Amphiphilic Degradable Polymer/Hydroxyapatite Composites as Smart Bone Tissue Engineering Scaffolds: A Dissertation

Over 600,000 bone-grafting operations are performed each year in the United States. The majority of the bone used for these surgeries comes from autografts that are limited in quantity or allografts with high failure rates. Current synthetic bone grafting materials have poor mechanical properties, handling characteristics, and bioactivity. The goal of this dissertation was to develop a clinically translatable bone tissue engineering scaffold with improved handling characteristics, bioactivity, and smart delivery modalities. We hypothesized that this could be achieved through the rational selection of Food and Drug Administration (FDA) approved materials that blend favorably with hydroxyapatite (HA), the principle mineral component in bone. This dissertation describes the development of smart bone tissue engineering scaffolds composed of the biodegradable amphiphilic polymer poly(D,L-lactic acid-co-ethylene glycol-co- D,L-lactic acid) (PELA) and HA. Electrospun nanofibrous HA-PELA scaffolds exhibited improved handling characteristics and bioactivity over conventional HApoly( D,L-lactic acid) composites. Electrospun HA-PELA was hydrophilic, elastic, stiffened upon hydration, and supported the attachment and osteogenic differentiation of rat bone marrow stromal cells (MSCs). These in vitro properties translated into robust bone formation in vivo using a critical-size femoral defect model in rats. Spiral-wrapped HA-PELA scaffolds, loaded with MSCs or a lowdose of recombinant human bone morphogenetic protein-2, templated bone formation along the defect. As an alternate approach, PELA and HA-PELA were viii rapid prototyped into three-dimensional (3-D) macroporous scaffolds using a consumer-grade 3-D printer. These 3-D scaffolds have differential cell adhesion characteristics, swell and stiffen upon hydration, and exhibit hydration-induced self-fixation in a simulated confined defect. HA-PELA also exhibits thermal shape memory behavior, enabling the minimally invasive delivery and rapid (\u3e3 sec) shape recovery of 3-D scaffolds at physiologically safe temperatures (~ 50ºC). Overall, this dissertation demonstrates how the rational selection of FDA approved materials with synergistic interactions results in smart biomaterials with high potential for clinical translation

High Performance Amphiphilic Polymer/Hydroxyapatite Composite Tissue Scaffolds

There is a critical clinical need for alternatives to autograft and allograft bone for over 500,000 bone grafting operations performed each year in the United States. Current synthetic bone grafts suffer from poor handling characteristics, brittle mechanical properties, and inconsistent bioactivity. By blending a biodegradable amphiphilic polymer with hydroxyapatite (HA), the main mineral component in bone, we developed an improved synthetic bone graft. The polymer/HA composites were fabricated in both 2-D and 3-D forms by electrospinning and 3-D printing. These materials exhibited unique handling characteristics such as high tensile elasticity (\u3e200% failure strain) and self-stiffening properties upon hydration, allowing their facile/stabile fixation around an open defect or within a confined defect. They are also superhydrophilic, enabling the absorption of aqueous cell suspensions and protein therapeutics. We showed that bone marrow-derived mesenchymal stem cells (MSCs) readily attached to these scaffolds and expressed increased levels of osteogenic genes with and without osteogenic induction in culture. These scaffolds also supported the retention and sustained release of rhBMP-2. These high-performance composite materials are being explored for guided bone regeneration and skeletal tissue repair in various formats

Rapid prototyping amphiphilic polymer/hydroxyapatite composite scaffolds with hydration-induced self-fixation behavior

Two major factors hampering the broad use of rapid prototyped biomaterials for tissue engineering applications are the requirement for custom-designed or expensive research-grade three-dimensional (3-D) printers and the limited selection of suitable thermoplastic biomaterials exhibiting physical characteristics desired for facile surgical handling and biological properties encouraging tissue integration. Properly designed thermoplastic biodegradable amphiphilic polymers can exhibit hydration-dependent hydrophilicity changes and stiffening behavior, which may be exploited to facilitate the surgical delivery/self-fixation of the scaffold within a physiological tissue environment. Compared to conventional hydrophobic polyesters, they also present significant advantages in blending with hydrophilic osteoconductive minerals with improved interfacial adhesion for bone tissue engineering applications. Here we demonstrated the excellent blending of biodegradable, amphiphilic PLA-PEG-PLA (PELA) triblock co-polymer with hydroxyapatite (HA) and the fabrication of high-quality rapid prototyped 3-D macroporous composite scaffolds using an unmodified consumer-grade 3-D printer. The rapid prototyped HA-PELA composite scaffolds and the PELA control (without HA) swelled (66% and 44% volume increases, respectively) and stiffened (1.38-fold and 4-fold increases in compressive modulus, respectively) in water. To test the hypothesis that the hydration-induced physical changes can translate into self-fixation properties of the scaffolds within a confined defect, a straightforward in vitro pull-out test was designed to quantify the peak force required to dislodge these scaffolds from a simulated cylindrical defect at dry vs. wet states. Consistent with our hypothesis, the peak fixation force measured for the PELA and HA-PELA scaffolds increased 6-fold and 15-fold upon hydration, respectively. Furthermore, we showed that the low-fouling 3-D PELA inhibited the attachment of NIH3T3 fibroblasts or MSCs while the HA-PELA readily supported cellular attachment and osteogenic differentiation. Finally, we demonstrated the feasibility of rapid prototyping biphasic PELA/HA-PELA scaffolds for potential guided bone regeneration where an osteoconductive scaffold interior encouraging osteointegration and a non-adhesive surface discouraging fibrous tissue encapsulation is desired. This work demonstrated that by combining facile and readily translatable rapid prototyping approaches with unique biomaterial designs, biodegradable composite scaffolds with well-controlled macroporosities, spatially defined biological microenvironment, and useful handling characteristics can be developed

Effect of hydration on conductivity of Ba4La x Ca2-X Nb2O11 + 0.5x (x = 0.5, 1, 1.5, 2) phases

Substitution of Ca by La in initial cubic double perovskite Ba 4(Ca2Nb2)O11[VO]1 allowed obtaining phases with a similar structure with a lower content of structural oxygen vacancies, Ba4(La x Ca2-x Nb 2)O11 + 0.5x [VO]1-0.5x (x = 0.5, 1, 1.5, 2). The impedance technique was used to measure the temperature dependences of conductivity in the atmosphere of dry and humid air. Transport numbers determined using the EMF method in an oxygen-air and water steam concentration cells point to the predominantly hole nature of conductivity in the high-temperature region (T > 600 C) and to predominance of proton conductivity in the low-temperature region. Activation energies of hole and proton conductivity were calculated. Thermogravimetric measurements were carried out under heating from 25 to 1000 C with simultaneous mass-spectrometric determination of evolved H2O and CO2. The properties of the studied Ba4(La x Ca2-x Nb 2)O11 + 0.5x (x = 0.5, 1, 1.5, 2) phases were compared with the earlier studied Ba4-x La x (Ca2Nb 2)O11 + 0.5x phases with similar lanthanum content. © 2013 Pleiades Publishing, Ltd

Stem cell labeling using polyethylenimine conjugated (alpha-NaYbF4:Tm3+)/CaF2 upconversion nanoparticles

We report on a polyethylenimine (PEI) covalently conjugated (alpha-NaYbF4:Tm3+)/CaF2 upconversion nanoparticle (PEI-UCNP) and its use for labeling rat mesenchymal stem cells (rMSCs). The PEI-UCNPs absorb and emit near-infrared light, allowing for improved in vivo imaging depth over conventional probes. We found that such covalent surface conjugation by PEI results in a much more stable PEI-UCNP suspension in PBS compared to conventional electrostatic layer by layer (LbL) self-assembling coating approach. We systematically examined the effects of nanoparticle dose and exposure time on rat mesenchymal stem cell (rMSC) cytotoxicity. The exocytosis of PEI-UCNPs from labeled rMSCs and the impact of PEI-UCNP uptake on rMSC differentiation was also investigated. Our data show that incubation of 100-microg/mL PEI-UCNPs with rMSCs for 4 h led to efficient labeling of the MSCs, and such a level of PEI-UCNP exposure imposed little cytotoxicity to rMSCs (95% viability). However, extended incubation of PEI-UCNPs at the 100 microg/mL dose for 24 hour resulted in some cytotoxicity to rMSCs (60% viability). PEI-UCNP labeled rMSCs also exhibited normal early proliferation, and the internalized PEI-UCNPs did not leak out to cause unintended labeling of adjacent cells during a 14-day transwell culture experiment. Finally, PEI-UCNP labeled rMSCs were able to undergo osteogenic and adipogenic differentiation upon in vitro induction, although the osteogenesis of labeled rMSCs appeared to be less potent than that of the unlabeled rMSCs. Taken together, PEI-UCNPs are promising agents for stem cell labeling and tracking

Деякі фармакоекономічні аспекти епілепсії у дорослих

According to some publications, epilepsy accounts for 0.6 % of the global burden of diseases, and the cost of treatment of one patient with epilepsy reaches 88.2 % of the gross national product per capita. All this determines a great importance of pharmacoeconomic aspects in the treatment and life of these patients.Aim. To perform pharmacoeconomic calculations for patients with epilepsy; to assess the following indices: cost of the disease, the human capital index, the realized human capital, a percentage of the human capital implementation, direct and indirect medical costs, a percentage of direct medical costs.Materials and methods. The analysis was carried out on the basis of a sample from 74 patients (including 31 male and 43 female patients) with the average age of 36.99 ± 1.10 years old, who lived in Kharkiv or the Kharkiv Region.Results. It has been found that the treatment of epilepsy inUkraine is rather expensive. The general cost of the disease is just over 100,000 UAH per year for a patient (approximately 3,500 US dollars). The most part of this cost consisted of indirect costs (73 % of the general cost of the disease on average). Indirect costs depend directly on the presence of seizures; patients, who have no seizures for a long time, have significantly lower indirect costs. All the indices of costs are the highest in a symptomatic form of epilepsy and the lowest in its idiopathic form. Among clinical parameters, the most significant ones for the formation of the cost of the disease were mental health disorders in epilepsy, especially dementia as a result of epilepsy.Conclusions. The study performed shows that it is appropriate to use high-cost drugs, which result in compensation of the disease, ever from the economic point of view, not to mention social and medical aspects. In addition, the successful therapy with achieving long-term control of seizures (in accordance with data from the International League Against Epilepsy, it is possible for 70 % of such patients) results in an increase of a part of the realized human capital, i.e. in an appropriate socialization of these patients.По некоторым данным, на эпилепсию приходится 0,6 % глобального бремени болезней, а стоимость лечения одного пациента с эпилепсией достигает 88,2 % валового национального продукта на душу населения. Это обусловливает большое значение фармакоэкономических аспектов лечения и жизни этих больных.Целью исследования было проведение фармакоэкономических расчетов для больных эпилепсией. Оценены такие показатели, как цена болезни, индекс человеческого капитала, реализованный человеческий капитал, процент реализации человеческого капитала, прямые и непрямые медицинские затраты, процент прямых медицинских затрат.Материалы и методы. Анализ был выполнен на основе выборки из медицинских карт 74 пациентов (включая 31 мужчину и 43 женщины), проживающих в г. Харькове или Харьковской области, средний возраст больных составляет 36,99 ± 1,10 лет.Результаты. Выявлено, что лечение эпилепсии в Украине является достаточно дорогим. Общая стоимость болезни составляет чуть более ста тысяч гривен в год у одного пациента, что эквивалентно примерно 3500 долларам США. Большую часть расходов составляют непрямые затраты (в среднем 73 % общей стоимости болезни). Размер косвенных затрат напрямую зависит от наличия приступов – у больных, не имеющих приступов длительное время, непрямые затраты значительно уменьшаются. Все затраты являются самыми высокими при симптоматической форме эпилепсии и наименьшими – при идиопатической форме болезни. Среди клинических показателей наиболее значимыми для формирования затрат вследствие болезни оказались психические расстройства при эпилепсии, особенно деменция вследствие эпилепсии.Выводы. Проведенное исследование свидетельствует о целесообразности применения препаратов высокой стоимости, которые приводят к компенсации заболевания даже с экономической точки зрения, не говоря уже о социальных и медицинских аспектах. Кроме того, именно успешное лечение с достижением длительного контроля приступов, которое, по данным Международной противоэпилептической лиги, возможно у 70 % этих пациентов, приводит к увеличению доли реализованного человеческого капитала, то есть к адекватной социализации этих больных.За окремими даними, на епілепсію припадає 0,6 % глобального тягаря хвороб, а вартість лікування одного пацієнта з епілепсією сягає 88,2 % валового національного продукту на душу населення. Це обумовлює велике значення фармакоекономічних аспектів лікування та життя цих хворих.Метою дослiдження було проведення фармакоекономічних розрахунків для хворих на епілепсію. Оцінені такі показники, як ціна хвороби, індекс людського капіталу, реалізований людський капітал, відсоток реалізації людського капіталу, прямі і непрямі медичні витрати, відсоток прямих медичних витрат.Матерiали та методи. Аналіз був виконаний на основі вибірки з медичних карток 74 пацієнтів (включаючи 31 чоловіка та 43 жінки) середнім віком 36,99 ± 1,10 років, які проживають у м. Харкові або Харківській області.Результати. Як виявилося, лікування епілепсії в Україні є достатньо дорогим. Загальна вартість хвороби становить трохи більше ста тисяч гривень на рік у одного пацієнта, що еквівалентно приблизно 3500 доларів США. Більшу частку витрат складають саме непрямі витрати (в середньому 73 % загальної вартості хвороби). Розмір непрямих витрат напряму залежить від наявності нападів – у хворих, які не мають нападів тривалий час, непрямі витрати значно зменшуються. Усі показники витрат є найбільшими при симптоматичній формі епілепсії і найменшими – при її ідіопатичній формі. Серед клінічних показників найбільш значущими для формування витрат на хворобу виявилися психічні розлади при епілепсії, особливо деменція внаслідок епілепсії.Висновки. Проведене дослідження свідчить про доцільність застосування препаратів високої вартості, які приводять до компенсації захворювання навіть з економічної точки зору, не кажучи вже про соціальні та медичні аспекти. Окрім того, саме успішне лікування з досягненням тривалого контролю нападів, яке за даними Міжнародної протиепілептичної ліги є можливим у 70 % таких хворих, приводить до збільшення частки реалізованого людського капіталу, тобто до адекватної соціалізації таких хворих

TRAFD1 (FLN29) Interacts with Plekhm1 and Regulates Osteoclast Acidification and Resorption

Plekhm1 is a large, multi-modular, adapter protein implicated in osteoclast vesicle trafficking and bone resorption. In patients, inactivating mutations cause osteopetrosis, and gain-of-function mutations cause osteopenia. Investigations of potential Plekhm1 interaction partners by mass spectrometry identified TRAFD1 (FLN29), a protein previously shown to suppress toll-like receptor signaling in monocytes/macrophages, thereby dampening inflammatory responses to innate immunity. We mapped the binding domains to the TRAFD1 zinc finger (aa 37-60), and to the region of Plekhm1 between its second pleckstrin homology domain and its C1 domain (aa 784-986). RANKL slightly increased TRAFD1 levels, particularly in primary osteoclasts, and the co-localization of TRAFD1 with Plekhm1 also increased with RANKL treatment. Stable knockdown of TRAFD1 in RAW 264.7 cells inhibited resorption activity proportionally to the degree of knockdown, and inhibited acidification. The lack of acidification occurred despite the presence of osteoclast acidification factors including carbonic anhydrase II, a3-V-ATPase, and the ClC7 chloride channel. Secretion of TRAP and cathepsin K were also markedly inhibited in knockdown cells. Truncated Plekhm1 in ia/ia osteopetrotic rat cells prevented vesicle localization of Plekhm1 and TRAFD1. We conclude that TRAFD1, in association with Plekhm1/Rab7-positive late endosomes-early lysosomes, has a previously unknown role in vesicle trafficking, acidification, and resorption in osteoclasts

Cadmium down-regulates expression of XIAP at the post-transcriptional level in prostate cancer cells through an NF-κB-independent, proteasome-mediated mechanism

<p>Abstract</p> <p>Background</p> <p>Cadmium has been classified as a human carcinogen, affecting health through occupational and environmental exposure. Cadmium has a long biological half-life (>25 years), due to the flat kinetics of its excretion. The prostate is one of the organs with highest levels of cadmium accumulation. Importantly, patients with prostate cancer appear to have higher levels of cadmium both in the circulation and in prostatic tissues.</p> <p>Results</p> <p>In the current report, we demonstrate for the first time that cadmium down-regulates expression of the X-linked inhibitor of apoptosis protein (XIAP) in prostate cancer cells. Cadmium-mediated XIAP depletion occurs at the post-transcriptional level via an NF-κB-independent, proteasome-mediated mechanism and coincides with an increased sensitivity of prostate cancer cells to TNF-α-mediated apoptosis. Prolonged treatment with cadmium results in selection of prostate cancer cells with apoptosis-resistant phenotype. Development of apoptosis-resistance coincides with restoration of XIAP expression in cadmium-selected PC-3 cells.</p> <p>Conclusions</p> <p>Selection of cadmium-resistant cells could represent an adaptive survival mechanism that may contribute to progression of prostatic malignancies.</p