Anti-Cancer Prodrug Cyclophosphamide Exerts Thrombogenic Effects on Human Venous Endothelial Cells Independent of CYP450 Activation-Relevance to Thrombosis.

Cancer patients are at a very high risk of serious thrombotic events, often fatal. The causes discussed include the detachment of thrombogenic particles from tumor cells or the adverse effects of chemotherapeutic agents. Cytostatic agents can either act directly on their targets or, in the case of a prodrug approach, require metabolization for their action. Cyclophosphamide (CPA) is a widely used cytostatic drug that requires prodrug activation by cytochrome P450 enzymes (CYP) in the liver. We hypothesize that CPA could induce thrombosis in one of the following ways: (1) damage to endothelial cells (EC) after intra-endothelial metabolization; or (2) direct damage to EC without prior metabolization. In order to investigate this hypothesis, endothelial cells (HUVEC) were treated with CPA in clinically relevant concentrations for up to 8 days. HUVECs were chosen as a model representing the first place of action after intravenous CPA administration. No expression of CYP2B6, CYP3A4, CYP2C9 and CYP2C19 was found in HUVEC, but a weak expression of CYP2C18 was observed. CPA treatment of HUVEC induced DNA damage and a reduced formation of an EC monolayer and caused an increased release of prostacyclin (PGI2) and thromboxane (TXA) associated with a shift of the PGI2/TXA balance to a prothrombotic state. In an in vivo scenario, such processes would promote the risk of thrombus formation.

In Vitro Cytotoxic Activity of an Aqueous Alkali Extract of the Tinder Conk Mushroom, Fomes fomentarius (Agaricomycetes), on Murine Fibroblasts, Human Colorectal Adenocarcinoma, and Cutaneous Melanoma Cells.

Bioactive complexes of medicinal mushrooms have become attractive as complementary anticancer remedies. Our in vitro study focused on the cytotoxicity of the polyphenol-reach and beta-glucan-containing aqueous alkali extract from Fomes fomentarius fruiting bodies (FFE) using murine fibroblasts (L929), human colon adenocarcinoma cells (Caco-2), and cutaneous melanoma cells (COLO-818). Dose-dependent FFE cytotoxicity with an half maximal inhibitory concentration of 0.44 mg/mL was observed for L929 cells upon analysis of the total number of adherent cells, degree of cell viability, cell morphology, and mitochondrial metabolic activity. Cytotoxic effects on cancer cells tested using cell impedance were dependent on FFE concentration, type of cells, and their density. As a routine in vitro model for predicting human intestinal absorption, Caco-2 cells did not react on FFE, which can indirectly support its safety for the human intestinal epithelium. Melanoma cells were affected in a dose-dependent manner, even at low FFE concentrations (0.01-0.05 mg/mL). The confluent cell layer, which resembles a fully formed tumor, was much more resistant than the incompletely formed, subconfluent cell layer, simulating tumor formation. FFE applied topically could be a promising candidate to prevent melanoma development in its early stages.

Impact of Multivalence and Self-Assembly in the Design of Polymeric Antimicrobial Peptide Mimics.

Antimicrobial resistance is an increasingly serious challenge for public health and could result in dramatic negative consequences for the health care sector during the next decades. To solve this problem, antibacterial materials that are unsusceptible toward the development of bacterial resistance are a promising branch of research. In this work, a new type of polymeric antimicrobial peptide mimic featuring a bottlebrush architecture is developed, using a combination of reversible addition-fragmentation chain transfer (RAFT) polymerization and ring-opening metathesis polymerization (ROMP). This approach enables multivalent presentation of antimicrobial subunits resulting in improved bioactivity and an increased hemocompatibility, boosting the selectivity of these materials for bacterial cells. Direct probing of membrane integrity of treated bacteria revealed highly potent membrane disruption caused by bottlebrush copolymers. Multivalent bottlebrush copolymers clearly outperformed their linear equivalents regarding bioactivity and selectivity. The effect of segmentation of cationic and hydrophobic subunits within bottle brushes was probed using heterograft copolymers. These materials were found to self-assemble under physiological conditions, which reduced their antibacterial activity, highlighting the importance of precise structural control for such applications. To the best of our knowledge, this is the first example to demonstrate the positive impact of multivalence, generated by a bottlebrush topology in polymeric antimicrobial peptide mimics, making these polymers a highly promising material platform for the design of new bactericidal systems.

Epigenetics-Shedding Light on the Path Ahead for Material Sciences.

The harmonious regulation of bodily function is a necessity for healthy individuals. Looking from the viewpoint of material sciences, one can only marvel at the cellular factories, their renewal, and the overall control of messaging and control of responses. As aging progresses and/or pathologies arise, clinicians may be forced to look for replacement of organs/tissues with medical devices. Since all devices are tailored, a detailed understanding of developmental processes, including aberrant processes leading to pathologies, is crucial to provide clinicians with a suitable device. Although research in the field of epigenetics has produced effective therapeutics and diagnostic markers, our currently fragmented understanding of epigenetic processes as they relate to material development is inherently limited, with logical implications for the success of medical procedures. Here, we illustrate how material sciences for clinical applications, critically depend on all aspects of biomedical sciences, including the field of epigenetics.

Functional Glyco-Nanogels for Multivalent Interaction with Lectins.

Interactions between glycans and proteins have tremendous impact in biomolecular interactions. They are important for cell-cell interactions, proliferation and much more. Here, we emphasize the glycan-mediated interactions between pathogens and host cells. Pseudomonas aeruginosa, responsible for a huge number of nosocomial infections, is especially the focus when it comes to glycan-derivatives as pathoblockers. We present a microwave assisted protecting group free synthesis of glycomonomers based on lactose, melibiose and fucose. The monomers were polymerized in a precipitation polymerization in the presence of NiPAm to form crosslinked glyco-nanogels. The influence of reaction parameters like crosslinker type or stabilizer amount was investigated. The gels were characterized in lectin binding studies using model lectins and showed size and composition-dependent inhibition of lectin binding. Due to multivalent presentation of glycans in the gel, the inhibition was clearly stronger than with unmodified saccharides, which was compared after determination of the glycan loading. First studies with Pseudomonas aeruginosa revealed a surprising influence on the secretion of virulence factors. Functional glycogels may be in the future potent alternatives or adjuvants for antibiotic treatment of infections based on glycan interactions between host and pathogen.

Facilitating a More Efficient Commercial Review Process for Pediatric Drugs and Biologics.

Over the past two decades, the biopharmaceutical industry has seen unprecedented expansion and innovation in concert with significant technological advancements. While the industry has experienced marked growth, the regulatory system in the United States still operates at a capacity much lower than the influx of new drug and biologic candidates. As a result, it has become standard for months or even years of waiting for commercial approval by the U.S. Food and Drug Administration. These regulatory delays have generated a system that stifles growth and innovation due to the exorbitant costs associated with awaiting approval from the nation's sole regulatory agency. The recent re-emergence of diseases that impact pediatric demographics represents one particularly acute reason for developing a regulatory system that facilitates a more efficient commercial review process. Herein, we present a range of initiatives that could represent early steps toward alleviating the delays in approving life-saving therapeutics.

Risk Evaluation Requires an Independent Mind.

Biomedical research pertaining to pathologies observed in adolescents can involve areas where patients can expect no immediate benefits. Here, Congress stipulates that local review boards are restricted to approving procedures posing no greater than minimal risk (45 CFR 46.404). An evaluation of risk embraces the current state of the art with regard to the safety and efficacy of procedures. A tendency of biomedical scholars to cite highly cited documents can introduce a bias in an argumentation in favor or against a given recommendation in the context that bias in citations can be correlated with an imprudent use of funds for research. We use choice examples to highlight the necessity of approaching any scholarly task with an independent mind.

An Abraded Surface of Doxorubicin-Loaded Surfactant-Containing Drug Delivery Systems Effectively Reduces the Survival of Carcinoma Cells.

An effective antitumor remedy is yet to be developed. All previous approaches for a targeted delivery of anticancer medicine have relied on trial and error. The goal of this study was to use structural insights gained from the study of delivery systems and malignant cells to provide for a systematic approach to the development of next-generation drugs. We used doxorubicin (Dox) liposomal formulations. We assayed for cytotoxicity via the electrical current exclusion method. Dialysis of the samples yielded information about their drug release profiles. Information about the surface of the delivery systems was obtained through synchrotron small-angle X-ray scattering (SAXS) measurements. SAXS measurements revealed that Dox-loading yielded an abraded surface of our Dox liposomal formulation containing soybean oil, which also correlated with an effective reduction of the survival of carcinoma cells. Furthermore, a dialysis assay revealed that a higher burst of Dox was released from soybean oil-containing preparations within the first five hours. We conclude from our results that an abraded surface of Dox-loaded drug delivery system increases their efficacy. The apparent match between surface geometry of drug delivery systems and target cells is suggested as a steppingstone for refined development of drug delivery systems. This is the first study to provide a systematic approach to developing next-generation drug carrier systems using structural insights to guide the development of next-generation drug delivery systems with increased efficacy and reduced side effects.

Nanomaterials-Tools, Technology and Methodology of Nanotechnology Based Biomedical Systems for Diagnostics and Therapy.

Nanomedicine helps to fight diseases at the cellular and molecular level by utilizing unique properties of quasi-atomic particles at a size scale ranging from 1 to 100 nm. Nanoparticles are used in therapeutic and diagnostic approaches, referred to as theranostics. The aim of this review is to illustrate the application of general principles of nanotechnology to select examples of life sciences, molecular medicine and bio-assays. Critical aspects relating to those examples are discussed.

Bone marrow cells and CD117-positive haematopoietic stem cells promote corneal wound healing.

PURPOSE: The present study was conducted to evaluate the therapeutic effects of topically applied bone marrow (BM) cells and CD117-positive haematopoietic stem (CD117(+)) cells on alkali-induced corneal ulcers. METHODS: Bone marrow cells and CD117(+) cells were isolated from syngenic mice and labelled with an intracellular cell tracer. Defined corneal wounds were produced in 89 eyes of syngenic mice and allowed to partially heal in vivo for 6 hr. The alkali-burned eyes were enucleated 6 hr postinjury and randomly divided into three groups. Control group (33 eyes) was incubated with medium only. The treatment groups received either BM cells (30 eyes) or CD117(+) cells (26 eyes) suspended in medium. Re-epithelialization process of corneal defects was qualitatively and quantitatively assessed and statistically analysed. The corneas were examined by histological and immunohistochemical methods. RESULTS: We found that the re-epithelialization of corneal wounds in both treatment groups was significantly accelerated as compared to the control group. During the follow-up period (85 hr), the corneal transparency was comparable in all groups. Morphological investigations of corneas from control and treatment group showed no evident differences in the phenotype of the regenerated epithelium. Additionally, corneas in the treatment groups were devoid of donor-derived BM cells and CD117(+) cells, respectively. CONCLUSIONS: This study provides evidence that topical application of BM cells or CD117(+) cells can be used to reconstruct corneal surfaces. Because neither BM cells nor CD117(+) cells were integrated into the corneal epithelium, we suggest that soluble factors could be responsible for the positive effect of BM cells and CD117(+) cells on corneal wound healing.

Oligophrenin-1 (Ophn1) is expressed in mouse retinal vessels.

The Rho GTPase activating protein (RhoGAP) Oligophrenin 1 (Ophn1) regulates numerous members of the Rho family that are involved in neuronal morphogenesis of the central and peripheral nervous system. In the present study we investigated the spatial and temporal expression of Ophn1 in the mouse eye. The expression of Ophn1 was analysed on both mRNA and protein level. To identify the Ophn1 transcripts, adult retina and cerebrum (positive control) of postnatal day (P) 158 was subjected to reverse transcription polymerase chain reaction (RT-PCR) and sequencing of the amplified cDNA. The Ophn1 protein was analyzed in adult retina by Western blotting and in developing eyes at embryonic day (E) 12, E14, E16, E18, P0, P3, P7, P14 and P158 by immunohistochemistry. Ophn1 transcripts were detected in adult retina by RT-PCR and confirmed by sequencing. Western blot analysis revealed the expression of Ophn1 protein in the adult retina. Immunohistochemical examination of developing eyes localized the protein to retinal vasculature with an onset of Ophn1 expression from P14 onwards. The specific expression pattern suggests that Ophn1 could have a physiological role in the retinal vasculatures. At P14, the vessel development in the retina is widely completed, implying that Ophn1 has either a function during adulthood or for the generation of the intermediate plexus during the late vessel development of the retina.

Unusual pattern of glistening formation on a 3-piece hydrophobic acrylic intraocular lens.

A 3-piece hydrophobic acrylic intraocular lens (IOL) was explanted from the left eye of a 68-year-old patient 26 days after implantation because of glistenings that impaired fundus visualization. The effect of the glistenings on the patient's visual function was unknown because of retinal issues. The fluid-filled vacuoles within the IOL optic were different from those described in clinical and experimental studies. They were initially observed 4 days after surgery and were very large. The glistening formation was reproduced experimentally by immersing the explanted IOL in balanced salt solution at 37 degrees C and analyzing the IOL at room temperature for up to 2 weeks. An IOL of the same design was used as a control. Although both IOLs were subjected to the same laboratory conditions, gross and light microscopic analyses confirmed that the pattern of the glistening formation differed between them. Analyses of the explanted IOL and the control IOL under differential scanning calorimetry, as well as by attenuated total reflection Fourier transform infrared spectroscopy, revealed slight differences between the IOLs.

Interaction forces between cellulose microspheres and ultrathin cellulose films monitored by colloidal probe microscopy-effect of wet strength agents.

Colloidal probe microscopy was employed to study forces between cellulose surfaces upon addition of a series of cationic copolymers in aqueous solution, as model compounds for wet strength agents. The content of quaternary ammonium groups and primary amines was systematically varied in the cationic polymers, to distinguish between the importance of electrostatical and H-bonding effects. Cellulose microspheres were glued at the apex of tipless microfabricated cantilevers and used as colloidal probes. Ultra thin cellulose films and cellulose fibres were employed as model surfaces. The cellulose films of a thickness of about 5 nm were spin-coated from cellulose solution onto silicon substrates. The root-mean-square-roughness (RMS) was 0.3-0.8 nm. The cationic model polymers were compared to Servamine, a polymer employed as standard wet strength resin in papermaking industries. Force versus separation measurements showed a detailed picture of adhesion and contact breaking. Relatively strong adhesion of the order of 0.3 mJ/m(2) was observed with Servamine within a range of approximately 10 nm. At larger distances weak bond breaking and elastic chain pulling were identified. When approaching the surface one to two small jump-in's possibly related to strong binding of Servamine and subsequent attraction could be found in the case of Servamine. In contrast, all the model copolymers showed only a weak adhesion of 8-30 micro/m(2), i.e., an order of magnitude less than that of Servamine and subsequent elastic rupture domains. The contour length, persistence length and characteristic rupture distances were calculated by means of applying the WLC model. Measurements against cellulose fibres obtained from the production process proved the relevance of the model systems.