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Bone implants with surface modifications that promote the physiological activities of osteoblasts are the first step for osseointegration in bone repair. Hydroxyapatite is the main inorganic component in mammal bones and teeth, and nanoscaled hydroxyapatite promotes the adhesion of osteoblastic cells. In this study, we created a nano/micro hierarchical structure using micro-arc oxidation coatings and hydrothermal treatments at 150 °C, 175 °C, and 200 °C for 2, 6, 12, and 24 h. After undergoing hydrothermal treatment for 24 h, CaTiO 3 began forming regular-shaped crystals at the surface at 175 °C. In order to decrease the CaTiO 3 formations and increase the apatite fabrication, a shorter time of hydrothermal treatment was required at 175 °C. There was still surface damage on samples treated for 6 h at 175 °C; however, the nano/micro hierarchical structures were formed in 2 h at 175 °C. The normalized alkaline phosphatase (ALP) activities of the MC3T3-E1 cells with micro-arc oxidation (MAO) coatings and nano/micro hierarchical bioceramics coatings were 4.51 ± 0.26 and 7.36 ± 0.51 μmol p-NP/mg protein (*** P value of <0.001), respectively. The MC3T3-E1 cells with coatings showed highly statistically significant results in terms of the ALP activity. This proposed nano/micro hierarchical structure promoted cell proliferation and osteogenic differentiation of the osteoblast MC3T3-E1 cells. This study realized a promising nano system for osseointegration via bone implant surface treatments, which can promote the physiological activities of osteoblasts.

Certain viruses have the ability to subvert the mammalian immune response, including interference in the chemokine system. Poxviruses produce the chemokine binding protein vCCI (viral CC chemokine inhibitor; also called 35K), which tightly binds to CC chemokines. To facilitate the study of vCCI, we first provide a protocol to produce folded vCCI from Escherichia coli ( E. coli .) It is shown here that vCCI binds with unusually high affinity to viral Macrophage Inflammatory Protein-II (vMIP-II), a chemokine analog produced by the virus, human herpesvirus 8 (HHV-8). Fluorescence anisotropy was used to investigate the vCCI:vMIP-II complex and shows that vCCI binds to vMIP-II with a higher affinity than most other chemokines, having a K d of 0.06 ± 0.006 nM. Nuclear magnetic resonance (NMR) chemical shift perturbation experiments indicate that key amino acids used for binding in the complex are similar to those found in previous work. Molecular dynamics were then used to compare the vCCI:vMIP-II complex with the known vCCI:Macrophage Inflammatory Protein-1β/CC-Chemokine Ligand 4 (MIP-1β/CCL4) complex. The simulations show key interactions, such as those between E143 and D75 in vCCI/35K and R18 in vMIP-II. Further, in a comparison of 1 μs molecular dynamics (MD) trajectories, vMIP-II shows more overall surface binding to vCCI than does the chemokine MIP-1β. vMIP-II maintains unique contacts at its N-terminus to vCCI that are not made by MIP-1β, and vMIP-II also makes more contacts with the vCCI flexible acidic loop (located between the second and third beta strands) than does MIP-1β. These studies provide evidence for the basis of the tight vCCI:vMIP-II interaction while elucidating the vCCI:MIP-1β interaction, and allow insight into the structure of proteins that are capable of broadly subverting the mammalian immune system., Competing Interests: The authors declare no conflict of interest.