Cell Adhesion Studies Research Articles

Synthesis, Characterization and Osteogenic Properties of Chitin-Polydioxanone Composite Gel.

In this study, we have developed a Chitin(Ch)-Poly(dioxanone)(PDO) gel system, which can be potentially used for tissue engineering. Hydrogel has been widely used in biomedical applications for its tuneable properties and biocompatibility. Chitin (Ch) is a natural biopolymer used for its ability to mimic the natural extracellular matrix due to its N-acetyl glucosamine structural units. Poly (dioxanone) (PDO) is a FDA-approved synthetic biopolymer known for its mechanical properties, good biocompatibility, and poor inflammatory response. Based on this, we have developed Ch-PDO composite gel using simple regeneration chemistry and characterized it using FT-IR and SEM. The developed composite gel showed improved gel strength, good swelling ability, and controlled degradation behaviour. It also showed good inject ability with shear thinning properties and hemocompatibility. Further, the biocompatibility and cell adhesion studies of the prepared gels were studied using dental follicle stem cells (DFSCs). The prepared Ch-PDO gel was biocompatible and showed DFSCs cell attachment. Osteogenic mineralization, RUNX2 and OPN expression of the prepared Ch and Ch-PDO gel was studied and Ch-PDO gel showed an enhanced mineralization and RUNX2 and OPN expression showed enhanced osteogenic activity in Ch-PDO. Therefore, the developed chitin-PDO gel could be potentially used for bone tissue engineering.

Comprehensive genomic analysis and evaluation of in vivo and in vitro safety of Heyndrickxia coagulans BC99

This study aimed to evaluate the safety profile and beneficial effects of Heyndrickxia coagulans strain BC99 (BC99) for potential use in functional foods and pharmaceuticals. We began with whole genome sequencing of BC99, followed by a comprehensive safety assessment comprising genome analysis, hemolysis, cytotoxicity, antibiotic susceptibility tests, and cell adhesion and tolerance studies, along with acute and subacute oral toxicity studies in animal models. BC99 was isolated from a well-characterized collection originating from the feces of a healthy infant. Our results indicated no hemolytic activity on Columbia blood agar plates and broad antibiotic sensitivity, including to gentamicin, ampicillin, chloramphenicol, ciprofloxacin, and others. Cytotoxicity testing confirmed no adverse effects on HT-29 cells and significant adhesive properties to intestinal epithelial cells. Tolerance tests demonstrated over 90% viability of BC99 under simulated gastrointestinal conditions. In vivo studies in mice and rats confirmed the absence of adverse effects following oral administration. Collectively, these findings support BC99’s robust tolerance to gastrointestinal environments, strong adhesion capabilities, and a broad spectrum of antibiotic resistance, underlining its potential as a safe and effective agent for gut microbiota modulation and host health enhancement.

Enhancement of corrosion, biocompatibility and drug delivery properties of nitinol implants surface by Al-Zn-LDH nanohybrids

This study investigates the enhancement of drug delivery and corrosion resistance by incorporating dexamethasone sodium phosphate into aluminum-zinc layered double hydroxide (LDH) coated nickel-titanium alloys. The nickel-titanium samples were fabricated from titanium and nickel powders using spark plasma sintering (SPS) and cold press sintering (CPS), followed by electrophoretic deposition of LDH nanoparticles. This composite construction aims to utilize the biocompatibility and mechanical properties of nickel-titanium, combined with the controlled drug release and corrosion resistance properties of LDH coatings. Structural and microstructural characterizations were performed using X-ray diffraction and scanning electron microscopy. The results indicated that SPS samples exhibited superior microstructural homogeneity and corrosion resistance compared to CPS samples. Dexamethasone sodium phosphate was successfully intercalated into the LDH layers, as evidenced by an increase in layer spacing from 7.14 Å to 20.130 Å. The LDH-coated nickel-titanium with intercalated dexamethasone sodium phosphate demonstrated a 5 % lower drug release rate and significantly improved corrosion resistance compared to uncoated samples. Cell adhesion studies confirmed good biocompatibility between cells and the coated surface. This composite material shows promise for enhanced performance in biomedical applications, particularly in drug delivery and corrosion resistance.

Eggshell derived scaffold of hydroxyapatite-ammonium bicarbonate nano-composite: Bioactivity and cytotoxicity studies

This work investigated the facile synthesis of porous scaffold eggshell derived hydroxylapatite (ESHAp) as a composite with ammonium bicarbonate (AMB) for potential biomaterial in tissue engineering application. The phase purity, composition, size, functional groups and morphology of the apatite were elucidated using high resolution transmission electron microscopy (HTEM), X-Ray Diffraction (XRD), Fourier Transform Infrared Spectroscopy (FT-IR) and scanning electron microscopy (SEM). The results showed that hydroxylapatite (HAp) nanoparticles have round morphologies with average diameters between 20 nm and 80 nm, FT-IR analysis confirmed significant hydroxylapatite functional groups like carbonate, phosphate, and hydroxyl groups, while XRD analysis revealed a well crystalline monophasic HAp powder. The scaffold samples containing 10, 20, 25 and 30 % of AMB withstood a compressive stress up to 5, 20, 30 and 42 N/mm2 respectively which indicates that the compressive stress increased with the AMB content introduced as the pore forming agent. MTT assay performed using MG63 osteosarcoma cell lines showed that on comparing the sample of ESTHAp which contained 0 % AMB with other samples in the range of 0.01–1 mM, viability of above 85 % MG63 cells was achieved except for ESTHAp with 40 % AMB, which showed some level of toxicity. The cell adhesion studies of sintered ESTHAp porous scaffold with different weight percent of the pore forming agents using inverted microscopic images of MG 63 cells incubated with ESTHAp samples and treated with heat at 1000 °C appeared to be unstable in the media used with particle leaching observed, and no cells observed near to the samples.

Unravelling the tissue regenerative nature of marine polysaccharide chitosan embedded halloysite reinforced poly (vinyl alcohol) nanocomposite films

The study focuses on developing the poly (vinyl alcohol) (PVA) nanocomposite films reinforced with Halloysite Nano-Tubes (HNT) which were surface functionalised with chitosan forming a Schiff base structure. The work was aimed to traverse its way in tissue engineering. Modification of halloysite with sea polysaccharide chitosan enhanced its ability to bind to the PVA matrix. The modification was assisted by FTIR, XRD, FESEM, TEM, AFM and DSC-TGA techniques. The films could withstand a higher temperature and exhibited high ranges of tensile strength and Young’s modulus. In addition, biocompatible studies such as in-vitro swelling, enzymatic degradation, water contact angle and hemolysis presented extremely well compatibilities proving it to be viable in physiological pH (in phosphate buffered saline). The cell adhesion and proliferation studies conducted on NIH3T3 mouse fibroblasts revealed the cell proliferation and tissue regeneration properties of the films. Both the tests performed for cell growth- Trypan blue dye exclusion and Acridine Orange Ethidium bromide assays showed a doubled rate of cell growth on the films which proves its biomedical nature.Graphical

Controlled release in sodium alendronate/halloysite/hydroxyapatite/gelatin nanocomposite scaffolds: A new insight into bone tissue engineering

The use of smart scaffolds with drug release capabilities is now widely recognized as a key approach in designing hard tissue substitutes. In this study, alendronate sodium (AL) was bound to halloysite nanotube (HNT) sheets through ionic and π-π interactions. Hydroxyapatite (HA) powder was synthesized using the co-precipitation method, and the drug/carrier complex was subsequently incorporated into a hydroxyapatite/gelatin (GEL) matrix. Composite scaffolds of GEL/HA/HNT/AL were then created using a freeze-drying technique. Different concentrations of HNT (0.5 %, 1 %, and 2 %) were tested to determine the optimal formulation. The scaffolds were subjected to extensive physical, chemical, and mechanical evaluations using scanning electron microscopy (SEM), X-ray diffraction (XRD), Fourier-transform infrared spectroscopy (FTIR), compressive stress tests, and measurements of density and porosity. The release profile of alendronate sodium, scaffold swelling behavior in water, bioactivity, and in vitro performance were thoroughly assessed. Cell viability was tested using the MTT assay, while alkaline phosphatase (ALP) activity and cell adhesion studies were performed to evaluate osteogenic potential. The results revealed that the GEL/HA/1 % HNT/AL nanocomposite scaffold exhibited excellent properties, including efficient drug release, optimal swelling rates, and enhanced bioactivity. The MTT assay confirmed high cell viability, and both ALP activity and cell adhesion studies indicated strong potential for bone cell differentiation and repair. Thus, the GEL/HA/1 % HNT/AL nanocomposite scaffold is a highly promising platform for bone tissue engineering.

Exploring marine-based compounds as cross-linkers to improve the biocompatibility and sustainability of chitosan-based hydrogels

Chitosan is one of the most promising natural polymers, its fundamental scientific research is growing uninterruptedly and has been applied in a wide and varied range of domains, including biomedical and water treatment applications. Therefore, the search and implementation of non-synthetic and non-toxic cross-linkers for chitosan-based hydrogels is crucial for the development of more sustainable and biocompatible materials. Herein, an alternative approach has been developed to explore and exploit methanolic and aqueous extracts from five red seaweed species as covalent cross-linkers for chitosan-based hydrogels. The formation of a gel could be denoted for all extracts, whereas the protein-rich methanolic extractions afforded instantaneous gel-forming ability and greater stiffness and stability. The obtained hydrogels present large porous system with high degrees of swelling up to ca. 3000 %, and were successfully applied as dye adsorbent to remove industrial dye methyl orange withing a circular process with adsorption capacities of 728.46 ± 66.17 mg/g. Furthermore, cytotoxicity and cell-adhesion studies revealed the biocompatibility of the hydrogels and their potential applicability for tissue-engineering. This work demonstrated that methanolic and aqueous extracts from different red seaweed species could replace toxic cross-linkers. Furthermore, the unexpected ability of some extracts could pave the way for the development of new formulations for additive manufacturing, in particular for 3D printing approaches.

Tissue Factor Maintains Lung Epithelial Barrier Integrity in Acute Lung Injury Through Cell Adhesion

Background. Acute Respiratory Distress Syndrome (ARDS) is a common cause of acute respiratory failure. While loss of lung epithelial barrier integrity is a pathologic mechanism of ARDS, little is known about the factors that regulate epithelial barrier integrity in the lung. Our previously published mouse studies showed that deletion of alveolar type II epithelial cell Tissue Factor (TF) caused impaired lung barrier integrity in models of Acute Lung Injury (ALI). TF is an integral membrane protein that both initiates the extrinsic coagulation cascade and serves several non-coagulant functions. Notably, TF promotes cell adhesion through interactions with cell surface integrin heterodimers, comprised of individual α and β integrin subunits. As such, we hypothesize that epithelial TF is necessary for maintaining lung barrier integrity and that its overexpression is protective in ALI through increased cell adhesion. Methods. To determine whether supraphysiologic overexpression of TF in the lung epithelium can enhance barrier integrity we created a novel transgenic mouse in which TF was inducibly overexpressed in the lung epithelium (TFEpi+) using a CMV-TetO promoter and crossed with SPC-rtTA59 mice. High alveolar epithelial TF expression compared to wild-type littermates (WT) was confirmed through immunohistochemistry and western blot analysis after one week of doxycycline in drinking water. To induce ALI, mice were intranasally infected with 2000 colony forming units of Klebsiella pneumoniae (KP) or PBS control. At 24-hours post infection, mice were euthanized, lung tissue was collected, and a bronchoalveolar lavage (BAL) was performed. BAL protein, clot time, and leukocyte influx were measured. Lung wet-to-dry weight ratios and bacterial burden were measured. To better understand TF regulation of β1 integrin function, TF-knockout A549 cells were created using CRISPR and cultured on specific β1 integrin ligands (Collagen I, IV, Laminin-511, and Fibronectin). On-cell western blots and crystal violet were used to assess cell surface β1 integrin expression and cell adhesion. Results. TFEpi+ mice showed higher lung TF protein expression (median 38031 [IQR 25033, 57948] vs 1801 [IQR 804.3, 2083] expression normalized to total protein; p=0.0016) compared to WT. TFEpi+ mice infected with KP showed lower BAL protein (mean 248.51 [SD 98.03] vs 366.3 [SD 159.8] μg/ml; p=0.0035) and lung wet-to-dry weight ratios (mean 4.27 [SD 0.37] vs 5.29 [SD 0.75]; p=0.0152) compared to WT. Bacterial burden and BAL inflammatory cell counts were higher, while BAL clot time was lower in infected WT mice compared to control. However, these outcomes did not differ between infected TFEpi+ and WT mice. Loss of TF in alveolar epithelial cells reduced cell surface β1 integrin expression and cell adhesion to Laminin-511. Conclusion: Alveolar epithelial TF overexpression is protective for maintaining lung barrier integrity independent of coagulation and inflammation. Further, cell adhesion studies suggest that this is potentially mediated through increased a6β1 epithelial cell adhesion. AHA Postdoctoral Fellowship, National Institute of Health Grants (HL150783, I01BX002288). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Core-Shell Structure of Photopolymer-Grafted Polyurethane as a Controlled Drug Delivery Vehicle for Biomedical Application.

Functionalized ultraviolet photocurable bisphenol A-glycerolate dimethacrylates with tailorable size have been synthesized as the core, which have further been grafted using the diisocyanate chain end of polyurethane (PU) as the shell to create a core-shell structure of tunable size for a controlled drug delivery vehicle. The core-shell structure has been elucidated through spectroscopic techniques like 1H NMR, FTIR, and UV-vis and their relative shape and size through TEM and AFM morphology. The greater cross-link density of the core is reflected in the higher glass transition temperature, and the improved thermal stability of the graft copolymer is proven from its thermogravimetric analyses. The flow behavior and enhanced strength of the graft copolymers have been revealed from rheological measurements. The graft copolymer exhibits sustained release of the drug, as compared to pure polyurethane and photopolymer, arising from its core-shell structure and strong interaction between the copolymer and drug, as observed through a significant shifting of absorption peaks in FTIR and UV-vis measurements. Biocompatibility has been tested for the real application of the novel graft copolymer in medical fields, as revealed from MTT assay, cell imaging, and cell adhesion studies. The efficacy of controlled release from a graft copolymer has been verified from the gradual cell killing and ∼70% killing in 3 days vs meager cell killing of ∼25% very quickly in 1 day, followed by the increased cell viability of the system treated with the pure drug. The mechanism of slow and controlled drug release from the core-shell structure has been explored. The fluorescence images support the higher cell-killing efficiency as opposed to a pure drug or a drug embedded in polyurethane. Cells seeded on 3D scaffolds have been developed by embedding a graft copolymer, and fluorescence imaging confirms the successful growth of cells within the scaffold, realizing the potential of the core-shell graft copolymer in the biomedical arena.

Role of steroid hormones in the maintenance of focal adhesions in bovine oviductal epithelial cells

The oviduct, the organ of the female reproductive system where fertilization and early embryonic development occur, provides an optimal environment for the final maturation of oocytes, storage, and sperm capacitation and transport of gametes and embryos. During the estrous cycle, the oviduct is affected by ovarian sex hormones, resulting in changes aimed at maintaining an appropriate microenvironment. Normal cell migration is tightly regulated, its role being essential for the development and maintenance of organ and tissue functions as well as for regeneration following injury. Due to their involvement in focal contact formations, focal adhesion kinase (PTK2) and paxillin (PXN) are key proteins in the study of cell migration and adhesion. The objective of this work was to compare the expression of PTK2 and PXN in oviductal cells along the estrous cycle and to determine if their expression is regulated by the presence of 17-β estradiol (E2) and/or progesterone (P4). No transcripts of PTK2 or of PXN were detected in cells corresponding to the luteal phase. Additionally, hormonal stimulation experiments on bovine oviductal cell cultures (BOECs) were carried out, where P4 inhibited the expression of both genes. Migration assays demonstrated that P4 reduced BOECs migration capacity. P4 treatment also reduced cell adhesion, while E2 increased the number of adhered cells. In conclusion, the presence of E2 and P4 regulates the expression of genes involved in the formation of focal contacts and modifies the migration and adhesion of BOECs. Understanding the effect of steroid hormones on BOECs is critical to grasp the impact of steroid control on oviductal function and its contribution to establishing successful pregnancies.

Biosurfactant-Assisted Cu Doping of Brushite Coatings: Enhancing Structural, Electrochemical, and Biofunctional Properties.

Stainless steel (316L SS) has been widely used in orthopedic, cardiovascular stents, and other biomedical implant applications due to its strength, corrosion resistance, and biocompatibility. To address the weak interaction between steel implants and tissues, it is a widely adopted strategy to enhance implant performance through the application of bioactive coatings. In this study, Cu-doped brushite coatings were deposited successfully through pulse electrodeposition on steel substrates facilitated with a biosurfactant (BS) (i.e., surfactin). Further, the combined effect of various concentrations of Cu ions and BS on the structural, electrochemical, and biological properties was studied. The X-ray diffraction (XRD) confirms brushite composition with Cu substitution causing lattice contraction and a reduced crystallite size. The scanning electron microscopy (SEM) and energy-dispersive spectroscopy (EDS) studies reveal the morphological changes of the coatings with the incorporation of Cu, which is confirmed by X-ray photoelectron spectroscopy (XPS) and elemental mapping. The Fourier transform infrared (FTIR) and Raman spectroscopy confirm the brushite and Cu doping in the coatings, respectively. Increased surface roughness and mechanical properties of Cu-doped coatings were analyzed by using atomic force microscopic (AFM) and nanohardness tests, respectively. Electrochemical assessments demonstrate corrosion resistance enhancement in Cu-doped coatings, which is further improved with the addition of biosurfactants. In vitro biomineralization studies show the Cu-doped coating's potential for osseointegration, with added stability. The cytocompatibility of the coatings was analyzed using live/dead and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assays; cell adhesion, proliferation, and migration studies were evaluated using SEM. Antibacterial assays highlight significant improvement in the antibacterial properties of Cu-doped coatings with BS. Thus, the developed Cu-doped brushite coatings with BS demonstrate their potential in the realm of biomedical implant technologies, paving the way for further exploration.

Synthesis of Superabsorbent Hydrogels with Predefined Geometries and Controlled Swelling Properties for Versatile 3D Cell Culture Scaffolds.

This research presents a simple but general method to prepare water-soluble-polymer-based superabsorbent hydrogels with predefined microscale geometries and controlled swelling properties. Unlike conventional hydrogel preparation methods based on bulk solution-phase cross-linking, poly(vinyl alcohol) is homogeneously mixed with polymer-based cross-linkers in the solution phase and thermally cross-linked in the solid phase after drying; the degree of cross-linking is modulated by controlling the cross-linker concentration, pH, and/or thermal annealing conditions. After the shape definition process, cross-linked films or electrospun nanofibers are treated with sulfuric acid to weaken hydrogen bonds and introduce sulfate functionality in polymer crystallites. The resultant superabsorbent hydrogels exhibit an isotropic expansion of the predefined geometry and tunable swelling properties. Particularly, hydrogel microfibers exhibit excellent optical transparency, good biocompatibility, large porosity, and controlled cell adhesion, leading to versatile 3D cell culture scaffolds that not only support immortalized cell lines and primary neurons but also enable stiffness-modulated cell adhesion studies.

Iron oxide nanoparticle-stabilized Pickering emulsion-templated porous scaffolds loaded with polyunsaturated fatty acids (PUFAs) for bone tissue engineering.

Dietary intake of ω-3-polyunsaturated fatty acids (PUFAs) can significantly improve the expression levels of alkaline phosphatase (ALP) and osteocalcin. However, PUFAs are hydrophobic and highly sensitive to temperature, oxygen concentration, pH, and ionic strength. Hence, it is challenging to use PUFAs as bioactive compounds for bone tissue engineering. Here, we encapsulated PUFAs in liposomes to improve their stability. The hydrodynamic size of the PUFA-loaded liposomes was found to be 121.3 ± 35 nm. GC-MS analysis showed that the encapsulation efficiency of the PUFAs was 19.9 ± 3.4%. These PUFA-loaded liposomes were loaded into porous scaffolds that were prepared by polymerizing glycidyl methacrylate and trimethylolpropane triacrylate monomers using the Pickering emulsion polymerization technique. Oleic acid-coated iron oxide nanoparticles were used as the stabilizing agent to prepare these acrylate-based scaffolds containing PUFA-loaded liposomes (P-Lipo-IO(GMA-TMPTA)). SEM micrographs confirmed the porous nature of the scaffolds and the presence of well-adhered liposomes. An in vitro cytotoxicity study conducted using MG63 cells confirmed that these scaffolds showed desirable cytocompatibility. Cell adhesion study showed a well-spread morphology, indicating firm adhesion of the cells. The alizarin red staining of P-Lipo-IO(GMA-TMPTA) scaffolds showed 3- and 2-fold higher calcium deposition compared to the control on days 7 and 14, respectively. ALP activity was also 2-fold higher than that of the control on day 14. RT-PCR analysis of cells exposed to P-Lipo-IO(GMA-TMPTA) scaffolds showed significantly higher expression of osteogenic markers compared to the control. An antibacterial study conducted on Staphylococcus aureus showed a higher percentage inhibition and reactive oxygen species generation in samples treated with P-Lipo-IO(GMA-TMPTA) scaffolds. These desirable biological properties indicate that the developed scaffolds are suitable for bone tissue engineering.

Fabrication of a novel biomaterial of mono-layer of halloysite nanotube-magnesium oxide/ β-tricalcium phosphate composite on implant substrate: Structural and biological properties

In the present study, a novel composite coating consisting of magnesium oxide on halloysite nanotube (HNT-MgO) and doping of zinc and strontium ions in beta-tricalcium phosphate (Mn+-β-TCP). The monolayers of biomaterials were developed by coating HNT-MgO-Mn+-β-TCP on Ti–6Al–4V alloy using the electrophoretic method. The physicochemical properties such as functional groups, phase composition, surface morphology, cross-section, and chemical composition of biomaterials were characterized by adopted analytical techniques such as Fourier transform infrared spectroscopy, X-ray diffraction analysis, Scanning electron microscopy-Energy dispersive spectroscopy, Atomic force microscopy, High-resolution transmission electron microscopy. The cytotoxic activity of the prepared composite was tested against osteosarcoma MG-63 cells using MTT assay, and AO/EB staining the results clearly showed, that by increasing the composite concentration (5–100 μg/ml) the cell viability was reduced. Further, a cell adhesion study was performed by FE-SEM analysis, which showed that the MG-63 cells could inhibit and restrict the HNT-MgO-Mn+-β-TCP monolayer composite. The composite coating confirms the significant anticancer activity against osteosarcoma cells suggesting it is a promising candidate for further studies for practical orthopedic applications.

Three-Dimensional Graphene Promotes the Proliferation of Cholinergic Neurons

Introduction: An early substantial loss of basal forebrain cholinergic neurons (BFCNs) is a common property of Alzheimer’s disease and the degeneration of functional BFCNs is related to learning and memory deficits. As a biocompatible and conductive scaffold for growth of neural stem cells, three-dimensional graphene foam (3D-GF) supports applications in tissue engineering and regenerative medicine. Although its effects on differentiation have been demonstrated, the effect of 3D-GF scaffold on the generation of BFCNs still remains unknown. Methods: In this study, we used 3D-GF as a culture substrate for neural progenitor cells (NPCs) and demonstrated that this scaffold material promotes the differentiation of BFCNs while maintaining excellent cell viability and proliferation. Results: Immunofluorescence analysis, real-time polymerase chain reaction, Western blotting, and ELISA revealed that the proportion of BFCNs at 21 days of differentiation reached approximately 30.5% on 3D-GF compared with TCPS group that only presented 9.7%. Furthermore, a cell adhesion study suggested that 3D-GF scaffold enhances the expression of adhesion proteins including vinculin, integrin, and N-cadherin. These findings indicate that 3D-GF scaffold materials are preferable candidates for the differentiation of BFCNs from NPCs. Conclusions: These results suggest new opportunities for the application of 3D-GF scaffold as a neural scaffold for cholinergic neurons therapies based on NPCs.

Programing Cell Assembly via Ink-Free, Label-Free Magneto-Archimedes Based Strategy.

Tissue engineering raised a high requirement to control cell distribution in defined materials and structures. In "ink"-based bioprintings, such as 3D printing and photolithography, cells were associated with inks for spatial orientation; the conditions suitable for one ink are hard to apply on other inks, which increases the obstacle in their universalization. The Magneto-Archimedes effect based (Mag-Arch) strategy can modulate cell locomotion directly without impelling inks. In a paramagnetic medium, cells were repelled from high magnetic strength zones due to their innate diamagnetism, which is independent of substrate properties. However, Mag-Arch has not been developed into a powerful bioprinting strategy as its precision, complexity, and throughput are limited by magnetic field distribution. By controlling the paramagnetic reagent concentration in the medium and the gaps between magnets, which decide the cell repelling scope of magnets, we created simultaneously more than a hundred micrometer scale identical assemblies into designed patterns (such as alphabets) with single/multiple cell types. Cell patterning models for cell migration and immune cell adhesion studies were conveniently created by Mag-Arch. As a proof of concept, we patterned a tumor/endothelial coculture model within a covered microfluidic channel to mimic epithelial-mesenchymal transition (EMT) under shear stress in a cancer pathological environment, which gave a potential solution to pattern multiple cell types in a confined space without any premodification. Overall, our Mag-Arch patterning presents an alternative strategy for the biofabrication and biohybrid assembly of cells with biomaterials featured in controlled distribution and organization, which can be broadly employed in tissue engineering, regenerative medicine, and cell biology research.

Dual-Action Effect of Gallium and Silver Providing Osseointegration and Antibacterial Properties to Calcium Titanate Coatings on Porous Titanium Implants.

Previously, functional coatings on 3D-printed titanium implants were developed to improve their biointegration by separately incorporating Ga and Ag on the biomaterial surface. Now, a thermochemical treatment modification is proposed to study the effect of their simultaneous incorporation. Different concentrations of AgNO3 and Ga(NO3)3 are evaluated, and the obtained surfaces are completely characterized. Ion release, cytotoxicity, and bioactivity studies complement the characterization. The provided antibacterial effect of the surfaces is analyzed, and cell response is assessed by the study of SaOS-2 cell adhesion, proliferation, and differentiation. The Ti surface doping is confirmed by the formation of Ga-containing Ca titanates and nanoparticles of metallic Ag within the titanate coating. The surfaces generated with all combinations of AgNO3 and Ga(NO3)3 concentrations show bioactivity. The bacterial assay confirms a strong bactericidal impact achieved by the effect of both Ga and Ag present on the surface, especially for Pseudomonas aeruginosa, one of the main pathogens involved in orthopedic implant failures. SaOS-2 cells adhere and proliferate on the Ga/Ag-doped Ti surfaces, and the presence of gallium favors cell differentiation. The dual effect of both metallic agents doping the titanium surface provides bioactivity while protecting the biomaterial from the most frequent pathogens in implantology.

Toward osteomimetic formation of calcium phosphate coatings with carbonated hydroxyapatite

Biomimetic production of coatings on various types of scaffolds is based mainly on simulated body fluid precipitation (SBF) of apatites, or, if the HCO3− is present, carbonated apatites. Recently, we proposed formation of calcium phosphates (CaP) precipitates by alkaline phosphatase (ALP) hydrolysing glycerophosphate in presence of calcium ions as an alternative to SBF. Since apatites synthesized in bone by the ALP activity contain carbonate anions, it was tempting to investigate whether the phosphatase method could be advanced into osteomimetic one. Therefore, taking example from the SBF studies, phosphatase incubation medium was enriched with carbonate ions at 4.2 and 27 mM concentration. X-ray diffraction of the precipitates disclosed peaks typical for hydroxyapatite (HAP). FTIR analysis showed that at both concentration of carbonate ions, apatites underwent both B and A substitution, more extensive at higher concentration. Thus, osteomimetic approach produced carbonated hydroxyapatites of the type encountered in bone tissue even at HCO3− concentration as low as 4.2 mM. Composite plates made of poly(ε-caprolactone) and mixture of β-tricalcium phosphate and hydroxyapatite at mass ratio of 1:0.5:0.5, respectively, were covered by CaP coatings, i.e., CaP-0, CaP-4.2, CaP-27, by incubation in phosphatase medium containing 0, 4.2 or 27 mM of NaHCO3, respectively. Pristine or coated PCL50 plates were used to study release of calcium and adsorption/desorption of proteins, or seeded with human bone marrow mesenchymal stem cells (hMSC) for study of cell adhesion, spreading and osteogenic differentiation. Introduction of carbonate into the CaP coatings significantly increased release of Ca2+ in a carbonate concentration-dependent manner; the release was up to 4 times higher, when compared to CaP-0 coating, and reached 0.41 ± 0.01 mM for CaP-27 after first 24 h. Coating CaP-4.2 yielded significantly higher adsorption of bovine serum albumin and cytochrome C than CaP-0. All of the CaP coatings improved significantly hMSC adhesion, however, only CaP-4.2 provided 2 times higher cell number than PCL50 after 2 weeks of culture. Interestingly, ALP activity calculated per cell number was the highest on pristine plates, presumably because hMSC differentiate preferentially into osteoblasts at lower seeding densities. It appears, therefore, that the osteomimetic approach may be useful for production of carbonated hydroxyapatite coatings, but requires further studies and replacing intestinal phosphatase used in this work with one originating from bone.

A model calibration procedure for count response

In recent years, there are numerous articles about computer model calibration. But these articles mainly focus on continuous output variables for the convenience of the derivation. However, there are some other types of output data in the application area, such as binary data and count data, the former of which has been investigated very well in the area of computer model calibration and applied to cell adhesion study successfully. Inspired by the existing methods for binary data, we propose a model calibration procedure for count response, and derive the consistency and asymptotic normality of the proposed estimator of the calibration parameters by the theory of empirical process. Some simulations are conducted to verify the good performance of the proposed method.

Synthesis and Biological Properties of Alanine-Grafted Hydroxyapatite Nanoparticles.

Hydroxyapatite attracts great attention as hard tissues implant material for bones and teeth. Its application in reconstructive medicine depends on its biocompatibility, which is in a function of composition and surface properties. The insertion of a protein element in the composition of implants can improve the cell adhesion and the osseointegration. Having this in mind, the proposal of this work was to develop L-alanine-grafted hydroxyapatite nanoparticles and to study their biocompatibility. Two L-alanine sources and three grafting methods were used for hydroxyapatite surface functionalization. The efficiency of grafting was determined based on X-ray powder diffraction, Fourier-transform infrared spectroscopy, thermal analyses, and field-emission scanning electron microscopy. The results indicated the formation of hydroxyapatite with 8-25 wt% of organic content, depending on the grafting method. Protein adsorption, cell adhesion, and viability studies were carried out to evaluate biological properties of grafted materials. The viability of MG-63 human osteoblastic cells following 24 h incubation with the alanine-grafted hydroxyapatite samples is well preserved, being in all cases above the viability of cells incubated with hydroxyapatite. The alanine-grafted hydroxyapatite prepared in situ and by simple mixture showed higher protein adsorption and cell adhesion, respectively, indicating their potential toward use in regenerative medicine.