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1.
J Neural Eng ; 19(1)2022 01 24.
Article in English | MEDLINE | ID: mdl-34942614

ABSTRACT

There is a need for the development of new cellular therapies for the treatment of many diseases, with the central nervous system (CNS) currently an area of specific focus. Due to the complexity and delicacy of its biology, there is currently a limited understanding of neurogenesis and consequently a lack of reliable test platforms, resulting in several CNS based diseases having no cure. The ability to differentiate pluripotent stem cells into specific neuronal sub-types may enable scalable manufacture for clinical therapies, with a focus also on the purity and quality of the cell population. This focus is targeted towards an urgent need for the diseases that currently have no cure, e.g. Parkinson's disease. Differentiation studies carried out using traditional 2D cell culture techniques are designed using biological signals and morphogens known to be important for neurogenesisin vivo.However, such studies are limited by their simplistic nature, including a general poor efficiency and reproducibility, high reagent costs and an inability to scale-up the process to a manufacture-wide design for clinical use. Biomimetic approaches to recapitulate a morein vivo-like environment are progressing rapidly within this field, with application of bio(chemical) gradients presented both as 2D surfaces and within a 3D volume. This review focusses on the development and application of these advanced extracellular environments particularly for the neural niche. We emphasise the progress that has been made specifically in the area of stem cell derived neuronal differentiation. Increasing developments in biomaterial approaches to manufacture stem cells will enable the improvement of differentiation protocols, enhancing the efficiency and repeatability of the process with a move towards up-scaling. Progress in this area brings these techniques closer to enabling the development of therapies for the clinic.


Subject(s)
Biomimetics , Neurogenesis , Cell Culture Techniques/methods , Cell Differentiation , Neurogenesis/physiology , Reproducibility of Results
2.
Front Cell Dev Biol ; 9: 722294, 2021.
Article in English | MEDLINE | ID: mdl-34527674

ABSTRACT

Digitally driven manufacturing technologies such as aerosol jet printing (AJP) can make a significant contribution to enabling new capabilities in the field of tissue engineering disease modeling and drug screening. AJP is an emerging non-contact and mask-less printing process which has distinct advantages over other patterning technologies as it offers versatile, high-resolution, direct-write deposition of a variety of materials on planar and non-planar surfaces. This research demonstrates the ability of AJP to print digitally controlled patterns that influence neuronal guidance. These consist of patterned poly(3,4-ethylenedioxythiophene)-poly(styrenesulfonate) (PEDOT:PSS) tracks on both glass and poly(potassium 3-sulfopropyl methacrylate) (PKSPMA) coated glass surfaces, promoting selective adhesion of SH-SY5Y neuroblastoma cells. The cell attractive patterns had a maximum height ≥0.2 µm, width and half height ≥15 µm, Ra = 3.5 nm, and RMS = 4.1. The developed biocompatible PEDOT:PSS ink was shown to promote adhesion, growth and differentiation of SH-SY5Y neuronal cells. SH-SY5Y cells cultured directly onto these features exhibited increased nuclei and neuronal alignment on both substrates. In addition, the cell adhesion to the substrate was selective when cultured onto the PKSPMA surfaces resulting in a highly organized neural pattern. This demonstrated the ability to rapidly and flexibly realize intricate and accurate cell patterns by a computer controlled process.

3.
Sci Rep ; 11(1): 11695, 2021 06 03.
Article in English | MEDLINE | ID: mdl-34083648

ABSTRACT

Investigations of the human neuromuscular junction (NMJ) have predominately utilised experimental animals, model organisms, or monolayer cell cultures that fail to represent the physiological complexity of the synapse. Consequently, there remains a paucity of data regarding the development of the human NMJ and a lack of systems that enable investigation of the motor unit. This work addresses this need, providing the methodologies to bioengineer 3D models of the human motor unit. Spheroid culture of iPSC derived motor neuron progenitors augmented the transcription of OLIG2, ISLET1 and SMI32 motor neuron mRNAs ~ 400, ~ 150 and ~ 200-fold respectively compared to monolayer equivalents. Axon projections of adhered spheroids exceeded 1000 µm in monolayer, with transcription of SMI32 and VACHT mRNAs further enhanced by addition to 3D extracellular matrices in a type I collagen concentration dependent manner. Bioengineered skeletal muscles produced functional tetanic and twitch profiles, demonstrated increased acetylcholine receptor (AChR) clustering and transcription of MUSK and LRP4 mRNAs, indicating enhanced organisation of the post-synaptic membrane. The number of motor neuron spheroids, or motor pool, required to functionally innervate 3D muscle tissues was then determined, generating functional human NMJs that evidence pre- and post-synaptic membrane and motor nerve axon co-localisation. Spontaneous firing was significantly elevated in 3D motor units, confirmed to be driven by the motor nerve via antagonistic inhibition of the AChR. Functional analysis outlined decreased time to peak twitch and half relaxation times, indicating enhanced physiology of excitation contraction coupling in innervated motor units. Our findings provide the methods to maximise the maturity of both iPSC motor neurons and primary human skeletal muscle, utilising cell type specific extracellular matrices and developmental timelines to bioengineer the human motor unit for the study of neuromuscular junction physiology.


Subject(s)
Bioengineering/methods , Motor Neurons/metabolism , Muscle, Skeletal/metabolism , Neuromuscular Junction/metabolism , Axons/metabolism , Humans , Muscle, Skeletal/physiology , Neuromuscular Junction/physiology , RNA, Messenger/metabolism
4.
J Cell Physiol ; 236(10): 7033-7044, 2021 10.
Article in English | MEDLINE | ID: mdl-33738797

ABSTRACT

Fatty acids (FA) exert physiological and pathophysiological effects leading to changes in skeletal muscle metabolism and function, however, in vitro models to investigate these changes are limited. These experiments sought to establish the effects of physiological and pathophysiological concentrations of exogenous FA upon the function of tissue engineered skeletal muscle (TESkM). Cultured initially for 14 days, C2C12 TESkM was exposed to FA-free bovine serum albumin alone or conjugated to a FA mixture (oleic, palmitic, linoleic, and α-linoleic acids [OPLA] [ratio 45:30:24:1%]) at different concentrations (200 or 800 µM) for an additional 4 days. Subsequently, TESkM morphology, functional capacity, gene expression and insulin signaling were analyzed. There was a dose response increase in the number and size of lipid droplets within the TESkM (p < .05). Exposure to exogenous FA increased the messenger RNA expression of genes involved in lipid storage (perilipin 2 [p < .05]) and metabolism (pyruvate dehydrogenase lipoamide kinase isozyme 4 [p < .01]) in a dose dependent manner. TESkM force production was reduced (tetanic and single twitch) (p < .05) and increases in transcription of type I slow twitch fiber isoform, myosin heavy chain 7, were observed when cultured with 200 µM OPLA compared to control (p < .01). Four days of OPLA exposure results in lipid accumulation in TESkM which in turn results in changes in muscle function and metabolism; thus, providing insight ito the functional and mechanistic changes of TESkM in response to exogenous FA.


Subject(s)
Fatty Acids/toxicity , Lipid Droplets/drug effects , Lipid Metabolism/drug effects , Muscle, Skeletal/drug effects , Myoblasts, Skeletal/drug effects , Animals , Cell Line , Dose-Response Relationship, Drug , Gene Expression Regulation , Insulin/pharmacology , Lipid Droplets/metabolism , Lipid Metabolism/genetics , Mice , Muscle Strength/drug effects , Muscle, Skeletal/metabolism , Muscle, Skeletal/physiopathology , Myoblasts, Skeletal/metabolism , Myoblasts, Skeletal/pathology , Tissue Engineering
5.
J Muscle Res Cell Motil ; 42(2): 149-159, 2021 06.
Article in English | MEDLINE | ID: mdl-32955689

ABSTRACT

Skeletal muscle atrophy as a consequence of acute and chronic illness, immobilisation, muscular dystrophies and aging, leads to severe muscle weakness, inactivity and increased mortality. Mechanical loading is thought to be the primary driver for skeletal muscle hypertrophy, however the extent to which mechanical loading can offset muscle catabolism has not been thoroughly explored. In vitro 3D-models of skeletal muscle provide a controllable, high throughput environment and mitigating many of the ethical and methodological constraints present during in vivo experimentation. This work aimed to determine if mechanical loading would offset dexamethasone (DEX) induced skeletal muscle atrophy, in muscle engineered using the C2C12 murine cell line. Mechanical loading successfully offset myotube atrophy and functional degeneration associated with DEX regardless of whether the loading occurred before or after 24 h of DEX treatment. Furthermore, mechanical load prevented increases in MuRF-1 and MAFbx mRNA expression, critical regulators of muscle atrophy. Overall, we demonstrate the application of tissue engineered muscle to study skeletal muscle health and disease, offering great potential for future use to better understand treatment modalities for skeletal muscle atrophy.


Subject(s)
Dexamethasone , Muscle Fibers, Skeletal , Animals , Cell Line , Dexamethasone/adverse effects , Mice , Muscle Fibers, Skeletal/pathology , Muscle, Skeletal/pathology , Muscular Atrophy/chemically induced , Muscular Atrophy/pathology
6.
J Tissue Eng ; 11: 2041731420967294, 2020.
Article in English | MEDLINE | ID: mdl-33194170

ABSTRACT

Herein, the cytotoxicity of a novel zwitterionic sulfobetaine hydrogel system with a nano-clay crosslinker has been investigated. We demonstrate that careful selection of the composition of the system (monomer to Laponite content) allows the material to be formed into controlled shapes using an extrusion based additive manufacturing technique with the ability to tune the mechanical properties of the product. Moreover, the printed structures can support their own weight without requiring curing during printing which enables the use of a printing-then-curing approach. Cell culture experiments were conducted to evaluate the neural cytotoxicity of the developed hydrogel system. Cytotoxicity evaluations were conducted on three different conditions; a control condition, an indirect condition (where the culture medium used had been in contact with the hydrogel to investigate leaching) and a direct condition (cells growing directly on the hydrogel). The result showed no significant difference in cell viability between the different conditions and cells were also found to be growing on the hydrogel surface with extended neurites present.

7.
Biotechnol J ; 15(1): e1900106, 2020 Jan.
Article in English | MEDLINE | ID: mdl-31468704

ABSTRACT

In vitro 3D tissue-engineered (TE) structures have been shown to better represent in vivo tissue morphology and biochemical pathways than monolayer culture, and are less ethically questionable than animal models. However, to create systems with even greater relevance, multiple integrated tissue systems should be recreated in vitro. In the present study, the effects and conditions most suitable for the co-culture of TE skeletal muscle and bone are investigated. High-glucose Dulbecco's modified Eagle medium (HG-DMEM) supplemented with 20% fetal bovine serum followed by HG-DMEM with 2% horse serum is found to enable proliferation of both C2C12 muscle precursor cells and TE85 human osteosarcoma cells, fusion of C2C12s into myotubes, as well as an upregulation of RUNX2/CBFa1 in TE85s. Myotube formation is also evident within indirect contact monolayer cultures. Finally, in 3D co-cultures, TE85 collagen/hydroxyapatite constructs have significantly greater expression of RUNX2/CBFa1 and osteocalcin/BGLAP in the presence of collagen-based C2C12 skeletal muscle constructs; however, fusion within these constructs appears reduced. This work demonstrates the first report of the simultaneous co-culture and differentiation of 3D TE skeletal muscle and bone, and represents a significant step toward a full in vitro 3D musculoskeletal junction model.


Subject(s)
Bone and Bones , Coculture Techniques/methods , Muscle, Skeletal , Tissue Engineering/methods , Animals , Bone and Bones/cytology , Bone and Bones/metabolism , Cell Line , Cell Proliferation/drug effects , Culture Media/chemistry , Culture Media/pharmacology , Humans , Mice , Muscle, Skeletal/cytology , Muscle, Skeletal/metabolism , Myoblasts/cytology , Myoblasts/drug effects , Myoblasts/metabolism , Osteoblasts/cytology , Osteoblasts/drug effects , Osteoblasts/metabolism
8.
Cytoskeleton (Hoboken) ; 76(6): 371-382, 2019 06.
Article in English | MEDLINE | ID: mdl-31376315

ABSTRACT

Skeletal muscle has a high regenerative capacity, injuries trigger a regenerative program which restores tissue function to a level indistinguishable to the pre-injury state. However, in some cases where significant trauma occurs, such as injuries seen in military populations, the regenerative process is overwhelmed and cannot restore full function. Limited clinical interventions exist which can be used to promote regeneration and prevent the formation of non-regenerative defects following severe skeletal muscle trauma. Robust and reproducible techniques for modelling complex tissue responses are essential to promote the discovery of effective clinical interventions. Tissue engineering has been highlighted as an alternative method, allowing the generation of three-dimensional in vivo like tissues without laboratory animals. Reducing the requirement for animal models promotes rapid screening of potential clinical interventions, as these models are more easily manipulated, genetically and pharmacologically, and reduce the associated cost and complexity, whilst increasing access to models for laboratories without animal facilities. In this study, an in vitro chemical injury using barium chloride is validated using the C2C12 myoblast cell line, and is shown to selectively remove multinucleated myotubes, whilst retaining a regenerative mononuclear cell population. Monolayer cultures showed limited regenerative capacity, with basement membrane supplementation or extended regenerative time incapable of improving the regenerative response. Conversely tissue engineered skeletal muscles, supplemented with basement membrane proteins, showed full functional regeneration, and a broader in vivo like inflammatory response. This work outlines a freely available and open access methodology to produce a cell line-based tissue engineered model of skeletal muscle regeneration.


Subject(s)
Basement Membrane/metabolism , Membrane Proteins/metabolism , Muscle, Skeletal/physiology , Regeneration/physiology , Tissue Engineering , Animals , Barium Compounds/pharmacology , Basement Membrane/drug effects , Cell Differentiation/drug effects , Cell Line , Chlorides/pharmacology , Collagen/pharmacology , Collagen Type I/metabolism , Drug Combinations , Gene Expression Regulation/drug effects , Laminin/pharmacology , Mice , Muscle Fibers, Skeletal/drug effects , Muscle Fibers, Skeletal/metabolism , Muscle, Skeletal/drug effects , Proteoglycans/pharmacology , Regeneration/drug effects , Regeneration/genetics
9.
J Cell Physiol ; 234(12): 23547-23558, 2019 12.
Article in English | MEDLINE | ID: mdl-31180593

ABSTRACT

Mechanical loading of skeletal muscle results in molecular and phenotypic adaptations typified by enhanced muscle size. Studies on humans are limited by the need for repeated sampling, and studies on animals have methodological and ethical limitations. In this investigation, three-dimensional skeletal muscle was tissue-engineered utilizing the murine cell line C2C12, which bears resemblance to native tissue and benefits from the advantages of conventional in vitro experiments. The work aimed to determine if mechanical loading induced an anabolic hypertrophic response, akin to that described in vivo after mechanical loading in the form of resistance exercise. Specifically, we temporally investigated candidate gene expression and Akt-mechanistic target of rapamycin 1 signalling along with myotube growth and tissue function. Mechanical loading (construct length increase of 15%) significantly increased insulin-like growth factor-1 and MMP-2 messenger RNA expression 21 hr after overload, and the levels of the atrophic gene MAFbx were significantly downregulated 45 hr after mechanical overload. In addition, p70S6 kinase and 4EBP-1 phosphorylation were upregulated immediately after mechanical overload. Maximal contractile force was augmented 45 hr after load with a 265% increase in force, alongside significant hypertrophy of the myotubes within the engineered muscle. Overall, mechanical loading of tissue-engineered skeletal muscle induced hypertrophy and improved force production.


Subject(s)
Cell Proliferation , Mechanotransduction, Cellular , Muscle Contraction , Muscle Fibers, Skeletal/physiology , Muscle Strength , Tissue Engineering , Adaptor Proteins, Signal Transducing/metabolism , Animals , Cell Cycle Proteins/metabolism , Cell Line , Hypertrophy , Insulin-Like Growth Factor I/genetics , Insulin-Like Growth Factor I/metabolism , Matrix Metalloproteinase 2/genetics , Matrix Metalloproteinase 2/metabolism , Mechanistic Target of Rapamycin Complex 1/metabolism , Mice , Muscle Fibers, Skeletal/metabolism , Phenotype , Phosphorylation , Proto-Oncogene Proteins c-akt/metabolism , Ribosomal Protein S6 Kinases, 70-kDa/metabolism , Stress, Mechanical , Time Factors
10.
Article in English | MEDLINE | ID: mdl-30838203

ABSTRACT

Tissue engineered skeletal muscle allows investigation of the cellular and molecular mechanisms that regulate skeletal muscle pathology. The fabricated model must resemble characteristics of in vivo tissue and incorporate cost-effective and high content primary human tissue. Current models are limited by low throughput due to the complexities associated with recruiting tissue donors, donor specific variations, as well as cellular senescence associated with passaging. This research presents a method using fused deposition modeling (FDM) and laser sintering (LS) 3D printing to generate reproducible and scalable tissue engineered primary human muscle, possessing aligned mature myotubes reminiscent of in vivo tissue. Many existing models are bespoke causing variability when translated between laboratories. To this end, a scalable model has been developed (25-500 µL construct volumes) allowing fabrication of mature primary human skeletal muscle. This research provides a strategy to overcome limited biopsy cell numbers, enabling high throughput screening of functional human tissue.

11.
J Mech Behav Biomed Mater ; 93: 130-142, 2019 05.
Article in English | MEDLINE | ID: mdl-30785078

ABSTRACT

Functionally graded materials (FGMs), with varying spatial, chemical and mechanical gradients (continuous or stepwise), have the potential to mimic heterogenous properties found across biological tissues. They can prevent stress concentrations and retain healthy cellular functions. Here, we show for the first time the fabrication of polydimethylsiloxane and poly(ether) ether ketone (PDMS-PEEK) composites. These were successfully manufactured as a bulk material and functionally graded (stepwise) without the use of hazardous solvents or the need of additives. Chemical, irreversible adhesion between layers (for the FGMs) was achieved without the formation of hard, boundary interfaces. The mechanical properties of PDMS-PEEK FGMs are proven to be further tailorable across the entirety of the build volume, mimicking the transition from soft to harder tissues. The introduction of 20 wt% PEEK particles into the PDMS matrix resulted in significant rises in the elastic modulus under tensile and compressive loading. Biological and thermal screenings suggested that these composites cause no adverse effects to human fibroblast cell lines and can retain physical state and mass at body temperature, which could make the composites suitable for a range of biomedical applications such as maxillofacial prosthetics, artificial blood vessels and articular cartilage replacement.


Subject(s)
Biocompatible Materials/chemistry , Dimethylpolysiloxanes/chemistry , Ketones/chemistry , Mechanical Phenomena , Polyethylene Glycols/chemistry , Benzophenones , Biocompatible Materials/toxicity , Cell Line , Cell Survival/drug effects , Fibroblasts/cytology , Fibroblasts/drug effects , Humans , Polymers
12.
ACS Biomater Sci Eng ; 5(10): 5525-5538, 2019 Oct 14.
Article in English | MEDLINE | ID: mdl-33464072

ABSTRACT

Bioengineered skeletal muscle tissues benefit from dynamic culture environments which facilitate the appropriate provision of nutrients and removal of cellular waste products. Biologically compatible perfusion systems hold the potential to enhance the physiological biomimicry of in vitro tissues via dynamic culture, in addition to providing technological advances in analytical testing and live cellular imaging for analysis of cellular development. To meet such diverse requirements, perfusion systems require the capacity and adaptability to incorporate multiple cell laden constructs of both monolayer and bioengineered tissues. This work reports perfusion systems produced using additive manufacturing technology for the in situ phenotypic development of myogenic precursor cells in monolayer and bioengineered tissue. Biocompatibility of systems 3D printed using stereolithography (SL), laser sintering (LS), and PolyJet outlined preferential morphological development within both SL and LS devices. When exposed to intermittent perfusion in the monolayer, delayed yet physiologically representative cellular proliferation, MyoD and myogenin transcription of C2C12 cells was evident. Long-term (8 days) intermittent perfusion of monolayer cultures outlined viable morphological and genetic in situ differentiation for the live cellular imaging of myogenic development. Continuous perfusion cultures (13 days) of bioengineered skeletal muscle tissues outlined in situ myogenic differentiation, forming mature multinucleated myotubes. Here, reductions in IL-1ß and TNF-α inflammatory cytokines, myostatin, and MuRF-1 atrophic mRNA expression were observed. Comparable myosin heavy chain (MyHC) isoform transcription profiles were evident between conditions; however, total mRNA expression was reduced in perfusion conditions. Decreased transcription of MuRF1 and subsequent reduced ubiquitination of the MyHC protein allude to a decreased requirement for transcription of MyHC isoform transcripts. Together, these data appear to indicate that 3D printed perfusion systems elicit enhanced stability of the culture environment, resulting in a reduced basal requirement for MyHC gene expression within bioengineered skeletal muscle tissue.

13.
ACS Omega ; 3(10): 12383-12391, 2018 Oct 31.
Article in English | MEDLINE | ID: mdl-30411006

ABSTRACT

Conventional in vitro cultures are useful to represent simplistic neuronal behavior; however, the lack of organization results in random neurite spreading. To overcome this problem, control over the directionality of SH-SY5Y cells was attained, utilizing photolithography to pattern the cell-repulsive anionic brush poly(potassium 3-sulfopropyl methacrylate) (PKSPMA) into tracks of 20, 40, 80, and 100 µm width. These data validate the use of PKSPMA brush coatings for a long-term culture of the SH-SY5Y cells, as well as providing a methodology by which the precise deposition of PKSPMA can be utilized to achieve a targeted control over the SH-SY5Y cells. Specifically, the PKSPMA brush patterns prevented cell attachment, allowing the SH-SY5Y cells to grow only on noncoated glass (gaps of 20, 50, 75, and 100 µm width) at different cell densities (5000, 10 000, and 15 000 cells/cm2). This research demonstrates the importance of achieving cell directionality in vitro, while these simplistic models could provide new platforms to study complex neuron-neuron interactions.

14.
Front Physiol ; 9: 483, 2018.
Article in English | MEDLINE | ID: mdl-29867538

ABSTRACT

There are several three-dimensional (3D) skeletal muscle (SkM) tissue engineered models reported in the literature. 3D SkM tissue engineering (TE) aims to recapitulate the structure and function of native (in vivo) tissue, within an in vitro environment. This requires the differentiation of myoblasts into aligned multinucleated myotubes surrounded by a biologically representative extracellular matrix (ECM). In the present work, a new commercially available 3D SkM TE culture chamber manufactured from polyether ether ketone (PEEK) that facilitates suitable development of these myotubes is presented. To assess the outcomes of the myotubes within these constructs, morphological, gene expression, and ECM remodeling parameters were compared against a previously published custom-built model. No significant differences were observed in the morphological and gene expression measures between the newly introduced and the established construct configuration, suggesting biological reproducibility irrespective of manufacturing process. However, TE SkM fabricated using the commercially available PEEK chambers displayed reduced variability in both construct attachment and matrix deformation, likely due to increased reproducibility within the manufacturing process. The mechanical differences between systems may also have contributed to such differences, however, investigation of these variables was beyond the scope of the investigation. Though more expensive than the custom-built models, these PEEK chambers are also suitable for multiple use after autoclaving. As such this would support its use over the previously published handmade culture chamber system, particularly when seeking to develop higher-throughput systems or when experimental cost is not a factor.

15.
Macromol Biosci ; 18(7): e1800113, 2018 07.
Article in English | MEDLINE | ID: mdl-29900676

ABSTRACT

The integration of additive manufacturing (AM) technology within biological systems holds significant potential, specifically when refining the methods utilized for the creation of in vitro models. Therefore, examination of cellular interaction with the physical/physicochemical properties of 3D-printed polymers is critically important. In this work, skeletal muscle (C2 C12 ), neuronal (SH-SY5Y) and hepatic (HepG2) cell lines are utilized to ascertain critical evidence of cellular behavior in response to 3D-printed candidate polymers: Clear-FL (stereolithography, SL), PA-12 (laser sintering, LS), and VeroClear (PolyJet). This research outlines initial critical evidence for a framework of polymer/AM process selection when 3D printing biologically receptive scaffolds, derived from industry standard, commercially available AM instrumentation. C2 C12 , SH-SY5Y, and HepG2 cells favor LS polymer PA-12 for applications in which cellular adherence is necessitated. However, cell type specific responses are evident when cultured in the chemical leachate of photopolymers (Clear-FL and VeroClear). With the increasing prevalence of 3D-printed biointerfaces, the development of rigorous cell type specific biocompatibility data is imperative. Supplementing the currently limited database of functional 3D-printed biomaterials affords the opportunity for experiment-specific AM process and polymer selection, dependent on biological application and intricacy of design features required.


Subject(s)
Biocompatible Materials/chemical synthesis , Polymers/chemical synthesis , Printing, Three-Dimensional , Animals , Biocompatible Materials/pharmacology , Cell Adhesion/drug effects , Cell Line , Cell Line, Tumor , Cell Survival/drug effects , Hep G2 Cells , Humans , Mice , Myoblasts/cytology , Myoblasts/drug effects , Myoblasts/metabolism , Neurons/cytology , Neurons/drug effects , Neurons/metabolism , Photochemical Processes , Polymers/pharmacology
16.
ACS Biomater Sci Eng ; 4(1): 98-106, 2018 Jan 08.
Article in English | MEDLINE | ID: mdl-33418681

ABSTRACT

The disordered environment found in conventional neural cultures impedes various applications where cell directionality is a key process for functionality. Neurons are highly specialized cells known to be greatly dependent on interactions with their surroundings. Therefore, when chemical cues are incorporated on the surface material, a precise control over neuronal behavior can be achieved. Here, the behavior of SH-SY5Y neurons on a variety of self-assembled monolayers (SAMs) and polymer brushes both in isolation and combination to promote cellular spatial control was determined. APTES and BIBB coatings promoted the highest cell viability, proliferation, metabolic activity, and neuronal maturation, while low cell survival was seen on PKSPMA and PMETAC surfaces. These cell-attractive and repulsive surfaces were combined to generate a binary BIBB-PKSPMA coating, whereby cellular growth was restricted to an exclusive neural region. The utility of these coatings when precisely combined could act as a bioactive/bioinert surface resulting in a biomimetic environment where control over neuronal growth and directionality can be achieved.

17.
Lab Chip ; 17(17): 2982-2993, 2017 08 22.
Article in English | MEDLINE | ID: mdl-28762415

ABSTRACT

The capability to 3D print bespoke biologically receptive parts within short time periods has driven the growing prevalence of additive manufacture (AM) technology within biological settings, however limited research concerning cellular interaction with 3D printed polymers has been undertaken. In this work, we used skeletal muscle C2C12 cell line in order to ascertain critical evidence of cellular behaviour in response to multiple bio-receptive candidate polymers; polylactic acid (PLA), acrylonitrile butadiene styrene (ABS), polyethylene terephthalate (PET) and polycarbonate (PC) 3D printed via fused deposition modelling (FDM). The extrusion based nature of FDM elicited polymer specific topographies, within which C2C12 cells exhibited reduced metabolic activity when compared to optimised surfaces of tissue culture plastic, however assay viability readings remained high across polymers outlining viable phenotypes. C2C12 cells exhibited consistently high levels of morphological alignment across polymers, however differential myotube widths and levels of transcriptional myogenin expression appeared to demonstrate response specific thresholds at which varying polymer selection potentiates cellular differentiation, elicits pre-mature early myotube formation and directs subsequent morphological phenotype. Here we observed biocompatible AM polymers manufactured via FDM, which also appear to hold the potential to simultaneously manipulate the desired biological phenotype and enhance the biomimicry of skeletal muscle cells in vitro via AM polymer choice and careful selection of machine processing parameters. When considered in combination with the associated design freedom of AM, this may provide the opportunity to not only enhance the efficiency of creating biomimetic models, but also to precisely control the biological output within such scaffolds.


Subject(s)
Biocompatible Materials/chemistry , Bioprinting/methods , Polymers/chemistry , Printing, Three-Dimensional , Animals , Biocompatible Materials/toxicity , Cell Culture Techniques/instrumentation , Cell Line , Cell Survival/drug effects , Mice , Myoblasts/cytology , Phenotype , Polymers/toxicity , RNA, Messenger/analysis , RNA, Messenger/metabolism , Surface Properties
18.
Beilstein J Org Chem ; 13: 111-119, 2017.
Article in English | MEDLINE | ID: mdl-28228852

ABSTRACT

Additive manufacturing or '3D printing' is being developed as a novel manufacturing process for the production of bespoke micro- and milliscale fluidic devices. When coupled with online monitoring and optimisation software, this offers an advanced, customised method for performing automated chemical synthesis. This paper reports the use of two additive manufacturing processes, stereolithography and selective laser melting, to create multifunctional fluidic devices with embedded reaction monitoring capability. The selectively laser melted parts are the first published examples of multifunctional 3D printed metal fluidic devices. These devices allow high temperature and pressure chemistry to be performed in solvent systems destructive to the majority of devices manufactured via stereolithography, polymer jetting and fused deposition modelling processes previously utilised for this application. These devices were integrated with commercially available flow chemistry, chromatographic and spectroscopic analysis equipment, allowing automated online and inline optimisation of the reaction medium. This set-up allowed the optimisation of two reactions, a ketone functional group interconversion and a fused polycyclic heterocycle formation, via spectroscopic and chromatographic analysis.

19.
Lab Chip ; 13(23): 4583-90, 2013 Dec 07.
Article in English | MEDLINE | ID: mdl-24100659

ABSTRACT

We review the use of additive manufacturing (AM) as a novel manufacturing technique for the production of milli-scale reactor systems. Five well-developed additive manufacturing techniques: stereolithography (SL), multi-jet modelling (MJM), selective laser melting (SLM), laser sintering (LS) and fused deposition modelling (FDM) were used to manufacture a number of miniaturised reactors which were tested using a range of organic and inorganic reactions.

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