Interaction of glia with a compliant,microstructured silicone surface

Soft bioengineered surfaces offer a route towards modulating the tissue responses to chronically implanted devices and may enhance their functionality. In this communication we fabricate microtopographically rich and mechanically compliant silicone surfaces for use in soft neural interfaces. We observe the interaction of primary rat microglia and astroglia with arrays of tall and short (4.7 and 0.5 μm) vertically oriented polydimethylsiloxane (PDMS) micropillars and a flat PDMS surface in vitro. With the pillar size and spacing that we use (1.3 μm diameter and 1.6 μm edge to edge), glia are found to engulf and bend tall pillars. The cytoskeleton of cells adhering to the pillar arrays lacks actin stress fibers; instead we observe actin ring formations around individual pillars. Tall, but not short pillar arrays are inhibitory to migration and spreading for both microglia and astrocytes. When compared to a flat PDMS surface and short pillar arrays, tall micropillar arrays cause nearly a 2-fold decrease in proliferation rates for both cell types. The antimitotic properties of tall pillar arrays may be useful for reducing the density of the glial capsule around brain-implanted devices.     

Surface engineering of titanium with potassium hydroxide and its effects on the growth behavior of mesenchymal stem cells

To improve the corrosion resistance and biological performance of commercially pure titanium (cp-Ti) substrates, potassium hydroxide was employed to modify the surfaces of titanium substrates, followed by biomimetic deposition of apatite on the substrates in a simulated body fluid. The morphologies of native and treated titanium substrates were characterized by field emission scanning electron microscopy (FE-SEM). Treatment with potassium hydroxide led to the formation of intermediate layers of potassium titanate on the surfaces of titanium substrates, while apatite was subsequently deposited onto the intermediate layer. The formation of potassium titanate and apatite was confirmed by thin-film X-ray diffraction and FE-SEM equipped with energy dispersive spectroscopy, respectively. Electrochemical impedance spectroscopy showed that the formed potassium titanate layer improved the corrosion-resistance properties of titanium substrates. The influence of modified titanium substrates on the biological behavior of mesenchymal stem cells (MSCs), including osteogenic differentiation, was investigated in vitro. Compared with cp-Ti substrates, MSCs cultured onto alkali- and heat-treated titanium substrates and apatite-deposited titanium substrates displayed significantly higher (P < 0.05 or P < 0.01) proliferation and differentiation levels of alkaline phosphatase and osteocalcin in 7 and 14 day cultures, respectively. More importantly, our results suggest that the modified titanium substrates have great potential for inducing MSCs to differentiate into osteoblasts. The approach presented here may be exploited to fabricate titanium-based implants.     

Poly(vinyl alcohol)/poly(acrylic acid) hydrogel coatings for improving electrode–neural tissue interface

A major problem which hinders the applications of neural prostheses is the inconsistent performance caused by tissue responses during long-term implantation. The study investigated a new approach for improving the electrode–neural tissue interface. Hydrogel poly(vinyl alcohol)/poly(acrylic acid) interpenetrating polymer networks (PVA/PAA IPNs) were synthesized and tailored as coatings for poly(dimethylsiloxane) (PDMS) based neural electrodes with the aid of plasma pretreatment. Changes in the electrochemical impedance and maximum charge injection (Q_inj) limits of the coated iridium oxide microelectrodes were negligible. Protein adsorption on PDMS was reduced by ～85% after coating. In the presence of nerve growth factor (NGF), neurite extension of rat pheochromocytoma (PC12) cells was clearly greater on PVA/PAA IPN films than on PDMS substrates. Furthermore, the tissue responses of PDMS implants coated with PVA/PAA IPN films were studied by 6-week implantation in the cortex of rats, which found that the glial fibrillary acidic protein (GFAP) immunoreactivity in animals (n = 8) receiving coated implants was significantly lower (p < 0.05) compared to that of uncoated implants (n = 7) along the entire distance of 150 μm from the outer skirt to the implant interface. The coated film remained on the surface of the explanted implants, confirmed by scanning electron microscopy (SEM). All of these suggest the hydrogel coating is feasible and favorable to neural electrode applications.     

Influence of a zirconia sandblasting treated surface on peri-implant bone healing: An experimental study in sheep

A sandblasting process with round zirconia (ZrO₂) particles might be an alternative surface treatment to enhance the osseointegration of titanium dental implants. Our previous study on sheep compared smooth surface titanium implants (control) with implant surfaces sandblasted with two different granulations of ZrO₂. As the sandblasted surfaces proved superior, the present study further compared the ZrO₂ surface implant with other surface treatments currently employed: machined titanium (control), titanium oxide plasma sprayed (TPS) and alumina sandblasted (Al-SL) at different times after insertion (2, 4 and 12 weeks). Twelve sheep were divided into three groups of four animals each and underwent implant insertion in tibia cortical bone under general anaesthesia. The implants with surrounding tissues were subjected to histology, histomorphometry, scanning electron microscopy and microhardness tests. The experimentation indicated that at 2 weeks Zr-SL implants had the highest significant bone ingrowth (p < 0.05) compared to the other implant surfaces, and a microhardness of newly formed bone inside the threads significantly higher than that of Ti. The present work shows that the ZrO₂ treatment produces better results in peri-implant newly formed bone than Ti and TPS processing, whereas its performance is similar to the Al-SL surface treatment.     

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.     

Clinicopathological and prognostic significance of p53, Ki-67, and Bcl-2 expression in Tunisian gastric adenocarcinomas

Gastric carcinoma (GC) is a highly aggressive malignancy with poor prognosis. It is widely accepted that malignancy results from abnormal cell growth due to dysregulation of the balance between cell proliferation and apoptosis. Our study aimed to investigate the clinicopathological and prognostic significance of p53, Ki-67, and Bcl-2 in Tunisian GC patients by immunohistochemistry. It was observed that the older patients showed p53 overexpression compared with the younger patients (p < 0.05). There was higher p53 expression in the intestinal-type compared with the diffuse-type (p < 0.05), and in well/moderate differentiated than in poor differentiated tumors. The expression of Ki-67 was positively associated with tumor size and venous invasion (p < 0.05). Bcl2 expression occurred in male patients and correlated with depth of invasion (p = 0.02). A Kaplan–Meier analysis indicated an inverse correlation between p53 and Ki-67 expression and the overall survival. Multivariate analysis revealed that the tumor site, Ki-67 and p53 expression were independent prognostic factors for gastric carcinomas (p < 0.05). Finally, combined expression of p53, Ki-67 and Bcl-2 showed that the group of patients with tumors p53+/Ki-67+/Bcl2− had aggressive behavior and poor prognosis (p log rank = 0.000). In summary, our data indicated that the expression of p53, Ki-67, and Bcl-2 may provide useful information for identifying patients with aggressive behavior and poor prognosis of GC.     

Metformin limits ceramide-induced senescence in C2C12 myoblasts

High lipid and ceramide concentrations are hallmarks of obese and/or insulin resistant skeletal muscle, yet little is known about its role on cell cycle and senescence. The purpose of this study was to examine the role of ceramide on muscle senescence, and whether metformin limited this response.MethodsLow passage, proliferating C2C12 myoblasts were treated with a control, 50 μM C2-ceramide (8 h), and/or 2 mM metformin, then examined for insulin sensitivity, cell senescence, cell proliferation, cell cycle, protein expression of cell cycle regulators.ResultsCeramide treatment caused a dephosphorylation (p < 0.05) of Akt and 4E-BP1, regardless of the presence of insulin. The ceramide treated myoblasts displayed higher β-galactosidase staining (p < 0.05), reduced BrDu incorporation and total number of cells (p < 0.05), and an increased proportion of cells in G2-phase (p < 0.05) versus control cultures. Ceramide treatment also upregulated (p < 0.05) p53 and p21 protein expression, that was reversed by either pifithrin-α or shRNA for p53. Metformin limited (p < 0.05) ceramide's effects on insulin signaling, senescence, and cell cycle regulation.ConclusionsHigh ceramide concentrations reduced myoblast proliferation that was associated with aberrant cell cycle regulation and a senescent phenotype, which could provide an understanding of skeletal muscle cell adaptation during conditions of high intramuscular lipid deposition and/or obesity.     

IL-10-592 polymorphism is associated with IL-10 expression and severity of enterovirus 71 infection in chinese children

BackgroundEnterovirus 71 (EV71) infection results in some severe complications with high mortality and disability in Hand, Foot and Mouth Disease (HFMD) in children. Recent studies have shown that cytokine genetic predispositions have associations with both the development of EV71 infection and severity of HFMD.ObjectiveThis study was designed to investigate whether the IL-10–592 polymorphism is associated with IL-10 levels and disease severity in Chinese children with EV71 infection.Study designIn patients selected, there were 378 cases with EV71 infection (including 291 mild cases, 70 severe cases and 17 critical cases), as well as 406 health controls. EV71 in serum was tested by RT-PCR, and IL-10-592 genotype was detected by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis techniques.ResultThe IL-10-592C allele was observed with higher frequency in patients with critical EV71 infection (70.59%) compared with severe EV71 infection (41.43%, P < 0.01), mild EV71 infection (43.81%, P < 0.01) and healthy children (44.46%, P < 0.01). The blood IL-10 levels of critical cases were significantly higher than severe cases, mild cases, and healthy children. Among all of the four groups, IL-10 levels in patients with genotype AA were significantly lower than those with genotypes AC + CC (t = 4.86, P < 0.05; t = 2.30, P < 0.05; t = 3.44, P < 0.05; t = 5.58, P < 0.05).ConclusionIL-10-592C allele is associated with IL-10 expressions and the severity of EV71 infection in Chinese patients.     

The induction of cell alignment by covalently immobilized gradients of the 6th Ig-like domain of cell adhesion molecule L1 in 3D-fibrin matrices

Concentration gradients of matrix-bound guidance cues in the extracellular matrix direct cell growth in native tissues and are of great interest for the design of biomedical scaffolds and on implant surfaces.This study describes effects of covalently immobilized gradients of the 6th Ig-like domain of cell adhesion molecule L1 (TG-L1Ig6) within 3D-fibrin matrices on cell alignment. Linear gradients of TG-L1Ig6 were established and shown to be stable for at least 24 h whereas soluble gradients disappeared completely. Fibroblast alignment along the gradients was observed when cultured on top and within TG-L1Ig6-gradient matrices. Fibroblasts responded to an increase of 0.2 μg TG-L1Ig6/ml per mm matrix, corresponding to a concentration change of <1% per cell. Significant differences were observed when fibroblasts were cultured within the TG-L1Ig6-gradient matrices as the number of aligned cells decreased by 20–30% in the middle of the gradient when compared to cells cultivated on top of the gradient. This finding might be explained by ～13% reduction in the average cell length of fibroblasts within compared to fibroblasts cultured on top of the gradient matrix. In contrast to fibroblasts endothelial cells did not show any alignment with TG-L1Ig6-gradient matrices. The study indicates that cells exposed to gradients of matrix-bound TG-L1Ig6 are able to respond differentially to 2D- or 3D-environments suggesting the use of gradients for cell guidance within 3D-scaffolds and on implant surfaces to improve their biomedical functions.     

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.     

Adsorption state of fibronectin on poly(dimethylsiloxane) surfaces with varied stiffness can dominate adhesion density of fibroblasts

The state of adsorbed fibronectin and the subsequent cell adhesion behavior on polydimethylsiloxane (PDMS) substrates with varied stiffness were investigated. The bulk elastic modulus as well as the macroscale and nanoscale surface repulsion forces on PDMS substrates with five different cross-linker concentrations (2.5, 5, 10, 20 and 40 wt.%) were evaluated by using tensile and compression tests as well as atomic force microscopy (AFM) indentation. The PDMS substrate with 10 wt.% cross-linker showed the maximum stiffness in the bulk elastic modulus and macroscale compression test. In contrast, PDMS substrates with 2.5 and 5 wt.% cross-linker concentration showed the maximum stiffness in the nanoscale compression test, which indicates that the physical properties of the nanoscale outermost surface are different from the bulk and macroscale surface properties. The fibronectin-treated PDMS substrates showed almost the same amount of fibronectin adsorption. However, the outermost surface density of fibronectin was related to the macroscale surface stiffness, and the exposure of the cell-binding motif was related to the nanoscale surface stiffness. Moreover, the different adsorption state of fibronectin was further confirmed by quartz crystal microbalance-dissipation (QCM-D) monitoring. The adhesion behavior of NIH3T3 mouse fibroblasts was in turn related to the exposure of the cell-binding motif. These results suggest that the well-known differences in cell adhesion behavior on PDMS substrates with varied stiffness are primarily induced by different responses of fibronectin to the PDMS substrates.     

Inflammatory predictors of ongoing pain 2 years following knee replacement surgery

IntroductionThe prevalence of unrelieved pain following total knee arthroplasty (TKA) is substantial.ObjectiveWe asked if cytokine markers of inflammation in preoperative serum or knee synovial fluid (SF) would predict pain 2 years following TKA.MethodsDemographic data and functional outcomes were recorded at baseline and 2 years with the WOMAC index. Serum and SF tissue samples were collected at the time of surgery. Linear regression modeling was used to determine the relationship between SF/serum inflammatory markers and a lesser improvement in self reported pain at two years follow-up.ResultsOf our 28 patient cohort, significant correlations between serum and SF levels were found for IL-1β (p < 0.002), MIP-1β (p < 0.001), adiponectin (p < 0.001) and leptin (p < 0.001). Adjusted analysis showed that greater SF concentrations of TNF-α, MMP-13 and IL-6 were independent predictors of less pain improvement at two years follow-up (p < 0.05).ConclusionsThose patients, having ongoing pain despite no clinical or radiological cause, may have an inflammatory profile characterizing a predisposition to ongoing pain after TKA.Level of EvidencePrognosis study, Level 2.     

Bone attachment to glass-fibre-reinforced composite implant with porous surface

A method has recently been developed for producing fibre-reinforced composites (FRC) with porous surfaces, intended for use as load-bearing orthopaedic implants. This study focuses on evaluation of the bone-bonding behaviour of FRC implants. Three types of cylindrical implants, i.e. FRC implants with a porous surface, solid polymethyl methacrylate (PMMA) implants and titanium (Ti) implants, were inserted in a transverse direction into the intercondular trabeculous bone area of distal femurs and proximal tibias of New Zealand White rabbits. Animals were sacrificed at 3, 6 and 12 weeks post operation, and push-out tests (n = 5–6 per implant type per time point) were then carried out. At 12 weeks the shear force at the porous FRC–bone interface was significantly higher (283.3 ± 55.3 N) than the shear force at interfaces of solid PMMA/bone (14.4 ± 11.0 N; p < 0.001) and Ti/bone (130.6 ± 22.2 N; p = 0.001). Histological observation revealed new bone growth into the porous surface structure of FRC implants. Solid PMMA and Ti implants were encapsulated mostly with fibrous connective tissue. Finite element analysis (FEA) revealed that porous FRC implants had mechanical properties which could be tailored to smooth the shear stress distribution at the bone–implant interface and reduce the stress-shielding effect.     

The influence of poly(ethylene oxide) grafting via siloxane tethers on protein adsorption

Amphiphilic PEO–silanes (a–c) having siloxane tethers of varying lengths with the general formula α-(EtO)₃Si–(CH₂)₂–oligodimethylsiloxane_n-block-poly(ethylene oxide)₈–OCH₃ [n = 0 (a), n = 4 (b), and n = 13 (c)] were grafted onto silicon wafers and resistance to adsorption of plasma proteins was measured. Distancing the PEO segment from the hydrolyzable triethoxysilane [(EtO)₃Si] grafting group by a oligodimethylsiloxane tether represents a new method of grafting PEO chains to surfaces. Properties of surfaces grafted with a–c were compared to surfaces grafted with a traditional PEO–silane containing a propyl spacer [(EtO)₃Si–(CH₂)₃–poly(ethylene oxide)₈–OCH₃, PEO control]. As the siloxane tether length increased, chain density of PEO–silanes grafted onto oxidized silicon wafers decreased and hydrophobicity of the PEO–silane increased which led to a decrease in surface hydrophilicity. Despite decreased surface hydrophilicity, resistance to the adsorption of bovine serum albumin (BSA) increased in the order: PEO control < a < b ≈ c and to human fibrinogen (HF) increased in the order: PEO control < a < b < c.     

Life-long endurance exercise in humans: Circulating levels of inflammatory markers and leg muscle size

Human aging is associated with a loss of skeletal muscle and an increase in circulating inflammatory markers. It is unknown whether endurance training (Tr) can prevent these changes. Therefore we studied 15 old trained (O-Tr) healthy males and, for comparison, 12 old untrained (O-Un), 10 Young-Tr (Y-Tr) and 12 Young-Un (Y-Un). Quadriceps size, VO₂ peak, CRP, IL-6, TNF-α and its receptors, suPAR, lipid profile, leucocytes and glucose homeostasis were measured. Tr was associated with an improved insulin profile (p < 0.05), and lower leucocyte (p < 0.05) and triglyceride levels (p < 0.05), independent of age. Aging was associated with poorer glucose control (p < 0.05), independent of training. The age-related changes in waist circumference, VO₂ peak, cholesterol, LDL, leg muscle size, CRP and IL-6 were counteracted by physical activity (p < 0.05). A significant increase in suPAR with age was observed (p < 0.05). Most importantly, life-long endurance exercise was associated with a lower level of the inflammatory markers CRP and IL-6 (p < 0.05), and with a greater thigh muscle area (p < 0.05), compared to age-matched untrained counterparts. These findings in a limited group of individuals suggest that regular physical endurance activity may play a role in reducing some markers of systemic inflammation, even within the normal range, and in maintaining muscle mass with aging.     

The in vitro corneal biocompatibility of hydroxyapatite coated carbon mesh

The purpose of this study was to consider the use of a hydroxyapatite (HA) coated porous carbon matrix as a synthetic dental laminate substitute in osteo-odonto-keratoprosthetic (OOKP) design. 3 types of carbon meshes were coated with HA by sonoelectrochemical deposition. The materials were characterised by scanning electron microscopy (SEM) and HA deposition was characterised by elemental analysis and X-ray diffractometry (XRD). In vitro assays were carried out to quantify the effects of HA coating on human keratocyte adhesion. Cellular cytokine production was used to assess inflammatory potential. HA coating significantly increased keratocyte adhesion to the carbon matrix (p < 0.01). The materials did not induce excessive cytokine production by the adherent keratocytes. In addition, the matrices themselves adsorbed significant levels of the cytokine IL-8 (p < 0.05). The results indicate that HA coated carbon matrices provide a suitable environment to enhance in-growth of corneal cells without inducing further inflammation. The materials may also suppress excessive inflammation by adsorption of the cytokine IL-8 into the porous, internal carbon structure.     

The effect of polyelectrolyte multilayer coated titanium alloy surfaces on implant anchorage in rats

Advances have been achieved in the design and biomechanical performance of orthopedic implants in the last decades. These include anatomically shaped and angle-stable implants for fracture fixation or improved biomaterials (e.g. ultra-high-molecular-weight polyethylene) in total joint arthroplasty. Future modifications need to address the biological function of implant surfaces. Functionalized surfaces can promote or reduce osseointegration, avoid implant-related infections or reduce osteoporotic bone loss. To this end, polyelectrolyte multilayer structures have been developed as functional coatings and intensively tested in vitro previously. Nevertheless, only a few studies address the effect of polyelectrolyte multilayer coatings of biomaterials in vivo. The aim of the present work is to evaluate the effect of polyelectrolyte coatings of titanium alloy implants on implant anchorage in an animal model. We test the hypotheses that (1) polyelectrolyte multilayers have an effect on osseointegration in vivo; (2) multilayers of chitosan/hyaluronic acid decrease osteoblast proliferation compared to native titanium alloy, and hence reduce osseointegration; (3) multilayers of chitosan/gelatine increase osteoblast proliferation compared to native titanium alloy, hence enhance osseointegration. Polyelectrolyte multilayers on titanium alloy implants were fabricated by a layer-by-layer self-assembly process. Titanium alloy (Ti) implants were alternately dipped into gelatine (Gel), hyaluronic acid (HA) and chitosan (Chi) solutions, thus assembling a Chi/Gel and a Chi/HA coating with a terminating layer of Gel or HA, respectively. A rat tibial model with bilateral placement of titanium alloy implants was employed to analyze the bones’ response to polyelectrolyte surfaces in vivo. 48 rats were randomly assigned to three groups of implants: (1) native titanium alloy (control), (2) Chi/Gel and (3) Chi/HA coating. Mechanical fixation, peri-implant bone area and bone contact were evaluated by pull-out tests and histology at 3 and 8 weeks. Shear strength at 8 weeks was statistically significantly increased (p < 0.05) in both Chi/Gel and Chi/HA groups compared to the titanium alloy control. No statistically significant difference (p > 0.05) in bone contact or bone area was found between all groups. No decrease of osseointegration of Chi/HA-coated implants compared to non-coated implants was found. The results of polyelectrolyte coatings in a rat model showed that the Chi/Gel and Chi/HA coatings have a positive effect on mechanical implant anchorage in normal bone.     

Biphasic calcium phosphate to repair nasal septum: The first in vitro and in vivo study

Our objective was to evaluate the cytocompatibility and biocompatibility of biphasic calcium phosphate (BCP) in the nasal respiratory airway. In vitro, the attachment rate was quantified on BCP disks with normal human epithelial cells at 1, 3 and 24 h by determining N-acetyl β-d-hexosaminidase activity. Proliferative activity of cells was indirectly assessed by MTT assay at 3, 9, 15 and 21 days. Plastic surfaces were used as positive control. In vivo, 15 rabbits underwent anterior nasal septum perforation and 10 septa were repaired with BCP disks. Five non-implanted animals were sacrificed at 3 months. Two groups of five implanted animals were sacrificed at 1 and 2 months. The surface of new airway mucosa covering BCP disks was evaluated macroscopically. During both steps, light microscopy, immunohistochemistry and scanning electron microscopy were performed. Statistical analysis was performed with the Mann–Whitney U-test. In vitro, at 1 and 3 h, the attachment rates were significantly better than on the plastic surface (p < 10^?2). Mitochondrial activity increased on both surfaces but began 6 days later than on plastic. After 21 days of culture, cells were confluent and formed a monolayer covering the implant even in the bottom of the pores. In vivo, no perforations in the control group closed spontaneously. The mean rate of closure was 63% in the 1 month group and 64% in the 2 month group (p > 0.05). Implants were invaded by inflammatory reaction covered by incomplete differentiated respiratory epithelium. Throughout the study, all immunohistochemical findings remained positive. These data suggest a good affinity between BCP and nasal epithelial cells. BCP could be used to rebuild nasal septa.     

Effects of phosphoric acid treatment of titanium surfaces on surface properties,osteoblast response and removal of torque forces

This study investigated the surface characteristics and biocompatibility of phosphate ion (P)-incorporated titanium (Ti) surfaces hydrothermally treated with various concentrations of phosphoric acid (H₃PO₄). 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). MC3T3-E1 cell attachment, spreading, proliferation and osteoblastic gene expression on different surfaces were evaluated. The degree of bony integration was biomechanically evaluated by removal torque testing after 4 weeks of healing in rabbit tibiae. The H₃PO₄ treatment produced micro-rough Ti surfaces with crystalline P-incorporated Ti oxide layers. High concentration H₃PO₄ treatment (1% and 2%) produced significantly higher hydrophilic surfaces compared with low H₃PO₄ treatment (0.5%) and untreated surfaces (P < 0.01). ICP-MS analysis showed P ions were released from P-incorporated surfaces. Significant increased cell attachment (P < 0.05) and notably higher mRNA expressions of Runx2, alkaline phosphatase, osteopontin and osteocalcin were observed in cells grown on P-incorporated surfaces compared with cells on untreated machined surfaces. P-incorporated surfaces showed significantly higher removal torque forces compared with untreated machined implants (P < 0.05). Ti surfaces treated with 2% H₃PO₄ showed increasing tendencies in osteoblastic gene expression and removal torque forces compared with those treated with lower H₃PO₄ concentrations or untreated surfaces. These results demonstrate that H₃PO₄ treatment may improve the biocompatibility of Ti implants by enhancing osteoblast attachment, differentiation and biomechanical anchorage.