Characteristics and in vitro response of thin hydroxyapatite-titania films produced by plasma electrolytic oxidation of Ti alloys in electrolytes with particle additions

The enhancement of the biological properties of Ti by surface doping with hydroxyapatite (HA) is of great significance, especially for orthodontic applications. This study addressed the effects of HA particle size in the electrolyte suspension on the characteristics and biological properties of thin titania-based coatings produced on Ti–6Al–4V alloy by plasma electrolytic oxidation (PEO). Detailed morphological investigation of the coatings formed by a single-stage PEO process with two-step control of the electrical parameters was performed using the Minkowski functionals approach. The surface chemistry was studied by glow discharge optical emission spectroscopy and Fourier transform infrared spectroscopy, whereas mechanical properties were evaluated using scratch tests. The biological assessment included in vitro evaluation of the coating bioactivity in simulated body fluid (SBF) as well as studies of spreading, proliferation and osteoblastic differentiation of MC3T3-E1 cells. The results demonstrated that both HA micro- and nanoparticles were successfully incorporated in the coatings but had different effects on their surface morphology and elemental distributions. The micro-particles formed an irregular surface morphology featuring interpenetrated networks of fine pores and coating material, whereas the nanoparticles penetrated deeper into the coating matrix which retained major morphological features of the porous TiO2 coating. All coatings suffered cohesive failure in scratch tests, but no adhesive failure was observed; moreover doping with HA increased the coating scratch resistance. In vitro tests in SBF revealed enhanced bioactivity of both HA-doped PEO coatings; furthermore, the cell proliferation/morphometric tests showed their good biocompatibility. Fluorescence microscopy revealed a well-organised actin cytoskeleton and focal adhesions in MC3T3-E1 cells cultivated on these substrates. The cell alkaline phosphatase activity in the presence of ascorbic acid and β-glycerophosphate was significantly increased, especially in HA nanoparticle-doped coatings

Экспериментальное исследование антибактериальной активности литического стафилококкового бактериофага ph20 и литического бактериофага синегнойной палочки ph57 при моделировании их импрегнации в ортопедические полимерные конструкции из полиметилметакрилата (костного цемента)

Background: The problem of bacterial colonization of implants used in medical practice continues to be relevant regardless of the material of the implant. Particular attention deserves polymeric implants, which are prepared ex tempore from polymethyl methacrylate, for example - duting orthopedic surgical interventions (so-called "bone cement"). The protection of such implants by antibiotic impregnation is subjected to multiple criticisms, therefore, as an alternative to antibiotics, lytic bacteriophages with a number of unique advantages can be used - however, no experimental studies have been published on the possibility of impregnating bacteriophages into polymethyl methacrylate and their antibacterial activity assessment under such conditions.Aims: to evaluate the possibility of physical placement of bacteriophages in polymethylmethacrylate and to characterize the lytic antibacterial effect of two different strains of bacteriophages when impregnated into polymer carrier ex tempore during the polymerization process in in vitro model.Materials and methods:  First stage - Atomic force microscopy (AFM) of polymethyl methacrylate samples for medical purposes was used to determine the presence and size of caverns in polymethyl methacrylate after completion of its polymerization at various reaction  temperatures (+6…+25°C and +18…+50°C).The second stage was performed in vitro and included an impregnation of two different bacteriophage strains (phage ph20 active against S. aureus and ph57 active against Ps. aeruginosa) into polymethyl methacrylate during the polymerization process, followed by determination of their antibacterial activity.Results: ACM showed the possibility of bacteriophages placement in the cavities of polymethyl methacrylate - the median of the section and the depth of cavities on the outer surface of the polymer sample polymerized at +18…+50°C were 100.0 and 40.0 nm, respectively, and on the surface of the transverse cleavage of the sample - 120.0 and 100.0 nm, respectively, which statistically did not differ from the geometric dimensions of the caverns of the sample polymerized at a temperature of +6…+25°C.The study of antibacterial activity showed that the ph20 bacteriophage impregnated in polymethyl methacrylate at +6…+25°C lost its effective titer within the first six days after the start of the experiment, while the phage ph57 retained an effective titer for at least 13 days.Conclusion: the study confirmed the possibility of bacteriophages impregnation into medical grade polymethyl methacrylate, maintaining the effective titer of the bacteriophage during phage emission into the external environment, which opens the way for the possible application of this method of bacteriophage delivery in clinical practice. It is also assumed that certain bacteriophages are susceptible to aggressive influences from the chemical components of "bone cement" and / or polymerization reaction products, which requires strict selection of bacteriophage strains that could be suitable for this method of delivery.Обоснование. Проблема бактериальной колонизации используемых в медицинской практике имплантатов из различных материалов продолжает оставаться актуальной, независимо от использованного для их изготовления материала. Отдельного внимания заслуживают имплантируемые в организм человека полимерные имплантаты, которые изготовляют ex tempore (по мере надобности) из полиметилметакрилата, например при ортопедических хирургических вмешательствах (так называемый костный цемент). Защита таких имплантатов путем импрегнации в них антибиотиков подвергается множественной критике, поэтому в качестве альтернативы антибиотикам могут быть использованы литические бактериофаги, обладающие рядом уникальных преимуществ, однако экспериментальных работ по изучению возможности импрегнации бактериофагов в полиметилметакрилат и антибактериальной активности в таких условиях в литературе не опубликовано. Цель исследования ― изучить возможность физического размещения бактериофагов в полиметилметакрилате и в модели in vitro охарактеризовать литический антибактериальный эффект двух различных штаммов бактериофагов при их импрегнации в изготавливаемый ex tempore полимерный носитель на этапе полимеризации. Методы. Первым этапом была проведена атомно-силовая микроскопия (АСМ) образцов полиметилметакрилата медицинского назначения для выяснения наличия и размеров каверн, образовавшихся после завершения полимеризации при различном диапазоне температур реакционной смеси (+6…+25 °C и +18…+50 °C). Вторым этапом in vitro было проведена импрегнация двух различных штаммов бактериофагов (ph20, активного в отношении Staphylococcus aureus, и ph57, активного в отношении Pseudomonas aeruginosa) в полиметилметакрилат на этапе полимеризации с последующим определением их антибактериальной активности. Результаты. В ходе выполнения АСМ установлена возможность размещения бактериофагов в кавернах полиметилметакрилата: медиана сечения и глубины каверн на внешней поверхности образца, полимеризованного при температуре +18…+50 °C, составила 100,0 и 40,0 нм соответственно, а на поверхности поперечного скола образца ― 120,0 и 100,0 нм соответственно, что статистически не отличалось от геометрических размеров каверн образца, полимеризованного при температуре +6…+25 °C. Изучение антибактериальной активности показало, что импрегнированный при +6…+25 °C в полиметилметакрилат стафилококковый бактериофаг ph20 утратил эффективный титр уже в течение первых шести суток с момента начала эксперимента, тогда как синегнойный бактериофаг ph57 сохранял эффективный титр как минимум в течение 13 сут. Заключение. В исследовании была подтверждена возможность импрегнации бактериофагов в полиметилметакрилат медицинского назначения с поддержанием эффективного титра бактериофага при его эмиссии во внешнюю среду, что открывает пути возможного применения такого способа доставки бактериофагов в клинической практике. Также сделаны предположения о вероятной подверженности некоторых бактериофагов агрессивным воздействиям со стороны химических компонентов «костного цемента» и/или продуктов реакции полимеризации, что требует строгого отбора пригодных для подобного способа доставки штаммов бактериофагов

Comparative investigation of Al- and Cr-doped TiSiCN coatings

The aim of this work was a comparative investigation of the structure and properties of Al- and Cr-doped TiSiCN coatings deposited by magnetron sputtering of composite TiAlSiCN and TiCrSiCN targets produced by self-propagating high-temperature synthesis method. Based on X-ray diffraction, scanning and transmission electron microscopy, X-ray photoelectron spectroscopy, and Raman spectroscopy data, the Al- and Cr-doped TiSiCN coatings possessed nanocomposite structures (Ti,Al)(C,N)/a-(Si,C) and (Ti,Cr)(C,N)/a-SiCxNy/a-C with cubic crystallites embedded in an amorphous matrix. To evaluate the thermal stability and oxidation resistance, the coatings were annealed either in vacuum at 1000, 1100, 1200, and 1300°C or in air at 1000°C for 1h. The results obtained show that the hardness of the Al-doped TiSiCN coatings increased from 41 to 46GPa, reaching maximum at 1000°C, and then slightly decreased to 38GPa at 1300°C. The Cr-doped TiSiCN coatings demonstrated high thermal stability up to 1100°C with hardness above 34GPa. Although both Al- and Cr-doped TiSiCN coatings possessed improved oxidation resistance up to 1000°C, the TiAlSiCN coatings were more oxidation resistant than their TiCrSiCN counterparts. The TiCrSiCN coatings showed better tribological characteristics both at 25 and 700°C and superior cutting performance compared with the TiAlSiCN coatings. © 2011 Elsevier B.V.The work was fulfilled due to financial support from the Ministry of Education and Science of the Russian Federation (Contracts 02.740.11.0859 and П1248). The authors thank A.V. Levanov (Moscow State University) for Raman spectroscopy investigations and T.B. Sagalova (MISIS) for help with XRD measurements.Peer Reviewe

Mechanical properties of decellularized extracellular matrix coated with TiCaPCON film

For the first time the surface of decellularized extracellular matrix (DECM) was modified via deposition of a multicomponent bioactive nanostructured film for improvement of the DECM's mechanical properties. TiCaPCON films were deposited onto the surface of intact and decellularized ulna, radius, and humerus bones by magnetron sputtering of TiC<SUB>0.5</SUB> + 10%Ca<SUB>3</SUB>(PO<SUB>4</SUB>)<SUB>2</SUB> and Ti targets in a gaseous mixture of Ar + N<SUB>2</SUB>. The film structure was studied using x-ray diffraction, scanning and transmission electron microscopy, and Raman spectroscopy. The films were characterized in terms of their wettability, as well as adhesion strength to the intact bone and DECM substrates. The mechanical properties of TiCaPCON-coated samples were investigated by compression testing. In addition, humerus bones were evaluated during three-point bending tests. The results indicate that the tightly adhered films, uniformly covering the DECM surfaces, possessed hydrophilic characteristics. A maximum improvement in mechanical properties (250%) was observed for coated humerus samples. In case of decellularized radius bones, the compressive strength also increased by 150% after coating. The positive role of TiCaPCON films was less noticeable for ulna bones because of large data scattering. These results clearly indicate that the films acted as a rigid frame that increased the material compressive strength. Compared with intact bones, fracture in the TiCaPCON-coated DECM samples was characterized by rarer and larger cracks generated under higher critical loads. As a result, the samples were crushed into several large pieces and numerous tiny fragments. Although the film deposition increased the bone stiffness, the bending tests revealed that the flexural strength of the coated samples became 20%–25% lower than the strength of the film-free samples