Evaluation of soft-tissue response around laser microgrooved titanium percutaneous devices.

Percutaneous devices are prone to epidermal downgrowth and sinus tract formation, which can serve as a nidus for bacterial colonization and increase the risk of peri-prosthetic infection. A laser microgrooved topography has been shown to limit gingival epidermal downgrowth around dental implants. However, the efficacy of this laser microgrooved topography to limit epidermal downgrowth around nongingival percutaneous devices is yet to be investigated. In this study, devices with a porous-coated subdermal component and a percutaneous post were designed and manufactured. The proximal 2 mm section of the percutaneous post were left smooth, or were textured with either a porous coating, or with the laser microgrooved topography. The smooth and porous topographies served as controls. The devices were tested in a hairless guinea pig back model, where 18 animals were randomly assigned into three groups, with each group receiving one implant type (n = 6/group). Four weeks postimplantation, the devices with surrounding soft-tissues were harvested and processed for histological analyses. Results indicated that the laser microgrooved topography failed to prevent epidermal downgrowth (23 ± 4%) around percutaneous posts in this model. Furthermore, no significant differences (p = 0.70) in epidermal downgrowth were present between the three topographies, with all the groups exhibiting similar measures of downgrowth. Overall, these findings suggest that the laser microgrooved topography may not halt downgrowth around percutaneous devices for dermal applications.

Biological therapeutics of Pongamia pinnata coated zinc oxide nanoparticles against clinically important pathogenic bacteria, fungi and MCF-7 breast cancer cells.

The overuse of antimicrobics and drugs has led to the development of resistance in a number of pathogens and parasites, which leads to great concerns for human health and the environment. Furthermore, breast cancer is the second most common cause of cancer death in women. MCF-7 is a widely used epithelial cancer cell line, derived from breast adenocarcinoma for in vitro breast cancer studies, since the cell line has retained several ideal characteristics particular to the mammary epithelium. In this scenario, the development of novel and eco-friendly drugs are of timely importance. Green synthesis of nanoparticles is cost effective, environmental friendly and does not involve the use of toxic chemicals or elevate energy inputs. This research focused on the anticancer activity of Pongamia pinnata seed extract-fabricated zinc oxide nanoparticles (Pp-ZnO NPs) on human MCF-7 breast cancer cells, antibiofilm activity against bacteria and fungi was also investigated. Nanoparticles were characterized by UV-Vis spectroscopy, X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FTIR), Scanning electron microscopy (SEM) and Energy dispersive X-ray spectroscopy (EDX). Pp-ZnO NPs effectively inhibited the growth of Gram positive Bacillus licheniformis (zone of inhibition: 17.3 mm) at 25 µg ml-1 followed by Gram negative Pseudomonas aeruginosa (14.2 mm) and Vibrio parahaemolyticus (12.2 mm). Pp-ZnO NPs also effectively inhibited the biofilm formation of C. albicans at 50 µg ml-1. Cytotoxicity studies revealed that a single treatment with Pp-ZnO NPs significantly reduced the cell viability of breast cancer MCF-7 cells at doses higher than 50 µg ml-1. Morphological changes in the Pp-ZnO NPs treated MCF-7 breast cancer cells were observed using phase contrast microscopy. This study concludes that the green synthesized Pp-ZnO NPs may be used as an effective antimicrobial and antibreast cancer agents.

Effect of nanocoating with rhamnogalacturonan-I on surface properties and osteoblasts response.

Long-term stability of titanium implants are dependent on a variety of factors. Nanocoating with organic molecules is one of the methods used to improve osseointegration. Therefore, the aim of this study is to evaluate the in vitro effect of nanocoating with pectic rhamnogalacturonan-I (RG-I) on surface properties and osteoblasts response. Three different RG-Is from apple and lupin pectins were modified and coated on amino-functionalized tissue culture polystyrene plates (aminated TCPS). Surface properties were evaluated by scanning electron microscopy, contact angle measurement, atomic force microscopy, and X-ray photoelectron spectroscopy. The effects of nanocoating on proliferation, matrix formation and mineralization, and expression of genes (real-time PCR) related to osteoblast differentiation and activity were tested using human osteoblast-like SaOS-2 cells. It was shown that RG-I coatings affected the surface properties. All three RG-I induced bone matrix formation and mineralization, which was also supported by the finding that gene expression levels of alkaline phosphatase, osteocalcin, and collagen type-1 were increased in cells cultured on the RG-I coated surface, indicating a more differentiated osteoblastic phenotype. This makes RG-I coating a promising and novel candidate for nanocoatings of implants.

Effects of dexamethasone-loaded PLGA microspheres on human fetal osteoblasts.

Integration of a drug delivery function into implantable medical devices enables local release of specific bioactives to control cells-surface interactions. One alternative to achieve this biofunctionality for bone implants is to incorporate particulate drug delivery systems (DDSs) into the rough or porous implant surfaces. The scope of this study was to assess the effects of a model DDS consisting of poly(D,L-lactide-co-glycolide) (PLGA) microspheres loaded with an anti-inflammatory drug, dexamethasone (DXM), on the response of Simian Virus-immortalized Human Fetal Osteoblast (SV-HFO) cells. The microspheres were prepared by the oil-in-water emulsion/solvent evaporation method, whereas cells response was investigated by Alamar Blue test for viability, alkaline phosphatase (ALP) activity for differentiation, and Alizarin Red staining for matrix mineralization. Cell viability was not affected by the presence of increased concentrations of polymeric microspheres in the culture media. Furthermore, in the cultures with DXM-loaded microspheres, ALP activity was expressed at levels similar with those obtained under osteogenic conditions, indicating that DXM released from the microsphere-stimulated cell differentiation. Matrix mineralization occurred preferentially around the DXM-loaded microspheres confirming that the released DXM could act as osteogenic supplement for the cells. These in vitro findings suggest that a particulate PLGA-DXM DDS may actually provide dual, anti-inflammatory and osteogenic functions when incorporated on the surface of bone implants.

The effect of zinc on hydroxyapatite-mediated activation of human polymorphonuclear neutrophils and bone implant-associated acute inflammation.

Hydroxyapatite (HA) is widely used as coating biomaterial for prosthesis metal parts and as bone substitute. The release of HA particles induces an inflammatory response and, if uncontrolled, could result in implant loss. At the inflamed site, the polymorphonuclear cells (PMNs) represent the earliest phagocytic cells that predominate the cellular infiltrate. We have recently proposed that HA wear debris activate polymorphonuclear cells (PMNs) initiating and/or amplifying thereby the acute inflammatory response. Previous studies have shown that activation of monocytes by HA could be modulated by supplementing this latter with the divalent cation, Zinc. The purpose of this work was to investigate the modulation of PMNs activation following exposure to zinc-substituted HA. Our study demonstrate that addition of zinc to HA particles resulted in decreased levels of the pro-inflammatory mediator interleukin-8 (IL-8) and the matrix metallo-proteinase-9. We also show that these changes involve IL-8 receptors (CXCR-1 and CXCR-2).

The effects of implant surface nanoscale features on osteoblast-specific gene expression.

This study investigated the influence of nanoscale implant surface features on osteoblast differentiation. Titanium disks (20.0 x 1.0 mm) with different nanoscale materials were prepared using sol-gel-derived coatings and characterized by scanning electron microscopy, atomic force microscopy and analyzed by X-ray Photoelectron Spectrometer. Human Mesenchymal Stem Cells (hMSCs) were cultured on the disks for 3-28 days. The levels of ALP, BSP, Runx2, OCN, OPG, and OSX mRNA and a panel of 76 genes related to osteogenesis were evaluated. Topographical and chemical evaluation confirmed nanoscale features present on the coated surfaces only. Bone-specific mRNAs were increased on surfaces with superimposed nanoscale features compared to Machined (M) and Acid etched (Ac). At day 14, OSX mRNA levels were increased by 2-, 3.5-, 4- and 3-fold for Anatase (An), Rutile (Ru), Alumina (Al), and Zirconia (Zr), respectively. OSX expression levels for M and Ac approximated baseline levels. At days 14 and 28 the BSP relative mRNA expression was significantly up-regulated for all surfaces with nanoscale coated features (up to 45-fold increase for Al). The PCR array showed an up-regulation on Al coated implants when compared to M. An improved response of cells adhered to nanostructured-coated implant surfaces was represented by increased OSX and BSP expressions. Furthermore, nanostructured surfaces produced using aluminum oxide significantly enhanced the hMSC gene expression representative of osteoblast differentiation. Nanoscale features on Ti implant substrates may improve the osseointegration response by altering adherent cell response.

Human osteoblastic cell response to a Ca- and P-enriched titanium surface obtained by anodization.

The aim of the present study was to evaluate the in vitro osteogenic potential of subcultured human osteoblastic cells derived from alveolar bone on a titanium (Ti) surface produced by an anodized alkali treatment (BSP-AK). Primary osteoblastic cells were subcultured on BSP-AK and machined Ti discs (control) and grown for periods of up to 21 days under osteogenic conditions. Morphologic and biochemical methods were used to assess important parameters of in vitro bone-like tissue formation. Although no major differences were observed between the BSP-AK and the control Ti surface in terms of cell attachment and mineralized matrix formation, a significant increase in cell population, ALP activity, and collagen content was detected in cultures on BSP-AK surface. Our results demonstrate that human osteoblastic cells are sensitive to the BSP-AK-modified Ti surface during the transitional stage between the end of the proliferative phase and the onset of the differentiation/matrix maturation ones. Together with the good mechanical properties exhibited by the Ca- and P- coating, our findings suggest that BSP-AK treatment could be useful for the development of a new surface for dental and orthopedic implants.

An in vitro comparison of possibly bioactive titanium implant surfaces.

The aim of the study was to compare Ca and P formation (CaP) and subsequent bone cell response of a blasted and four different possibly bioactive commercially pure (cp) titanium surfaces; 1. Fluoride etched (Fluoride), 2. Alkali-heat treated (AH), 3. Magnesium ion incorporated anodized (TiMgO), and 4. Nano HA coated and heat treated (nano HA) in vitro. Furthermore, to evaluate the significance of the SBF formed CaP coat on bone cell response. The surfaces were characterized by Optical Interferometry, Scanning Electron Microscopy (SEM) and X-ray Photoelectron Spectroscopy (XPS). CaP formation was evaluated after 12, 24 and 72 h in simulated body fluid (SBF). Primary human mandibular osteoblast-like cells were cultured on the various surfaces subjected to SBF for 72 h. Cellular attachment, differentiation (osteocalcin) and protein production (TGF-beta(1)) was evaluated after 3 h and 10 days respectively. Despite different morphological appearances, the roughness of the differently modified surfaces was similar. The possibly bioactive surfaces gave rise to an earlier CaP formation than the blasted surface, however, after 72 h the blasted surface demonstrated increased CaP formation compared to the possibly bioactive surfaces. Subsequent bone cell attachment was correlated to neither surface roughness nor the amount of formed CaP after SBF treatment. In contrast, osteocalcin and TGF-beta(1) production were largely correlated to the amount of CaP formed on the surfaces. However, bone response (cell attachment, osteocalcin and TGF-F production) on the blasted controls were similar or increased compared to the SBF treated fluoridated, AH and TiMgO surface.

Using thermal energy produced by irradiation of Mn-Zn ferrite magnetic nanoparticles (MZF-NPs) for heat-inducible gene expression.

One of the main advantages of gene therapy over traditional therapy is the potential to target the expression of therapeutic genes in desired cells or tissues. To achieve targeted gene expression, we developed a novel heat-inducible gene expression system in which thermal energy generated by Mn-Zn ferrite magnetic nanoparticles (MZF-NPs) under an alternating magnetic field (AMF) was used to activate gene expression. MZF-NPs, obtained by co-precipitation method, were firstly surface modified with cation poly(ethylenimine) (PEI). Then thermodynamic test of various doses of MZF-NPs was preformed in vivo and in vitro. PEI-MZF-NPs showed good DNA binding ability and high transfection efficiency. In AMF, they could rise to a steady temperature. To analyze the heat-induced gene expression under an AMF, we combined P1730OR vector transfection with hyperthermia produced by irradiation of MZF-NPs. By using LacZ gene as a reporter gene and Hsp70 as a promoter, it was demonstrated that expression of a heterogeneous gene could be elevated to 10 to 500-fold over background by moderate hyperthermia (added 12.24 or 25.81 mg MZF-NPs to growth medium) in tissue cultured cells. When injected with 2.6 or 4.6 mg MZF-NPs, the temperature of tumor-bearing nude mice could rise to 39.5 or 42.8 degrees C, respectively, and the beta-gal concentration could increase up to 3.8 or 8.1 mU/mg proteins accordingly 1 day after hyperthermia treatment. Our results therefore supported hyperthermia produced by irradiation of MZF-NPs under an AMF as a feasible approach for targeted heat-induced gene expression. This novel system made use of the relative low Curie point of MZF-NPs to control the in vivo hyperthermia temperature and therefore acquired safe and effective heat-inducible transgene expression.

High concentration of residual aluminum oxide on titanium surface inhibits extracellular matrix mineralization.

In the present study we characterized titanium (Ti) surfaces submitted to different treatments and evaluated the response of osteoblasts derived from human alveolar bone to these surfaces. Five different surfaces were evaluated: ground (G), ground and chemical etched (G1-HF for 60 s), sand blasted (SB-Al(2)O(3) particles 65 mum), sand blasted and chemical etched (SLA1-HF for 60 s and SLA2-HF for 13 s). Surface morphology was evaluated under SEM and roughness parameters by contact scanning instrument. The presence of Al(2)O(3) was detected by EDS and the amount calculated by digital analyses. Osteoblasts were cultured on these surfaces and it was evaluated: cell adhesion, proliferation, and viability, alkaline phosphatase activity, total protein content, and matrix mineralization formation. Physical and chemical treatments produced very different surface morphologies. Al(2)O(3) residues were detected on SB and SLA2 surfaces. Only matrix mineralization formation was affected by different surface treatments, being increased on rough surface (SLA1) and reduced on surface with high amount of Al(2)O(3) residues (SB). On the basis of these findings, it is possible to conclude that high concentration of residual Al(2)O(3) negatively interfere with the process of matrix mineralization formation in contact with Ti implant surfaces.

Biocompatibility of nanoporous alumina membranes for immunoisolation.

Cellular immunoisolation using semi-permeable barriers has been investigated over the past several decades as a promising treatment approach for diseases such as Parkinson's, Alzheimer's, and Type 1 diabetes. Typically, polymeric membranes are used for immunoisolation applications; however, recent advances in technology have led to the development of more robust membranes that are able to more completely meet the requirements for a successful immunoisolation device, including well controlled pore size, chemical and mechanical stability, nonbiodegradability, and biocompatibility with both the graft tissue as well as the host. It has been shown previously that nanoporous alumina biocapsules can act effectively as immunoisolation devices, and support the viability and functionality of encapsulated beta cells. The aim of this investigation was to assess the biocompatibility of the material with host tissue. The cytotoxicity of the capsule, as well as its ability to activate complement and inflammation was studied. Further, the effects of poly(ethylene glycol) (PEG) modification on the tissue response to implanted capsules were studied. Our results have shown that the device is nontoxic and does not induce significant complement activation. Further, in vivo work has demonstrated that implantation of these capsules into the peritoneal cavity of rats induces a transient inflammatory response, and that PEG is useful in minimizing the host response to the material.

Prolonged duration local anesthesia with lipid-protein-sugar particles containing bupivacaine and dexamethasone.

Glucocorticoids prolong block duration from polymeric microspheres containing bupivacaine, but not from unencapsulated drug. Here we investigate this effect applies to particles with much more rapid drug release and improved long-term biocompatibility. Male Sprague-Dawley rats were given sciatic nerve blocks with 75 mg of 3% or 60% (w/w) dipalmitoylphosphatidylcholine (DPPC) spray-dried lipid-protein-sugar particles (LPSPs) containing 10% (w/w) bupivacaine and 0%, 0.05%, or 0.1% (w/w) dexamethasone. Sensory nerve block from bupivacaine-containing 3% and 60% (w/w) DPPC particles without dexamethasone yielded blocks lasting 301 +/- 56 and 321 +/- 127 min, respectively. Addition of 0.05% (w/w) dexamethasone increased block durations to 610 +/- 182 and 538 +/- 222 min, respectively; increasing dexamethasone loading to 0.1% did not further increase duration. One day after injection, dexamethasone-containing particles resulted in lower inflammation scores and capsule thickness than dexamethasone-free particles, but the difference was gone by day 4. Excipient composition had prominent effects at all time points. For all groups, inflammation was largely resolved by 2 weeks after injection. Dexamethasone approximately doubled the duration of nerve block from bupivacaine-loaded LPSPs, while maintaining excellent biocompatibility. Such formulations could be useful in clinical applications when nerve blockade is needed for 24 hours or less.