Hydrogels for biomedical applications.

Increased use of reconstruction procedures in orthopedics, due to trauma, tumor, deformity, degeneration and an aging population, has caused a blossom, not only in surgical advancement, but also in the development of bone implants. Traditional synthetic porous scaffolds are made of metals, polymers, ceramics or even composite biomaterials, in which the design does not consider the native structure and properties of cells and natural tissues. Thus, these synthetic scaffolds often poorly integrate with the cells and surrounding host tissue, thereby resulting in unsatisfactory surgical outcomes due to poor corrosion and wear, mechanical mismatch, unamiable surface environment, and other unfavorable properties. Musculoskeletal tissue reconstruction is the ultimate objective in orthopedic surgery. This objective can be achieved by (i) prosthesis or fixation device implantation, and (ii) tissue engineered bone scaffolds. These devices focus on the design of implants, regardless of the choice of new biomaterials. Indeed, metallic materials, e.g. 316L stainless steel, titanium alloys and cobalt chromium alloys, are predominantly used in bone surgeries, especially in the load-bearing zone of prostheses. The engineered scaffolds take biodegradability, cell biology, biomolecules and material mechanical properties into account, in which these features are ideally suited for bone tissue repair and regeneration. Therefore, the design of the scaffold is extremely important to the success of clinical outcomes in musculoskeletal surgeries. The ideal scaffolds should mimic the natural extracellular matrix (ECM) as much as possible, since the ECM found in natural tissues supports cell attachment, proliferation, and differentiation, indicating that scaffolds should consist of appropriate biochemistry and nano/micro-scale surface topographies, in order to formulate favorable binding sites to actively regulate and control cell and tissue behavior, while interacting with host cells. In addition, scaffolds should also possess a similar macro structure to what is found in natural bone. This feature may provide space for the growth of cells and new tissues, as well as for the carriers of growth factors. Another important concern is the mechanical properties of scaffolds. It has been reported that the mechanical features can significantly influence the osteointegration between implants and surrounding tissues, as well as cell behaviors. Since natural bone exhibits super-elastic biomechanical properties with a Young's modulus value in the range of 1–27 GPa, the ideal scaffolds should mimic strength, stiffness and mechanical behavior, so as to avoid possible post-operation stress shielding effects, which induce bone resorption and consequent implant failure. In addition, the rate of degradation and the by-products of biodegradable materials are also critical in the role of bone regeneration. Indeed, the mechanical integrity of a scaffold will be significantly reduced if the degradation rate is rapid, thereby resulting in a pre-matured collapse of the scaffold before the tissue is regenerated. Another concern is that the by-products upon degradation may alter the tissue microenvironment and then challenge the biocompatibility of the scaffold and the subsequent tissue repair. Therefore, these two factors should be carefully considered when designing new biomaterials for tissue regeneration. To address the aforementioned questions, an overview of the design of ideal biomimetic porous scaffolds is presented in this paper. Hence, a number of original engineering processes and techniques, including the production of a hierarchical structure on both the macro- and nano-scales, the adjustment of biomechanical properties through structural alignment and chemical components, the control of the biodegradability of the scaffold and its by-products, the change of biomimetic surface properties by altering interfacial chemistry, and micro- and nano-topographies will be discussed. In general, the concepts and techniques mentioned above provide insights into designing superior biomimetic scaffolds for bone tissue engineering.

Biomimetic porous scaffolds for bone tissue engineering

Generation of thick vascularized tissues that fully match the patient still remains an unmet challenge in cardiac tissue engineering. Here, a simple approach to 3D-print thick, vascularized, and perfusable cardiac patches that completely match the immunological, cellular, biochemical, and anatomical properties of the patient is reported. To this end, a biopsy of an omental tissue is taken from patients. While the cells are reprogrammed to become pluripotent stem cells, and differentiated to cardiomyocytes and endothelial cells, the extracellular matrix is processed into a personalized hydrogel. Following, the two cell types are separately combined with hydrogels to form bioinks for the parenchymal cardiac tissue and blood vessels. The ability to print functional vascularized patches according to the patient's anatomy is demonstrated. Blood vessel architecture is further improved by mathematical modeling of oxygen transfer. The structure and function of the patches are studied in vitro, and cardiac cell morphology is assessed after transplantation, revealing elongated cardiomyocytes with massive actinin striation. Finally, as a proof of concept, cellularized human hearts with a natural architecture are printed. These results demonstrate the potential of the approach for engineering personalized tissues and organs, or for drug screening in an appropriate anatomical structure and patient-specific biochemical microenvironment.

https://onlinelibrary.wiley.com/doi/pdf/10.1002/advs.201900344

3D Printing of Personalized Thick and Perfusable Cardiac Patches and Hearts

Bone biomaterials play a vital role in bone repair by providing the necessary substrate for cell adhesion, proliferation, and differentiation and by modulating cell activity and function. In past decades, extensive efforts have been devoted to developing bone biomaterials with a focus on the following issues: (1) developing ideal biomaterials with a combination of suitable biological and mechanical properties; (2) constructing a cell microenvironment with pores ranging in size from nanoscale to submicro- and microscale; and (3) inducing the oriented differentiation of stem cells for artificial-to-biological transformation. Here we present a comprehensive review of the state of the art of bone biomaterials and their interactions with stem cells. Typical bone biomaterials that have been developed, including bioactive ceramics, biodegradable polymers, and biodegradable metals, are reviewed, with an emphasis on their characteristics and applications. The necessary porous structure of bone biomaterials for the cell microenvironment is discussed, along with the corresponding fabrication methods. Additionally, the promising seed stem cells for bone repair are summarized, and their interaction mechanisms with bone biomaterials are discussed in detail. Special attention has been paid to the signaling pathways involved in the focal adhesion and osteogenic differentiation of stem cells on bone biomaterials. Finally, achievements regarding bone biomaterials are summarized, and future research directions are proposed.

/pdf/bone-biomaterials-and-interactions-with-stem-cells-4c9eg43ssz.pdf

Bone biomaterials and interactions with stem cells.

The inadequacy of animal models in correctly predicting drug and biothreat agent toxicity in humans has resulted in a pressing need for in vitro models that can recreate the in vivo scenario. One of the most important organs in the assessment of drug toxicity is liver. Here, we report the development of a liver-on-a-chip platform for long-term culture of three-dimensional (3D) human HepG2/C3A spheroids for drug toxicity assessment. The bioreactor design allowed for in situ monitoring of the culture environment by enabling direct access to the hepatic construct during the experiment without compromising the platform operation. The engineered bioreactor could be interfaced with a bioprinter to fabricate 3D hepatic constructs of spheroids encapsulated within photocrosslinkable gelatin methacryloyl (GelMA) hydrogel. The engineered hepatic construct remained functional during the 30 days culture period as assessed by monitoring the secretion rates of albumin, alpha-1 antitrypsin, transferrin, and ceruloplasmin, as well as immunostaining for the hepatocyte markers, cytokeratin 18, MRP2 bile canalicular protein and tight junction protein ZO-1. Treatment with 15 mM acetaminophen induced a toxic response in the hepatic construct that was similar to published studies on animal and other in vitro models, thus providing a proof-of-concept demonstration of the utility of this liver-on-a-chip platform for toxicity assessment.

/pdf/a-liver-on-a-chip-platform-with-bioprinted-hepatic-spheroids-sqnirb0v77.pdf

A liver-on-a-chip platform with bioprinted hepatic spheroids.

Bone extracellular matrix (ECM) is a natural composite made of collagen and mineral hydroxyapatite (HA). Dynamic cell-ECM interactions play a critical role in regulating cell differentiation and function. Understanding the principal ECM cues promoting osteogenic differentiation would be pivotal for both bone tissue engineering and regenerative medicine. Altering the mineral content generally modifies the stiffness as well as other physicochemical cues provided by composite materials, complicating the “cause-effect” analysis of resultant cell behaviour. To isolate the contribution of mechanical cues from other HA-derived signals, we developed and characterised composite HA/gelatin scaffolds with different mineral contents along with a set of stiffness-matched HA-free gelatin scaffolds. Samples were seeded with human periosteal derived progenitor cells (PDPCs) and cultured over 7 days, analysing their resultant morphology and gene expression. Our results show that both stiffness and HA contribute to directing PDPC osteogenic differentiation, highlighting the role of stiffness in triggering the expression of osteogenic genes and of HA in accelerating the process, particularly at high concentrations.

/pdf/correction-corrigendum-decoupling-the-role-of-stiffness-from-4zmnhpbyqv.pdf

Correction: Corrigendum: Decoupling the role of stiffness from other hydroxyapatite signalling cues in periosteal derived stem cell differentiation

In this paper, we discuss the basic design requirements for the development of physiologically meaningful in vitro systems comprising cells, scaffolds and bioreactors, through a bottom up approach. Very simple micro- and milli-fluidic geometries are first used to illustrate the concepts, followed by a real device case-study. At each step, the fluidic and mass transport parameters in biological tissue design are considered, starting from basic questions such as the minimum number of cells and cell density required to represent a physiological system and the conditions necessary to ensure an adequate nutrient supply to tissues. At the next level, we consider the use of three-dimensional scaffolds, which are employed both for regenerative medicine applications and for the study of cells in environments which better recapitulate the physiological milieu. Here, the driving need is the rate of oxygen supply which must be maintained at an appropriate level to ensure cell viability throughout the thickness of a scaffold. Scaffold and bioreactor design are both critical in defining the oxygen profile in a cell construct and are considered together. We also discuss the oxygen-shear stress trade-off by considering the levels of mechanical stress required for hepatocytes, which are the limiting cell type in a multi-organ model. Similar considerations are also made for glucose consumption in cell constructs. Finally, the allometric approach for generating multi-tissue systemic models using bioreactors is described.

https://www.mdpi.com/2227-9717/2/3/548/pdf

Design Criteria for Generating Physiologically Relevant In Vitro Models in Bioreactors

Measuring the viscoelastic behavior of highly hydrated biological materials is challenging because of their intrinsic softness and labile nature. In these materials, it is difficult to avoid prestress and therefore to establish precise initial stress and strain conditions for lumped parameter estimation using creep or stress-relaxation (SR) tests. We describe a method ( or epsilon dot method) for deriving the viscoelastic parameters of soft hydrated biomaterials which avoids prestress and can be used to rapidly test degradable samples. Standard mechanical tests are first performed compressing samples using different strain rates. The dataset obtained is then analyzed to mathematically derive the material's viscoelastic parameters. In this work a stable elastomer, polydimethylsiloxane, and a labile hydrogel, gelatin, were first tested using the, in parallel SR was used to compare lumped parameter estimation. After demonstrating that the elastic parameters are equivalent and that the estimation of short-time constants is more precise using the proposed method, the viscoelastic behavior of porcine liver was investigated using this approach. The results show that the constitutive parameters of hepatic tissue can be quickly quantified without the application of any prestress and before the onset of time-dependent degradation phenomena. © 2013 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 102A: 3352–3360, 2014

https://onlinelibrary.wiley.com/doi/pdf/10.1002/jbm.a.34914

Strain rate viscoelastic analysis of soft and highly hydrated biomaterials

Mechanostructure and composition of highly reproducible decellularized liver matrices.

The aim of this study was to identify a method for modifying the time-dependent viscoelastic properties of gels without altering the elastic component. To this end, two hydrogels commonly used in biomedical applications, agarose and acrylamide, were prepared in aqueous solutions of dextran with increasing concentrations (0%, 2% and 5% w/v) and hence increasing viscosities. Commercial polyurethane sponges soaked in the same solutions were used as controls, since, unlike in hydrogels, the liquid in these sponge systems is poorly bound to the polymer network. Sample viscoelastic properties were characterised using the epsilon-dot method, based on compression tests at different constant strain-rates. Experimental data were fitted to a standard linear solid model. While increasing the liquid viscosity in the controls resulted in a significant increase of the characteristic relaxation time (τ), both the instantaneous (Einst) and the equilibrium (Eeq) elastic moduli remained almost constant. However, in the hydrogels a significant reduction of both Einst and τ was observed. On the other hand, as expected, Eeq - an indicator of the equilibrium elastic behaviour after the occurrence of viscoelastic relaxation dynamics - was found to be independent of the liquid phase viscosity. Therefore, although the elastic and viscous components of hydrogels cannot be completely decoupled due to the interaction of the liquid and solid phases, we show that their viscoelastic behaviour can be modulated by varying the viscosity of the aqueous phase. This simple-yet-effective strategy could be useful in the field of mechanobiology, particularly for studying cell response to substrate viscoelasticity while keeping the elastic cue (i.e. equilibrium modulus, or quasi-static stiffness) constant.

https://arpi.unipi.it/bitstream/11568/930491/9/Engineering%20hydrogel%20viscoelasticity%20Manuscript%20-preprint.pdf

Giorgio Mattei

Papers

Correction: Corrigendum: Decoupling the role of stiffness from other hydroxyapatite signalling cues in periosteal derived stem cell differentiation

Design Criteria for Generating Physiologically Relevant In Vitro Models in Bioreactors

Strain rate viscoelastic analysis of soft and highly hydrated biomaterials

Mechanostructure and composition of highly reproducible decellularized liver matrices.

Engineering hydrogel viscoelasticity.