Osteoblast response and osseointegration of a Ti–6Al–4V alloy implant incorporating strontium

This study investigated the surface characteristics, in vitro and in vivo biocompatibility of Ti–6Al–4V alloy implants incorporating strontium ions (Sr), produced by hydrothermal treatment using a Sr-containing solution, for future biomedical applications. The surface characteristics were evaluated by scanning electron microscopy, thin-film X-ray diffractometry, X-ray photoelectron spectroscopy, optical profilometry, contact angle and surface energy measurement and inductively coupled plasma-mass spectroscopy (ICP-MS). Human osteoblast-like cell (MG63) attachment, proliferation, alkaline phosphatase (ALP) activity, and quantitative analysis of osteoblastic gene expression on Sr-containing Ti–6Al–4V surfaces were compared with untreated Ti–6Al–4V surfaces. Fifty-six screw implants (28 control and 28 experimental) were placed in the tibiae and femoral condyles of seven New Zealand White rabbits. The osteoconductivity of Sr-containing Ti–6Al–4V implants was evaluated by removal torque testing and histomorphometric analysis after 4 weeks implantation. Hydrothermal treatment produced a crystalline SrTiO₃ layer. ICP-MS analysis showed that Sr ions were released from treated surfaces into the solution. Significant increases in ALP activity (P = 0.000), mRNA expressions of key osteoblast genes (osterix, bone sialoprotein, and osteocalcin), removal torque values (P < 0.05) and bone–implant contact percentages (P < 0.05) in both cortical and cancellous bone were observed for Sr-containing Ti–6Al–4V surfaces. The results indicate that the Sr-containing oxide layer produced by hydrothermal treatment may be effective in improving the osseointegration of Ti–6Al–4V alloy implants by enhancing differentiation of osteoblastic cells, removal torque forces and bone apposition in both cortical and cancellous bone.     

A biaxial rotating bioreactor for the culture of fetal mesenchymal stem cells for bone tissue engineering

The generation of effective tissue engineered bone grafts requires efficient exchange of nutrients and mechanical stimulus. Bioreactors provide a manner in which this can be achieved. We have recently developed a biaxial rotating bioreactor with efficient fluidics through in-silico modeling. Here we investigated its performance for generation of highly osteogenic bone graft using polycaprolactone–tricalcium phosphate (PCL–TCP) scaffolds seeded with human fetal mesenchymal stem cell (hfMSC). hfMSC scaffolds were cultured in either bioreactor or static cultures, with assessment of cellular viability, proliferation and osteogenic differentiation in vitro and also after transplantation into immunodeficient mice. Compared to static culture, bioreactor-cultured hfMSC scaffolds reached cellular confluence earlier (day 7 vs. day 28), with greater cellularity (2×, p < 0.01), and maintained high cellular viability in the core, which was 2000 μm from the surface. In addition, bioreactor culture was associated with greater osteogenic induction, ALP expression (1.5× p < 0.01), calcium deposition (5.5×, p < 0.001) and bony nodule formation on SEM, and in-vivo ectopic bone formation in immunodeficient mice (3.2×, p < 0.001) compared with static-cultured scaffolds. The use of biaxial bioreactor here allowed the maintenance of cellular viability beyond the limits of conventional diffusion, with increased proliferation and osteogenic differentiation both in vitro and in vivo, suggesting its utility for bone tissue engineering applications.     

Corrosion protection and improved cytocompatibility of biodegradable polymeric layer-by-layer coatings on AZ31 magnesium alloys

Composite coatings of electrostatically assembled layer-by-layer anionic and cationic polymers combined with an Mg(OH)₂ surface treatment serve to provide a protective coating on AZ31 magnesium alloy substrates. These ceramic conversion coating and layer-by-layer polymeric coating combinations reduced the initial and long-term corrosion progression of the AZ31 alloy. X-ray diffraction and Fourier transform infrared spectroscopy confirmed the successful application of coatings. Potentiostatic polarization tests indicate improved initial corrosion resistance. Hydrogen evolution measurements over a 2 week period and magnesium ion levels over a 1 week period indicate longer range corrosion protection and retention of the Mg(OH)₂ passivation layer in comparison to the uncoated substrates. Live/dead staining and DNA quantification were used as measures of biocompatibility and proliferation while actin staining and scanning electron microscopy were used to observe the cellular morphology and integration with the coated substrates. The coatings simultaneously provided improved biocompatibility, cellular adhesion and proliferation in comparison to the uncoated alloy surface utilizing both murine pre-osteoblast MC3T3 cells and human mesenchymal stem cells. The implementation of such coatings on magnesium alloy implants could serve to improve the corrosion resistance and cellular integration of these implants with the native tissue while delivering vital drugs or biological elements to the site of implantation.     

Effect of extracts from Phyllanthus watsonii Airy Shaw on cell apoptosis in cultured human breast cancer MCF-7 cells

Species of Phyllanthus have traditionally been used for hundreds of years for treating many ailments including diabetes, anemia, bronchitis and hepatitis. The present study aims to investigate the cytotoxic and apoptotic effects of methanol (PWM), hexane (PWH) and ethyl acetate (PWE) extracts from the leaves of the endemic plant Phyllanthus watsonii Airy Shaw (Phyllanthaceae) on MCF-7 human breast cancer cells. We observed that the PWM, PWH and PWE extracts were cytotoxic and selectively inhibited the growth and proliferation of MCF-7 cells compared to untreated control in a dose dependent manner with an IC₅₀ of 12.7 ± 4.65, 7.9 ± 0.60 and 7.7 ± 0.29 μg/ml, respectively. However, the extracts were not toxic at these concentrations to normal human lung fibroblast MRC-5 cells. Cell death induced by PWM, PWH and PWE extracts were mainly due to apoptosis which was characterized by apoptotic morphological changes and a nuclear DNA fragmentation. Caspase-3 activation following P. watsonii extracts treatment was also evident for apoptotic cell death which was preceded by an S phase cell cycle perturbation. The results suggested that the cytotoxic activity of P. watsonii extracts was related to an early event of cell cycle perturbation and a later event of apoptosis. Hence, P. watsonii displays potential to be further exploited in the discovery and development of new anticancer agents.     

UV photoactivation of 7-dehydrocholesterol on titanium implants enhances osteoblast differentiation and decreases Rankl gene expression

Vitamin D plays a central role in bone regeneration, and its insufficiency has been reported to have profound negative effects on implant osseointegration. The present study aimed to test the in vitro biological effect of titanium (Ti) implants coated with UV-activated 7-dehydrocholesterol (7-DHC), the precursor of vitamin D, on cytotoxicity and osteoblast differentiation. Fourier transform infrared spectroscopy confirmed the changes in chemical structure of 7-DHC after UV exposure. High-pressure liquid chromatography analysis determined a 16.5 ± 0.9% conversion of 7-DHC to previtamin D₃ after 15 min of UV exposure, and a 34.2 ± 4.8% of the preD₃ produced was finally converted to 25-hydroxyvitamin D₃ (25-D₃) by the osteoblastic cells. No cytotoxic effect was found for Ti implants treated with 7-DHC and UV-irradiated. Moreover, Ti implants treated with 7-DHC and UV-irradiated for 15 min showed increased 25-D₃ production, together with increased ALP activity and calcium content. Interestingly, Rankl gene expression was significantly reduced in osteoblasts cultured on 7-DHC-coated Ti surfaces when UV-irradiated for 15 and 30 min to 33.56 ± 15.28% and 28.21 ± 4.40%, respectively, compared with the control. In conclusion, these findings demonstrate that UV-activated 7-DHC is a biocompatible coating of Ti implants, which allows the osteoblastic cells to produce themselves active vitamin D, with demonstrated positive effects on osteoblast differentiation in vitro.     

Study of cellular dynamics on polarized CoCrMo alloy using time-lapse live-cell imaging

The physico-chemical processes and phenomena occurring at the interface of metallic biomedical implants and the body dictate their successful integration in vivo. Changes in the surface potential and the associated redox reactions at metallic implants can significantly influence several aspects of biomaterial/cell interactions such as cell adhesion and survival in vitro. Accordingly, there is a voltage viability range (voltages which do not compromise cellular viability of the cells cultured on the polarized metal) for metallic implants. We report on cellular dynamics (size, polarity, movement) and temporal changes in the number and total area of focal adhesion complexes in transiently transfected MC3T3-E1 pre-osteoblasts cultured on CoCrMo alloy surfaces polarized at the cathodic and anodic edges of its voltage viability range (?400 and +500 mV (Ag/AgCl), respectively). Nucleus dynamics (size, circularity, movement) and the release of reactive oxygen species (ROS) were also studied on the polarized metal at ?1000, ?400 and +500 mV (Ag/AgCl). Our results show that at ?400 mV, where reduction reactions dominate, a gradual loss of adhesion occurs over 24 h while cells shrink in size during this time. At +500 mV, where oxidation reactions dominate (i.e. metal ions form, including Cr⁶⁺), cells become non-viable after 5 h without showing any significant changes in adhesion behavior right before cell death. Nucleus size of cells at ?1000 mV decreased sharply within 15 min after polarization, which rendered the cells completely non-viable. No significant amount of ROS release by cells was detected on the polarized CoCrMo at any of these voltages.     

Autoimmune disease development based on possible mislocalization of intracellular and extracellular proteins

BackgroundT helper cells can differentiate into several subsets of T lymphocytes, including Th1, Th2, and regulatory T (T_reg) cells. As a result of this ability to differentiate, the corresponding T cell receptor (TCR) spectra display considerable cellular plasticity and interchangeability. In contrast, T lymphocyte differentiation and separation into CD4⁺ and/or CD8⁺ T cell lines creates stable populations over a person’s lifetime, which abrogates the plasticity and interchange between these cell types and their corresponding TCR spectra but results in considerable stability regarding the corresponding TCR sequences and spectra. This separation of TCR spectra agrees with the well-known concept of major histocompatibility complex class (MHC) restriction. Therefore, CD4⁺ and CD8⁺ T cell populations possess different (but stable) TCR spectra, which present differences in antigens between intra- and extracellular space. Thus, mislocalization can lead to autoimmunization and the development of autoimmune disease.MethodsTo test this hypothesis, human intra- and extracellular proteins and intra- and extracellular extracts were incubated overnight with whole-blood samples from the same subject, and the following day, a cell proliferation assay based on bromodeoxyuridine (BrdU) incorporation was performed.ResultsThe BrdU assay showed that the addition of intracellular proteins and extracts to the mixture resulted in significantly greater cell proliferation after overnight incubation, whereas significantly less proliferation was obtained with addition of extracellular proteins and extracts (plasma).ConclusionsThese results support the proposed hypothesis and show that hidden antigens are present in and released with intracellular proteins. Furthermore, both albumin and insulin activated CD4⁺ and CD8⁺ lymphocytes in a concentration-dependent manner. At low concentrations (<0.1 μg/ml), both proteins showed the ability to inhibit CD4⁺ and CD8⁺, whereas at high concentrations (>1000 μg/ml), both proteins activated CD4⁺ and CD8⁺ T lymphocytes.     

In vivo corrosion and corrosion protection of magnesium alloy LAE442

The aim of this study was to investigate whether the extruded magnesium alloy LAE442 reacts in vivo with an appropriate host response and to investigate how an additional magnesium fluoride (MgF₂) coating influences the in vivo corrosion rate. Forty cylinders were machined from extruded LAE442 and 20 of these were coated additionally with MgF₂ and implanted into the medial femur condyle of adult rabbits. Synchrotron-radiation-based X-ray computed micro-tomography (SRμCT) was used to quantitatively analyse corrosion non-destructively in vivo and comparisons were made to magnesium degradation rates based on area measurements of the remaining metal on uncalcified sections. Blood concentrations of the alloying elements were measured below toxicological limits. The MgF₂ layer was no longer detected after 4 weeks of implantation by particle-induced gamma emission, and the MgF₂ coating reduced the blood content of alloying elements during the first 6 weeks of implantation with no elevated fluoride concentration in the adjacent bone. Histopathological examinations of liver showed in 9 out of 40 cases minimal infiltrations of heterophil granulocytes of unknown origin (5 LAE442, 4 LAE442 + MgF₂). The kidneys were mainly regular in structure. The synovial tissue showed a granular cell infiltration as a temporary observation in the LAE442 + MgF₂ group after 2 weeks. No subcutaneous gas cavities were observed clinically and on postoperative X-rays in all animals. All specimens were scanned by SRμCT at 2, 4, 6 and 12 weeks postoperatively before uncalcified sections were performed. All magnesium implants have been observed in direct bone contact and without a fibrous capsule. Localized pitting corrosion occurred in coated and uncoated magnesium implants. This study shows that the extruded magnesium alloy LAE442 provides low corrosion rates and reacts in vivo with an acceptable host response. The in vivo corrosion rate can be further reduced by additional MgF₂ coating.     

Polarization of hydroxyapatite: Influence on osteoblast cell proliferation

Hydroxyapatite (HA) has been used clinically to treat bone defects. However, modifications of the surface properties of HA could improve and control bone matrix deposition and localized host tissue integration. The aim of this study was to investigate the effect of developing a surface charge on HA discs with respect to osteoblast activity in vitro. HA discs (12 mm × 2 mm) were sintered in either air or water vapour. The HA discs were then electrically polarized (positive and negative surfaces) or non-polarized (controls) and seeded with MC3T3-E1 cells. Polarized HA sintered in water vapour was shown to retain six times more charge than polarized HA sintered in air. Picogreen analysis demonstrated that at 4 h cell number was significantly higher on the negatively and positively charged HA surface (water sintered) in comparison to the non-charged water and air-sintered HA controls. At 7 days there was a significant increase in cell number on the negatively charged HA (air sintered) sample in comparison to the negatively charged water vapour sintered HA sample and the non-charged water vapour sintered control sample. Also at 7 days, the picogreen data showed a significant increase in cell number on the positively charged water-treated HA sample in comparison to both the air- and water-treated HA non-charged control HA samples. An alamarBlue assay at 7 days demonstrated significant cell metabolic activity on the charged surfaces (both positive and negative) in comparison to the non-charged HA and the tissue culture plastic controls. This study demonstrated that all of the HA discs tested supported cell viability/attachment. However, cell attachment/proliferation/metabolic activity was significantly increased as a result of developing a charge on the HA surface.     

The size of surface microstructures as an osteogenic factor in calcium phosphate ceramics

The microporosity of calcium phosphate (CaP) ceramics has been shown to have an essential role in osteoinduction by CaP ceramics after ectopic implantation. Here we show that it is not the microporosity but the size of surface microstructural features that is the most likely osteogenic factor. Two tricalcium phosphate (TCP) ceramics, namely TCP-S and TCP-B, were fabricated with equivalent chemistry and similar microporosity but different sizes of surface microstructural features. TCP-S has a grain size of 0.99 ± 0.20 μm and a micropore size of 0.65 ± 0.25 μm, while TCP-B displays a grain size of 3.08 ± 0.52 μm and a micropore size of 1.58 ± 0.65 μm. In vitro, both cell proliferation and osteogenic differentiation were significantly enhanced when human bone marrow stromal cells were cultured on TCP-S without any osteogenic growth factors, compared to TCP-B ceramic granules. The possible involvement of direct contact between cells and the TCP ceramic surface in osteogenic differentiation is also shown with a trans-well culture model. When the ceramic granules were implanted in paraspinal muscle of dogs for 12 weeks, abundant bone was formed in TCP-S (21 ± 10% bone in the available space), whereas no bone was formed in any of the TCP-B implants. The current in vitro and in vivo data reveal that the readily controllable cue, i.e. the size of the surface microstructure, could be sufficient to induce osteogenic differentiation of mesenchymal stem cells, ultimately leading to ectopic bone formation in calcium phosphate ceramics.     

In vitro cellular response and in vivo primary osteointegration of electrochemically modified titanium

Anodic spark deposition (ASD) is an attractive technique for improving the implant–bone interface that can be applied to titanium and titanium alloys. This technique produces a surface with microporous morphology and an oxide layer enriched with calcium and phosphorus. The aim of the present study was to investigate the biological response in vitro using primary human osteoblasts as a cellular model and the osteogenic primary response in vivo within a short experimental time frame (2 and 4 weeks) in an animal model (rabbit). Responses were assessed by comparing the new electrochemical biomimetic treatments to an acid-etching treatment as control. The in vitro biological response was characterized by cell morphology, adhesion, proliferation activity and cell metabolic activity. A complete assessment of osteogenic activity in vivo was achieved by estimating static and dynamic histomorphometric parameters at several time points within the considered time frame. The in vitro study showed enhanced osteoblast adhesion and higher metabolic activity for the ASD-treated surfaces during the first days after seeding compared to the control titanium. For the ASD surfaces, the histomorphometry indicated a higher mineral apposition rate within 2 weeks and a more extended bone activation within the first week after surgery, leading to more extensive bone–implant contact after 2 weeks. In conclusion, the ASD surface treatments enhanced the biological response in vitro, promoting an early osteoblast adhesion, and the osteointegrative properties in vivo, accelerating the primary osteogenic response.     

Biofunctional alendronate–Hydroxyapatite thin films deposited by Matrix Assisted Pulsed Laser Evaporation

We applied Matrix Assisted Pulsed Laser Evaporation (MAPLE) in order to synthesize alendronate-hydroxyapatite thin films on titanium substrates. Alendronate-hydroxyapatite composite nanocrystals with increasing bisphosphonate content (0, 3.9, 7.1% wt) were synthesized in aqueous medium. Then, they were suspended in deionised water, frozen at liquid nitrogen temperature and used as targets for MAPLE experiments. The depositions were conducted with a KrF* excimer laser source (l = 248 nm, t_FWHM = 25 ns) in mild conditions of temperature and pressure. The obtained thin films had a good crystallinity, which slightly decreases with the increase of alendronate content, and exhibited a porous-like structure. Osteoblast-like MG63 cells and human osteoclasts were cultured on the thin films up to 14 days. In the presence of alendronate, MG63 cells displayed a normal morphology, increased proliferation and higher values of differentiation parameters, namely type I collagen, osteocalcin, and osteoprotegerin/TNF-related activation-induced cytokine receptor ratio. In contrast, osteoclasts showed significantly reduced proliferation, and increased level of Caspase 3. Moreover, the coatings synthesized from hydroxyapatite at relatively high bisphosphonate content (7.1% wt) displayed a reduced production of Tumour Necrosis Factor alpha (TNF-α) and Interleukin 6 (IL-6), suggesting a down-regulatory role of alendronate on the inflammatory reaction. The successful deposition of alendronate modified hydroxyapatite thin films yields coatings with enhanced bioactivity, able to promote osteoblast differentiation and to inhibit osteoclast proliferation.     

Perfused culture of gingival fibroblasts in a degradable/polar/hydrophobic/ionic polyurethane (D-PHI) scaffold leads to enhanced proliferation and metabolic activity

Periodontal diseases cause the breakdown of the tooth-supporting gingival tissue. In treatments aimed at gingival tissue regeneration, tissue engineering is preferred over the common treatments such as scaling. Perfused (dynamic) culture has been shown to increase cell growth in tissue-engineered scaffolds. Since gingival tissues are highly vascularized, it was desired to investigate the influence of perfusion on the function of human gingival fibroblasts (HGF) when cultured in a degradable/polar/hydrophobic/ionic polyurethane scaffold during the early culture phase (4 weeks) of engineering gingival tissues. It was observed that the growth of HGF was continuous over 28 days in dynamic culture (3-fold increase, p < 0.05), while it was reduced after 14 days in static culture (i.e. no flow condition). Cell metabolic activity, as measured by a WST-1 assay, and total protein production show that HGF were in different metabolic states in the dynamic vs. static cultures. Observations from scanning electron microscopy and type I collagen (Col I) production measured by Western blotting suggest that medium perfusion significantly promoted collagen production in HGF after the first 4 weeks of culture (p < 0.05). The different proliferative and metabolic states for HGF in the perfused scaffolds suggest a different cell phenotype which may favour tissue regeneration.     

Electrically polarized HAp-coated Ti: In vitro bone cell–material interactions

Electrically polarized bulk sintered hydroxyapatite (HAp) compacts have been shown to accelerate mineralization and bone tissue ingrowth in vivo. In this work, a comprehensive study has been carried out to investigate the influence of surface charge and polarity on in vitro bone cell adhesion, proliferation and differentiation on electrically polarized HAp-coated Ti. Uniform and crack free sol–gel derived HAp coatings of 20 ± 1.38 μm thickness were polarized by application of an external d.c. field of 2.0 kV cm^?1 at 400 °C for 1h. In vitro bioactivity of polarized HAp coatings was evaluated by soaking in simulated body fluid, and bone cell–material interactions were studied by culturing with human fetal osteoblast cells (hFOB) for a maximum period of 11 days. Scanning electron microscopic observation showed that accelerated mineralization on negatively charged surfaces favored rapid cell attachment and faster tissue ingrowth over non-polarized HAp coating surfaces, while positive charge on HAp coating surfaces restricted apatite nucleation with limited cellular response. Immunochemistry and confocal microscopy confirmed that the cell adhesion and early stage differentiation were more pronounced on negatively charged coating surfaces as hFOB cells expressed higher vinculin and alkaline phosphatase proteins on negatively charged surface compared to cells grown on all other surfaces. Our results in this study are process independent and potentially applicable to any other commercially available coating techniques.     

Analyses and comparisons of telomerase activity and telomere length in human T and B cells: Insights for epidemiology of telomere maintenance

Telomeres are the DNA–protein complexes that protect the ends of eukaryotic chromosomes. The cellular enzyme telomerase counteracts telomere shortening by adding telomeric DNA. A growing body of literature links shorter telomere length and lower telomerase activity with various age-related diseases and earlier mortality. Thus, leukocyte telomere length (LTL) and telomerase activity are emerging both as biomarkers and contributing factors for age-related diseases. However, no clinical study has directly examined telomerase activity and telomere length in different lymphocyte subtypes isolated from the same donors, which could offer insight into the summary measure of leukocyte telomere maintenance.We report the first quantitative data in humans examining both levels of telomerase activity and telomere length in four lymphocyte subpopulations from the same donors—CD4+, CD8+CD28+ and CD8+CD28? T cells and B cells, as well as total PBMCs—in a cohort of healthy women. We found that B cells had the highest telomerase activity and longest telomere length; CD4+ T cells had slightly higher telomerase activity than CD8+CD28+ T cells, and similar telomere length. Consistent with earlier reports that CD8+CD28? T cells are replicatively senescent cells, they had the lowest telomerase activity and shortest telomere length. In addition, a higher percentage of CD8+CD28? T cells correlated with shorter total PBMC TL (r = ? 0.26, p = 0.05). Interestingly, telomerase activities of CD4+ and CD8+CD28+ T cells from the same individual were strongly correlated (r = 0.55, r < 0.001), indicating possible common mechanisms for telomerase activity regulation in these two cell subtypes. These data will facilitate the understanding of leukocyte aging and its relationship to human health.     

Electrospun nanostructured scaffolds for bone tissue engineering

The current challenge in bone tissue engineering is to fabricate a bioartificial bone graft mimicking the extracellular matrix (ECM) with effective bone mineralization, resulting in the regeneration of fractured or diseased bones. Biocomposite polymeric nanofibers containing nanohydroxyapatite (HA) fabricated by electrospinning could be promising scaffolds for bone tissue engineering. Nanofibrous scaffolds of poly-l-lactide (PLLA, 860 ± 110 nm), PLLA/HA (845 ± 140 nm) and PLLA/collagen/HA (310 ± 125 nm) were fabricated, and the morphology, chemical and mechanical characterization of the nanofibers were evaluated using scanning electron microscopy, Fourier transform infrared spectroscopy and tensile testing, respectively. The in vitro biocompatibility of different nanofibrous scaffolds was also assessed by growing human fetal osteoblasts (hFOB), and investigating the proliferation, alkaline phosphatase activity (ALP) and mineralization of cells on different nanofibrous scaffolds. Osteoblasts were found to adhere and grow actively on PLLA/collagen/HA nanofibers with enhanced mineral deposition of 57% higher than the PLLA/HA nanofibers. The synergistic effect of the presence of an ECM protein, collagen and HA in PLLA/collagen/HA nanofibers provided cell recognition sites together with apatite for cell proliferation and osteoconduction necessary for mineralization and bone formation. The results of our study showed that the biocomposite PLLA/collagen/HA nanofibrous scaffold could be a potential substrate for the proliferation and mineralization of osteoblasts, enhancing bone regeneration.     

Development and functional evaluation of biomimetic silicone surfaces with hierarchical micro/nano-topographical features demonstrates favourable in vitro foreign body response of breast-derived fibroblasts

Reproducing extracellular matrix topographical cues, such as those present within acellular dermal matrix (ADM), in synthetic implant surfaces, may augment cellular responses, independent of surface chemistry. This could lead to enhanced implant integration and performance while reducing complications. In this work, the hierarchical micro and nanoscale features of ADM were accurately and reproducibly replicated in polydimethylsiloxane (PDMS), using an innovative maskless 3D grayscale fabrication process not previously reported. Human breast derived fibroblasts (n = 5) were cultured on PDMS surfaces and compared to commercially available smooth and textured silicone implant surfaces, for up to one week. Cell attachment, proliferation and cytotoxicity, in addition to immunofluorescence staining, SEM imaging, qRT-PCR and cytokine array were performed. ADM PDMS surfaces promoted cell adhesion, proliferation and survival (p= < 0.05), in addition to increased focal contact formation and spread fibroblast morphology when compared to commercially available implant surfaces. PCNA, vinculin and collagen 1 were up-regulated in fibroblasts on biomimetic surfaces while IL8, TNFα, TGFβ1 and HSP60 were down-regulated (p= < 0.05). A reduced inflammatory cytokine response was also observed (p= < 0.05). This study represents a novel approach to the development of functionalised biomimetic prosthetic implant surfaces which were demonstrated to significantly attenuate the acute in vitro foreign body reaction to silicone.     

Controlling initial biodegradation of magnesium by a biocompatible strontium phosphate conversion coating

A simple strontium phosphate (SrP) conversion coating process was developed to protect magnesium (Mg) from the initial degradation post-implantation. The coating morphology, deposition rate and resultant phases are all dependent on the processing temperature, which determines the protective ability for Mg in minimum essential medium (MEM). Coatings produced at 80 °C are primarily made up of strontium apatite (SrAp) with a granular surface, a high degree of crystallinity and the highest protective ability, which arises from retarding anodic dissolution of Mg in MEM. Following 14 days’ immersion in MEM, the SrAp coating maintained its integrity with only a small fraction of the surface corroded. The post-degradation effect of uncoated Mg and Mg coated at 40 and 80 °C on the proliferation and differentiation of human mesenchymal stem cells was also studied, revealing that the SrP coatings are biocompatible and permit proliferation to a level similar to that of pure Mg. The present study suggests that the SrP conversion coating is a promising option for controlling the early rapid degradation rate, and hence hydrogen gas evolution, of Mg implants without adverse effects on surrounding cells and tissues.     

Mesenchymal stem cell proliferation and differentiation on load-bearing trabecular Nitinol scaffolds

Bone tissue regeneration in load-bearing regions of the body requires high-strength porous scaffolds capable of supporting angiogenesis and osteogenesis. 70% porous Nitinol (NiTi) scaffolds with a regular 3-D architecture resembling trabecular bone were produced from Ni foams using an original reactive vapor infiltration technique. The “trabecular Nitinol” scaffolds possessed a high compressive strength of 79 MPa and high permeability of 6.9 × 10^?6 cm². The scaffolds were further modified to produce a near Ni-free surface layer and evaluated in terms of Ni ion release and human mesenchymal stem cell (hMSC) proliferation (AlamarBlue), differentiation (alkaline phosphatase activity, ALP) and mineralization (Alizarin Red S staining). Scanning electron microscopy was employed to qualitatively corroborate the results. hMSCs were able to adhere and proliferate on both as-produced and surface-modified trabecular NiTi scaffolds, to acquire an osteoblastic phenotype and produce a mineralized extracellular matrix. Both ALP activity and mineralization were increased on porous scaffolds compared to control polystyrene plates. Experiments in a model coculture system of microvascular endothelial cells and hMSCs demonstrated the formation of prevascular structures in trabecular NiTi scaffolds. These data suggest that load-bearing trabecular Nitinol scaffolds could be effective in regenerating damaged or lost bone tissue.