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The promotion of osteogenic differentiation by photothermal signaling in the ECM microenvironment was achieved, through ECM films which were photothermally functionalized with graphene during cell cultivation.

Macrophages have two functionalized phenotypes, M1 and M2, and the regulation of M1/M2 polarization of macrophages is critical for tissue repair. Tissue-derived immune factors are considered the major drivers of macrophage polarization. Based on the main cytokine-induced polarization pathways, we tested the effect of electrical stimulation (ES) of macrophages on the regulation of M1/M2 polarization and a possible synergistic effect with the cytokines. Indium tin oxide (ITO) planar microelectrodes were used to produce ES under different voltages, frequencies and waveforms. We evaluated the influence of ES on the cytokine-induced M1/M2 polarization using mouse bone marrow-derived macrophages cultured with both lipopolysaccharide (LPS)/IFN-γ factors and IL-4 factors for M1 and M2, respectively. The results showed that ES promoted the cytokine-induced macrophage polarization. Importantly, we found that stimulation with a square waveform selectively promoted LPS/IFN-γ-induced M1 polarization, while stimulation with a sinusoidal waveform promoted both LPS/IFN-γ-induced M1, and IL-4-induced M2 polarization. Mechanistically, stimulation with a square waveform affected the intracellular ion concentration, whereas stimulation with a sinusoidal waveform promoted both the intracellular ion concentration and membrane receptors. We hereby establish an ES-mediated strategy for immunomodulation via macrophage polarization.

Piezoelectric poly(vinylidene fluoride-trifluoroethylene) has demonstrated an ability to promote osteogenesis, and biomaterials with a chirality-patterned topological surface could enhance cellular osteogenic differentiation. In this work, we created a chirality-patterned surface potential distribution of CoFe

Surface potential of biomaterials is found to be important for wound healing. Here, poly(vinylidenefluoride-co-trifluoroethylene) (P(VDF-TrFE)) films with different surface potentials and piezoelectric responses were prepared and explored for the effect of surface potential on wound healing. The crystalline state of P(VDF-TrFE) films were characterized with X-ray diffraction (XRD), differential scanning calorimetry (DSC) and Fourier-transformed infrared spectroscopy (FTIR), illustrated that the electric polarization will promote the crystallization of the β phase of P(VDF-TrFE), in which the content of β phase increased from 82.9 % to 86.8 % compared with the control. Then, Kelvin potential and piezoelectric coefficient d

Bone tissue engineering requires a material that can simultaneously promote osteogenic differentiation and anti-inflammatory effects at specific times in response to a series of problems after bone implantation. In this study, the porous network-like titanium matrix was constructed and polypyrrole/dexamethasone (Ppy/Dex) composite coatings with three-dimensional nano-network structure were prepared by electrochemical deposition. The biocompatibility of the composite coatings was further improved by the composite of the extracellular matrix (ECM). The Ppy/Dex/ECM composite coatings released Dex by changing the redox state of Ppy under the electrical stimulation of negative pulses, achieving a drug release controlled by electric field. In terms of osteogenic differentiation, the Ppy/Dex/ECM composite coatings exhibited the best osteogenic activity under electrical controlled release, indicating the synergistic effect of Dex and ECM on osteogenic differentiation. In terms of anti-inflammatory properties, ECM exhibited simultaneous inhibition of both pro- and anti-inflammatory process, while Dex demonstrated significant promotion of anti-inflammatory processes. In this work, the effect of electrical controlled drug release on osteogenic differentiation and inflammation in the ECM cell microenvironment was achieved by preparing Ppy/Dex/ECM composite coatings, which is of great significance for bone tissue engineering and regenerative medicine.

In this work, nanocomposite membranes based on graphene and polylactide were evaluated for mechanical properties and biocompatibility. Single-layer graphene (SLG), graphene nanosheets (GNS), and poly L-lactic acid (PLLA) were prepared through layer-by-layer deposition and homogeneous mixing. The results revealed that PLLA/SLG nanocomposites and PLLA/GNS nanocomposites could show enhanced mechanical properties and biocompatibility. The addition of a tiny amount of SLG significantly improved Young’s modulus and tensile strength of the PLLA matrix by 15.9% and 32.8% respectively, while the addition of the same mass ratio of GNS boosted the elongation at break of the PLLA matrix by 79.7%. These results were ascribed to the crystallinity and interfacial interaction differences resulting from graphene incorporation. Also, improved biocompatibility was observed with graphene incorporation. Such nanocomposites membranes showed a lot of potential as environment-friendly and biomedical materials.

Titanium dioxide (TiO2) photofunctionalization has been demonstrated as an effective surface modification method for the osseointegration of implants. However, the insufficient understanding of the mechanism underlying photofunctionalization limits its clinical applications. Here, we report an ultraviolet (UV) radiant energy-dependent functionalization on TiO2 nanodots (TN) surfaces. We found the cell adhesion, proliferation, and osteogenic differentiation gradually increased with the accumulation of UV radiant energy (URE). The optimal functionalizing treatment energy was found to be 2000 mJ/cm2, which could regulate cell-specific behaviors on TN surfaces. The enhanced cell behaviors were regulated by the adsorption and functional site exposure of the extracellular matrix (ECM) proteins, which were the result of the surface physicochemical changes induced by the URE. The correlation between the URE and the reconstruction of surface hydroxyl groups was considered as an alternative mechanism of this energy-dependent functionalization. We also demonstrated the synergistic effects of FAK-RHOA and ERK1/2 signaling pathways on mediating the URE-dependent cell behaviors. Overall, this study provides a novel insight into the mechanisms of photofunctionalization, guiding the design of implants and the clinical practice of photofunctionalization.

Light-induced cell harvest shows much potential in in vitro cell culture. In this work, a light-responsive monolayer graphene (Gr)/titanium dioxide nanodot (TN) film is designed and used for light-induced cell harvest. It is found that after 20 min of 365 nm UV or 450 nm visible light illumination, different types of cells could be detached from the surface effectively. The highest cell detachment ratio reaches about 95%. The mechanism of such a cell detachment is contributed to light illumination generates charge accumulation, which, in turn, changes the conformation of the extracellular matrix protein molecules adsorbed to a more disordered state, and eventually leads to the cells detachment. Such UV and visible light responsive Gr/TiO2 film could be a good candidate for a surface with light-induced cell detachment property.

Macrophages polarization in bone immune microenvironment is crucial in bone regeneration. In this work, mineralized collagen (MC) coatings with photo-thermal effect were prepared through incorporation of polydopamine (PDA). MC coatings with different thicknesses were deposited on titanium substrate through electrochemical deposition. PDA preformed on the substrate, acting as a photo-thermal agent. The effects of light illumination, i.e., different thermal effects, on the polarization of mouse bone marrow-derived macrophages were explored. It was found that heat can promote the M1 polarization of macrophages and inhibit the M2 polarization. Also, gene expression results revealed that such photo illumination based macrophage modulation is effective and safe. It provides a possible way for the design of functional materials to regulate the bone immune microenvironment.

Mesenchymal stem cells (MSCs), which are undifferentiated stem cells with the property of stemness and the potential to differentiate into multiple lineages, including osteoblasts, have attracted a great deal of attention in bone tissue engineering. Consistent with the heterogeneity of MSCs, various surface markers have been used. However, it is still unclear which markers of MSCs are best for cell amplification in vitro and later bone regeneration in vivo. Krüppel-like Factor 2 (KLF2) is an important indicator of the stemness of human MSCs (hMSCs) and as early vascularization is also critical for bone regeneration, we used KLF2 as a novel in vitro marker for MSCs and investigated the angiogenesis and osteogenesis between KLF2+ MSCs and endothelial cells (ECs). We found a synergistic interaction between hMSCs and human umbilical vein ECs (HUVECs) in that KLF2+ stemness-maintained hMSCs initially promoted the angiogenesis of HUVECs, which in turn more efficiently stimulated the osteogenesis of hMSCs. In fact, KLF2+ hMSCs secreted angiogenic factors initially, with some of the cells then differentiating into pericytes through the PDGF-BB/PDGFR-β signaling pathway, which improved blood vessel formation. The matured HUVECs in turn synergistically enhanced the osteogenesis of KLF2+ hMSCs through upregulated vascular endothelial growth factor. A three-dimensional coculture model using cell-laden gelatin methacrylate (GelMA) hydrogel further confirmed these results. This study provides insight into the stemness-directed synergistic interaction between hMSCs and HUVECs, and our results will have a profound impact on further strategies involving the application of KLF2+ hMSC/HUVEC-laden GelMA hydrogel in vascular network bioengineering and bone regeneration.

Mechanical stimulation has been considered to be critical to cellular response and tissue regeneration. However, harnessing the direction of mechanical stimulation during osteogenesis still remains a challenge. In this study, we designed a series of novel magnetized collagen coatings (MCCs) (randomly or parallel-oriented collagen fibers) to exert the anisotropic mechanical stimulation using oriented magnetic actuation during osteogenesis. Strikingly, we found the osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs) were significantly up-regulated when the direction of magnetic actuation was parallel to the randomly-oriented collagen coating surface, in contrast to the down-regulated capacity under the perpendicular magnetic actuation. Moreover, further exerting a parallel mechanical stimulation along the parallel-oriented collagen coating, which cells have been oriented by the oriented collagens, were not only able to up-regulate the osteogenic differentiation of BMSCs but also promote the new bone formation during osteogenesis in vivo. We also demonstrated the anisotropic magneto-mechanical stimulation for the osteogenic differences might be attributed to the stretching or bending tensile status of collagen fibers controlled by the direction of magnetic actuation, driving the α5β1-dependent integrin signaling cascade. This study therefore got insight of understanding the directional mechanical stimulation on osteogenesis, and also paved a way for sustaining regulation of the biomaterials-host interface.

Light-induced surface potential have been demonstrated as an effective bone marrow mesenchymal stem cells (BMSCs) osteogenic differentiation regulator. However, traditional bone repair implants almost were weak or no light-responsive. Fortunately, surface modification was a feasible strategy to realize its light functionalization for bone implants. Herein, a graphene oxide (GO)/titanium dioxide (TiO

Mechanical stimulus has been demonstrated to be critical to stem cell fate commitment and tissue repair. However, it still remains a challenge to remote control of the mechanical stimulus acting on cells. Here, we designed a magnetic Fe3O4/mineralized collagen coating on titanium substrate to regulate the osteogenic differentiation of mesenchymal stem cells (MSCs). The mode and intensity of the mechanical stimulus acting on cells could be controlled by adjusting the remote applied magnetic field. We demonstrated that the adhesion, proliferation, and differentiation of MSCs were strongly dependent on the mode and intensity of the mechanical stimuli. Strikingly, the periodic mechanical stimulus (12 h every other day, PMS) showed the significantly up-regulated expression of osteogenesis-related markers, ALP, compared to that of the static mechanical stimulus mode. The reason is proposed as (1) initially, PMS mode enables the coatings to have appropriate surface mechanical properties for promoting focal adhesion, integrin expression, and cytoskeleton development of MSCs, letting MSCs have good capability of accepting as well as transferring mechanical stimuli; (2) during MSCs growth, PMS mode may effectively manipulate MSCs cytoskeleton development and movement, and mechanotranduction mechanism could be well activated; thus, MSCs osteogenic differentiation is enhanced. This work therefore provides a novel strategy to engineer bioactive coatings with remote control over the intensity and mode of the mechanical stimulus acting on cells, and would have an impact on the design of smart biomaterial surfaces for orthopedic applications.

Naringin, a Chinese herbal medicine, has been demonstrated to concentration-dependently promote osteogenic differentiation of mesenchymal stem cells (MSCs). However, it remains a challenge to load naringin on coatings for osteogenesis and further control the release kinetics. Here, we demonstrated that the release behavior of naringin on rutile nanorod films could be controlled by either mixing naringin with gelatin methacryloyl (GelMA) before spinning onto the films or soaking the obtained GelMA-incorporated films with the naringin solution to achieve the distinct degradation-type release and diffusion-type release, respectively. We further revealed that the naringin-loaded coatings facilitated adhesion, proliferation and late differentiation, and mineralization of MSCs. Our findings provided a novel strategy to engineer the coatings with controlled release of naringin and emphasized the bioactivity of naringin for the osteogenic differentiation of MSCs.

Natural extracellular matrices (ECMs) are three-dimensional (3D) and multi-scale hierarchical structure. However, coatings used as ECM-mimicking structures for osteogenesis are typically two-dimensional or single-scaled. Here, we design a distinct quasi-three-dimensional hierarchical topography integrated of density-controlled titania nanodots and nanorods. We find cellular pseudopods preferred to anchor deeply across the distinct 3D topography, dependently of the relative density of nanorods, which promote the osteogenic differentiation of osteoblast but not the viability of fibroblast. The in vivo experimental results further indicate that the new bone formation, the relative bone-implant contact as well as the push-put strength, are significantly enhanced on the 3D hierarchical topography. We also show that the exposures of HFN7.1 and mAb1937 critical functional motifs of fibronectin for cellular anchorage are up-regulated on the 3D hierarchical topography, which might synergistically promote the osteogenesis. Our findings suggest the multi-dimensions and multi-scales as vital characteristic of cell-ECM interactions and as an important design parameter for bone implant coatings.

A facile strategy is needed for accurate time-space supply of suitable growth factors or drugs. Polypyrrole (PPy) was able to carry almost all kinds of negatively charged biomolecules through anodizing method, which made it an appropriate way for codeposition of multiple molecules. The difference in the conjugation between different molecules and PPy makes it possible for selective release when the redox state of PPy changes. In this work, bovine serum albumin (BSA) and heparin (Hep) were chosen to be the model molecules in view of their differences in the level of electronegativity and molecular weight. Double-layer deposition method was used to improve the biocompatibility of PPy/BSA/Hep film. It was found the content of BSA and Hep in the film can be controlled by regulating deposition current and time. BSA release was facilitated under positive voltage and then promote the proliferation of preosteoblasts, while Hep release was promoted under negative voltage and enhance cell differentiation. Our work provides a dual-molecule model in PPy for selective release and further explores the mechanism of release selectivity, this discovery has potential applications in tissue engineering and regenerative medicine.

The surface electric potential of biomaterials has been extensively proven to play a critical role in stem cells' fate. However, there are ambiguous reports on the relation of stem cells' osteogenic capacity to surface potential characteristics (potential polarity and intensity). To address this, we adopted a surface with a wide potential range and both positive/negative polarity in a comprehensive view to get insight into surface potential-regulating cellular osteogenic differentiation. Tb

In this work, hydrothermal treatment and ultraviolet illumination (254 nm) were utilized to regulate the surface hydroxyls of titanium dioxide (TiO2) nanodots films. The effects on light-induced cell detachment behaviors were also investigated and discussed. Scanning electron microscopy, Raman, X-ray photoelectron spectroscopy and Circular dichroism were used to characterize the surface morphology, crystalline phase, surface composition and adsorption conformation of protein molecules. It was found that a combined process of ultraviolet illumination followed by hydrothermal treatment not only promoted cells adhesion but also showed the highest efficiency in light induced cell detachment. The reason is ascribed to that such process promotes the formation of terminal hydroxyl groups in TiO2 surface, and subsequently affect the adsorption conformation of protein molecules. This study indicated that such treatment could effectively modulate the contents of surface hydroxyl groups in different states. That shows much potential in optimizing the biological performance of TiO2 based biomaterials.

This study presents a convenient and versatile way to prepare functionalized composite polydopamine/titanium dioxide (PDA/TiO2) film on polystyrene (PS). First, polystyrene substrate was immersed in dopamine chloride solution, and then collosol containing TiO2 and water was spun on it, to produce uniform, continuous PDA/TiO2 composite films. The thickness of film was controllable by adjustment of the spin speed. It was found that the films were strongly adhered on the PS substrate, with peel strength and shear strength of 2.78 MPa and 37.78 MPa, respectively. After 20 min of ultraviolet (365 nm) illumination, over 90% of fibroblasts and 77% of osteoblasts detached from the PDA/TiO2 composite film. Additionally, the detached cells showed good viability, allowing further culture and applications. This preparation method could be widely applied for cell and cell sheet harvesting directly from PS-based culture wares.

Electrical stimulation has been proved to be critical to regulate cell behavior. But, cell behavior is also susceptible to multiple parameters of the adverse interferences such as surface current, electrochemical reaction products, and non-uniform compositions, which often occur during direct electric stimulation. To effectively prevent the adverse interferences, a novel piezoelectric poly(vinylidene fluoride-trfluoroethylene)(P(VDF-TrFE)) layer was designed to coat onto the indium tin oxide (ITO) planar microelectrode. We found the electrical stimulation was able to regulate the osteogenic differentiation of mesenchymal stem cells (MSCs) through calcium-mediated PKC signaling pathway. Meanwhile, the surface charge of the designed P(VDF-TrFE) coating layer could be easily controlled by the pre-polarization process, which was demonstrated to trigger integrin-mediated FAK signaling pathway, finally up-regulating the osteogenic differentiation of MSCs. Strikingly, the crosstalk in the downstream of the two signaling cascades further strengthened the ERK pathway activation for osteogenic differentiation of MSCs. This P(VDF-TrFE) layer coated electrical stimulation microelectrodes therefore provide a distinct strategy to manipulate multiple-elements of biomaterial surface to regulate stem cell fate commitment.

Light-induced cell harvest shows much potential in in vitro cell culture. In this work, a light-responsive monolayer graphene (Gr)/titanium dioxide nanodot (TN) film is designed and used for light-induced cell harvest. It is found that after 20 min of 365 nm UV or 450 nm visible light illumination, different types of cells could be detached from the surface effectively. The highest cell detachment ratio reaches about 95%. The mechanism of such a cell detachment is contributed to light illumination generates charge accumulation, which, in turn, changes the conformation of the extracellular matrix protein molecules adsorbed to a more disordered state, and eventually leads to the cells detachment. Such UV and visible light responsive Gr/TiO

Well-aligned collagen nanofibers are crucial in engineering bioinspired regenerative strategies, such as bone, muscle and cornea. However, keeping the natural bioactive of collagen and controlling its orientation in a coating still remain a challenge. Here we present a novel magnetically assisted electrochemical technique to deposit type-I collagen nanofibers with high alignment onto titanium. The magnetic assistance involved mainly the incorporation of iron oxide nanoparticles (IOPs) and the application of an external magnetic field during the electrochemical deposition. The combination of IOPs with the collagen nanofibrils in electrolyte endowed the nanofibrils with magnetism, which forced the collagen nanofibrils to be straightened and assembled into aligned nanofibers under magnetic field during electrodeposition. The influence of the applied magnetic field on orientational order of the collagen nanofibers in the coatings extended to drying stage. The aligned collagen coatings demonstrated to favorably guide the bone marrow mesenchymal stem cells (BMSCs) grow in the form of elongated morphology, which promoted the cellular osteogenic differentiation dramatically. The present magnetically assisted electrodeposition could emerge as an attractive approach to fabrication of aligned nanofibers on substrates for subsequent uses such as bone tissue engineering.

Cell responses to oxide biomaterials depend on the protein adsorption behavior of the biomaterial surface. Thus, the inherent properties of oxide biomaterial surfaces play a key role in this process. However, commonly used biomaterials, such as calcium phosphate and titanium dioxide, have surfaces with strong mineralization, which may interfere with the ability to clarify the key aspects of the oxide biomaterial regarding protein adsorption and cellular processes. Here, nonmineralized crystalline and vitreous silica were selected as model oxide biomaterials to explore the inherent properties of these materials on the absorption behavior of the functional protein fibronectin (Fn) and on the osteogenic differentiation of mesenchymal stem cells (MSCs). We demonstrated that due to the smaller O

The immune response of bone implants is closely related to the interaction between macrophages and biomaterial surfaces. In this work, the polarization behavior of mouse bone marrow-derived macrophages (BMDMs), including their morphology and expression of phenotypic markers, genes and cytokines, on charged surfaces with different potential intensities was systematically explored. We found that the charged surface could effectively promote BMDM polarization, and a higher potential intensity was conducive to the upregulation of the polarization of BMDMs into the M2 phenotype. Based on the analysis of the signaling pathways involved in integrins (αMβ2 and α5β1) and the potassium ion channel (Kv1.3), a possible underlying mechanism revealed that the integrin originated signaling pathways could more dominantly regulate macrophage polarization to the M2 phenotype. The present work therefore demonstrates that the surface charge, as a physicochemical property of material surfaces, could effectively regulate macrophage polarizations, which may provide an immunoregulation view for the surface design of biomaterials.

Loading of an appropriate amount of rhBMP-2 and avoiding its “burst-release” are key challenges for upgrading the biological performance of thin bioactive coatings on metal implants. In this study, we adopted incorporation of chitosan nanospheres into thin mineralized collagen coatings to enhance rhBMP-2 loading and improve releasing behavior based on the good affinity of chitosan for proteins and the large surface area of nanospheres. We realized the incorporation process by electrophoretic injection. Using chitosan nanospheres, we were able to increase the rhBMP-2 loading amount in the thin coating by 2.7-fold (from 446 ng cm−2 to 1186 ng cm−2), and showed that the rhBMP-2 exhibited sustained release behavior. MC3T3-E1 cells cultured on the rhBMP-2/chitosan nanosphere-incorporated thin coatings (Col/Cs/BMP) showed good cell attachment and proliferative behavior, and high levels of differentiation and mineralization. In in vivo tests, spiral CT analysis and histological observations demonstrated that Col/Cs/BMP coatings on metal implants were able to increase bone density and accelerate new bone growth after implantation for 4 to 8 weeks, and the boundary between host bones and new bones disappeared after implantation for 8 weeks. The pull-out tests further confirmed that the Col/Cs/BMP coatings could significantly enhance osseointegration. The present results indicate that incorporation of chitosan nanospheres into thin coatings is an effective way to enhance the loading amount properly, improve the release behavior of rhBMP-2 and finally accelerate the osseointegration process.

Surface-mediated gene delivery has attracted more and more attentions in biomedical research and applications because of its characteristics of low toxicity and localized delivery. Herein, a novel visible-light-regulated, surface-mediated gene-delivery platform is exhibited, arising from the photoinduced surface-charge accumulation on silicon. Silicon with a pn junction is used and tested subsequently for the behavior of surface-mediated gene delivery under visible-light illumination. It is found that positive-charge accumulation under light illumination changes the surface potential and then facilitates the delivery of gene-loaded carriers. As a result, the gene-expression efficiency shows a significant improvement from 6% to 28% under a 10 min visible-light illumination. Such improvement is ascribed to the increase in surface potential caused by light illumination, which promotes both the release of gene-loaded carriers and the cellular uptake. This work suggests that silicon with photovoltaic effect could offer a new strategy for surface-mediated, gene-delivery-related biomedical research and applications.

Invited for this month's cover is the interdisciplinary research group of Prof. Kui Cheng from Zhejiang University, China. The cover picture shows a schematic figure of carbon quantum dots incorporated as a sensitizer into TiO

Graphene (Gr)/poly(vinylidenefluoride-co-trifluoroethylene) (P(VDF-TrFE)) composite films were annealed by the photothermal effect that produced between near-infrared laser and graphene. Raman spectra, X-ray Diffraction (XRD), Fourier-Transformed Infrared Spectroscopy (FTIR), Differential scanning calorimetry (DSC) are used to characterize the crystalline state of Gr/P(VDF-TrFE) films. Further, piezoelectric coefficient d33 were to test polarization performance. Finally, the Bovine serum albumin (BSA) adsorption shows that annealing and polarization of Gr/P(VDF-TrFE) films are important for protein adsorption.

Polydopamine (PDA) is capable of adhering on nearly all kinds of surfaces and shows good biocompatibility. Moreover, its surface state could be switched between oxidation and reduction states under different electrical stimulation. In this work, the effects of PDA redox states on protein adsorption, as well as the subsequent effects on cellular responses were characterized and evaluated. It was found that the electrical treatment changed the redox states of PDA, which in turn changed the state of preadsorbed protein molecules and eventually affected the cellular responses. BSA preadsorbed PDA film was found to be beneficial for cell proliferation when PDA was changed into reduction state (RPDA), while BMP-2 preadsorbed PDA film showed promotion on cell differentiation when PDA was changed into oxidation state (OPDA). It was found that the transitions of PDA to RPDA and OPDA greatly changed the secondary structure of protein preadsorbed on it. This work provides a deeper insight on changes of protein molecules between cells and PDA surfaces with different redox states, which is important for both optimization of cell-material interactions and application of PDA as a functional coating for biomedical engineering.

Surface potential of biomaterials can dramatically influence cellular osteogenic differentiation. In this work, a wide range of surface potential on ferroelectric polyvinylidene fluoride trifluoroethylene (P(VDF-TrFE)) films was designed to get insight into the interfacial interaction of cell-charged surface. The P(VDF-TrFE) films poled by contact electric poling at various electric fields obtained well stabilized surface potential, with wide range from −3 to 915 mV. The osteogenic differentiation level of cells cultured on the films was strongly dependent on surface potential and reached the optimum at 391 mV in this system. Binding specificity assay indicated that surface potential could effectively govern the binding state of the adsorbed fibronectin (FN) with integrin. Molecular dynamic (MD) simulation further revealed that surface potential brought a significant difference in the relative distance between RGD and synergy PHSRN sites of adsorbed FN, resulting in a distinct integrin-FN binding state. These results suggest that the full binding of integrin α5β1 with both RGD and PHSRN sites of FN possesses a strong ability to activate osteogenic signaling pathway. This work sheds light on the underlying mechanism of osteogenic differentiation behavior on charged material surfaces, and also provides a guidance for designing a reasonable charged surface to enhance osteogenic differentiation. Statement of Significance The ferroelectric P(VDF-TrFE) films with steady and a wide range of surface potential were designed to understand underlying mechanism of cell-charged surface interaction. The results showed that the charged surface well favored upregulation of osteogenic differentiation of MC3T3-E1 cells, and more importantly, a highest level occurred on the film with a moderate surface potential. Experiments and molecular dynamics simulation demonstrated that the surface potential could govern fibronectin conformation and then the integrin-fibronectin binding. We propose that a full binding state of integrin α5β1 with fibronectin induces effective activation of integrin-mediated FAK/ERK signaling pathway to upregulate cellular osteogenic differentiation. This work provides a guidance for designing a reasonable charged surface to enhance osteogenic differentiation.

Titanium and tantalum have been widely used for orthopedic and dental implant applications. However, how their inherent surface features regulate cellular osteogeneses still remains elusive. In this study, we engineered two distinct TiO2 and Ta2O5 nanorod films as the two model oxidized surfaces to investigate their intrinsic osteogenic behaviors. The results indicated that the distinctive gradient on zeta potential against pH, corresponding to the deprotonation rate, but not the hydroxyl amount or hydroxylation polarity played a critical role on the cellular osteogenic performance. TiO2 nanorod film with a higher deprotonation rate significantly upregulated the expression of osteogeneses-related gene and protein, comparing to that of Ta2O5 nanorod film. These results might be attributed to that surface with higher deprotonation rateprovided more Bronsted acid-base surface sites to react with protein residues, leading to a mild change in conformation of the absorbed proteins, and subsequently facilitating to trigger the integrin-focal adhesion cytoskeleton actin transduction pathway. This study, therefore, provides a new insight into the understanding the role of material surface hydroxylation on cellular osteogenic responses.

Three-dimensional (3D) printing technology has been widely used to fabricate of various titanium and its alloy implants. However, engineering the 3D printing nanoscaled feature to realize a hierarchical micronano structured surface topography still remains a challenge. On one hand, enhanced bioactivity is always expected on micronano-hybrid biomimetic topography; on the other hand, a typical functional protein in extracellular matrix (ECM) is nanoscaled; therefore, nanoscaled features might affect its binding to specific receptor and subsequent cell response. Here, we engineered a novel hierarchical structure with microparticles and rutile TiO2 nanorods topography that fabricated by 3D printing of pure titanium followed by a hydrothermal process. Although there was no difference on the microscaled feature before/after nanonization, cellular behaviors including adhesion, proliferation, and osteogenic differentiation of mesenchymal stem cells (MSCs) were significantly upregulated on the hierarchical microna...

Mechanical stimuli at the bone-implant interface are considered to activate the mechanotransduction pathway of the cell to improve the initial osseointegration establishment and to guarantee clinical success of the implant. However, control of the mechanical stimuli at the bone–implant interface still remains a challenge. In this study, we have designed a strategy of a magnetically responsive coating on which the mechanical stimuli is controlled because of coating deformation under static magnetic field (SMF). The iron oxide nanoparticle/mineralized collagen (IOP-MC) coatings were electrochemically codeposited on titanium substrates in different quantities of IOPs and distributions; the resulting coatings were verified to possess swelling behavior with flexibility same as that of hydrogel. The relative quantity of IOP to collagen and the IOP distribution in the coatings were demonstrated to play a critical role in mediating cell behavior. The cells present on the outer layer of the distributed IOP-MC (O-IOP-MC) coating with a mass ratio of 0.67 revealed the most distinct osteogenic differentiation activity being promoted, which could be attributed to the maximized mechanical stimuli with exposure to SMF. Furthermore, the enhanced osteogenic differentiation of the stimulated MC3T3-E1 cells originated from magnetically actuated mechanotransduction signaling pathway, embodying the upregulated expression of osteogenic-related and mechanotransduction-related genes. This work therefore provides a promising strategy for implementing mechanical stimuli to activate mechanotransduction on the bone–implant interface and thus to promote osseointegration. Statement of Significance The magnetically actuated coating is designed to produce mechanical stimuli to cells for promoting osteogenic differentiation based on the coating deformation. Iron oxide nanoparticles (IOPs) were incorporated into the mineralized collagen coatings (MC) forming the composite coatings (IOP-MC) with spatially distributed IOPs, and the IOP-MC coatings with outer distributed IOPs (O-IOPs-MC) shows the maximized mechanical stimuli to cells with enhanced osteogenic differentiation under static magnetic field. The upregulated expression of the associated genes reveals that the enabled mechanotransduction signaling pathway is responsible for the promoted cellular osteogenic differentiation. This work therefore provides a promising strategy for implementing mechanical stimuli to activate mechanotransduction on the bone–implant interface to promote osseointegration.

In the present study, TiO2/ZnO composite nanodot films were prepared and the effects of Zn incorporation on light-induced cell detachment were investigated. The nanodots films, which were successfully synthesized by phase-separation-induced self-assembly method, were characterized on the morphology, composition, microstructure, and other properties, and evaluated on cell compatibility and cell detachment performances as well. Live-dead staining was used to study the viability of cell sheet detached by light illumination. Results shows that with the increasing of introduced Zn, the band gap widened and the absorbance in UV region increased, while the crystallinity and performance of light-induced hydrophilicity weakened. All the nanodots films showed good cell compatibility and cell detachment performance induced by light. The nanodots film which had a Zn/Ti molar ratio of 0.03 showed the highest detachment ratio of 91.0% after 20 min ultraviolet illumination. The prepared TiO2/ZnO composite nanodots films could be helpful in optimizing light-induced cell detachment behavior.

In electrical stimulation (ES), daily stimulation time means the interacting duration with cells per day, and is a vital factor for mediating cellular function. In the present study, the effect of stimulation time on osteogenic differentiation of MC3T3-E1 cells was investigated under ES on polypyrrole (Ppy) planar interdigitated electrodes (IDE). The results demonstrated that only a suitable daily stimulation time supported to obviously upregulate the expression of ALP protein and osteogenesis-related genes (ALP, Col-I, Runx2 and OCN), while a short or long daily stimulation time showed no significant outcomes. These might be attributed to the mechanism that an ES induced transient change in intracellular calcium ion concentration, which was responsible for activating calcium ion signaling pathway to enhance cellular osteogenic differentiation. A shorter daily time could lead to insufficient duration for the transient change in intracellular calcium ion concentration, and a longer daily time could give rise to cellular fatigue with no transient change. This work therefore provides new insights into the fundamental understanding of cell responses to ES and will have an impact on further designing materials to mediate cell behaviors.

Light-induced surface potential have been demonstrated as an effective bone marrow mesenchymal stem cells (BMSCs) osteogenic differentiation regulator. However, traditional bone repair implants almost were weak or no light-responsive. Fortunately, surface modification was a feasible strategy to realize its light functionalization for bone implants. Herein, a graphene oxide (GO)/titanium dioxide (TiO2) nanodots composite coating on the surface of titanium (Ti) implant was constructed, and GO was reduced to reduced graphene oxide (rGO) with the method of UV-assisted photocatalytic reduction. After rGO deposited on the surface of TiO2, a heterojunction formed at the interface of rGO and TiO2. With visible light illumination, positive charges accumulated on the surface of rGO/TiO2 film, and performed as a positive surface potential change. The light-induced surface potential which was generated under proper light intensity is harmless to the cell adhesion and proliferation behavior, but presented a good BMSCs osteogenic differentiation promoting effect, and the activation of the voltage-gated calcium channels through surface potential and the promotion of the adsorption of osteogenic growth factors could be the reason. This work given a new insight of the modification for Ti implant with a light-induced surface potential, and shows potential application for bone regeneration on the clinical practice through light stimulation.

The combination of bone marrow-derived mesenchymal stem cells (BMSCs) and biological scaffolds has been demonstrated to be a promising strategy for bone regeneration. However, this method does not result in satisfactory bone regeneration, because the BMSCs are dispersed in the biological scaffolds. The current study developed a new bone regeneration system, which combines synthetic porous three-dimensional scaffolds of β-TCP/COL-I composite with cultured osteogenic sheets of BMSCs. Activity of alkaline phosphatase (ALP), a marker of bone regeneration, was assayed in vitro using enzyme-linked immunosorbent assays and quantitative real-time polymerase chain reaction. In vivo bone regeneration was assayed in male nude mice. The study samples were BMSC sheet, scaffold/scattered BMSCs, scaffold/BMSC sheet, and scaffold alone. The samples were implanted dorsally in the mice. In vitro analysis showed that β-TCP/COL-I scaffold combined with BMSC sheets significantly upregulated both gene expression and protein levels of ALP, osteocalcin, and osteopontin. Histological and micro-computed tomography showed that the only implants that demonstrated new bone formation after 4 weeks were scaffold/BMSC sheet implants. These results underscore the crucial requirement of a synergistic effect of β-TCP/COL-I scaffolds and BMSC sheets. This could be a promising novel strategy for bone tissue engineering. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 106B: 2037-2045, 2018.

The topography at the micro/nanoscale level for biomaterial surfaces has been thought to play vital roles in their interactions with cells. However, discovering the interdisciplinary mechanisms underlying how cells respond to micro-nanostructured topography features still remains a challenge. In this work, ∼37 μm 3D printing used titanium microspheres and their further hierarchical micro-nanostructured spheres through hydrothermal treatment were adopted to construct typical model surface topographies to study the preosteoblastic cell responses (adhesion, proliferation, and differentiation). We here demonstrated that not only the hierarchical micro-nanostructured surface topography but also their distribution density played critical role on cell cytocompatibility. The microstructured topography feature surface with middle-density distributed titanium microspheres showed significantly enhanced cell responses, which might be attributed to the better cellular interaction due to the cell aggregates. However, the hierarchical micro-nanostructured topography surface, regardless of the distribution density of titanium microspheres, improved the cell-surface interactions because of the enhanced initial protein adsorption, thereby reducing the cell aggregates and consequently their responses. This work, therefore, provides new insights into the fundamental understanding of cell-material interactions and will have a profound impact on further designing micro-nanostructured topography surfaces to control cell responses.