In situ 4D tomography image analysis framework to follow sintering within 3D-printed glass scaffolds.

We propose a novel image analysis framework to automate analysis of X-ray microtomography images of sintering ceramics and glasses, using open-source toolkits and machine learning. Additive manufacturing (AM) of glasses and ceramics usually requires sintering of green bodies. Sintering causes shrinkage, which presents a challenge for controlling the metrology of the final architecture. Therefore, being able to monitor sintering in 3D over time (termed 4D) is important when developing new porous ceramics or glasses. Synchrotron X-ray tomographic imaging allows in situ, real-time capture of the sintering process at both micro and macro scales using a furnace rig, facilitating 4D quantitative analysis of the process. The proposed image analysis framework is capable of tracking and quantifying the densification of glass or ceramic particles within multiple volumes of interest (VOIs) along with structural changes over time using 4D image data. The framework is demonstrated by 4D quantitative analysis of bioactive glass ICIE16 within a 3D-printed scaffold. Here, densification of glass particles within 3 VOIs were tracked and quantified along with diameter change of struts and interstrut pore size over the 3D image series, delivering new insights on the sintering mechanism of ICIE16 bioactive glass particles in both micro and macro scales.

Laser-Guided Corrosion Control: A New Approach to Tailor the Degradation of Mg-Alloys.

Despite corrosion being commonly seen as a problem to be avoided, applications such as batteries or biodegradable implants do benefit from corrosion-like phenomena. However, current strategies address corrosion control from a global perspective for a whole component, without considering local adaptations to functionality specifications or inhomogeneous environments. Here, a novel concept is presented: the local control and guidance of corrosion through a laser surface treatment. Immersion tests in saline solution of AZ31 magnesium alloy samples show degradation rates reduced up to 15 times with the treatment, owing to a fast passivation after the induced microstructural modifications. By controlling the treatment conditions, the degradation can be restricted to delimited regions and driven towards specific directions. The applicability of the method for the design of tailored degradation biomedical implants is demonstrated and uses for cathodic protection systems and batteries can also be anticipated.

Rheological Characterization of Biomaterials Directs Additive Manufacturing of Strontium-Substituted Bioactive Glass/Polycaprolactone Microfibers.

Additive manufacturing via melt electrowriting (MEW) can create ordered microfiber scaffolds relevant for bone tissue engineering; however, there remain limitations in the adoption of new printing materials, especially in MEW of biomaterials. For example, while promising composite formulations of polycaprolactone with strontium-substituted bioactive glass have been processed into large or disordered fibres, from what is known, biologically-relevant concentrations (>10 wt%) have never been printed into ordered microfibers using MEW. In this study, rheological characterization is used in combination with a predictive mathematical model to optimize biomaterial formulations and MEW conditions required to extrude various PCL and PCL/SrBG biomaterials to create ordered scaffolds. Previously, MEW printing of PCL/SrBG composites with 33 wt% glass required unachievable extrusion pressures. The composite formulation is modified using an evaporable solvent to reduce viscosity 100-fold to fall within the predicted MEW pressure, temperature, and voltage tolerances, which enabled printing. This study reports the first fabrication of reproducible, ordered high-content bioactive glass microfiber scaffolds by applying predictive modeling.

Osteogenic potential of sol-gel bioactive glasses containing manganese.

Bioactive glasses (BGs) are widely used for bone regeneration, and allow the incorporation of different ions with therapeutic properties into the glass network. Amongst the different ions with therapeutic benefits, manganese (Mn) has been shown to influence bone metabolism and activate human osteoblasts integrins, improving cell adhesion, proliferation and spreading. Mn has also been incorporated into bioceramics as a therapeutic ion for improved osteogenesis. Here, up to 4.4 mol% MnO was substituted for CaO in the 58S composition (60 mol% SiO2, 36 mol% CaO, 4 mol% P2O5) and its effects on the glass properties and capability to influence the osteogenic differentiation were evaluated. Mn-containing BGs with amorphous structure, high specific surface area and nanoporosity were obtained. The presence of Mn2+ species was confirmed by X-ray photoelectron spectroscopy (XPS). Mn-containing BGs presented no cytotoxic effect on human mesenchymal stem cells (hMSCs) and enabled sustained ion release in culture medium. hMSCs osteogenic differentiation stimulation and influence on the mineralisation process was also confirmed through the alkaline phosphatase (ALP) activity, and expression of osteogenic differentiation markers, such as collagen type I, osteopontin and osteocalcin, which presented higher expression in the presence of Mn-containing samples compared to control. Results show that the release of manganese ions from bioactive glass provoked human mesenchymal stem cell (hMSC) differentiation down a bone pathway, whereas hMSCs exposed to the Mn-free glass did not differentiate. Mn incorporation offers great promise for obtaining glasses with superior properties for bone tissue regeneration.

Construction of DNAzyme-Encapsulated Fibermats Using the Precursor Network Polymer of Poly(γ-glutamate) and 4-Glycidyloxypropyltrimethoxysilane.

Here, we developed functional nucleic acid (FNA)-encapsulated electrospun fibermats. To facilitate stable FNA encapsulation in the γ-PGA/GPTMS fibermats, we used the FNA as an FNA/streptavidin complex, and as a representative FNA, we selected a DNAzyme, the DNA/hemin complex, which is composed of G-quadraplex-forming single-stranded DNA and hemin and exhibits oxidation activity with the aid of a cocatalyst, H2O2. Scanning electron microscopy and Fourier-transform infrared spectroscopy measurements revealed that encapsulation of the DNA/hemin complex (∼1 wt % against the γ-PGA/GPTMS hybrid) in the nanofibers of the γ-PGA/GPTMS fibermats did not affect the structure of the original nanofibers. However, because a unique MW-dependent molecular permeability originated from the 3D network structure of the γ-PGA/GPTMS hybrid, low-MW substrates such as 4-aminoantipyrine, N-ethyl-N-(2-hydroxy-3-sulfopropyl)-3-methylaniline, and luminol were able to reach the encapsulated DNA/hemin complex by permeating to the inside of the nanofibers from an immersion buffer and then underwent catalytic oxidation. Conversely, nucleases, which are proteins featuring high MWs (>5 kDa), could not penetrate the γ-PGA/GPTMS nanofibers, and the encapsulated DNA/hemin complex was therefore effectively protected against nuclease digestion. Thus, encapsulating FNAs on the inside of the nanofibers of fibermats offers clear advantages for the practical application of FNAs in sensors and drugs, particularly for use in the in vivo circumstances.

Effect of Comonomers on Physical Properties and Cell Attachment to Silica-Methacrylate/Acrylate Hybrids for Bone Substitution.

Hybrids with a silica network covalently bonded to a polymer are promising materials for bone repair. Previous work on synthesizing methyl methacrylate (MMA) based copolymers by reversible addition-fragmentation chain transfer (RAFT) polymerization gives high tailorability of mechanical properties since sophisticated polymer structures can be designed. However, more flexible hybrids would be beneficial. Here, n-butyl methacrylate (BMA) and methyl acrylate (MA) based hybrids are produced. Unlike MMA, BMA and MA hybrids do not show plastic deformation, and BMA hybrid has strain to failure of 33%. Although the new hybrids are more flexible, preosteoblast cells do not adhere on their surfaces, due to higher hydrophobicity and lower stiffness. Comonomer choice is crucial for bone regenerative hybrids.

A correlative imaging based methodology for accurate quantitative assessment of bone formation in additive manufactured implants.

A correlative imaging methodology was developed to accurately quantify bone formation in the complex lattice structure of additive manufactured implants. Micro computed tomography (µCT) and histomorphometry were combined, integrating the best features from both, while demonstrating the limitations of each imaging modality. This semi-automatic methodology registered each modality using a coarse graining technique to speed the registration of 2D histology sections to high resolution 3D µCT datasets. Once registered, histomorphometric qualitative and quantitative bone descriptors were directly correlated to 3D quantitative bone descriptors, such as bone ingrowth and bone contact. The correlative imaging allowed the significant volumetric shrinkage of histology sections to be quantified for the first time (~15 %). This technique demonstrated the importance of location of the histological section, demonstrating that up to a 30 % offset can be introduced. The results were used to quantitatively demonstrate the effectiveness of 3D printed titanium lattice implants.

Toward Hybrid Materials: Group Transfer Polymerization of 3-(Trimethoxysilyl)propyl Methacrylate.

In this study, the group transfer polymerization (GTP) of the functional monomer 3-(trimethoxysilyl)propyl methacrylate (TMSPMA) is reported to produce polymers of different architectures and topologies. TMSPMA is successfully polymerized and copolymerized with GTP to produce well-defined (co)polymers that can be used to fabricate functional hybrid materials like hydrogels and films.

RAFT Polymerization of N-[3-(Trimethoxysilyl)-propyl]acrylamide and Its Versatile Use in Silica Hybrid Materials.

Reversible addition-fragmentation chain transfer (RAFT) polymerization and characterization of an alkoxysilane acrylamide monomer using a trithiocarbonate chain transfer agent are described. Poly(N-[3-(trimethoxysilyl)propyl]acrylamide) (PTMSPAA) homopolymers are obtained with good control over the polymerization. A linear increase in the molecular weight is observed whereas the polydispersity values do not exceed 1.2 regardless of the monomer conversion. Moreover, PTMSPAA is used as a macro-RAFT agent to polymerize N-isopropylacrylamide (NIPAM). By varying the degree of polymerization of NIPAM within the block copolymer, different sizes of thermoresponsive particles are obtained. These particles are stabilized by the condensation of the alkoxysilane moieties of the polymers. Furthermore, a co-network of silica and PTMSPAA is prepared using the sol-gel process. After drying, transparent mesoporous hybrids are obtained with a surface area of up to 400 m(2) g(-1).

A multinuclear solid state NMR spectroscopic study of the structural evolution of disordered calcium silicate sol-gel biomaterials.

Disordered sol-gel prepared calcium silicate biomaterials show significant, composition dependent ability to bond with bone. Bone bonding is attributed to rapid hydroxycarbonate apatite (HCA) formation on the glass surface after immersion in body fluid (or implantation). Atomic scale details of the development of the structure of (CaO)x(SiO2)1-x (x = 0.2, 0.3 and 0.5) under heat treatment and subsequent dissolution in simulated body fluid (SBF) are revealed through a multinuclear solid state NMR approach using one-dimensional (17)O, (29)Si, (31)P and (1)H. Central to this study is the combination of conventional static and magic angle spinning (MAS) and two-dimensional (2D) triple quantum (3Q) (17)O NMR experiments that can readily distinguish and quantify the bridging (BOs) and non-bridging (NBOs) oxygens in the silicate network. Although soluble calcium is present in the sol, the (17)O NMR results reveal that the sol-gel produced network structure is initially dominated by BOs after gelation, aging and drying (e.g. at 120 °C), indicating a nanoscale mixture of the calcium salt and a predominantly silicate network. Only once the calcium salt is decomposed at elevated temperatures do the Ca(2+) ions become available to break BO. Apatite forming ability in SBF depends strongly on the surface OH and calcium content. The presence of calcium aids HCA formation via promotion of surface hydration and the ready availability of Ca(2+) ions. (17)O NMR shows the rapid loss of NBOs charge balanced by calcium as it is leached into the SBF. The formation of nanocrystalline, partially ordered HCA can be detected via(31)P NMR. This data indicates the importance of achieving the right balance of BO/NBO for optimal biochemical response and network properties.

A structural and physical study of sol-gel methacrylate-silica hybrids: intermolecular spacing dictates the mechanical properties.

Sol-gel hybrids are inorganic/organic co-networks with nanoscale interactions between the components leading to unique synergistic mechanical properties, which can be tailored, via a selection of the organic moiety. Methacrylate based polymers present several benefits for class II hybrids (which exhibit formal covalent bonding between the networks) as they introduce great versatility and can be designed with a variety of chemical side-groups, structures and morphologies. In this study, the effect of high cross-linking density polymers on the structure-property relationships of hybrids generated using poly(3-trimethoxysilylpropyl methacrylate) (pTMSPMA) and tetraethyl orthosilicate (TEOS) was investigated. The complexity and fine scale of the co-network interactions requires the development of new analytical methods to understand how network evolution dictates the wide-ranging mechanical properties. Within this work we developed data manipulation techniques of acoustic-AFM and solid state NMR output that provide new approaches to understand the influence of the network structure on the macroscopic elasticity. The concentration of pTMSPMA in the silica sol affected the gelation time, ranging from 2 h for a hybrid made with 75 wt% inorganic with pTMSPMA at 2.5 kDa, to 1 minute for pTMSPMA with molecular weight of 30 kDa without any TEOS. A new mechanism of gelation was proposed based on the different morphologies derived by AC-AFM observations. We established that the volumetric density of bridging oxygen bonds is an important parameter in structure/property relationships in SiO2 hybrids and developed a method for determining it from solid state NMR data. The variation in the elasticity of pTMSPMA/SiO2 hybrids originated from pTMSPMA acting as a molecular spacer, thus decreasing the volumetric density of bridging oxygen bonds as the inorganic to organic ratio decreased.

A unified in vitro evaluation for apatite-forming ability of bioactive glasses and their variants.

The aim of this study was to propose and validate a new unified method for testing dissolution rates of bioactive glasses and their variants, and the formation of calcium phosphate layer formation on their surface, which is an indicator of bioactivity. At present, comparison in the literature is difficult as many groups use different testing protocols. An ISO standard covers the use of simulated body fluid on standard shape materials but it does not take into account that bioactive glasses can have very different specific surface areas, as for glass powders. Validation of the proposed modified test was through round robin testing and comparison to the ISO standard where appropriate. The proposed test uses fixed mass per solution volume ratio and agitated solution. The round robin study showed differences in hydroxyapatite nucleation on glasses of different composition and between glasses of the same composition but different particle size. The results were reproducible between research facilities. Researchers should use this method when testing new glasses, or their variants, to enable comparison between the literature in the future.

Poly(γ-glutamic acid)/silica hybrids with calcium incorporated in the silica network by use of a calcium alkoxide precursor.

Current materials used for bone regeneration are usually bioactive ceramics or glasses. Although they bond to bone, they are brittle. There is a need for new materials that can combine bioactivity with toughness and controlled biodegradation. Sol-gel hybrids have the potential to do this through their nanoscale interpenetrating networks (IPN) of inorganic and organic components. Poly(γ-glutamic acid) (γ-PGA) was introduced into the sol-gel process to produce a hybrid of γ-PGA and bioactive silica. Calcium is an important element for bone regeneration but calcium sources that are used traditionally in the sol-gel process, such as Ca salts, do not allow Ca incorporation into the silicate network during low-temperature processing. The hypothesis for this study was that using calcium methoxyethoxide (CME) as the Ca source would allow Ca incorporation into the silicate component of the hybrid at room temperature. The produced hybrids would have improved mechanical properties and controlled degradation compared with hybrids of calcium chloride (CaCl2 ), in which the Ca is not incorporated into the silicate network. Class II hybrids, with covalent bonds between the inorganic and organic species, were synthesised by using organosilane. Calcium incorporation in both the organic and inorganic IPNs of the hybrid was improved when CME was used. This was clearly observed by using FTIR and solid-state NMR spectroscopy, which showed ionic cross-linking of γ-PGA by Ca and a lower degree of condensation of the Si species compared with the hybrids made with CaCl2 as the Ca source. The ionic cross-linking of γ-PGA by Ca resulted in excellent compressive strength and reduced elastic modulus as measured by compressive testing and nanoindentation, respectively. All hybrids showed bioactivity as hydroxyapatite (HA) was formed after immersion in simulated body fluid (SBF).

3D printed hybrid scaffolds do not induce adverse inflammation in mice and direct human BM-MSC chondrogenesis in vitro.

Biomaterials that can improve the healing of articular cartilage lesions are needed. To address this unmet need, we developed novel 3D printed silica/poly(tetrahydrofuran)/poly(ε-caprolactone) (SiO2/PTHF/PCL-diCOOH) hybrid scaffolds. Our aim was to carry out essential studies to advance this medical device towards functional validation in pre-clinical trials. First, we show that the chemical composition, microarchitecture and mechanical properties of these scaffolds were not affected by sterilisation with gamma irradiation. To evaluate the systemic and local immunogenic reactivity of the sterilised 3D printed hybrid scaffolds, they were implanted subcutaneously into Balb/c mice. The scaffolds did not trigger a systemic inflammatory response over one week of implantation. The interaction between the host immune system and the implanted scaffold elicited a local physiological reaction with infiltration of mononuclear cells without any signs of a chronic inflammatory response. Then, we investigated how these 3D printed hybrid scaffolds direct chondrogenesis in vitro. Human bone marrow-derived mesenchymal stem/stromal cells (hBM-MSCs) seeded within the 3D printed hybrid scaffolds were cultured under normoxic or hypoxic conditions, with or without chondrogenic supplements. Chondrogenic differentiation assessed by both gene expression and protein production analyses showed that 3D printed hybrid scaffolds support hBM-MSC chondrogenesis. Articular cartilage-specific extracellular matrix deposition within these scaffolds was enhanced under hypoxic conditions (1.7 or 3.7 fold increase in the median of aggrecan production in basal or chondrogenic differentiation media). Our findings show that 3D printed SiO2/PTHF/PCL-diCOOH hybrid scaffolds have the potential to support the regeneration of cartilage tissue.

Ovine Mesenchymal Stem Cell Chondrogenesis on a Novel 3D-Printed Hybrid Scaffold In Vitro.

This study evaluated the use of silica/poly(tetrahydrofuran)/poly(ε-caprolactone) (SiO2/PTHF/PCL-diCOOH) 3D-printed scaffolds, with channel sizes of either 200 (SC-200) or 500 (SC-500) µm, as biomaterials to support the chondrogenesis of sheep bone marrow stem cells (oBMSC), under in vitro conditions. The objective was to validate the potential use of SiO2/PTHF/PCL-diCOOH for prospective in vivo ovine studies. The behaviour of oBMSC, with and without the use of exogenous growth factors, on SiO2/PTHF/PCL-diCOOH scaffolds was investigated by analysing cell attachment, viability, proliferation, morphology, expression of chondrogenic genes (RT-qPCR), deposition of aggrecan, collagen II, and collagen I (immunohistochemistry), and quantification of sulphated glycosaminoglycans (GAGs). The results showed that all the scaffolds supported cell attachment and proliferation with upregulation of chondrogenic markers and the deposition of a cartilage extracellular matrix (collagen II and aggrecan). Notably, SC-200 showed superior performance in terms of cartilage gene expression. These findings demonstrated that SiO2/PTHF/PCL-diCOOH with 200 µm pore size are optimal for promoting chondrogenic differentiation of oBMSC, even without the use of growth factors.

Epoxide opening versus silica condensation during sol-gel hybrid biomaterial synthesis.

Hybrid organic-inorganic solids represent an important class of engineering materials, usually prepared by sol-gel processes by cross-reaction between organic and inorganic precursors. The choice of the two components and control of the reaction conditions (especially pH value) allow the synthesis of hybrid materials with novel properties and functionalities. 3-Glycidoxypropyltrimethoxysilane (GPTMS) is one of the most commonly used organic silanes for hybrid-material fabrication. Herein, the reactivity of GPTMS in water at different pH values (pH 2-11) was deeply investigated for the first time by solution-state multinuclear NMR spectroscopic and mass spectrometric analysis. The extent of the different and competing reactions that take place as a function of the pH value was elucidated. The NMR spectroscopic and mass spectrometric data clearly indicate that the pH value determines the kinetics of epoxide hydrolysis versus silicon condensation. Under slighly acidic conditions, the epoxy-ring hydrolysis is kinetically more favourable than the formation of the silica network. In contrast, under basic conditions, silicon condensation is the main reaction that takes place. Full characterisation of the formed intermediates was carried out by using NMR spectroscopic and mass spectrometric analysis. These results indicate that strict control of the pH values allows tuning of the reactivity of the organic and inorganic moities, thus laying the foundations for the design and synthesis of sol-gel hybrid biomaterials with tuneable properties.

Cotton wool-like poly(lactic acid)/vaterite composite scaffolds releasing soluble silica for bone tissue engineering.

Cotton wool-like poly(L-lactic acid) and siloxane-doped vaterite (SiV) composite scaffolds were prepared with a modified electrospinning system for bone tissue engineering applications. The effects of changing the SiV content in the materials from 10 to 30 wt% on elasticity and the ability to release calcium ions and soluble silica were evaluated. The elasticity of the cotton wool-like composites was almost the same as that of the PLLA from the results of compressibility and recovery tests. The materials released calcium ions for more than 56 days and soluble silica for 28-56 days in a tris buffer solution (pH 7.4). Mouse osteoblast-like cells (MC3T3-E1 cells) were cultured on/in the cotton wool-like materials or the fibremats out of the same composite materials as that used for the cotton wool-like materials. The cells penetrated into and proliferated inside the cotton wool-like materials, although they mainly adhered on the fibremat surface.

Bioactive glass-based organic/inorganic hybrids: an analysis of the current trends in polymer design and selection.

Bioactive glass-based organic/inorganic hybrids are a family of materials holding great promise in the biomedical field. Developed from bioactive glasses following recent advances in sol-gel and polymer chemistry, they can overcome many limitations of traditional composites typically used in bone repair and orthopedics. Thanks to their unique molecular structure, hybrids are often characterized by synergistic properties that go beyond a mere combination of their two components; it is possible to synthesize materials with a wide variety of mechanical and biological properties. The polymeric component, in particular, can be tailored to prepare tough, load-bearing materials, or rubber-like elastomers. It can also be a key factor in the determination of a wide range of interesting biological properties. In addition, polymers can also be used within hybrids as carriers for therapeutic ions (although this is normally the role of silica). This review offers a brief look into the history of hybrids, from the discovery of bioactive glasses to the latest developments, with a particular emphasis on polymer design and chemistry. First the benefits and limitations of hybrids will be discussed and compared with those of alternative approaches (for instance, nanocomposites). Then, key advances in the field will be presented focusing on the polymeric component: its chemistry, its physicochemical and biological advantages, its drawbacks, and selected applications. Comprehensive tables summarizing all the polymers used to date to fabricate sol-gel hybrids for biomedical applications are also provided, to offer a handbook of all the available candidates for hybrid synthesis. In addition to the current trends, open challenges and possible avenues of future development are proposed.

Nanocomposite Hydrogels with Polymer Grafted Silica Nanoparticles, Using Glucose Oxidase.

Nanocomposite hydrogels offer remarkable potential for applications in bone tissue engineering. They are synthesized through the chemical or physical crosslinking of polymers and nanomaterials, allowing for the enhancement of their behaviour by modifying the properties and compositions of the nanomaterials involved. However, their mechanical properties require further enhancement to meet the demands of bone tissue engineering. Here, we present an approach to improve the mechanical properties of nanocomposite hydrogels by incorporating polymer grafted silica nanoparticles into a double network inspired hydrogel (gSNP Gels). The gSNP Gels were synthesised via a graft polymerization process using a redox initiator. gSNP Gels were formed by grafting 2-acrylamido-2-methylpropanesulfonic acid (AMPS) as the first network gel followed by a sequential second network acrylamide (AAm) onto amine functionalized silica nanoparticles (ASNPs). We utilized glucose oxidase (GOx) to create an oxygen-free atmosphere during polymerization, resulting in higher polymer conversion compared to argon degassing. The gSNP Gels showed excellent compressive strengths of 13.9 ± 5.5 MPa, a strain of 69.6 ± 6.4%, and a water content of 63.4% ± 1.8. The synthesis technique demonstrates a promising approach to enhance the mechanical properties of hydrogels, which can have significant implications for bone tissue engineering and other soft tissue applications.

The effect of Si species released from bioactive glasses on cell behaviour: A quantitative review.

Despite over 50 years of silicate bioactive glass (SBG) research, commercial success, and 6000+ published articles, there remains a lack of understanding of how soluble silicate (Si) species released from SBGs influences cellular responses. Using a systematic approach, this article quantitatively compares the in vitro responses of cells to SBG dissolution products reported in the literature and determines if there is a Si concentration ([Si]) dependent effect on cell behaviour. Cell behavioural responses to SBGs [Si] in dissolution products included metabolic activity (reported in 52 % of articles), cell number (24 %), protein production (22 %), gene expression (22 %) and biomineralization (24 %). There was a difference in the [Si] reported to cause increased (desirable) cellular responses (median = 30.2 ppm) compared to the [Si] reported to cause decreased (undesirable) cellular responses (median = 52.0 ppm) (P ≤ 0.001). The frequency of undesirable outcomes increased with increasing [Si], with ∼3 times more negative outcomes reported above 52 ppm. We also investigated the effect of [Si] on specific cellular outcomes (e.g., metabolic activity, angiogenesis, osteogenesis), if cell type/species influenced these responses and the impact of other ions (Ca, P, Na) within the SBG dissolution media on cell behaviour. This review has, for the first time, quantitatively compared the cellular responses to SBGs from the literature, providing a quantitative overview of SBG in vitro practices and presents evidence of a range of [Si] where desirable cellular responses may be more likely (30-52 ppm). This review also demonstrates the need for greater standardisation of in vitro methodological approaches and recommends some minimum reporting standards. STATEMENT OF SIGNIFICANCE: This systematic review investigates the relationship between the concentration of Si released from Si-bioactive glasses (SBG) and in vitro cellular responses. Si releasing materials continue to be of considerable scientific, commercial, and medical interest (with 1500+ articles published in the last 3 years) but there is considerable variation in the reported biologically effective Si concentrations and on the importance of Si on cell behaviour. Despite the variation in methodological approaches, this article demonstrated statistical commonalities in the Si concentrations that cause desirable and undesirable cellular behaviours, suggesting a window where positive cellular outcomes are more likely. This review also provides a quantitative analysis of in vitro practices within the bioactive glass field and highlights the need for greater standardisation.