργατική ή ανταγωνιστική επίδραση της διατμητικής τάσης και της τοπογραφίας στη συμπεριφορά των νευρικών κυττάρων. Τα αποτελέσματα της κυτταρικής καλλιέργειας συμπληρώθηκαν με υπολογιστικές προσομοιώσεις ροής με σκοπό τον ακριβή υπολογισμό των αντίστοιχων τιμών διατμητικής τάσης.

scholarly journals Collagen-Based Electrospun Materials for Tissue Engineering: A Systematic Review

2021 ◽  
Vol 8 (3) ◽  
pp. 39
Author(s):  
Britani N. Blackstone ◽  
Summer C. Gallentine ◽  
Heather M. Powell
Keyword(s):  
Systematic Review ◽  
Tissue Engineering ◽  
Collagen Type I ◽  
The Body ◽  
Pool Data ◽  
Type I ◽  
High Concentrations ◽  
Increased Strength

Collagen is a key component of the extracellular matrix (ECM) in organs and tissues throughout the body and is used for many tissue engineering applications. Electrospinning of collagen can produce scaffolds in a wide variety of shapes, fiber diameters and porosities to match that of the native ECM. This systematic review aims to pool data from available manuscripts on electrospun collagen and tissue engineering to provide insight into the connection between source material, solvent, crosslinking method and functional outcomes. D-banding was most often observed in electrospun collagen formed using collagen type I isolated from calfskin, often isolated within the laboratory, with short solution solubilization times. All physical and chemical methods of crosslinking utilized imparted resistance to degradation and increased strength. Cytotoxicity was observed at high concentrations of crosslinking agents and when abbreviated rinsing protocols were utilized. Collagen and collagen-based scaffolds were capable of forming engineered tissues in vitro and in vivo with high similarity to the native structures.

scholarly journals Validation of in vitro assays in three-dimensional human dermal constructs

2018 ◽  
Vol 41 (11) ◽  
pp. 779-788 ◽  
Author(s):  
Ayesha Idrees ◽  
Valeria Chiono ◽  
Gianluca Ciardelli ◽  
Siegfried Shah ◽  
Richard Viebahn ◽  
...  
Keyword(s):  
Three Dimensional ◽  
Dermal Fibroblasts ◽  
In Vitro Assays ◽  
Dimensional System ◽  
Type I ◽  
Human Dermal Fibroblasts ◽  
Cell Viability Assay ◽  
Normal Human ◽  
Normal Human Dermal Fibroblasts

Three-dimensional cell culture systems are urgently needed for cytocompatibility testing of biomaterials. This work aimed at the development of three-dimensional in vitro dermal skin models and their optimization for cytocompatibility evaluation. Initially “murine in vitro dermal construct” based on L929 cells was generated, leading to the development of “human in vitro dermal construct” consisting of normal human dermal fibroblasts in rat tail tendon collagen type I. To assess the viability of the cells, different assays CellTiter-Blue®, RealTime-Glo™ MT, and CellTiter-Glo® (Promega) were evaluated to optimize the best-suited assay to the respective cell type and three-dimensional system. Z-stack imaging (Live/Dead and Phalloidin/DAPI-Promokine) was performed to visualize normal human dermal fibroblasts inside matrix revealing filopodia-like morphology and a uniform distribution of normal human dermal fibroblasts in matrix. CellTiter-Glo was found to be the optimal cell viability assay among those analyzed. CellTiter-Blue reagent affected the cell morphology of normal human dermal fibroblasts (unlike L929), suggesting an interference with cell biological activity, resulting in less reliable viability data. On the other hand, RealTime-Glo provided a linear signal only with a very low cell density, which made this assay unsuitable for this system. CellTiter-Glo adapted to three-dimensional dermal construct by optimizing the “shaking time” to enhance the reagent penetration and maximum adenosine triphosphate release, indicating 2.4 times higher viability value by shaking for 60 min than for 5 min. In addition, viability results showed that cells were viable inside the matrix. This model would be further advanced with more layers of skin to make a full thickness model.

scholarly journals An Assessment of Cell Culture Plate Surface Chemistry for in Vitro Studies of Tissue Engineering Scaffolds

2015 ◽  
Vol 6 (4) ◽  
pp. 1054-1063 ◽  
Author(s):  
Alexander Röder ◽  
Elena García-Gareta ◽  
Christina Theodoropoulos ◽  
Nikola Ristovski ◽  
Keith Blackwood ◽  
...  
Keyword(s):  
Tissue Engineering ◽  
Cell Culture ◽  
Surface Chemistry ◽  
In Vitro Studies ◽  
Plate Surface ◽  
Tissue Engineering Scaffolds ◽  
Culture Plate

Preliminary testing of silicone-urethane elastomers as substrates in human cell culture

e-Polymers ◽  
2007 ◽  
Vol 7 (1) ◽  
Author(s):  
Malgorzata Lewandowska-Szumieł ◽  
Janusz Kozakiewicz ◽  
Piotr Mrówka ◽  
Agnieszka Jurkowska ◽  
Edyta Sienkiewicz-Łatka ◽  
...  
Keyword(s):  
Tissue Engineering ◽  
Cell Culture ◽  
Blood Platelets ◽  
Human Fibroblasts ◽  
Medical Implants ◽  
Tissue Engineering Scaffolds ◽  
Xtt Assay ◽  
In Vitro Biocompatibility ◽  
Different Types

AbstractSilicone-urethanes, polymers combining the characteristics of two widely used biomaterials, i.e. polyurethanes and silicones, are highly valued in many applications, including medical implants. To assess properties of these materials in contact with living cells, a set of different silicone-urethane materials, candidates for tissue engineering scaffolds, was synthesized and characterized. Two different oligomeric siloxane diols: Tegomer-2111 (Teg) and KF-6001 (KF), and two different types of diisocyanate, MDI and IPDI, were used in synthesis. Blood platelets adhesion to surfaces of selected materials showed a higher thrombogenicity of material based on Teg. Human fibroblasts were used in in vitro biocompatibility tests. The viability of cells cultured on silicone-urethanes was tested by XTT assay. Teg-based silicone-urethanes showed a significantly higher biocompatibility than those based on KF. Materials based on MDI compared to IPDI were found to be significantly more favoured by cells, not necessarily due to the type of diisocyanate but maybe also because of the necessity of using potentially toxic catalyst which accompanies the use of IPDI. Our studies indicate that silicone-urethanes are potent materials for tissue engineering products development. On the basis of the observations performed in cell culture, Tegomer- 2111 as oligomeric siloxane diol and MDI as diisocyanate are recommended as starting materials for silicone-urethane scaffolds synthesis.

Mechanically Enhanced Precipitation of Phase-Inversion Sprayed Polyurethane Scaffold May Be Used to Match Tissue Specific Anisotropy

2009 ◽  
Author(s):  
James P. Kennedy ◽  
Robert W. Hitchcock
Keyword(s):  
Mechanical Properties ◽  
Tissue Engineering ◽  
Cell Culture ◽  
Pore Size ◽  
Phase Inversion ◽  
Anisotropy Ratio ◽  
Material Anisotropy ◽  
Polyurethane Scaffold ◽  
Stiffness Anisotropy

Methods of creating a scaffold for tissue engineering that allow for modification of properties such as pore size, porosity, and anisotropy are essential for tissue engineering applications. For example the pore size and material anisotropy have been shown to affect cardiomyocyte elongation and alignment [1]. Phase-inversion spray polymerization (PISP) is a method for rapidly precipitating polymers onto a surface by depositing the polymer solution simultaneously with a nonsolvent, and may be used to create biocompatible scaffolds of engineered morphological and mechanical properties by varying the solubility of the polymer in the nonsolvent [2]. We report here on the fabrication of scaffolds using different nonsolvents and methods of in-process elongation that allow for control of stiffness, anisotropy ratio, porosity, and in vitro cell culture.

Human Mesenchymal Stromal Cells are Mechanosensitive to Vibration Stimuli

2012 ◽  
Vol 91 (12) ◽  
pp. 1135-1140 ◽  
Author(s):  
I.S. Kim ◽  
Y.M. Song ◽  
B. Lee ◽  
S.J. Hwang
Keyword(s):  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Type I Collagen ◽  
Bone Cells ◽  
Type I ◽  
Related Factors ◽  
Matrix Mineralization ◽  
Vibration Signals ◽  
Human Mesenchymal Stromal Cells

Low-magnitude high-frequency (LMHF) vibrations have the ability to stimulate bone formation and reduce bone loss. However, the anabolic mechanisms that are mediated by vibration in human bone cells at the cellular level remain unclear. We hypothesized that human mesenchymal stromal cells (hMSCs) display direct osteoblastic responses to LMHF vibration signals. Daily exposure to vibrations increased the proliferation of hMSCs, with the highest efficiency occurring at a peak acceleration of 0.3 g and vibrations at 30 to 40 Hz. Specifically, these conditions promoted osteoblast differentiation through an increase in alkaline phosphatase activity and in vitro matrix mineralization. The effect of vibration on the expression of osteogenesis-related factors differed depending on culture method. hMSCs that underwent vibration in a monolayer culture did not exhibit any changes in the expressions of these genes, while cells in three-dimensional culture showed increased expression of type I collagen, osteoprotegerin, or VEGF, and VEGF induction appeared in 2 different hMSC lines. These results are among the first to demonstrate a dose-response effect upon LMHF stimulation, thereby demonstrating that hMSCs are mechanosensitive to LMHF vibration signals such that they could facilitate the osteogenic process.

Improvement of Bioactivity of Collagen/Alginate Scaffolds by Hydroxyapatite for Tissue Engineering Cartilage

2008 ◽  
Vol 396-398 ◽  
pp. 445-448 ◽  
Author(s):  
J. Sun ◽  
R. Wang ◽  
L. Zheng ◽  
Yan Fei Tan ◽  
Yu Mei Xiao ◽  
...  
Keyword(s):  
Mechanical Property ◽  
Tissue Engineering ◽  
Cell Culture ◽  
Composite Film ◽  
Cartilage Tissue Engineering ◽  
Cartilage Tissue ◽  
The Poor ◽  
Pure Collagen ◽  
Good Biocompatibility

With good biocompatibility, collagen is often used in cartilage tissue engineering. Collagen/alginate composite was hoped to improve the poor mechanical property of pure collagen but the biocompatibity was decreased. In this study, hydroxyapatite (HA) particles were used to get collagen/alginate/HA (CAHA) composite film to enhance the bioactivity properties. The bioactivity of the composite was investigated by in vitro co-culture with chondrocytes. During the 6-day cell culture in vitro, the composite showed a significant improvement in promoting proliferation and maintaining morphology/phenotype of the chondrocytes over collagen/alginate composite by MTT, SEM, fluorescent and immunohistochemical assays. Cytocompatibility and cytoviablility of CAHA even come up to that of collagen film alone. The results indicated that the composite film may provide an appropriate environment for the proliferation and maintaining the morphology and phenotype of chondrocytes and have a potential clinical application in the cartilage tissue engineering field.

scholarly journals Craniofacial Bone Tissue Engineering: Current Approaches and Potential Therapy

Cells ◽  
2021 ◽  
Vol 10 (11) ◽  
pp. 2993
Author(s):  
Arbi Aghali
Keyword(s):  
Tissue Engineering ◽  
Growth Factors ◽  
Bone Tissue ◽  
Stromal Cells ◽  
Critical Size ◽  
Tumor Resection ◽  
Bone Morphogenetic Protein 2 ◽  
Craniofacial Bone

Craniofacial bone defects can result from various disorders, including congenital malformations, tumor resection, infection, severe trauma, and accidents. Successfully regenerating cranial defects is an integral step to restore craniofacial function. However, challenges managing and controlling new bone tissue formation remain. Current advances in tissue engineering and regenerative medicine use innovative techniques to address these challenges. The use of biomaterials, stromal cells, and growth factors have demonstrated promising outcomes in vitro and in vivo. Natural and synthetic bone grafts combined with Mesenchymal Stromal Cells (MSCs) and growth factors have shown encouraging results in regenerating critical-size cranial defects. One of prevalent growth factors is Bone Morphogenetic Protein-2 (BMP-2). BMP-2 is defined as a gold standard growth factor that enhances new bone formation in vitro and in vivo. Recently, emerging evidence suggested that Megakaryocytes (MKs), induced by Thrombopoietin (TPO), show an increase in osteoblast proliferation in vitro and bone mass in vivo. Furthermore, a co-culture study shows mature MKs enhance MSC survival rate while maintaining their phenotype. Therefore, MKs can provide an insight as a potential therapy offering a safe and effective approach to regenerating critical-size cranial defects.

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