Investigation of the Influence of Machining Parameters and Surface Roughness on the Wettability of the Al6082 Surfaces Produced with WEDM.

Electrical Discharge Machining (EDM) is a non-conventional machining technique, capable of processing any kind of conductive material. Recently, it has been successfully utilized for producing hydrophobic characteristics in inherently hydrophilic metallic materials. In this work, Wire Electrical Discharge Machining (WEDM) was utilized for producing hydrophobic characteristics on the surface of the aluminum alloy 6082, and various parameters that can affect wettability were investigated. Adopting an orthogonal Taguchi approach, the effects of the process parameter values of peak current, pulse-on time, and gap voltage on the contact angles of the machined surfaces were investigated. After machining, all samples were observed to have obtained hydrophobic properties, reaching contact angles up to 132°. The peak current was identified as the most influential parameter regarding the contact angle, while the gap voltage was the less influential parameter. A contact angle variation of 30° was observed throughout different combinations of machining parameters. Each combination of the machining parameters resulted in a distinct surface morphology. The samples with moderate roughness values (3.4 µm > Sa > 5.7 µm) were found to be more hydrophobic than the samples with high or low values, where the contact angle was measured under 115°. In addition, the finite element modeling of the experimental setup, with parametric surfaces of uniform random and Perlin noise types of roughness, was implemented. Time dependent simulations coupling phase field and laminar flow for the modelingof the wetting of surfaces with different surface roughness characteristics showed that an increase in the Sa roughness and total wetted area can lead to an increase in the contact angle. The combination of experimental and computational results suggests that the complexity of the wettability outcomes of aluminum alloy surfaces processed with WEDM lies in the interplay between variations of the surface chemical composition, roughness, micro/nano morphology, and the surface capability of forming a composite air/water interface.

Controlled Delivery of Celecoxib-ß-Cyclodextrin Complexes from the Nanostructured Titanium Dioxide Layers.

Considering the potential of nanostructured titanium dioxide layers as drug delivery systems, it is advisable to indicate the possibility of creating a functional drug delivery system based on anodic TiO2 for celecoxib as an alternative anti-inflammatory drug and its inclusion complex with ß-cyclodextrin. First, the optimal composition of celecoxib-ß-cyclodextrin complexes was synthesized and determined. The effectiveness of the complexation was quantified using isothermal titration calorimetry (ITC), differential scanning calorimetry (DSC), infrared spectroscopy (FT-IR) nuclear magnetic resonance (1H NMR), and scanning electron microscopy (SEM). Then, nanostructured titanium dioxide layers (TiO2) were synthesized using the electrochemical oxidation technique. The TiO2 layers with pore diameters of 60 nm and layer thickness of 1.60 µm were used as drug delivery systems. The samples were modified with pure celecoxib and the ß-cyclodextrin-celecoxib complex. The release profiles shown effective drug release from such layers during 24 h. After the initial burst release, the drug was continuously released from the pores. The presented results confirm that the use of nanostructured TiO2 as a drug delivery system can be effectively used in more complicated systems composed of ß-cyclodextrin-celecoxib complexes, making such drugs available for pain treatment, e.g., for orthopedic surgeries.

Multidisciplinary management of chronic pain in elderly oncology patients.

Chronic pain is one of the most common and most bothersome symptoms in cancer patients, which occurs especially often in the elderly population. Although methods of pain treatment are well known, it is not uncommon for individuals with chronic or terminal illnesses to remain underdiagnosed or untreated. Effective pain management has become the measure of success in oncology therapy. For this reason, effective pain management has become an indispensable success factor of multidisciplinary oncological therapy. Along with the growing interest in the holistic approach in medicine, and hence in interdisciplinary treatment, the management of cancer pain in older patients was presented.

Estrogen Receptors Mediated Negative Effects of Estrogens and Xenoestrogens in Teleost Fishes-Review.

Estrogen receptors (ERs) play a key role in many biochemical and physiological processes, that are involved in maintaining organism homeostasis. At the most basic level, they can be divided into nuclear estrogen receptors and membrane estrogen receptors that imply their effect in two ways: slower genomic, and faster non-genomic. In these ways, estrogens and xenoestrogens can negatively affect animal health and welfare. Most of the available literature focuses on human and mammalian physiology, and clearly, we can observe a need for further research focusing on complex mutual interactions between different estrogens and xenoestrogens in aquatic animals, primarily fishes. Understanding the mechanisms of action of estrogenic compounds on the ERs in fishes and their negative consequences, may improve efforts in environmental protection of these animals and their environment and benefit society in return. In this review, we have summarized the ER-mediated effects of xenoestrogens and estrogens on teleost fishes metabolism, their carcinogenic potential, immune, circulatory, and reproductive systems.

The Impacts of Crystalline Structure and Different Surface Functional Groups on Drug Release and the Osseointegration Process of Nanostructured TiO2.

In implantable materials, surface topography and chemistry are the most important in the effective osseointegration and interaction with drug molecules. Therefore, structural and surface modifications of nanostructured titanium dioxide (TiO2) layers are reported in the present work. In particular, the modification of annealed TiO2 samples with -OH groups and silane derivatives, confirmed by X-ray photoelectron spectroscopy, is shown. Moreover, the ibuprofen release process was studied regarding the desorption-desorption-diffusion (DDD) kinetic model. The results proved that the most significant impact on the release profile is annealing, and further surface modifications did not change its kinetics. Additionally, the cell adhesion and proliferation were examined based on the MTS test and immunofluorescent staining. The obtained data showed that the proposed changes in the surface chemistry enhance the samples' hydrophilicity. Moreover, improvements in the adhesion and proliferation of the MG-63 cells were observed.

Growth of Lactic Acid Bacteria on Gold-Influence of Surface Roughness and Chemical Composition.

The main focus of this work was to establish a correlation between surface topography and chemistry and surface colonization by lactic acid bacteria. For this reason, we chose gold substrates with different surface architectures (i.e., smooth and nanorough) that were characterized by atomic force microscopy (AFM), electron scanning microscopy (SEM), and X-ray diffractometry (XRD). Moreover, to enhance biocompatibility, we modified gold substrates with polymeric monolayers, namely cationic dextran derivatives with different molar masses. The presence of those layers was confirmed by AFM, infrared spectroscopy (IR), and X-ray photoelectron spectroscopy (XPS). In order to determine the adhesion abilities of non-modified and modified gold surfaces, we tested three lactic acid bacteria (LAB) strains (i.e., Lactobacillus rhamnosus GG, Lactobacillus acidophilus, and Lactobacillus plantarum 299v). We have shown that surface roughness influences the surface colonization of bacteria, and the most significant impact on the growth was observed for the Lactobacillus rhamnosus GG strain. What is more, covering the gold surface with a molecular polymeric film by using the layer-by-layer (LbL) method allows additional changes in the bacterial growth, independently on the used strain. The well-being of the bacteria cells on tested surfaces was confirmed by using selective staining and fluorescence microscopy. Finally, we have determined the bacterial metabolic activity by measuring the amount of produced lactic acid regarding the growth conditions. The obtained results proved that the adhesion of bacteria to the metallic surface depends on the chemistry and topography of the surface, as well as the specific bacteria strain.

Plant Elongator-Protein Complex of Diverse Activities Regulates Growth, Development, and Immune Responses.

Contrary to the conserved Elongator composition in yeast, animals, and plants, molecular functions and catalytic activities of the complex remain controversial. Elongator was identified as a component of elongating RNA polymerase II holoenzyme in yeast, animals, and plants. Furthermore, it was suggested that Elonagtor facilitates elongation of transcription via histone acetyl transferase activity. Accordingly, phenotypes of Arabidopsis elo mutants, which show development, growth, or immune response defects, correlate with transcriptional downregulation and the decreased histone acetylation in the coding regions of crucial genes. Plant Elongator was also implicated in other processes: transcription and processing of miRNA, regulation of DNA replication by histone acetylation, and acetylation of alpha-tubulin. Moreover, tRNA modification, discovered first in yeast and confirmed in plants, was claimed as the main activity of Elongator, leading to specificity in translation that might also result indirectly in a deficiency in transcription. Heterologous overexpression of individual Arabidopsis Elongator subunits and their respective phenotypes suggest that single Elongator subunits might also have another function next to being a part of the complex. In this review, we shall present the experimental evidence of all molecular mechanisms and catalytic activities performed by Elongator in nucleus and cytoplasm of plant cells, which might explain how Elongator regulates growth, development, and immune responses.

Hierarchical Nanoporous Sn/SnOx Systems Obtained by Anodic Oxidation of Electrochemically Deposited Sn Nanofoams.

A simple two-step electrochemical method for the fabrication of a new type of hierarchical Sn/SnOx micro/nanostructures is proposed for the very first time. Firstly, porous metallic Sn foams are grown on Sn foil via hydrogen bubble-assisted electrodeposition from an acidulated tin chloride electrolyte. As-obtained metallic foams consist of randomly distributed dendrites grown uniformly on the entire metal surface. The estimated value of pore diameter near the surface is ~35 µm, while voids with a diameter of ~15 µm appear in a deeper part of the deposit. Secondly, a layer of amorphous nanoporous tin oxide (with a pore diameter of ~60 nm) is generated on the metal surface by its anodic oxidation in an alkaline electrolyte (1 M NaOH) at the potential of 4 V for various durations. It is confirmed that if only optimal conditions are applied, the dendritic morphology of the metal foam does not change significantly, and an open-porous structure is still preserved after anodization. Such kinds of hierarchical nanoporous Sn/SnOx systems are superhydrophilic, contrary to those obtained by thermal oxidation of metal foams which are hydrophobic. Finally, the photoelectrochemical activity of the nanostructured metal/metal oxide electrodes is also presented.

Electrochemical Oxidation of Ti15Mo Alloy-The Impact of Anodization Parameters on Surface Morphology of Nanostructured Oxide Layers.

It is well-known that the structure and composition of the material plays an important role in the processes occurring at the surface. In this paper, a surface morphology of nanostructured oxide layers electrochemically grown on Ti15Mo, tuned by applying different anodization parameters, was investigated in detail. The one-step anodization of Ti15Mo alloy was performed at room temperature in an ethylene glycol-based electrolyte containing 0.11 M NH4F and 1.11 M H2O. Different anodization times (ranging from 5 to 60 min) and applied potentials (40-100 V) were tested, and the surface morphology, elemental content, and crystalline structure were monitored by scanning electron microscopy (SEM), energy dispersive X-ray spectrometry (EDS), and X-ray diffractometry (XRD), respectively. The results showed that contrary to the multistep anodization of titanium foil, the surface morphology of anodic oxide obtained via the one-step process contains the nanoporous outer layer covering the nanotubular structure. What is more, the pore diameter (Dp) and interpore distance (Dint) of such layers exhibit different trends than those observed for anodization of pure titanium. In particular, at a certain potential range, a decrease in both Dp and Dint with increasing potential was observed. However, independently on the used anodization conditions, the elemental content of oxide layers remained similar, showing the amount of molybdenum at c.a. 15 wt.%. Finally, the amorphous nature of as-anodized layers was confirmed, and their optical band-gap was determined from the diffuse reflectance UV-Vis spectra. It was found that Eg is tunable to some extent by changing the anodizing potential. However, further thermal treatment in air at 400 °C resulted in the anatase phase formation that was accompanied by a significant Eg reduction. Therefore, we believe that the presented results will greatly contribute to the understanding of anodic formation of nanostructured functional oxide layers with tunable properties that can be applied in various fields.

Tunicamycin induced endoplasmic reticulum changes in endothelial cells investigated in vitro by confocal Raman imaging.

This paper describes how tunicamycin (Tu), the most widely used pharmacological agent for inducing endoplasmic reticulum (ER) stress, interacts with endothelial cells. Our results show that tunicamycin enters the cells and accumulates within the ER area. ER stress takes place when improperly folded or damaged proteins begin to accumulate; however, spectroscopic markers of these changes have not been identified as yet. In this work, Raman spectroscopy and scanning electron microscopy imaging of individual endothelial cells treated with Tu were performed. The changes in the biochemical composition of endothelial cells induced by Tu attributed to ER stress were studied in detail. A main feature of the Tu impact on the cells was a decrease of the phospholipid content in the area of ER, and the most abundant lipid with phosphorus groups found there, was identified as sphingomyelin.

Elderly People's Acceptance of Death: A Study of a Polish Cohort.

Introduction: Old age is usually the natural time for people to prepare for death, which may evoke various emotions ranging from acceptance to hostility. Aim of the work: The study aimed at specifying various degrees to which elderly people accept death. Material and method: The study employed the diagnostic poll method and an Inventory of the Attitude towards Death (IAD) poll questionnaire. The investigation was administered in a cohort of 150 people over 65 years of age living in Poland. Results: The highest results were noted both for males and females on the "Value" scale (M = 4.94 and M = 4.96) and on the "Necessity" scale (M = 4.79 and M = 4.95). These two scales also had the highest values in the cohorts of city dwellers and country dwellers. A statistically significant difference (Z = 2.339, p = 0.019) was found in the "Necessity" dimension between investigated people with higher education and others. Furthermore, statistically significant differences were found in the following dimensions: "Mysteriousness", "Value", "Dread", "Tragedy", and "Absurdity". Comparing death dimensions in people with chronic illnesses and in those without such illnesses, meaningful statistical differences were noted in the "Necessity" dimension (t = 1.983, p = 0.049). However, analysing death dimensions in people who suffered because of a severe illness in a family member and respondents whose families were healthy, statistically significant differences were noted in the "Absurdity" dimension (t = 2.057, p = 0.041). Conclusions: Sex, the place of residence, and death of a close person did not affect elderly people's acceptance of death. On the other hand, those suffering from chronic diseases were more aware of the inevitability of death. People without higher education were also more aware of the inevitability of death. Suffering of a serious disease of a close one considerably affected acceptance of death in the elderly.

Long-lasting antidepressant-like activity of the GPR39 zinc receptor agonist TC-G 1008.

BACKGROUND: The discovery of the zinc-sensing receptor, has provided new possibilities for explaining the neurobiology of zinc. Recent studies indicate that the GPR39 zinc receptor may play an important role in the pathogenesis of depression as well as in the antidepressant mechanism of action. METHODS: In this study we evaluated the time-course of the antidepressant response of the GPR39 agonist (TC-G 1008), imipramine, ZnCl2 and MK-801 in the forced swim test in mice 30 min, 3 h, 6 h and 24 h after acute drug administration as well as after 14-day treatment. Zinc level was measured in serum of mice. BDNF protein level was evaluated in hippocampus following both acute and chronic TC-G 1008 treatment. RESULTS: A single administration of the GPR39 agonist caused an antidepressant-like effect lasting up to 24 h following the injection, which is longer than the effect of imipramine, ZnCl2 and MK-801. Chronic treatment with these compounds caused a decrease in immobility time in the FST. Serum zinc concentrations showed an increased level following chronic ZnCl2 administration, but not following administration of TC-G 1008, imipramine or MK-801. We also observed some tendencies for increased BDNF following acute TC-G 1008 treatment. LIMITATIONS: TC-G 1008 is new drug designed to study GPR39 therefore additional pharmacodynamic and pharmacokinetic properties in preclinical studies are required. CONCLUSION: This study shows for the first time the long-lasting antidepressant effect of the GPR39 agonist in comparison with imipramine, ZnCl2 and MK-801. Our findings suggest that GPR39 should be considered as a target in efforts to develop new antidepressant drugs.

Main communication barriers in the process of delivering bad news to oncological patients - medical perspective.

Delivering bad news is a major aspect of a doctor's work. The literature most often refers to patient's expectations or needs, and methods of delivering bad news, while medical perspective is often skipped. The purpose of this paper is to examine competencies (knowledge, skills and experience) in delivering bad news by medical specialists in the areas related to the causal and symptomatic treatment of oncological patients; identification of major communication problems and obstacles in this specific situation and evaluation of teaching needs for delivering bad news. The study was performed on a group of 61 medical specialists in the areas related to the causal and symptomatic treatment of oncological patients, using a self-generated questionnaire based on other studies in the literature. Topics that are considered most demanding are: delivering news on the termination of causal treatment and preparing the patient/ close ones for death. The most difficult aspect of such discussions for the respondents was associated with the emotions manifested by the patient. On the other hand, doctors were mostly distressed by the feeling of taking the patient's hope away. The study points to the need for education of doctors in the eld of techniques for delivering bad news, particularly in the area of dealing with the emotions manifested by the patient and giving them real hope. The results encourage to conduct studies on a larger group of doctors.

Evaluation of anti-inflammatory and ulcerogenic potential of zinc-ibuprofen and zinc-naproxen complexes in rats.

Because of numerous indications and high availability, non-steroidal anti-inflammatory drugs (NSAIDs) are among the most commonly prescribed and used medicines in the world. However, long-term therapy with and improper use of NSAIDs may lead to gastrointestinal damage. Therefore, improving the therapeutic index of the existing drugs has become a priority over the past decades. Considerable attention in the field has been concentrated on metal complexes of non-steroidal anti-inflammatory drugs. The aim of this study is to evaluate the effect of complexation with zinc on the anti-inflammatory and ulcerogenic effects of ibuprofen and naproxen after single and triple intragastric administration to rats. The anti-inflammatory effect was assessed in carrageenan-induced inflammatory edema in the hind paw of male albino Wistar rats. The mucosal lesions were inspected and evaluated for gross pathology. Single administration of both the investigated complexes, namely zinc-ibuprofen and zinc-naproxen (20 mg/kg equivalent to ibuprofen and naproxen, respectively) and their parent drugs and physical mixtures with zinc hydroaspartate (ZHA doses: 16.05 and 14.37 mg/kg), caused a significant reduction of the edema after the same time from the carrageenan injection in comparison to the control groups. However, no statistically significant differences between the investigated drugs were observed after their single administration. The mean ulceration score for the mixture of ibuprofen and ZHA was statistically lower than the mean score achieved in rats after treatment with ibuprofen alone. On the other hand, triple intragastric administration of the ZHA-ibuprofen and ZHA-naproxen combination showed substantial enhancement of the anti-inflammatory activity against control groups, as well as against the parent NSAIDs. The most potent anti-inflammatory activity was demonstrated after 2 h from the carrageenan injection in animals receiving ZHA together with naproxen. The edema growth was reduced in these animals by 80.9% as compared to the control group. This result was significantly higher than the results achieved in animals receiving zinc-naproxen (50.2%) or naproxen alone (47.9%). Both NSAID complexes with zinc and mixtures with ZHA alleviated ulcerations caused by parent NSAIDs; however, the mixtures of both ibuprofen and naproxen with ZHA after triple administration were the least damaging. In view of the above results, zinc supplementation during NSAID therapy may have a beneficial effect on ulcer prevention and healing by reducing the effective dose of the parent drug and increasing its potency.

Antioxidant and anti-inflammatory effects of zinc. Zinc-dependent NF-κB signaling.

Zinc is a nutritionally fundamental trace element, essential to the structure and function of numerous macromolecules, including enzymes regulating cellular processes and cellular signaling pathways. The mineral modulates immune response and exhibits antioxidant and anti-inflammatory activity. Zinc retards oxidative processes on a long-term basis by inducing the expression of metallothioneins. These metal-binding cysteine-rich proteins are responsible for maintaining zinc-related cell homeostasis and act as potent electrophilic scavengers and cytoprotective agents. Furthermore, zinc increases the activation of antioxidant proteins and enzymes, such as glutathione and catalase. On the other hand, zinc exerts its antioxidant effect via two acute mechanisms, one of which is the stabilization of protein sulfhydryls against oxidation. The second mechanism consists in antagonizing transition metal-catalyzed reactions. Zinc can exchange redox active metals, such as copper and iron, in certain binding sites and attenuate cellular site-specific oxidative injury. Studies have demonstrated that physiological reconstitution of zinc restrains immune activation, whereas zinc deficiency, in the setting of severe infection, provokes a systemic increase in NF-κB activation. In vitro studies have shown that zinc decreases NF-κB activation and its target genes, such as TNF-α and IL-1ß, and increases the gene expression of A20 and PPAR-α, the two zinc finger proteins with anti-inflammatory properties. Alternative NF-κB inhibitory mechanism is initiated by the inhibition of cyclic nucleotide phosphodiesterase, whereas another presumed mechanism consists in inhibition of IκB kinase in response to infection by zinc ions that have been imported into cells by ZIP8.

Co-delivery of ibuprofen and gentamicin from nanoporous anodic titanium dioxide layers.

Although single-drug therapy may prove insufficient in treating bacterial infections or inflammation after orthopaedic surgeries, complex therapy (using both an antibiotic and an anti-inflammatory drug) is thought to address the problem. Among drug delivery systems (DDSs) with prolonged drug release profiles, nanoporous anodic titanium dioxide (ATO) layers on Ti foil are very promising. In the discussed research, ATO samples were synthesized via a three-step anodization process in an ethylene glycol-based electrolyte with fluoride ions. The third step lasted 2, 5 and 10min in order to obtain different thicknesses of nanoporous layers. Annealing the as-prepared amorphous layers at the temperature of 400°C led to obtaining the anatase phase. In this study, water-insoluble ibuprofen and water-soluble gentamicin were used as model drugs. Three different drug loading procedures were applied. The desorption-desorption-diffusion (DDD) model of the drug release was fitted to the experimental data. The effects of crystalline structure, depth of TiO2 nanopores and loading procedure on the drug release profiles were examined. The duration of the drug release process can be easily altered by changing the drug loading sequence. Water-soluble gentamicin is released for a long period of time if gentamicin is loaded in ATO as the first drug. Additionally, deeper nanopores and anatase phase suppress the initial burst release of drugs. These results confirm that factors such as morphological and crystalline structure of ATO layers, and the procedure of drug loading inside nanopores, allow to alter the drug release performance of nanoporous ATO layers.

Effects of nanoporous anodic titanium oxide on human adipose derived stem cells.

The aim of current bone biomaterials research is to design implants that induce controlled, guided, successful, and rapid healing. Titanium implants are widely used in dental, orthopedic, and reconstructive surgery. A series of studies has indicated that cells can respond not only to the chemical properties of the biomaterial, but also, in particular, to the changes in surface topography. Nanoporous materials remain in focus of scientific queries due to their exclusive properties and broad applications. One such material is nanostructured titanium oxide with highly ordered, mutually perpendicular nanopores. Nanoporous anodic titanium dioxide (TiO2) films were fabricated by a three-step anodization process in propan-1,2,3-triol-based electrolyte containing fluoride ions. Adipose-derived stem cells offer many interesting opportunities for regenerative medicine. The important goal of tissue engineering is to direct stem cell differentiation into a desired cell lineage. The influence of nanoporous TiO2 with pore diameters of 80 and 108 nm on cell response, growth, viability, and ability to differentiate into osteoblastic lineage of human adipose-derived progenitors was explored. Cells were harvested from the subcutaneous abdominal fat tissue by a simple, minimally invasive, and inexpensive method. Our results indicate that anodic nanostructured TiO2 is a safe and nontoxic biomaterial. In vitro studies demonstrated that the nanotopography induced and enhanced osteodifferentiation of human adipose-derived stem cells from the abdominal subcutaneous fat tissue.

Nanoporous anodic titanium dioxide layers as potential drug delivery systems: Drug release kinetics and mechanism.

Nanoporous anodic titanium dioxide (ATO) layers on Ti foil were prepared via a three step anodization process in an electrolyte based on an ethylene glycol solution with fluoride ions. Some of the ATO samples were heat-treated in order to achieve two different crystallographic structures - anatase (400°C) and a mixture of anatase and rutile (600°C). The structural and morphological characterizations of ATO layers were performed using a field emission scanning electron microscope (SEM). The hydrophilicity of ATO layers was determined with contact angle measurements using distilled water. Ibuprofen and gentamicin were loaded effectively inside the ATO nanopores. Afterwards, an in vitro drug release was conducted for 24h under a static and dynamic flow conditions in a phosphate buffer solution at 37°C. The drug concentrations were determined using UV-Vis spectrophotometry. The absorbance of ibuprofen was measured directly at 222nm, whether gentamicin was determined as a complex with silver nanoparticles (Ag NPs) at 394nm. Both compounds exhibited long term release profiles, despite the ATO structure. A new release model, based on the desorption of the drug from the ATO top surface followed by the desorption and diffusion of the drug from the nanopores, was derived. The proposed release model was fitted to the experimental drug release profiles, and kinetic parameters were calculated.

D-K6L 9 peptide combination with IL-12 inhibits the recurrence of tumors in mice.

D-K6L9 peptide is bound by phosphatidylserine and induces necrosis in cancer cells. In our therapeutic experience, this peptide, when administered directly into B16-F10 murine melanoma tumors, inhibited their growth. Cessation of therapy results, however, in tumor relapse. We aimed at developing a combined therapy involving D-K6L9 and additional factors that would yield complete elimination of tumor cells in experimental animals. To this purpose, we employed glycyrrhizin, an inhibitor of HMGB1 protein, BP1 peptide and interleukin (IL)-12. Glycyrrhizin or BP1, when combined with D-K6L9, inhibits growth of primary tumors only during the period of their administration. A long-term tumor growth inhibitory effect was obtained only in combining D-K6L9 with IL-12. At 2 months following therapy cessation, 60 % of animals were alive. Prolonged survival was noted in mice bearing B16-F10 tumors as well as in mice bearing C26 colon carcinoma tumors.

Antitumor effects of recombinant antivascular protein ABRaA-VEGF121 combined with IL-12 gene therapy.

Development and neoplastic progression strongly rely on tumor microenvironment cells. Various kinds of cells that form such tumor milieu play substantial roles in angiogenesis and immunosuppression. Attempts to inhibit tumor vascularization alter tumor milieu and enhance immune response against the tumor. Anticancer therapeutic strategy bringing together antiangiogenic and immunostimulating agents has emerged as a promising approach. We here investigated whether therapy directed against preexisting vessels, combined with an immunomodulatory factor would be equally effective in arresting tumor growth. To this goal, we investigated the effectiveness of ABRaA-vascular endothelial growth factor isoform 121 (VEGF121), an antivascular drug constructed by us. It is a fusion protein composed of VEGF121, and abrin A chain (translation-inhibiting toxin). We used it in combination with interleukin (IL-12) gene therapy and tried to inhibit B16-F10 melanoma tumor growth. ABRaA-VEGF121 is a chimeric recombinant protein capable of destroying tumor vasculature and triggering necrosis in the vicinity of damaged vessels. IL-12 cytokine, in turn, activates both specific and non-specific immune responses. Our results demonstrate that combination of ABRaA-VEGF121 antivascular agent with immunostimulatory cytokine IL-12 indeed inhibits tumor growth more effectively than either agent alone, leading to complete cure of ca. 20 % mice. Post-therapeutic analysis of tumors excised from mice treated with combination therapy showed decreased numbers of blood microvessels in the tumor microenvironment, lowered numbers of regulatory T lymphocytes, as well as showed higher levels of CD4(+) and CD8(+) as compared to control mice. It seems that bringing together antivascular strategy and the action of immunostimulating agents indeed inhibits growth of tumors.