Low-intensity pulsed ultrasound as a useful adjuvant during distraction osteogenesis: a prospective, randomized controlled trial.

BACKGROUND: Low-intensity pulsed ultrasound (LIPUS) was proven to have a positive impact on bone healing in animal and clinical studies. METHODS: In this prospective, randomized controlled trial the effect of LIPUS during distraction osteogenesis was investigated. Thirty-six patients who underwent distraction osteogenesis (>2 cm) were enrolled. Sixteen patients in the treatment group received LIPUS, and 20 patients as control group did not. Ultrasound treatment device was transcutaneously applied at the distraction gap for 20 minutes daily (frequency 1.5 MHz, signal burst with 200 µs, signal repetition frequency 1.0 kHz, intensity 30 mW/cm(2)). Evaluation of patients was performed by standard radiographs every 3 weeks to 4 weeks. RESULTS: Average transport distance was 7.0 cm in the ultrasound group and 6.3 cm in the control group. Mean Paley index for the ultrasound group was 1.09 mo/cm and 1.49 mo/cm for the control group. Mean distraction consolidation index for the ultrasound group was 32.8 d/cm and 44.6 d/cm for the control group. The calculated indices indicated no significant statistical difference between the two groups (p < 0.116) but the fixator gestation period could be decreased for 43.6 days in the treatment group. CONCLUSIONS: Therapeutic application of LIPUS during callus distraction constitutes a useful adjuvant treatment during distraction osteogenesis and has a positive effect on healing time with no negative effects.

Sterilization of heat-sensitive silicone implant material by low-pressure gas plasma.

BACKGROUND: In recent years, plasma treatment of medical devices and implant materials has gained more and more acceptance. Inactivation of microorganisms by exposure to ultraviolet (UV) radiation produced by plasma discharges and sterilization of medical implants and instruments is one possible application of this technique. The aim of this study was to evaluate the effectiveness of this sterilization technique on silicone implant material. METHODS: Bacillus atrophaeus spores (10(6) colony-forming units [CFUs]) were sprayed on the surfaces of 12 silicone implant material samples. Four plasma sets with different gas mixtures (argon [Ar], argon-oxygen [Ar:O(2)], argon-hydrogen [Ar:H(2)] and argon-nitrogen [Ar:N(2)]) were tested for their antimicrobial properties. Post-sterilization mechanical testing of the implant material was performed in order to evaluate possible plasma-induced structural damage. RESULTS: The inductively coupled low-pressure plasma technique can achieve fast and efficient sterilization of silicone implant material without adverse materials effects. All four gas mixtures led to a significant spore reduction, and no structural damage to the implant material could be observed.

Plasma mediated collagen-I-coating of metal implant materials to improve biocompatibility.

This study describes the collagen-I coating of titanium and steel implants via cold low-pressure gas plasma treatment. To analyze the coatings in terms of biocompatibility osteoblast-like osteosarcoma cells and human leukocytes were cultivated on the metal surfaces. Two different implant materials were assessed (Ti6Al4V, X2CrNiMo18) and four different surface properties were evaluated: (a) plasma pretreated and collagen-I coated implant materials; (b) collagen-I dip-coated without plasma pretreatment; (c) plasma treated but not collagen-I coated; (d) standard implant materials served as control. The different coating characteristics were analyzed by scanning electron microscopy (SEM). For adhesion and viability tests calcein-AM staining of the cells and Alamar blue assays were performed. The quantitative analysis was conducted by computer assisted microfluorophotography and spectrometer measurements. SEM analysis revealed that stable collagen-I coatings could not be achieved on the dip-coated steel and titanium alloys. Only due to pretreatment with low-pressure gas plasma a robust deposition of collagen I on the surface could be achieved. The cell viability and cell attachment rate on the plasma pretreated, collagen coated surfaces was significantly (p < 0.017) increased compared to the non coated surfaces. Gas plasma treatment is a feasible method for the deposition of proteins on metal implant materials resulting in an improved biocompatibility in vitro. (c) 2010 Wiley Periodicals, Inc. J Biomed Mater Res, 2010.

[Surface modification of metal implant materials by low-pressure plasma treatment]. / Oberflächenmodifikation metallischer Implantatmaterialien durch Niederdruckplasmabehandlung / Surface modification of metal implant materials by low-pressure plasma treatment.

BACKGROUND: Plasma treatment leads to a significant change of surface free energy of medical implant materials. These changes strongly influence protein and cell adhesion on the material surface. The aim of the study was to quantify the plasma-induced surface changes and to analyse whether the change of treatment parameters, such as pressure, gas mixture, energy and treatment time, influences the surface free energy of the implant materials. To improve the biocompatibility of the surfaces, polyamino acid coating experiments were performed. MATERIALS AND METHODS: Three different metal implant materials (X2CrNiMo18-15-3, Ti6Al4V, Ti6Al7Nb) were treated with a double-inductively coupled low-pressure plasma. The influence of treatment parameter variation on the surface free energy was evaluated by drop shape analysis. The plasma treated and non-treated materials were incubated in collagen I solution. Afterwards, the coatings were analysed by electron microscopy in terms of structure and adhesion. RESULTS: Drop shape analysis revealed that plasma treatment leads to a significant increase of surface free energy in all groups. Long plasma treatment times and low treatment pressures lead to a significant (p<0.05) extension of the detectable surface free energy increase. Coating experiments showed that only on plasma-treated samples solid and adherent collagen layers could be achieved.

Ultrasound-induced modifications of cytoskeletal components in osteoblast-like SAOS-2 cells.

In clinical and experimental studies an acceleration of fracture healing and increased callus formation induced by low-intensity pulsed ultrasound (LIPUS) has been demonstrated. The exact molecular mechanisms of ultrasound treatment are still unclear. In this study ultrasound transmitted cytoskeletal and growth rate changes of SAOS-2 cells were examined. Osteoblast-like cell lines (SAOS-2) were treated using low-intensity pulsed ultrasound. Cytoskeletal changes were analyzed using rhodamine phalloidine for f-actin staining and indirect immunofluorescence techniques with different monoclonal antibodies against several tubulin modifications. To examine changes of cell number after ultrasound treatment cell counts were done. Significant changes in cytoskeleton structure were detected compared to controls, including an enhancement of stress fiber formation combined with a loss of cell migration after ultrasound application. We further observed that sonication altered the proportion of the more stable microtubules to the more labile microtubule subclass. The labile tyrosinated microtubules appeared highly enhanced, whereas the amount of the more stable acetylated microtubules was remarkably diminished. All these observations were quantified by fluorometric measurements. The centrosomal gamma-tubulin was frequently scattered throughout the cell's cytoplasm, giving rise to additional polyglu-positive microtubular asters, which induced multipolar spindles, leading either to aneuploid mini-or giant cells. Moreover, a significant increase of cell number was noticed in the sonicated group. These experiments demonstrate that ultrasound treatment increases cell number and leads to significant changes of the cytoskeletal structure and composition in vitro.

[A double inductively coupled low-pressure plasma for sterilization of medical implant materials]. / Doppelt induktiv gekoppeltes Niederdruckplasma zur Sterilisation von medizinischen Implantatmaterialien.

The potential of plasma treatment in medicine is only slowly gaining acceptance. Inactivation of germs through exposure to UV radiation produced by plasma discharges and sterilization of medical implant devices and instruments is one possible application of this technique. In addition, due to the manifold possibilities of coating through plasma processes, quick sterilization-coating combinations of medical implant devices are possible. To analyze the effectiveness of this sterilization process on different material surfaces, three different alloys (X2CrNiMo18-15-3, Ti6Al7Nb and Ti6Al4V) and one thermoplastic material (ultra-high molecular weight polyethylene, UHMWPE), commonly used in medical implant devices, were examined in the presented study. After spraying Bacillus atrophaeus spores (10(6) CFU) on the surfaces of four different implant materials tested in this study (X2CrNiMo18-15-3, UHMWPE, Ti6Al7Nb and Ti6Al4V), it was demonstrated in each of four gas mixtures used (Ar, Ar:O2, Ar:H2 and Ar:N2) that due to the application of inductively coupled low-pressure plasma technique, plain medical implant materials can be sterilized rapidly, and can be protective and efficient.

Morphological characterization and in vitro biocompatibility of a porous nickel-titanium alloy.

Disks consisting of macroporous nickel-titanium alloy (NiTi, Nitinol, Actipore) are used as implants in clinical surgery, e.g. for fixation of spinal dysfunctions. The morphological properties were studied by scanning electron microscopy (SEM) and by synchrotron radiation-based microtomography (SRmuCT). The composition was studied by X-ray diffractometry (XRD), differential scanning calorimetry (DSC), and energy-dispersive X-ray spectroscopy (EDX). The mechanical properties were studied with temperature-dependent dynamical mechanical analysis (DMA). Studies on the biocompatibility were performed by co-incubation of porous NiTi samples with isolated peripheral blood leukocyte fractions (polymorphonuclear neutrophil granulocytes, PMN; peripheral blood mononuclear leukocytes, PBMC) in comparison with control cultures without NiTi samples. The cell adherence to the NiTi surface was analyzed by fluorescence microscopy and scanning electron microscopy. The activation of adherent leukocytes was analyzed by measurement of the released cytokines using enzyme-linked immunosorbent assay (ELISA). The cytokine response of PMN (analyzed by the release of IL-1ra and IL-8) was not significantly different between cell cultures with or without NiTi. There was a significant increase in the release of IL-1ra (p<0.001), IL-6 (p<0.05), and IL-8 (p<0.05) from PBMC in the presence of NiTi samples. In contrast, the release of TNF-alpha by PBMC was not significantly elevated in the presence of NiTi. IL-2 was released from PBMC only in the range of the lower detection limit in all cell cultures. The material, clearly macroporous with an interconnecting porosity, consists of NiTi (martensite; monoclinic, and austenite; cubic) with small impurities of NiTi2 and possibly NiC(x). The material is not superelastic upon manual compression and shows a good biocompatibility.

Inhibition of PMN apoptosis after adherence to dip-coated calcium phosphate surfaces on a NiTi shape memory alloy.

Nickel-titanium shape-memory alloys (NiTi-SMA) were coated with calcium phosphate by dipping in oversaturated calcium phosphate solution (CaP-coating). Polymorphonuclear neutrophil granulocytes (PMN) belong to the first cells which will adhere to implant materials. We analyzed the apoptosis of isolated human PMN after cell culture on non-coated and CaP-coated NiTi-SMA by light and scanning electron microscopy (cell morphology) and by flow cytometry (DNA-fragmentation). In contrast to PMN adherent to non-coated NiTi-SMA, the apoptosis of PMN adherent CaP-coated samples was inhibited. Cell culture media obtained from cultured leukocytes with CaP-coatings (conditioned media, CM) were able to transfer the apoptosis inhibiting activities to freshly isolated PMN. There was a significant (p<0.01) increase in GM-CSF, IL-8, IL-6, and TNF-alpha within CM obtained from coated versus non-coated NiTi-SMA as was determined by ELISA.

Calcium phosphate coating of nickel-titanium shape-memory alloys. Coating procedure and adherence of leukocytes and platelets.

Nickel-titanium shape-memory alloys (NiTi-SMA) were coated with calcium phosphate by dipping in oversaturated calcium phosphate solution. The layer thickness (typically 5-20 micrometer) can be varied by choice of the immersion time. The porous nature of the layer of microcrystals makes it mechanically stable enough to withstand both the shape-memory transition upon cooling and heating and also strong bending of the material (superelastic effect). This layer may improve the biocompatibility of NiTi-SMA, particulary for osteosynthetic devices by creating a more physiological surface and by restricting a potential nickel release. The adherence of human leukocytes (peripheral blood mononuclear cells and polymorphonuclear neutrophil granulocytes) and platelets to the calcium phosphate layer was analyzed in vitro. In comparison to non-coated NiTi-SMA, leukocytes and platelets showed a significantly increased adhesion to the coated NiTi-SMA.