Bioactive Glasses Modulate Anticancer Activity and Other Polyphenol-Related Properties of Polyphenol-Loaded PCL/Bioactive Glass Composites.

In this work, bioactive glass (BG) particles obtained by three different methods (melt-quenching, sol-gel, and sol-gel-EISA) were used as modifiers of polyphenol-loaded PCL-based composites. The composites were loaded with polyphenolic compounds (PPh) extracted from sage (Salvia officinalis L.). It was hypothesized that BG particles, due to their different textural properties (porosity, surface area) and surface chemistry (content of silanol groups), would act as an agent to control the release of polyphenols from PCL/BG composite films and other significant properties associated with and affected by the presence of PPh. The polyphenols improved the hydrophilicity, apatite-forming ability, and mechanical properties of the composites and provided antioxidant and anticancer activity. As the BG particles had different polyphenol-binding capacities, they modulated the kinetics of polyphenol release from the composites and the aforementioned properties to a great extent. Importantly, the PPh-loaded materials exhibited multifaceted and selective anticancer activity, including ROS-mediated cell cycle arrest and apoptosis of osteosarcoma (OS) cells (Saos-2) via Cdk2-, GADD45G-, and caspase-3/7-dependent pathways. The materials showed a cytotoxic and antiproliferative effect on cancerous osteoblasts but not on normal human osteoblasts. These results suggest that the composites have great potential as biomaterials for treating bone defects, particularly following surgical removal of OS tumors.

In Vitro and In Vivo Studies of Antibacterial Coatings on Titanium Alloy Implants for Veterinary Application.

The aim of this work was the evaluation of biological properties of hybrid coatings modified with Ag, Cu, and Zn nanoparticles (NPs) applied on TPLO medical implants by the sol-gel process. The implant coatings enriched with various concentrations of metallic NPs were investigated in the in vitro bactericidal efficacy tests against Gram+ and Gram- bacteria and pathogenic yeast. Next, the designed materials were tested on human osteosarcoma cell lines. The cells adhesion, proliferation, viability, and differentiation were investigated. The cell growth wasevaluated using SEM, and the metallic ion release was measured. The results revealed that the NPs concentration in the hybrid layers decreased with the incubation time. In the last stage, the implants were tested in vivo on six canine patients. Three months after the operation, the radiological evaluation of the performed anastomosis was carried out as well as the histopathological evaluation of tissue regeneration. The strongest bactericidal efficacy was observed for the layers containing AgNPs. Along with an increased concentration of metallic additives, a growing toxic effect was clearly observed. The most pronounced toxic effect was especially evident with the AgNPs concentration exceeding 1 mol %. In all the operated patients, no deviations were found during the follow-up examinations in the postoperative period. The low dose of AgNPs in the hybrid layer facilitated the tissue healing process. It was proven that silver nanoparticles may accelerate the bone healing process. The correct tissue reparation was observed.

Polyphenol-Enriched Composite Bone Regeneration Materials: A Systematic Review of In Vitro Studies.

One of the possible alternatives for creating materials for the regeneration of bone tissue supporting comprehensive reconstruction is the incorporation of active substances whose controlled release will improve this process. This systematic review aimed to identify and synthesize in vitro studies that assess the suitability of polyphenolics as additives to polymer-ceramic composite bone regeneration materials. Data on experimental studies in terms of the difference in mechanical, wettability, cytocompatibility, antioxidant and anti-inflammatory properties of materials were synthesized. The obtained numerical data were compiled and analyzed in search of percentage changes of these parameters. The results of the systematic review were based on data from forty-six studies presented in nineteen articles. The addition of polyphenolic compounds to composite materials for bone regeneration improved the cytocompatibility and increased the activity of early markers of osteoblast differentiation, indicating a high osteoinductive potential of the materials. Polyphenolic compounds incorporated into the materials presumably give them high antioxidant properties and reduce the production of reactive oxygen species in macrophage cells, implying anti-inflammatory activity. The evidence was limited by the number of missing data and the heterogeneity of the data.

Newly crosslinked chitosan- and chitosan-pectin-based hydrogels with high antioxidant and potential anticancer activity.

Monoaldehydes, due to natural origin and therapeutic activity, have attracted great attention for their ability to crosslink chitosan hydrogels for biomedical applications. However, most studies have focused on single-component hydrogels. In this work, chitosan-based hydrogels, crosslinked for the first time with 2,3,4-trihydroxybenzaldehyde (THBA), were modified with pectin (PC), bioactive glass (BG), and rosmarinic acid (RA). All of these were not only involved in the crosslinking, but also modulated properties or imparted completely new ones. THBA functioned as a crosslinker, resulting in improved mechanical properties, high swelling capacity and delayed degradation and also imparted high antioxidant activity and antiproliferative effect on cancer cells without cytotoxicity for normal cells. Hydrogels containing PC showed enhanced mechanical strength, while the combination with BG gave improved stability in PBS. All hydrogels modified with BG exhibited the ability to mineralise in SBF. The addition of RA enhanced antioxidant and anticancer activities and promoting the mineralisation process.

The Changes in Bioactive Compounds and Antioxidant Activity of Chia (Salvia hispanica L.) Herb under Storage and Different Drying Conditions: A Comparison with Other Species of Sage.

Studies on herb chia (Salvia hispanica L.) are very limited. Therefore, the aim of this study was to assess how different drying methods and periods of storage affect the bioactive properties of the herb Salvia hispanica and to compare it with other species of sage (Salvia officinalis L. and Salvia sclarea L.). In fresh herbs, directly after drying (freeze-drying, natural drying, and drying at 30, 40, and 50 °C), and after storage (3, 6, and 12 months), the following analyses were performed: content of total carotenoids and total polyphenols, polyphenol profile (including 25 compounds), and antioxidant activity. Additionally, the basic chemical compositions of the herbs were analyzed. To the best of our knowledge, the content of total carotenoids and the quantitative polyphenol profile in Salvia hispanica and Salvia sclarea were evaluated for the first time. The obtained results showed that the barely investigated herb Salvia hispanica is rich in polyphenolic compounds and shows high antioxidant activity. In all the tested species, rosmarinic acid was the most abundant polyphenolic compound. The use of different drying methods allowed us to determine that freeze-drying was the most effective for preserving polyphenols and carotenoids. Long-term storage up to 12 months resulted in a gradual reduction in antioxidant activity and in the content of polyphenols and carotenoids.

Chemical Compounds Released from Specific Osteoinductive Bioactive Materials Stimulate Human Bone Marrow Mesenchymal Stem Cell Migration.

In this work, a poly(L-lactide-co-glycolide) (PLGA)-based composite was enriched with one of the following sol-gel bioactive glasses (SBG) at 50 wt.%: A1-40 mol% SiO2, 60 mol% CaO, CaO/SiO2 ratio of 1.50; S1-80 mol% SiO2, 20 mol% CaO, CaO/SiO2 ratio of 0.25; A2-40 mol% SiO2, 54 mol% CaO, 6 mol% P2O5, CaO/SiO2 ratio of 1.35; S2-80 mol% SiO2,16 mol% CaO, 4 mol% P2O5, CaO/SiO2 ratio of 0.20. The composites and PLGA control sheets were then soaked for 24 h in culture media, and the obtained condition media (CM) were used to treat human bone marrow stromal cells (hBMSCs) for 72 h. All CMs from the composites increased ERK 1/2 activity vs. the control PLGA CM. However, expressions of cell migration-related c-Fos, osteopontin, matrix metalloproteinase-2, C-X-C chemokine receptor type 4, vascular endothelial growth factor, and bone morphogenetic protein 2 were significantly increased only in cells treated with the CM from the A1/PLGA composite. This CM also significantly increased the rate of human BMSC migration but did not affect cell metabolic activity. These results indicate important biological markers that are upregulated by products released from the bioactive composites of a specific chemical composition, which may eventually prompt osteoprogenitor cells to colonize the bioactive material and accelerate the process of tissue regeneration.

Modification of TiAlV Alloys with Hybrid Layers Containing Metallic Nanoparticles Obtained by the Sol-Gel Method: Surface and Structural Properties.

The aim of the work was to obtain hybrid coatings containing silver, copper, and zinc nanoparticles on the TiAlV medical alloy via a sol-gel process. The developed layers were designed to bring about a bactericidal and fungicidal effect, as well as for protection against surgical scratches during the implantation of implants used in veterinary medicine. In this work, the authors focused on evaluating the microstructure (SEM + EDS); the structure (XRD, FTIR); and the surface properties, such as wettability, free surface energy, and roughness of layers with various concentrations of metallic nanoparticles (2 and 5 mol %). Our results confirmed that the sol-gel method enables the easy manufacturing of hybrid layers endowed with different porosity values as well as various shapes and sizes of metallic nanoparticles. A higher concentration of nanoparticles was observed on the surface containing 5 mol % of metallic salts. The highest degree of homogeneity was obtained for the layers containing silver nanoparticles. In addition, the silver nanoparticles were round and had the smallest dimensions, even below 20 nm. The FTIR and XRD structural studies confirmed the presence of an organosilicon matrix containing all three types of the metallic particles. We conclude that the higher concentration of nanoparticles influenced the alloy surface parameters.

Molecular Indicators of Biomaterials Osteoinductivity - Cell Migration, BMP Production and Signalling Turns a Key.

In this work we dissected the osteoinductive properties of selected, PLGA-based scaffolds enriched with gel-derived bioactive glasses (SBGs) of either binary SiO2-CaO or ternary SiO2-CaO-P2O5 system, differing in CaO/SiO2 ratio (i.e. high -or low-calcium SBGs). To assess the inherent ability of the scaffolds to induce osteogenesis of human bone marrow stromal cells (BMSC), the study was designed to avoid any osteogenic stimuli beyond the putative osteogenic SBG component of the studied scaffolds. The bioactivity and porosity of scaffolds were confirmed by SBF test and porosimetry. Condition media (CM) from BMSC-loaded scaffolds exhibited increased Ca and decreased P content corresponding to SBGs CaO/SiO2 ratio, whereas Si content was relatively stable and overall lower in CM from scaffolds containing binary SBGs. CM from cell-loaded scaffolds containing high-calcium, binary SBGs promoted migration of BMSC and BMP-response in reporter osteoblast cell line. BMSC culture on these scaffolds or the ones containing ternary, low-calcium SBGs resulted in the activation of BMP-related signaling and expression of several osteogenic markers. Ectopic bone formation was induced by scaffolds containing binary SBGs, but high-calcium ones produced significantly more osteoid. Scaffolds containing ternary SBGs negatively influenced the expression of osteogenic transcription factors and Cx43, involved in cell-cell interactions. High-calcium scaffolds stimulated overall higher Cx43 expression. We believe the initial cell-cell communication may be crucial to induce and maintain osteogenesis and high BMP signaling on the studied scaffolds. The presented scaffolds' biological properties may also constitute new helpful markers to predict osteoinductive potential of other bioactive implant materials.

PCL and PCL/bioactive glass biomaterials as carriers for biologically active polyphenolic compounds: Comprehensive physicochemical and biological evaluation.

In this work, polymeric and bioactive glass (BG)-modified composite films were successfully loaded with polyphenols (PPh) extracted from sage. It was hypothesized that PPh, alone and in combination with BGs particles, would affect physicochemical and biological properties of the films. Furthermore, sol-gel-derived BG particles would serve as an agent for control the release of the polyphenolic compounds, and other important properties related to the presence of PPh. The results showed that polyphenolic compounds significantly modified numerous material properties and also acted as biologically active substances. On the one hand, PPh can be considered as plasticizers for PCL, on the other hand, they can act as coupling agent in composite materials, improving their mechanical performance. The presence of PPh in materials improved their hydrophilicity and apatite-forming ability, and also provided antioxidant activity. What is important is that the aforementioned properties and kinetics of PPh release can be modulated by the use of various concentrations of PPh, and by the modification of PCL matrix with sol-gel-derived BG particles, capable of binding PPh. The films containing the lowest concentration of PPh exhibited cytocompatibility, significantly increased alkaline phosphatase activity and the expression of bone extracellular matrix proteins (osteocalcin and osteopontin) in human normal osteoblasts, while they reduced intracellular reactive oxygen species production in macrophages. Furthermore, materials loaded with PPh showed antibiofilm properties against Gram positive and Gram negative bacteria. The results suggest that obtained materials represent potential multifunctional biomaterials for bone tissue engineering with a wide range of tunable properties.

Laser cladding of bioactive glass coating on pure titanium substrate with highly refined grain structure.

Free from toxic elements biomaterial potentially applicable for load bearing biomedical implants was obtained for the first time by laser cladding of S520 bioactive glass onto ultrafine-grained commercially pure titanium. The cladding process affected the refined structure of the substrate inducing martensitic transformation near its surface. The α' acicular martensite gradually passes into relatively large grains with increasing distance from the substrate surface, which subsequently are transformed into smaller grains of about 2 µm in diameter. Both the melted zone, where the martensite crystalline structure was found, and the HAZ are characterised by relatively lower hardness in comparison with that of the substrate core indicating increased ductility. Such a combination of zones with different properties may have a synergistic effect and is beneficial for the obtained biomaterial. A characteristic region in the form of about 3 µm width band was formed in the melted zone at about 10 µm below the titanium surface. The results of EDS analysis indicate that several glass elements moved into the region while the titanium content in the same area was decreased. High bioactivity of the coated S520 glass was revealed by in vitro testing with SBF solution and almost complete reduction of P concentration occurred after 14 days.

Antibacterial composite hybrid coatings of veterinary medical implants.

The aim of the work was to develop innovative antibacterial hybrid coatings applied on implants that are used for anastomoses of animals' long bones and to assess their physicochemical and biological properties. Plates made of the titanium alloy were covered with composite hybrid layers so as to protect the implant surface against corrosion and to enhance it with antibacterial properties.The hybrid coatings were obtained electrochemical oxidation and sol-gel. First, a layer of titanium nanotubes was applied to the implants surface through anodization. Next, the sol-gel method was used to create the second layer with silver nanoparticles. The microstructure examination of the materials was performed with the SEM. The phase composition analysis was carried out via the X-ray diffraction. The surface parameters (roughness, contact angle and free surface energy) were assessed. Biological studies of implants were conducted, including the analysis of degradation processes, cell response and bactericidal activity. The results confirmed that the hybrid antibacterial layers effectively protected the implant surface against scratches and corrosion and eliminated bacteria, which in turn would promote bone healing. The advantageous physicochemical and biological properties of metallic implants with hybrid composite layers raise hopes for their applicability in the veterinary treatment of bone fractures.

The role of CaO/SiO2 ratio and P2O5 content in gel-derived bioactive glass-polymer composites in the modulation of their bioactivity and osteoinductivity in human BMSCs.

We obtained a range of PLGA-based composites containing sol-gel bioactive glasses (SBG) from the SiO2-CaO and SiO2-CaO-P2O5 systems. Eight SBGs with different CaO/SiO2 ratios with and without P2O5 were incorporated at 50% w/w to PLGA matrix and structured into thin films suitable for cell culture. The SBG/PLGA composites were examined for their bioactivity in simulated body fluid (SBF), ion release profile in culture media with and without cells, and osteoinductivity in standard human bone marrow stromal cell (hBMSC) cultures without osteogenic growth factors. Our results indicate different surface activity of composites depending on the presence/absence of P2O5 in SBG composition. Furthermore, ion release profile to culture medium differed depending on the presence/absence of cells. Direct culture of hBMSC on the SiO2-CaO/PLGA composite films resulted in elevated Runx-2 mRNA, opposite to low Runx-2 mRNA levels on SiO2-CaO-P2O5/PLGA films. All studied composites increased Osx mRNA levels. Whereas some of SiO2-CaO/PLGA composites did not elevate BMP-2 and -6 proteins in hBMSC cultures, high levels of these BMPs were present in all cultures on SiO2-CaO-P2O5/PLGA composites. All composites induced BMP-related Tak1 signalling, whereas Smad1 signalling was restricted mostly to composites containing three-component SBGs. ALP activity of hBMSC and BMP-related luciferase activity of mouse BRITE cells differed depending on whether the cells were stimulated with culture medium conditioned with SBG/PLGA composites or the cells were directly cultured on the composite surfaces. Altogether, beyond bioactivity and osteoinductivity of SBG/PLGA composites, our studies show key differences in the biological response to both the bioactive material dissolution products and upon direct cell-material contacts.

Novel multicomponent organic-inorganic WPI/gelatin/CaP hydrogel composites for bone tissue engineering.

The present work focuses on the development of novel multicomponent organic-inorganic hydrogel composites for bone tissue engineering. For the first time, combination of the organic components commonly used in food industry, namely whey protein isolate (WPI) and gelatin from bovine skin, as well as inorganic material commonly used as a major component of hydraulic bone cements, namely α-TCP in various concentrations (0-70 wt%) was proposed. The results showed that α-TCP underwent incomplete transformation to calcium-deficient hydroxyapatite (CDHA) during preparation process of the hydrogels. Microcomputer tomography showed inhomogeneous distribution of the calcium phosphate (CaP) phase in the resulting composites. Nevertheless, hydrogels containing 30-70 wt% α-TCP showed significantly improved mechanical properties. The values of Young's modulus and the stresses corresponding to compression of a sample by 50% increased almost linearly with increasing concentration of ceramic phase. Incomplete transformation of α-TCP to CDHA during preparation process of composites provides them high reactivity in simulated body fluid during 14-day incubation. Preliminary in vitro studies revealed that the WPI/gelatin/CaP composite hydrogels support the adhesion, spreading, and proliferation of human osteoblast-like MG-63 cells. The WPI/gelatin/CaP composite hydrogels obtained in this work showed great potential for the use in bone tissue engineering and regenerative medicine applications.

Electrospun polycaprolactone membranes with Zn-doped bioglass for nasal tissues treatment.

In this work, composite membranes were investigated as future components of a layered implant for the reconstruction of nasal septum. Incorporation of zinc ions into nasal implants could potentially provide antibacterial properties to decrease or eliminate bacterial infections and subsequent surgical complications. Two types of membranes were prepared using an electrospinning method: PCL with bioglass and PCL with bioglass doped with Zn. The aim of this work was to investigate the influence of bioglass addition on the morphology, fiber diameter and composition of the membranes. The apatite-forming ability was examined in Simulated Body Fluid (SBF). The cytotoxicity of the membranes, ALP activity and in vitro mineralization were evaluated in cell culture. The mineralization and ALP activity was higher for polycaprolactone membranes modified with Zn doped bioglass than compared to pure PCL membranes or control material. The results proved that the presence of Zn2+ in the electrospun membranes = influence the osteogenic differentiation of cells.

Pectin-bioactive glass self-gelling, injectable composites with high antibacterial activity.

The present work focuses on the development of novel injectable, self-gelling composite hydrogels based on two types of low esterified amidated pectins from citrus peels and apple pomace. Sol-gel-derived, calcium-rich bioactive glass (BG) fillers in a particle form are applied as delivery vehicles for the release of Ca2+ ions to induce internal gelation of pectins. Composites were prepared by a relatively simple mixing technique, using 20% w/v BG particles of two different sizes (2.5 and <45 µm). Smaller particles accelerated pectin gelation slightly faster than bigger ones, which appears to result from the higher rate of Ca2+ ion release. µCT showed inhomogeneous distribution of the BG particles within the hydrogels. All composite hydrogels exhibited strong antibacterial activity against methicilin-resistant Staphylococcus aureus. The mineralization process of pectin-BG composite hydrogels occurred upon incubation in simulated body fluid for 28 days. In vitro studies demonstrated cytocompatibility of composite hydrogels with MC3T3-E1 osteoblastic cells.

Novel injectable gellan gum hydrogel composites incorporating Zn- and Sr-enriched bioactive glass microparticles: High-resolution X-ray microcomputed tomography, antibacterial and in vitro testing.

Mineralization of hydrogel biomaterials is desirable to improve their suitability as materials for bone regeneration. In this study, gellan gum (GG) hydrogels were formed by simple mixing of GG solution with bioactive glass microparticles of 45S5 composition, leading to hydrogel formation by ion release from the amorphous bioactive glass microparticles. This resulted in novel injectable, self-gelling composites of GG hydrogels containing 20% bioactive glass. Gelation occurred within 20 min. Composites containing the standard 45S5 bioactive glass preparation were markedly less stiff. X-ray microcomputed tomography proved to be a highly sensitive technique capable of detecting microparticles of diameter approximately 8 µm, that is, individual microparticles, and accurately visualizing the size distribution of bioactive glass microparticles and their aggregates, and their distribution in GG hydrogels. The widely used melt-derived 45S5 preparation served as a standard and was compared with a calcium-rich, sol-gel derived preparation (A2), as well as A2 enriched with zinc (A2Zn5) and strontium (A2Sr5). A2, A2Zn, and A2Sr bioactive glass particles were more homogeneously dispersed in GG hydrogels than 45S5. Composites containing all four bioactive glass preparations exhibited antibacterial activity against methicillin-resistant Staphylococcus aureus. Composites containing A2Zn5 and A2Sr5 bioactive glasses supported the adhesion and growth of osteoblast-like cells and were considerably more cytocompatible than 45S5. All composites underwent mineralization with calcium-deficient hydroxyapatite upon incubation in simulated body fluid. The extent of mineralization appeared to be greatest for composites containing A2Zn5 and 45S5. The results underline the importance of the choice of bioactive glass when preparing injectable, self-gelling composites.

Ectopic bone formation by gel-derived bioactive glass-poly-L-lactide-co-glycolide composites in a rabbit muscle model.

In this study we aimed to assess the in vivo osteoinductive properties of two composite scaffolds made of PLGA (poly-L-lactide-co-glycolide) and two types of gel-derived bioactive glasses, namely a high silica S2 bioactive glass (S2-PLGA composites) or high lime A2 bioactive glass (A2-PLGA composites). To achieve that, the potential of the composites to induce ectopic bone formation in a rabbit muscle has been examined along with the control PLGA scaffold. Cylinder-like scaffolds of 7 × 3 mm (width × height) were implanted into pouches created in the latissimus dorsi muscle of 18 New Zealand rabbits. The tissue sections were obtained at 6, 12 or 24 weeks post-surgery (six rabbits per each time point) and stained with hematoxylin-eosin. The process of wound healing, the formation of collagen-rich connective tissue and its transition to cartilage were examined by Sirius red and Alcian blue histological stainings. We also performed immunohistochemical verification of the presence of osteoblast- and osteoclast- like cells in the vicinity of the scaffolds. A typical foreign body reaction and wound healing process was observed for all implanted scaffolds. Osteoblast- and osteoclast-like cells were observed in the vicinity of the scaffolds as determined by the immunohistochemical staining for Osteocalcin, BMP-2 and Cathepsin K. Compared to plain PLGA scaffolds, numerous osteoblast-like cells were observed 12 weeks post implantation near the composites and the scaffolds gradually degraded as bone formation proceeded. S2-PLGA and A2-PLGA composites display osteoinductive properties in vivo. Furthermore, they are more effective at inducing ectopic bone formation in a rabbit muscle compared to plain PLGA. Thus these SBG-PLGA composite scaffolds have potential for clinical applications in dental and/or orthopedic-bone tissue engineering.

Biodegradable ceramic-polymer composites for biomedical applications: A review.

The present work focuses on the state-of-the-art of biodegradable ceramic-polymer composites with particular emphasis on influence of various types of ceramic fillers on properties of the composites. First, the general needs to create composite materials for medical applications are briefly introduced. Second, various types of polymeric materials used as matrices of ceramic-containing composites and their properties are reviewed. Third, silica nanocomposites and their material as well as biological characteristics are presented. Fourth, different types of glass fillers including silicate, borate and phosphate glasses and their effect on a number of properties of the composites are described. Fifth, wollastonite as a composite modifier and its effect on composite characteristics are discussed. Sixth, composites containing calcium phosphate ceramics, namely hydroxyapatite, tricalcium phosphate and biphasic calcium phosphate are presented. Finally, general possibilities for control of properties of composite materials are highlighted.

High-resolution synchrotron X-ray analysis of bioglass-enriched hydrogels.

Enrichment of hydrogels with inorganic particles improves their suitability for bone regeneration by enhancing their mechanical properties, mineralizability, and bioactivity as well as adhesion, proliferation, and differentiation of bone-forming cells, while maintaining injectability. Low aggregation and homogeneous distribution maximize particle surface area, promoting mineralization, cell-particle interactions, and homogenous tissue regeneration. Hence, determination of the size and distribution of particles/particle agglomerates in the hydrogel is desirable. Commonly used techniques have drawbacks. High-resolution techniques (e.g., SEM) require drying. Distribution in the dry state is not representative of the wet state. Techniques in the wet state (histology, µCT) are of lower resolution. Here, self-gelling, injectable composites of Gellan Gum (GG) hydrogel and two different types of sol-gel-derived bioactive glass (bioglass) particles were analyzed in the wet state using Synchrotron X-ray radiation, enabling high-resolution determination of particle size and spatial distribution. The lower detection limit volume was 9 × 10(-5) mm(3) . Bioglass particle suspensions were also studied using zeta potential measurements and Coulter analysis. Aggregation of bioglass particles in the GG hydrogels occurred and aggregate distribution was inhomogeneous. Bioglass promoted attachment of rat mesenchymal stem cells (rMSC) and mineralization.