3D Printing Hydrogel Scaffolds with Nanohydroxyapatite Gradient to Effectively Repair Osteochondral Defects in Rats

Osteochondral (OC) defects pose an enormous challenge with no entirely satisfactory repair strategy to date. Herein, a 3D printed gradient hydrogel scaffold with a similar structure to that of OC tissue is designed, involving a pure hydrogel-based top cartilage layer, an intermediate layer for calcified cartilage with 40% (w w⁻¹) nanohydroxyapatite (nHA) and 60% (w w⁻¹) hydrogel, and a 70/30% (w w⁻¹) nHA/hydrogel-based bottom subchondral bone layer. This study is conducted to evaluate the efficacy of the scaffold with nHA gradients in terms of its ability to promote OC defect repair. The fabricated composites are evaluated for physicochemical, mechanical, and biological properties, and then implanted into the OC defects in 56 rats. Overall, bone marrow stromal cells (BMSCs)-loaded gradient scaffolds exhibit superior repair results as compared to other scaffolds based on gross examination, micro-computed tomography (micro-CT), as well as histologic and immunohistochemical analyses, confirming the ability of this novel OC graft to facilitate simultaneous regeneration of cartilage-subchondral bone.     

Intermetallic Compound Formation and Evolution in Solid-State Sn/Immersion-Ag/Cu Trilayer Interfacial Reactions on a Flexible Polymer Board

Immersion Ag is a promising candidate Pb-free surface finish on printed circuit boards (PCBs). For flexible PCB and optoelectronic packaging, solid-state bonding rather than reflow is commonly used to join the chips to the PCB with Sn-based solders, after which the immersion Ag layer remains at the joint interface and participates in the interfacial reactions at the solder joints. Solder joint samples composed of a Sn/Ag/Cu trilayer on flexible PCBs were prepared to study the interfacial reactions at 150°C and 200°C. Three phases, Ag₃Sn, Cu₆Sn₅, and Cu₃Sn, were sequentially formed at the interface. Remarkable change of the morphology of the Ag₃Sn phase was observed during thermal aging. The thickness of the immersion Ag layer was found to have significant effects on the growth rates of the Cu₆Sn₅ and Cu₃Sn phases and the void formation in the Cu₃Sn phase.     

Solid-state interfacial reactions at the solder joints employing Au/Pd/Ni and Au/Ni as the surface finish metallizations

A tri-layer of nickel/palladium/gold (Au/Pd/Ni) is a promising candidate to replace the conventional Au/Ni bi-layer as the surface finish metallization for lead-free packaging. A surface finish metallization (Au/Pd/Ni or Au/Ni) and a Sn layer are sequentially deposited on a Cu substrate and then are subjected to thermal aging at 150 and 200 °C to investigate the interfacial reactions in the stacking multilayer structure made by low-temperature solid-state bonding. Because of the absence of the reflow process, the Pd and Au layers do not dissolve in the Sn matrix but remain at the interface and participate in the interfacial reaction to form the (Pd,Ni,Au)Sn₄ and (Au,Ni)Sn₄ phases at the Au/Pd/Ni- and Au/Ni-based interfaces, respectively. Though the Pd layer was only 0.4 μm, its resulting (Pd,Ni,Au)Sn₄ phase is much thicker than the (Au,Ni)Sn₄ phase. These two intermetallic compounds exhibit very different microstructural evolution which significantly affects the interfacial microstructures and growth rate of other intermetallic compound formed at the same interfaces.     

Lamellar Mesostructured Silicas with Chemically Significant Hierarchical Morphologies

We describe the hierarchical structures of mesostructured silicas assembled from electrically neutral and unsymmetrical Gemini surfactants of the type C_nH_2n+1NH(CH₂)_mNH₂ with n = 10, 12, 14 and m = 3, 4. As expected for Gemini surfactants with an all anti‐chain configuration and a packing parameter near 1.0, lamellar framework structures are formed, regardless of the length of the alkyl chain (n) and the number of carbon atoms (m) linking the two amino group centers. However, different layer curvatures and levels of hierarchical structure are observed depending on the delicate balance between the hydrophilic interactions at the surfactant head group–silica interface and the hydrophobic interactions between the surfactant alkyl groups. For Gemini derivatives with n = 12 or 14 and m = 3 or 4, well‐expressed hierarchical vesicles are formed that are analogous to those assembled previously from Gemini surfactants with m = 2. However, for n = 10, a new coiled slab structure (m = 3) and an onion‐like core–shell structure (m = 4) are formed. In addition, a previously unobserved stripe‐like silica structure is obtained from a C⁰₁₂₊₂₊₀ Gemini surfactant in combination with an α,ω‐diamine co‐surfactant. The relative stability of these hierarchical structures depends on the delicate competition between the long‐range elastic forces occurring in the hydrophobic region of the assembled surfactant and the short‐range chemical forces in the hydrophilic moiety. Lamellar silicas with hierarchical vesicular structures, the new coiled slab, and stripe‐like phases promise to be chemically significant morphologies, because they can minimize the framework pore length and provide optimal access to the framework walls under diffusion‐limited conditions.     

Screening self-assembled monolayers as Cu diffusion barriers

Self-assembled monolayers (SAMs) are investigated as potential Cu diffusion barriers for application in back-end-of-line (BEOL) interconnections. A screening of SAMs derived from molecules with different head group (SiCl₃, Si(OCH₃)₃, Si(OCH₃)Cl₂) bonding to the dielectric substrate, chain lengths (n = 3-21) and terminal group (CH₃, Br, CN, NH₂, C₅H₄N and SH) bonding to the Cu overlayer are compared in terms of inhibition of interfacial Cu diffusion and promotion of Cu-SiO₂ adhesion. SAM barrier properties against Cu silicide formation are examined upon annealing from 200 to 400 °C by visual inspection, sheet resistance measurements (Rs) and X-ray Diffraction Spectroscopy (XRD). Cu/SAM/SiO₂ adhesion is evaluated by tape test and four-point probe measurements. Results indicate that NH₂-SAM derived from 3-aminopropyltrimethoxysilane is the most promising for Cu diffusion barrier application. Silicide formation is inhibited to at least 400 °C, essential stability for BEOL integration. However, the 2.9 Gc (J/m²) adhesion of the layer compared with 3.1 Gc (J/m²) on SiO₂ does need improvement.     

Interfacial Reactions in Cu/Ga and Cu/Ga/Cu Couples

Cu-to-Cu bonding to connect through-silicon vias in three-dimensional integrated-circuit packaging is the most important interconnection technology in the next-generation semiconductor industry. Soldering is an economic and fast process in comparison with diffusion bonding methods. Ga has high solubility of up to 20 at.% in the Cu-rich face-centered cubic (FCC) phase and high mobility at moderate temperatures. In this work, an attempt has been made to evaluate Ga-based Cu-to-Cu interconnection by transient liquid-phase (TLP) bonding. The Cu/Ga interfacial reactions at temperatures ranging from 160°C to 300°C were examined. For reactions at temperatures lower than 240°C, the reaction path is Cu/γ ₃-Cu₉Ga₄/θ-CuGa₂/liquid, where the γ ₃-Cu₉Ga₄ and θ-CuGa₂ phases are thin planar and thick scalloped layers, respectively, while for the reactions at 280°C and 300°C, the scalloped γ ₃-Cu₉Ga₄ phase is the only reaction product. The phase transformation kinetics, reaction mechanisms, and microstructural evolution in the Cu/Ga couples are elaborated. In addition, reactions of Cu/Ga/Cu sandwich couples at 160°C were investigated. The original Cu/liquid/Cu couples isothermally transformed to Cu/γ ₃-Cu₉Ga₄/ θ-CuGa₂/γ ₃-Cu₉Ga₄/Cu couples as the reaction progressed. However, cracks were observed in the θ-CuGa₂ phase regions after metallographic processing. The brittle θ-CuGa₂ phase is undesirable for Ga-based TLP bonding.     

A Hermetic Seal Using Composite Thin-Film In/Sn Solder as an Intermediate Layer and Its Interdiffusion Reaction with Cu

A bonding joint between Cu metallization and evaporated In/Sn composite solder is produced at a temperature lower than 200°C in air. The effects of bonding temperature and duration on the interfacial bonding strength are studied herein. Cross sections of bonding joints processed at different bonding conditions were examined by scanning electron microscopy (SEM). The optimal condition, i.e., bonding temperature of 180°C for 20 min, was chosen because it gave rise to the highest average bonding strength of 6.5 MPa, and a uniform bonding interface with minimum voids or cracks. Good bond formation was also evidenced by scanning acoustic imaging. For bonding couples of patterned dies, a helium leak rate of 5.8 × 10⁻⁹ atm cc/s was measured, indicating a hermetic seal. The interfacial reaction between Cu and In/Sn was also studied. Intermetallic compounds (IMCs) such as AuIn₂, Cu₆Sn₅, and Cu₁₁In₉ were detected by means of x-ray diffraction analysis (XRD), and transmission electron microscopy (TEM) accompanied by energy-dispersive x-ray (EDX) spectroscopy. Chemical composition analysis also revealed that solder interlayers, Sn, and In were completely converted into IMCs by reaction with Cu. All the IMCs formed in the joints have remelting temperatures above 300°C according to the Cu-In, Cu-Sn, and Au-In phase diagrams. Therefore, the joint is able to sustain high service temperatures due to the presence of these IMCs.     

Effects of Pd addition on Au stud bumps/Al pads interfacial reactions and bond reliability

The main purposes for developing low-alloyed Au bonding wires were to increase wire stiffness and to control the wire loop profile and heat-affected zone length. For these reasons, many alloying elements have been used for the various Au bonding wires. Although there have been many studies reported on wire strengthening mechanisms by adding alloying elements, few studies were performed on their effects on Au bonding wires and Al pad interfacial reactions. Palladium has been used as one of the important alloying elements of Au bonding wires. In this study, Au-1wt.%Pd wire was used to make Au stud bumps on Al pads, and effects of Pd on Au/Al interfacial reactions, at 150°C, 175°C, and 200°C for 0 to 1200 h thermal aging, were investigated. Cross-sectional scanning electron microscopy (SEM), energy-dispersive spectroscopy (EDS), and electron probe microanalysis (EMPA) were performed to identify intermetallic compound (IMC) phases and Pd behavior at the Au/Al bonding interface. According to experimental results, the dominant IMC was Au₅Al₂, and a Pd-rich layer was at the Au wire and Au-Al IMC. Moreover, Au-Al interfacial reactions were significantly affected by the Pd-rich layer. Finally, bump shear tests were performed to investigate the effects of Pd-rich layers on Au wire bond reliability, and there were three different failure modes. Cracks, accompanied with IMC growth, formed above a Pd-rich layer. Furthermore, in longer aging times, fracture occurred along the crack, which propagated from the edges of a bonding interface to the center along a Pd-rich layer.     

Low-Temperature,Strong SiO<Subscript>2</Subscript>-SiO<Subscript>2</Subscript> Covalent Wafer Bonding for III–V Compound Semiconductors-to-Silicon Photonic Integrated Circuits

We report a low-temperature process for covalent bonding of thermal SiO₂ to plasma-enhanced chemical vapor deposited (PECVD) SiO₂ for Si-compound semiconductor integration. A record-thin interfacial oxide layer of 60 nm demonstrates sufficient capability for gas byproduct diffusion and absorption, leading to a high surface energy of 2.65 J/m² after a 2-h 300°C anneal. O₂ plasma treatment and surface chemistry optimization in dilute hydrofluoric (HF) solution and NH₄OH vapor efficiently suppress the small-size interfacial void density down to 2 voids/cm², dramatically increasing the wafer-bonded device yield. Bonding-induced strain, as determined by x-ray diffraction measurements, is negligible. The demonstration of a 50 mm InP epitaxial layer transferred to a silicon-on-insulator (SOI) substrate shows the promise of the method for wafer-scale applications.     

Physically Crosslinked Biocompatible Silk‐Fibroin‐Based Hydrogels with High Mechanical Performance

Developing hydrogel which combines superior mechanical performance and biocompatibility attracts researchers’ attention in recent years. Here, a novel biocompatible hydrogel with excellent mechanical performance, comprised of regenerated silk fibroin (RSF) and hydroxypropyl methyl cellulose (HPMC), is fabricated by simply mixing and heating. It is found that both of compressive modulus and tensile modulus of the optimal RSF/HPMC hydrogel are over 1.0 MPa. Meanwhile, the break energy is up to 3500 J m^?2, which is higher than that of some natural elastomers, such as cartilage, cork, and skin. The investigation of gelation mechanism reveals that more uniformly dispersed crosslinks dominated by smaller β‐sheet structures, which is attributed to the synergistic effects of hydrogen bonding and hydrophobic interaction between HPMC and RSF molecules, contribute to the superior mechanical performance of RSF/HPMC hydrogel. This biocompatible high strength silk protein based hydrogel diversifies the robust hydrogels and holds a great promise as candidates for load‐bearing materials in biomedical field.     

Phase Transitions in the BiVO<Subscript>4</Subscript>-<Emphasis Type="Italic">n</Emphasis>PbO (<Emphasis Type="Italic">n</Emphasis> = 1, 2) System: Structural–Electrical Properties Relationships

In a general survey of nPbO-BiVO₄ compounds, interesting phases corresponding to n = 1: PbBiVO₅, and n = 2: Pb₂BiVO₆ are described. A phase transition has been unambiguously characterized for PbBiVO₅. The crystal structures were solved from twinned crystals at room temperature (α phase, triclinic, S.G. P-1) and at 530°C (β phase, monoclinic, C2/m). Powder neutron diffraction experiments confirmed these settings and both room-temperature (RT) and high-temperature (HT) refinements corroborated space group choices, clearing up a literature controversy about the centrosymmetry of the α phase, and identifying structural modifications occurring under the α → β transition. Cationic substitutions for V were tested and PbBi(V_1−x M _x )O₅ (M = P) solid solutions identified. Pb₂BiVO₆ (n = 2) is a compound showing several successive structural transitions, i.e., α → β → δ. Structures of α and δ forms have been previously described from powder diffraction data (x-ray and neutron). In this work, we have refined these structures from single-crystal data, and the resolution of the intermediate β form, so far unsolved, was possible through a stabilization thermal cycle; its complete structural understanding required a 4D formalism. Two new polymorphic phases, α′ and δ′, were obtained by substituting Mn or P for V; their structures are closely related to, respectively, the α phase at room temperature, and the δ phase at 680°C. Electrical conductivities of all structurally characterized compositions were investigated, and correlations were drawn between their conduction properties and structural characteristics. Conductivity properties measured under variable O₂ partial pressures for Pb₂Bi(V_0.75P_0.25)O₆ were interpreted as a mixed ionic–electronic (p-type) conduction mechanism.     

Schottky Heterojunction Facilitates Osteosarcoma Ferroptosis and Enhances Bone Formation in a Switchable Mode

Effective antitumor agents with concurrent osteogenic properties are essential for comprehensive osteosarcoma (OS) treatment. However, the current clinical therapeutic strategies of OS fail to completely eradicate tumors while simultaneously encouraging bone formation. To address this issue, a switchable strategy for dynamic OS ablation and static bone regeneration is developed by integrating piezoelectric BaTiO₃ (BTO) with atomic-thin Ti₃C₂ (TC) through a Schottky heterojunction, resulting in the formation of TC@BTO. Under sequential ultrasound and near-infrared irradiation, the optimized carrier transport of TC@BTO, based on Schottky heterojunction, exhibits excellent characteristics of photothermal conversion and reactive oxygen species generation. This results in ferroptosis of tumor cells and eventual elimination of OS. Moreover, in the static state, the interfacial Schottky heterojunction facilitates the carriers’ directed transfer from the semiconductor to the metal. The Schottky heterojunction-enhanced static electrical stimulation enhances the osteogenic differentiation of bone marrow-derived mesenchymal stem cells and repair of bone defects. Furthermore, RNA-sequencing analysis reveals that static TC@BTO promotes bone regeneration by activating Wnt signaling pathway, and remarkably, pharmacological inhibition of Wnt signaling suppresses the TC@BTO-induced osteogenesis. Overall, this work broadens the biomedical potential of Schottky heterojunction-based therapies and provides a comprehensive strategy for overall OS ablation and bone regeneration.     

Interfacial reaction between multicomponent lead-free solders and Ag,Cu, Ni,and Pd substrates

The interfacial reaction between two prototype multicomponent lead-free solders, Sn-3.4Ag-1Bi-0.7Cu-4In and Sn-3.4Ag-3Bi-0.7Cu-4In (mass%), and Ag, Cu, Ni, and Pd substrates are studied at 250°C and 150°C. The microstructural characterization of the solder bumps is carried out by scanning electron microscopy (SEM) coupled with energy dispersive x-ray analysis. Ambient temperature, isotropic elastic properties (bulk, shear, and Young’s moduli and Poisson’s ratio) of these solders along with eutectic Sn-Ag, Sn-Bi, and Sn-Zn solders are measured. The isotropic elastic moduli of multicomponent solders are very similar to the eutectic Sn-Ag solder. The measured solubility of the base metal in liquid solders at 250°C agrees very well with the solubility limits reported in assessed Sn-X (X=Ag, Cu, Ni, Pd) phase diagrams. The measured contact angles were generally less than 15° on Cu and Pd substrates, while they were between 25° and 30° on Ag and Ni substrates. The observed intermediate phases in Ag/solder couples were Ag₃Sn after reflow at 250°C and Ag₃Sn and ζ (Ag-Sn) after solid-state aging at 150°C. In Cu/solder and Ni/solder couples, the interfacial phases were Cu₆Sn₅ and (Cu,Ni)₆Sn₅, respectively. In Pd/solder couples, only PdSn₄ after 60-sec reflow, while both PdSn₄ and PdSn₃ after 300-sec reflow, were observed.     

A Study on the Breakdown Mechanism of an Electroless-Plated Ni(P) Diffusion Barrier for Cu/Sn/Cu 3D Interconnect Bonding Structures

This study examined the thermal stability of an electroless-plated Ni(P) barrier layer inserted between Sn and Cu in the bonding structure of Cu/Sn/Cu for three-dimensional (3D) interconnect applications. A combination of transmission electron microscopy (TEM) and scanning electron microscopy allowed us to fully characterize the bonding morphology of the Cu/Ni(P)/Sn/Ni(P)/Cu joints bonded at various temperatures. The barrier suppressed Cu and Sn interdiffusion very effectively up to 300°C; however, an interfacial reaction between Ni(P) and Sn led to gradual decomposition into Ni₃P and Ni₃Sn₄. Upon 350°C bonding, the interfacial reaction brought about complete disintegration of the barrier in local areas, which allowed unhindered interdiffusion between Cu and Sn.     

Cross-Interaction Between Au/Sn and Cu/Sn Interfacial Reactions

The cross-interaction between Sn/Cu and Sn/Au interfacial reactions in an Au/Sn/Cu sandwich structure was studied. Field-emission electron probe microanalysis (FE-EPMA) revealed that the Cu content in the three Au-Sn phases (AuSn, AuSn₂, and AuSn₄) was very low, less than 1 at.%. This means␣that Cu from the opposite Cu foil did not participate in the interfacial reaction at the Sn/Au interface. On the opposite Sn/Cu side, Au-substituted (Cu,Au)₆Sn₅ formed within the initial 1 min of reflow. With prolonged reflow, the Au content in the Au-substituted (Cu,Au)₆Sn₅ increased and it transformed into a Cu-substituted (Au,Cu)Sn phase with 25 at.% Cu after 1 min of reflow at 250°C. The x-ray diffraction (XRD) pattern confirmed the phase transformation of Au-substituted (Cu,Au)₆Sn₅ to Cu-substituted (Au,Cu)Sn phase. In addition, there was greater Au consumption in the Au/Sn/Cu sandwich joint structure than in the single Au/Sn reaction case, due to some of the Au participating in the opposite Sn/Cu interfacial reaction.     

Exact solutions to the problem of wave diffraction from a Bragg grating with an apodized asymmetric, symmetric, or antisymmetric coupling factor

The coupling-wave model is applied to obtain an exact analytic solution to the problem of diffraction of a plane electromagnetic wave from a nonharmonic Bragg grating with a spatially modulated refractive index n(z) = n ₀ + Δn(z)cos(2πz/d + ?). Apodization for such gratings is described by continuous functions of the form n(z) = ±Δn/[1 ? n(z ? L/2)] (d, L, ?, and Δn = const are the period, length, phase, and amplitude of a grating, respectively) or piecewise-continuous symmetric and antisymmetric analogues constructed from these functions. Conditions ensuring suppression of oscillations in the reflection and transmission spectra of these gratings are found. It is shown that, when the coupling factor of a grating is antisymmetric, a narrow transparency band is observed within the forbidden transmission band located near a Bragg resonance. An analytic expression is obtained for the dependence of the width of the transparency band on the grating’s parameters. Gratings with an antisymmetric coupling factor can be used in narrowband frequency-selective Bragg transmission filters.     

The influence of minor titanium addition on thermal properties of diamond/copper composites via in situ reactive sintering

A minor amount of titanium addition is proposed to improve interfacial bonding between diamond particles and copper matrix for diamond/copper composites. The volume fractions of diamond and minor titanium in the sintering process are optimized. The microstructures, thermal properties, interface reaction production, and the effects of minor amount of titanium on the properties of the composites are investigated. The results show that the interfacial bonding of the composites could be strengthened by changing the volume fraction of titanium and diamond. The 45 vol%-diamond/copper composites with 3 vol% titanium at 945 °C for 5 min exhibits thermal conductivity as high as 670 W/(m K), which is 90% of the theoretical prediction value. High thermal conductivity is achieved by forming the titanium carbide (TiC) and intermetallic compounds (Cu₃Ti₂) at the diamond/copper interface to obtain a good interface.     

Liquid-State Interfacial Reactions of Sn-Zn/Co Couples at 250°C

This study examined the interfacial reactions between liquid Sn-Zn alloys and Co substrates at 250°C with different Zn compositions from 1?wt.% to 9?wt.%. The reaction phases formed at the interface were significantly affected by the Zn content. One irregular reaction phase, γ₂-CoZn₁₃, was observed in the Sn-9?wt.%Zn/Co reaction. Decreasing to 7?wt.% Zn, the interfacial product changed to the γ₁ phase. After a longer reaction time, a new ternary compound with composition Sn-20.2?at.%Zn-31.4?at.%Co, the T phase, was formed between the γ₁ phase and Co substrate. Only one very thin and continuous T phase layer was found, and it was stable at the Sn-4?wt.%Zn/Co interface, even after 120?h reaction. However, the Sn-3?wt.%Zn/Co couples revealed significant microstructural evolution in both the T and CoSn₃ layers. Moreover, the dominant phase became CoSn₃ as the Zn content decreased below 2?wt.%. Based on these interfacial results, a predicted Sn-Zn-Co phase diagram (less than 30?at.% Co) at 250°C is proposed. The corresponding reaction paths of these couples are illustrated on the phase diagram.     

Effect of plasma treatments on interface adhesion between SiOCH ultra-low-k film and SiCN etch stop layer

The effect of different plasma treatments on the interfacial bonding configurations and adhesion strengths between porous SiOCH ultra-low-dielectric-constant film and SiCN etch stop layer have been investigated in this study. From X-ray photoelectron spectroscopic analyses, interlayer regions of about 10 nm thick with complicated mixing bonds were found at SiOCH/SiCN interfaces. With plasma treatments, especially H₂/NH₃ two-step plasma, a carbon-depletion region of about 30 nm thick with more Si-O related bonds of high binding energy formed at the interface. Furthermore, the adhesion strengths of the SiOCH/SiCN interfaces were measured by nanoscratch and microscratch tests. For the untreated interface, the adhesion energy was obtained as about 0.22 and 0.44 J/m² by nanoscratch and microscratch tests, respectively. After plasma treatments, especially the H₂/NH₃ treatment, the interfacial adhesion energy was effectively improved to 0.41 and 0.89 J/m² because more Si-O bonds of high binding energy formed at the interfaces.     

Differentiation and function of osteoclasts cultured on bone and cartilage

We examined the differentiation and resorptive function of osteoclasts (OC) cultured on the slices of calcified bone, decalcified bone and hyaline cartilage, and found that OC differentiation depends on the co-cultured substratum, as well as osteoblast-derived factors. Bone marrow-derived macrophages (BMM) were formed from marrow cells of 5 week old ddY mice and cultured for 3 days on freeze-dried slices of calcified bone, decalcified bone or cartilage, all prepared from rabbit costal bone. BMM cultured on calcified bone slices exhibited tartrate-resistant acid phosphatase (TRAP) activity and were structurally characterized by multinucleation and ruffled border development. However, on decalcified bone slices, BMM seldom became multinucleated and exhibited weak TRAP activity. BMM cultured on cartilage slices were mononuclear, devoid of TRAP activity and structurally resembled mononuclear phagocytes. In SEM observations of co-cultured slices, resorption lacunae were formed only on calcified bone slices, and not on slices of decalcified bone and cartilage. Our results, therefore, indicated that BMM could differentiate into functional OC only on calcified bone slices, suggesting a key role of calcified components in the bone matrix for the terminal OC differentiation. Then, we cultured BMM on the same slices with yeast particles. In cultures with yeast particles, BMM exhibited intense TRAP activity, developed a ruffled border-like structure and formed resorption lacunae even on decalcified bone and cartilage slices. Vacuolar-type H+-ATPase was strongly expressed along the ruffled border membranes of these OC. Only the BMM that had not incorporated yeast particles developed a ruffled border, whereas the BMM that had incorporated yeast particles did not become multinucleated and lacked a ruffled border structure. Thus, our results further suggest that, even on uncalcified substrata, the terminal differentiation of BMM into functional OC is induced by an unidentified external stimulus, which may be contained in the cell membrane of yeast particles.