Activating macrophages for enhanced osteogenic and bactericidal performance by Cu ion release from micro/nano-topographical coating on a titanium substrate.

In the field of orthopaedics, inflammation-modulatory biomaterials are receiving increasing attentions due to their abilities to regulate innate immune response and mediate wound healing. In the current work, a Cu-containing micro/nano-topographical bio-ceramic surface (Cu-Hier-Ti surface) was employed as material model to explore the role played by Cu2+ release or material surface in regulating macrophage polarization as well as macrophage-mediated osteogenic and bactericidal effect. A Cu-free micro-topographical surface (Micro-Ti surface) generated by micro-arc oxidation was employed as control. The results showed that Cu2+ supplemented directly into the culture medium or released from Cu-Hier-Ti surface could polarize macrophages to pro-inflammatory M1 phenotype by activating Cu-transport signaling (copper transporter 1 (CTR1) and ATP7A) in macrophages, while the material characteristics exhibited anti-inflammatory effect to some extent by regulating integrin (α5, αM, β1 and β2) and TLR (TLR-3, TLR-4, Myd88 and Ticam-1/2) signaling. Macrophages grown on Cu-Hier-Ti surface or treated by Cu2+ could create a favorable inflammatory microenvironment for osteoblast-like SaOS-2 cell proliferation and differentiation. Moreover, Cu-Hier-Ti surface promoted macrophage capacity to engulf and kill bacteria, even though it did not show direct bactericidal effect against Staphylococcus aureus. In vivo results showed that Cu-Hier-Ti surface could lead to promoted osteointegraion and enhanced expression levels of M1 surface marker CD11c, growth factor BMP-6 and osteogenic makers including osteocalcin (OCN) and Runx-2 at the biomaterial/bone tissue interface in a rat model. The results indicate that Cu could be employed as a promising inflammation-modulatory agent to activate macrophages for enhanced osteogenic and bactericidal effect. STATEMENT OF SIGNIFICANCE: The next generation of bone biomaterials should be active to regulate the local inflammatory environment such that it favors bone regeneration. For the design and development of Cu-containing inflammation-modulatory biomaterials, it is of great importance to recognize the exact role played by Cu2+ release or material surface characteristics. So far, relatively little is known about the regulatory role of Cu2+ or micro/nano-topographical surface on macrophages. The results in the current work suggest that Cu2+ release and material surface characteristics of Cu-containing micro/nano-topographical coating could activate distinct signaling pathways in macrophages. The activated M1 macrophages exhibited stimulatory effect on osteoblast maturation and enhanced bactericidal capacity against Staphylococcus aureus. This study might provide new thoughts for the development of multi-functional Cu-containing biomaterials.

Huang Q ，Ouyang Z ，Tan Y ，Wu H ，Liu Y ... -  《-》

The Cu-containing TiO(2) coatings with modulatory effects on macrophage polarization and bactericidal capacity prepared by micro-arc oxidation on titanium substrates.

The implant materials with both osteogenic and anti-bacterial properties are promising for orthopedic and dental applications. Moreover, the inflammatory response induced by biomaterials has been recently recognized as one of the critical factors in determining implantation fate. A new generation of implant materials should have modulatory effects on the local inflammatory environment such that it favors osteogenesis and osteointegration instead of being bio-inert. In this study, the micro-arc oxidation (MAO) technique was employed to fabricate Cu-containing ceramic coatings on titanium substrates. The macrophages cultured on Cu-containing MAO-fabricated surfaces were polarized to M1 phenotype, evidenced by the high expression levels of inducible nitric oxide synthase (iNOS), low expression levels of arginase1 (Arg1), enhanced pro-inflammatory cytokine interleukin-6 (IL-6) release and inhibited IL-4 and IL-10 (anti-inflammatory cytokines) release. The MAO-treated surface incorporated with larger amounts of Cu (referred as Cu(h)-MAO) could modulate a favorable inflammatory microenvironment for osteoblast-like cell differentiation. Moreover, the macrophages cultured on Cu(h)-MAO surface exhibited enhanced bacteria uptake and killing rate, indicating that the Cu(h)-MAO surface promoted the bactericidal capacity of macrophages. Together, Cu could be used as a promising modulatory agent for macrophage functions. The integration of Cu in biomaterials could lead to enhanced macrophage-mediated osteogenesis and bactericidal capacity.

Huang Q ，Li X ，Elkhooly TA ，Liu X ，Zhang R ，Wu H ，Feng Q ，Liu Y ... -  《-》

Evaluating the osteoimmunomodulatory properties of micro-arc oxidized titanium surface at two different biological stages using an optimized in vitro cell culture strategy.

It is known that introducing a porous ceramic coating on titanium (Ti) implant surface fabricated by micro-arc oxidation (MAO) could enhance the differentiation of osteoblasts. However, the osteogenic capacity of MAO-fabricated coating still remains unknown when immune cells especially macrophages are involved. The influence of the inflammatory microenvironment and the co-influence of the inflammatory microenvironment and surface characteristics of MAO-fabricated coating on osteoblast response need to be explored. In this study, a new in vitro cell culture strategy is proposed by mimicking the biological events happened after implantation based on the recruitment of osteoblasts to biomaterial surfaces to investigate biological performances of MAO-modified Ti surface. It is found that macrophages grown on MAO-modified Ti surface were switched to M1-like phenotype, evidenced by the promoted expressions of inflammatory genes (tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and IL-1β) and production of pro-inflammatory cytokine TNF-α. Moreover, the inflammatory microenvironment created by macrophage/MAO-modified Ti surface interactions could promote the collagen syntheses and matrix mineralization of osteoblast-like cells grown tissue culture plate. When osteoblasts were cultured on MAO-modified Ti surface and cultured by macrophage/MAO-modified Ti surface conditioned medium (CM), the alkaline phosphatase (ALP) activity and collagen synthesis of osteoblast-like cells were promoted. This study suggests that MAO-modified Ti surface is beneficial for osteogenesis at both stages after implantation (before and after osteoblast recruitment to biomaterial surfaces).

Li X ，Huang Q ，Hu X ，Wu D ，Li N ，Liu Y ，Li Q ，Wu H ... -  《-》

Application of a Strontium-Loaded, Phase-Transited Lysozyme Coating to a Titanium Surface to Enhance Osteogenesis and Osteoimmunomodulation.

Lu X ，Zhang W ，Liu Z ，Ma S ，Sun Y ，Wu X ，Zhang X ，Gao P ... -  《MEDICAL SCIENCE MONITOR》

The osteogenic, inflammatory and osteo-immunomodulatory performances of biomedical Ti-Ta metal-metal composite with Ca- and Si-containing bioceramic coatings.

It is known that good mechanical properties, low modulus to reduce stress-shielding effect, favorable osteogenic activity and limited inflammatory response are critical factors for orthopedic implants to induce excellent osteointegration. In this study, Ti-20% Ta metal-metal composite (referred as Ti-Ta) which consisted of Ti- and Ta-rich phases was fabricated via the strategy of powder metallurgy. Micro-arc oxidation (MAO) was employed to modify the surface of Ti-Ta composite. The surfaces of Ti-Ta composite after MAO treatment at an applied voltage of 250 (referred as MAO-250 V) or 300 V (referred as MAO-300 V) exhibited three distinct zones with significantly different morphological features and surface chemistry. Osteoblast-like SaOS-2 cells were found to be preferential to attach on the Ta-rich phase and its surrounding areas, exhibiting an area-dependent adhesion tendency. However, the attachment of Raw 264.7 macrophages was found to be insensitive to the surface characteristics. The proliferation and differentiation of SaOS-2 cells cultured on various surfaces basically followed the trend: MAO-modified surfaces > Ti-Ta surface > Ti surface. The Ti-Ta and MAO-modified surfaces were found to inhibit the inflammatory response and polarize macrophages to anti-inflammatory M2 phenotype compared to Ti surface. Moreover, the microenvironments created by Ti-Ta, MAO-250 V and MAO-300 V/macrophage interactions promoted the proliferation and differentiation of SaOS-2 cells compared to that created by Ti/macrophage interactions. MAO-300 V surface exhibited further enhanced positive osteo-immunomodulatory effects compared to Ti-Ta surface. Together, the Ti-20% Ta metal-metal composite modified by MAO at an applied voltage of 300 V is considered as a promising implant material for orthopedic applications.

Huang Q ，Li X ，Elkhooly TA ，Xu S ，Liu X ，Feng Q ，Wu H ，Liu Y ... -  《-》