• Kinetics of mRNA delivery and protein translation in dendritic cells using lipid-coated PLGA nanoparticles.

      Yasar, Hanzey; Biehl, Alexander; De Rossi, Chiara; Koch, Marcus; Murgia, Xabi; Loretz, Brigitta; Lehr, Claus-Michael; HIPS, Helmholtz-Institut für pharmazeutische Forschung Saarland, Universitätscampus 8.1, 66123 Saarbrücken, Germany. (2018-09-19)
      Messenger RNA (mRNA) has gained remarkable attention as an alternative to DNA-based therapies in biomedical research. A variety of biodegradable nanoparticles (NPs) has been developed including lipid-based and polymer-based systems for mRNA delivery. However, both systems still lack in achieving an efficient transfection rate and a detailed understanding of the mRNA transgene expression kinetics. Therefore, quantitative analysis of the time-dependent translation behavior would provide a better understanding of mRNA's transient nature and further aid the enhancement of appropriate carriers with the perspective to generate future precision nanomedicines with quick response to treat various diseases. A lipid-polymer hybrid system complexed with mRNA was evaluated regarding its efficiency to transfect dendritic cells (DCs) by simultaneous live cell video imaging of both particle uptake and reporter gene expression. We prepared and optimized NPs consisting of poly (lactid-co-glycolid) (PLGA) coated with the cationic lipid 1, 2-di-O-octadecenyl-3-trimethylammonium propane abbreviated as LPNs. An earlier developed polymer-based delivery system (chitosan-PLGA NPs) served for comparison. Both NPs types were complexed with mRNA-mCherry at various ratios. While cellular uptake and toxicity of either NPs was comparable, LPNs showed a significantly higher transfection efficiency of ~ 80% while chitosan-PLGA NPs revealed only ~ 5%. Further kinetic analysis elicited a start of protein translation after 1 h, with a maximum after 4 h and drop of transgene expression after 48 h post-transfection, in agreement with the transient nature of mRNA. Charge-mediated complexation of mRNA to NPs enables efficient and fast cellular delivery and subsequent protein translation. While cellular uptake of both NP types was comparable, mRNA transgene expression was superior to polymer-based NPs when delivered by lipid-polymer NPs.
    • Local pulmonary drug delivery in the preterm rabbit: feasibility and efficacy of daily intratracheal injections.

      Salaets, Thomas; Gie, André; Jimenez, Julio; Aertgeerts, Margo; Gheysens, Olivier; Vande Velde, Greetje; Koole, Michel; Murgia, Xabi; Casiraghi, Costanza; Ricci, Francesca; et al. (American Physiological Society, 2019-01-24)
      Recent clinical trials in newborns have successfully used surfactant as a drug carrier for an active compound, to minimize systemic exposure. To investigate the translational potential of surfactant-compound mixtures and other local therapeutics, a relevant animal model is required in which intratracheal administration for maximal local deposition is technically possible and well tolerated. Preterm rabbit pups (born at 28 days of gestation) were exposed to either hyperoxia or normoxia and randomized to receive daily intratracheal surfactant, daily intratracheal saline, or no injections for 7 days. At day 7, the overall lung function and morphology were assessed. Efficacy in terms of distribution was assessed by micro-PET-CT on both day 0 and day 7. Lung function as well as parenchymal and vascular structure were altered by hyperoxia, thereby reproducing a phenotype reminiscent of bronchopulmonary dysplasia (BPD). Neither intratracheal surfactant nor saline affected the survival or the hyperoxia-induced BPD phenotype of the pups. Using PET-CT, we demonstrate that 82.5% of the injected radioactive tracer goes and remains in the lungs, with a decrease of only 4% after 150 min. Surfactant and saline can safely and effectively be administered in spontaneously breathing preterm rabbits. The described model and method enable researchers to evaluate intratracheal pharmacological interventions for the treatment of BPD.
    • Lymphatic endothelial cells are a replicative niche for Mycobacterium tuberculosis.

      Lerner, Thomas R; de Souza Carvalho-Wodarz, Cristiane; Repnik, Urska; Russell, Matthew R G; Borel, Sophie; Diedrich, Collin R; Rohde, M; Wainwright, Helen; Collinson, Lucy M; Wilkinson, Robert J; et al. (2016-03-01)
      In extrapulmonary tuberculosis, the most common site of infection is within the lymphatic system, and there is growing recognition that lymphatic endothelial cells (LECs) are involved in immune function. Here, we identified LECs, which line the lymphatic vessels, as a niche for Mycobacterium tuberculosis in the lymph nodes of patients with tuberculosis. In cultured primary human LECs (hLECs), we determined that M. tuberculosis replicates both in the cytosol and within autophagosomes, but the bacteria failed to replicate when the virulence locus RD1 was deleted. Activation by IFN-γ induced a cell-autonomous response in hLECs via autophagy and NO production that restricted M. tuberculosis growth. Thus, depending on the activation status of LECs, autophagy can both promote and restrict replication. Together, these findings reveal a previously unrecognized role for hLECs and autophagy in tuberculosis pathogenesis and suggest that hLECs are a potential niche for M. tuberculosis that allows establishment of persistent infection in lymph nodes.
    • Macrophage uptake of cylindrical microparticles investigated with correlative microscopy.

      Tscheka, Clemens; Hittinger, Marius; Lehr, Claus-Michael; Schneider-Daum, Nicole; Schneider, Marc; Helmholtz Institute for Pharmaceutical Research Saarland (HIPS);Saarland University, Building A4.1, 66123 Saarbruecken, Germany. (2015-09)
      Cylindrical particles offer the opportunity to develop controlled and sustained release systems for the respiratory tract. One reason is that macrophages can phagocyte such particles only from either of the two ends. We investigated the uptake behaviour of murine alveolar macrophages incubated with elongated submicron-structured particles. For that purpose, fluorescent model silica nanoparticles were interconnected with the biocompatible polysaccharide agarose, building up cylindrical particles within the pores of track-etched membranes. In contrast to common approaches we determined the uptake at different time points with scanning electron microscopy, fluorescence microscopy, and the combination of both techniques - correlative microscopy (CLEM). As a consequence, we could securely identify uptake events and observe in detail the engulfment of particles and confirm, that phagocytosis could only be observed from the tips of the cylinders. CLEM allowed a comparison of the uptake measured with different techniques at identical macrophages. Qualitative and quantitative evaluation of this cylindrical particle uptake showed substantial differences between fluorescence microscopy, electron microscopy and the combination of both (CLEM) within 24h.
    • Mastering the Gram-negative bacterial barrier - Chemical approaches to increase bacterial bioavailability of antibiotics.

      Ropponen, Henni-Karoliina; Richter, Robert; Hirsch, Anna K H; Lehr, Claus Michael; HIPS, Helmholtz-Institut für Pharmazeutische Forschung Saarland, Universitätscampus E8.1 66123 Saarbrücken, Germany. (Elsevier, 2021-03-08)
      To win the battle against resistant, pathogenic bacteria, novel classes of anti-infectives and targets are urgently needed. Bacterial uptake, distribution, metabolic and efflux pathways of antibiotics in Gram-negative bacteria determine what we here refer to as bacterial bioavailability. Understanding these mechanisms from a chemical perspective is essential for anti-infective activity and hence, drug discovery as well as drug delivery. A systematic and critical discussion of in bacterio, in vitro and in silico assays reveals that a sufficiently accurate holistic approach is still missing. We expect new findings based on Gram-negative bacterial bioavailability to guide future anti-infective research.
    • Medium throughput breathing human primary cell alveolus-on-chip model.

      Stucki, Janick D; Hobi, Nina; Galimov, Artur; Stucki, Andreas O; Schneider-Daum, Nicole; Lehr, Claus-Michael; Huwer, Hanno; Frick, Manfred; Funke-Chambour, Manuela; Geiser, Thomas; et al. (2018-09-25)
      Organs-on-chips have the potential to improve drug development efficiency and decrease the need for animal testing. For the successful integration of these devices in research and industry, they must reproduce in vivo contexts as closely as possible and be easy to use. Here, we describe a 'breathing' lung-on-chip array equipped with a passive medium exchange mechanism that provide an in vivo-like environment to primary human lung alveolar cells (hAEpCs) and primary lung endothelial cells. This configuration allows the preservation of the phenotype and the function of hAEpCs for several days, the conservation of the epithelial barrier functionality, while enabling simple sampling of the supernatant from the basal chamber. In addition, the chip design increases experimental throughput and enables trans-epithelial electrical resistance measurements using standard equipment. Biological validation revealed that human primary alveolar type I (ATI) and type II-like (ATII) epithelial cells could be successfully cultured on the chip over multiple days. Moreover, the effect of the physiological cyclic strain showed that the epithelial barrier permeability was significantly affected. Long-term co-culture of primary human lung epithelial and endothelial cells demonstrated the potential of the lung-on-chip array for reproducible cell culture under physiological conditions. Thus, this breathing lung-on-chip array, in combination with patients' primary ATI, ATII, and lung endothelial cells, has the potential to become a valuable tool for lung research, drug discovery and precision medicine.
    • Micro-rheological properties of lung homogenates correlate with infection severity in a mouse model of Pseudomonas aeruginosa lung infection.

      Murgia, Xabier; Kany, Andreas M; Herr, Christian; Ho, Duy-Khiet; de Rossi, Chiara; Bals, Robert; Lehr, Claus-Michael; Hirsch, Anna K H; Hartmann, Rolf W; Empting, Martin; et al. (Nature publishing group (NPG), 2020-10-05)
      Lung infections caused by Pseudomonas aeruginosa pose a serious threat to patients suffering from, among others, cystic fibrosis, chronic obstructive pulmonary disease, or bronchiectasis, often leading to life-threatening complications. The establishment of a chronic infection is substantially related to communication between bacteria via quorum-sensing networks. In this study, we aimed to assess the role of quorum-sensing signaling molecules of the Pseudomonas quinolone signal (PQS) and to investigate the viscoelastic properties of lung tissue homogenates of PA-infected mice in a prolonged acute murine infection model. Therefore, a murine infection model was successfully established via intra-tracheal infection with alginate-supplemented Pseudomonas aeruginosa NH57388A. Rheological properties of lung homogenates were analyzed with multiple particle tracking (MPT) and quorum-sensing molecules were quantified with LC-MS/MS. Statistical analysis of bacterial load and quorum-sensing molecules showed a strong correlation between these biomarkers in infected lungs. This was accompanied by noticeable changes in the consistency of lung homogenates with increasing infection severity. Furthermore, viscoelastic properties of the lung homogenates strongly correlated with bacterial load and quorum sensing molecules. Considering the strong correlation between the viscoelasticity of lung homogenates and the aforementioned biomarkers, the viscoelastic properties of infected lungs might serve as reliable new biomarker for the evaluation of the severity of P. aeruginosa infections in murine models.
    • Microstructure of calcium stearate matrix pellets: a function of the drying process.

      Schrank, Simone; Kann, Birthe; Windbergs, Maike; Glasser, Benjamin J; Zimmer, Andreas; Khinast, Johannes; Roblegg, Eva; Institute for Process and Particle Engineering, Graz University of Technology, Graz, Austria; Research Center Pharmaceutical Engineering GmbH, Graz, Austria; Institute of Pharmaceutical Sciences, Department of Pharmaceutical Technology, University of Graz, Graz, Austria. (2013-11)
      Drying is a common pharmaceutical process, whose potential to modify the final drug and/or dosage form properties is often underestimated. In the present study, pellets consisting of the matrix former calcium stearate (CaSt) incorporating the active pharmaceutical ingredient ibuprofen were prepared via wet extrusion and spheronization. Subsequent drying was performed by either desiccation, fluid-bed drying, or lyophilization, and the final pellets were compared with respect to their microstructure. To minimize the effect of solute ibuprofen molecules on the shrinking behavior of the CaSt, low ibuprofen loadings were used, as ibuprofen is soluble in the granulation liquid. Pellet porosity and specific surface area increased during desiccation, fluid-bed drying, and lyophilization. The inlet-air temperature during fluid-bed drying affected the specific surface area, which increased at lower inlet-air temperatures rather than the pellet porosity. The in vitro dissolution profiles were found to be a nonlinear function of the specific surface area. Overall, the microstructure, including porosity, pore size, and specific surface area, of CaSt pellets was a strong function of the drying conditions.
    • A miRNA181a/NFAT5 axis links impaired T cell tolerance induction with autoimmune type 1 diabetes.

      Serr, Isabelle; Scherm, Martin G; Zahm, Adam M; Schug, Jonathan; Flynn, Victoria K; Hippich, Markus; Kälin, Stefanie; Becker, Maike; Achenbach, Peter; Nikolaev, Alexei; et al. (2018-01-03)
      Molecular checkpoints that trigger the onset of islet autoimmunity or progression to human type 1 diabetes (T1D) are incompletely understood. Using T cells from children at an early stage of islet autoimmunity without clinical T1D, we find that a microRNA181a (miRNA181a)-mediated increase in signal strength of stimulation and costimulation links nuclear factor of activated T cells 5 (NFAT5) with impaired tolerance induction and autoimmune activation. We show that enhancing miRNA181a activity increases NFAT5 expression while inhibiting FOXP3+regulatory T cell (Treg) induction in vitro. Accordingly, Treginduction is improved using T cells from NFAT5 knockout (NFAT5ko) animals, whereas altering miRNA181a activity does not affect Treginduction in NFAT5ko T cells. Moreover, high costimulatory signals result in phosphoinositide 3-kinase (PI3K)-mediated NFAT5, which interferes with FoxP3+Treginduction. Blocking miRNA181a or NFAT5 increases Treginduction in murine and humanized models and reduces murine islet autoimmunity in vivo. These findings suggest targeting miRNA181a and/or NFAT5 signaling for the development of innovative personalized medicines to limit islet autoimmunity.
    • miRNA92a targets KLF2 and the phosphatase PTEN signaling to promote human T follicular helper precursors in T1D islet autoimmunity.

      Serr, Isabelle; Fürst, Rainer W; Ott, Verena B; Scherm, Martin G; Nikolaev, Alexei; Gökmen, Füsun; Kälin, Stefanie; Zillmer, Stephanie; Bunk, Melanie; Weigmann, Benno; et al. (2016)
      Aberrant immune activation mediated by T effector cell populations is pivotal in the onset of autoimmunity in type 1 diabetes (T1D). T follicular helper (TFH) cells are essential in the induction of high-affinity antibodies, and their precursor memory compartment circulates in the blood. The role of TFH precursors in the onset of islet autoimmunity and signaling pathways regulating their differentiation is incompletely understood. Here, we provide direct evidence that during onset of islet autoimmunity, the insulin-specific target T-cell population is enriched with a C-X-C chemokine receptor type 5 (CXCR5)+CD4+ TFH precursor phenotype. During onset of islet autoimmunity, the frequency of TFH precursors was controlled by high expression of microRNA92a (miRNA92a). miRNA92a-mediated TFH precursor induction was regulated by phosphatase and tension homolog (PTEN) - phosphoinositol-3-kinase (PI3K) signaling involving PTEN and forkhead box protein O1 (Foxo1), supporting autoantibody generation and triggering the onset of islet autoimmunity. Moreover, we identify Krueppel-like factor 2 (KLF2) as a target of miRNA92a in regulating human TFH precursor induction. Importantly, a miRNA92a antagomir completely blocked induction of human TFH precursors in vitro. More importantly, in vivo application of a miRNA92a antagomir to nonobese diabetic (NOD) mice with ongoing islet autoimmunity resulted in a significant reduction of TFH precursors in peripheral blood and pancreatic lymph nodes. Moreover, miRNA92a antagomir application reduced immune infiltration and activation in pancreata of NOD mice as well as humanized NOD Scid IL2 receptor gamma chain knockout (NSG) human leucocyte antigen (HLA)-DQ8 transgenic animals. We therefore propose that miRNA92a and the PTEN-PI3K-KLF2 signaling network could function as targets for innovative precision medicines to reduce T1D islet autoimmunity.
    • A Model for the Transient Subdiffusive Behavior of Particles in Mucus.

      Ernst, Matthias; John, Thomas; Guenther, Marco; Wagner, Christian; Schaefer, Ulrich F; Lehr, Claus-Michael; HIPS, Helmholtz-Institut für pharmazeutische Forschung Saarland, Universitätscampus 8.1, 66123 Saarbrücken, Germany. (2017-01-10)
      In this study we have applied a model to explain the reported subdiffusion of particles in mucus, based on the measured mean squared displacements (MSD). The model considers Brownian diffusion of particles in a confined geometry, made from permeable membranes. The applied model predicts a normal diffusive behavior at very short and long time lags, as observed in several experiments. In between these timescales, we find that the "subdiffusive" regime is only a transient effect, MSD∝τ
    • Modeling the human skin barrier--towards a better understanding of dermal absorption.

      Hansen, Steffi; Lehr, Claus-Michael; Schaefer, Ulrich F (2013-02)
    • Modulating the Barrier Function of Human Alveolar Epithelial (hAELVi) Cell Monolayers as a Model of Inflammation.

      Metz, Julia Katharina; Wiegand, Birgit; Schnur, Sabrina; Knoth, Katharina; Schneider-Daum, Nicole; Groß, Henrik; Croston, Glenn; Reinheimer, Torsten Michael; Lehr, Claus-Michael; Hittinger, Marius; et al. (SAGE Publications, 2021-01-29)
      The incidence of inflammatory lung diseases such as acute respiratory distress syndrome (ARDS) remains an important problem, particularly in the present time with the Covid-19 pandemic. However, an adequate in vitro test system to monitor the barrier function of the alveolar epithelium during inflammation and for assessing anti-inflammatory drugs is urgently needed. Therefore, we treated human Alveolar Epithelial Lentivirus-immortalised cells (hAELVi cells) with the pro-inflammatory cytokines TNF-α (25 ng/ml) and IFN-γ (30 ng/ml), in the presence or absence of hydrocortisone (HC). While TNF-α and IFN-γ are known to reduce epithelial barrier properties, HC could be expected to protect the barrier function and result in an anti-inflammatory effect. We investigated the impact of anti-inflammatory/inflammatory treatment on transepithelial electrical resistance (TEER) and the apparent permeability coefficient (P app ) of the low permeability marker sodium fluorescein (NaFlu). After incubating hAELVi cells for 48 hours with a combination of TNF-α and IFN-γ, there was a significant decrease in TEER and a significant increase in the P app . The presence of HC maintained the TEER values and barrier properties, so that no significant P app change was observed. By using hAELVi cells to study anti-inflammatory drugs in vitro, the need for animal experiments could be reduced and pulmonary drug development accelerated.
    • Nanocarriers for optimizing the balance between interfollicular permeation and follicular uptake of topically applied clobetasol to minimize adverse effects.

      Mathes, C; Melero, A; Conrad, P; Vogt, T; Rigo, L; Selzer, D; Prado, W A; De Rossi, C; Garrigues, T M; Hansen, S; et al. (2016-02-10)
      The treatment of various hair disorders has become a central focus of good dermatologic patient care as it affects men and women all over the world. For many inflammatory-based scalp diseases, glucocorticoids are an essential part of treatment, even though they are known to cause systemic as well as local adverse effects when applied topically. Therefore, efficient targeting and avoidance of these side effects are of utmost importance. Optimizing the balance between drug release, interfollicular permeation, and follicular uptake may allow minimizing these adverse events and simultaneously improve drug delivery, given that one succeeds in targeting a sustained release formulation to the hair follicle. To test this hypothesis, three types of polymeric nanocarriers (nanospheres, nanocapsules, lipid-core nanocapsules) for the potent glucocorticoid clobetasol propionate (CP) were prepared. They all exhibited a sustained release of drug, as was desired. The particles were formulated as a dispersion and hydrogel and (partially) labeled with Rhodamin B for quantification purposes. Follicular uptake was investigated using the Differential Stripping method and was found highest for nanocapsules in dispersion after application of massage. Moreover, the active ingredient (CP) as well as the nanocarrier (Rhodamin B labeled polymer) recovered in the hair follicle were measured simultaneously, revealing an equivalent uptake of both. In contrast, only negligible amounts of CP could be detected in the hair follicle when applied as free drug in solution or hydrogel, regardless of any massage. Skin permeation experiments using heat-separated human epidermis mounted in Franz Diffusion cells revealed equivalent reduced transdermal permeability for all nanocarriers in comparison to application of the free drug. Combining these results, nanocapsules formulated as an aqueous dispersion and applied by massage appeare to be a good candidate to maximize follicular targeting and minimize drug penetration into the interfollicular epidermis. We conclude that such nanotechnology-based formulations provide a viable strategy for more efficient drug delivery to the hair follicle. Moreover, they present a way to minimize adverse effects of potent glucocorticoids by releasing the drug in a controlled manner and simultaneously decreasing interfollicular permeation, offering an advantage over conventional formulations for inflammatory-based skin/scalp diseases.
    • Nanoencapsulation of a glucocorticoid improves barrier function and anti-inflammatory effect on monolayers of pulmonary epithelial cell lines.

      Rigo, Lucas A; Carvalho-Wodarz, Cristiane S; Pohlmann, Adriana R; Guterres, Silvia S; Schneider-Daum, Nicole; Lehr, Claus Michael; Beck, Ruy C R; Helmholtz-Institut für Pharmazeutische Forschung Saarland [HIPS], Universitätscampus E8.1, 66123 Saarbrücken, Germany. (2017-05-13)
      The anti-inflammatory effect of polymeric deflazacort nanocapsules (NC-DFZ) was investigated, and possible improvement of epithelial barrier function using filter grown monolayers of A549 and Calu-3 using as models was assessed. NC prepared from poly(ε-caprolactone) (PCL) had a mean size around 200 nm, slightly negative zeta potential (∼ - 8 mV), and low polydispersity index (< 0.10). Encapsulation of DFZ had an efficiency of 85%. No cytotoxic effects were observed at particle concentration of 9.85 x 10(11) NC/ml, which was therefore chosen to evaluate the effect of NC-DFZ at 1% (w/v) of PCL and 0.5% (w/v) of DFZ on the epithelial barrier function of Calu-3 monolayers. Nanoencapsulated drug at 0.5% (w/v) increased transepithelial electrical resistance and decrease permeability of the paracellular marker sodium fluorescein, while non-encapsulated DFZ failed to improve these parameters. Moreover, NC-DFZ reduced the lipopolysaccharide (LPS) mediated secretion of the inflammatory marker IL-8. In vitro dissolution testing revealed controlled release of DFZ from nanocapsules, which may explain the improved effect of DFZ on the cells. These data suggest that nanoencapsulation of pulmonary delivered corticosteroids could be advantageous for the treatment of inflammatory conditions, such as asthma and chronic obstructive pulmonary diseases.
    • Nanomedicines for the treatment of inflammatory bowel diseases

      Ali, Hussain; Collnot, Eva-Maria; Windbergs, Maike; Lehr, Claus-Michael (2013-09-20)
    • A New PqsR Inverse Agonist Potentiates Tobramycin Efficacy to Eradicate Pseudomonas aeruginosa Biofilms

      Schütz, Christian; Ho, Duy‐Khiet; Hamed, Mostafa Mohamed; Abdelsamie, Ahmed Saad; Röhrig, Teresa; Herr, Christian; Kany, Andreas Martin; Rox, Katharina; Schmelz, Stefan; Siebenbürger, Lorenz; et al. (Wiley and Sons Inc., 2021-03-18)
      Pseudomonas aeruginosa (PA) infections can be notoriously difficult to treat and are often accompanied by the development of antimicrobial resistance (AMR). Quorum sensing inhibitors (QSI) acting on PqsR (MvfR) – a crucial transcriptional regulator serving major functions in PA virulence – can enhance antibiotic efficacy and eventually prevent the AMR. An integrated drug discovery campaign including design, medicinal chemistry‐driven hit‐to‐lead optimization and in‐depth biological profiling of a new QSI generation is reported. The QSI possess excellent activity in inhibiting pyocyanin production and PqsR reporter‐gene with IC50 values as low as 200 and 11 × 10−9 m, respectively. Drug metabolism and pharmacokinetics (DMPK) as well as safety pharmacology studies especially highlight the promising translational properties of the lead QSI for pulmonary applications. Moreover, target engagement of the lead QSI is shown in a PA mucoid lung infection mouse model. Beyond that, a significant synergistic effect of a QSI‐tobramycin (Tob) combination against PA biofilms using a tailor‐made squalene‐derived nanoparticle (NP) formulation, which enhance the minimum biofilm eradicating concentration (MBEC) of Tob more than 32‐fold is demonstrated. The novel lead QSI and the accompanying NP formulation highlight the potential of adjunctive pathoblocker‐mediated therapy against PA infections opening up avenues for preclinical development.
    • Non-animal models of epithelial barriers (skin, intestine and lung) in research, industrial applications and regulatory toxicology

      Gordon, Sarah; Helmholtz Institute for Pharmaceutical Research Saarland (HIPS), Campus C23, D-66123 Saarbrücken, Germany. (2015)
    • Non-animal models of epithelial barriers (skin, intestine and lung) in research, industrial applications and regulatory toxicology.

      Gordon, Sarah; Daneshian, Mardas; Bouwstra, Joke; Caloni, Francesca; Constant, Samuel; Davies, Donna E; Dandekar, Gudrun; Guzman, Carlos A; Fabian, Eric; Haltner, Eleonore; et al. (2015)
      Models of the outer epithelia of the human body - namely the skin, the intestine and the lung - have found valid applications in both research and industrial settings as attractive alternatives to animal testing. A variety of approaches to model these barriers are currently employed in such fields, ranging from the utilization of ex vivo tissue to reconstructed in vitro models, and further to chip-based technologies, synthetic membrane systems and, of increasing current interest, in silico modeling approaches. An international group of experts in the field of epithelial barriers was convened from academia, industry and regulatory bodies to present both the current state of the art of non-animal models of the skin, intestinal and pulmonary barriers in their various fields of application, and to discuss research-based, industry-driven and regulatory-relevant future directions for both the development of new models and the refinement of existing test methods. Issues of model relevance and preference, validation and standardization, acceptance, and the need for simplicity versus complexity were focal themes of the discussions. The outcomes of workshop presentations and discussions, in relation to both current status and future directions in the utilization and development of epithelial barrier models, are presented by the attending experts in the current report.
    • Novel anti-ulcer phytosomal formulation of ethanol extract of pentaclethra macrophylla stem-bark

      Nnamani, Petra O.; Kenechukwu, Franklin C.; Asogwa, Francis O.; Momoh, Mumuni A.; Lehr, Claus Michael; Attama, Anthony A.; HIPS, Helmholtz-Institut für Pharmazeutische Forschung Saarland, Universitätscampus E8.1 66123 Saarbrücken, Germany. (, Faculty of Pharmacy, University of Benin, Benin City, Nigeria., 2020-01-01)
      Pentaclethra macrophylla, a medicinal plant widely used for ulcer treatment in Nigeria by herbal practitioners, is limited by poor lipid solubility, resulting in poor absorption and bioavailability. Phytosomes, a novel dosage form that are better absorbed and produce better results than conventional herbal extracts, could be employed to enhance its antiulcer activity. The objective of this study was to formulate Pentaclethra macrophylla stem-bark extract as phytosomes by forming complexes with phospholipid and compare the antiulcer activity with omeprazole, a standard anti-ulcer drug. Phytosomal formulations of ethanol extract of Pentaclethra macrophylla stem-bark and Phospholipon® 90G (P90G) (extract:P90G ratios of 1:1, 1:3, 1:5) were prepared following established method. Their physicochemical properties, in vitro drug release in simulated intestinal fluid (SIF, pH=7.4) and simulated gastric fluid (SGF, pH=1.2) and anti-ulcer properties on aspirin-induced ulcer using Wistar rats were determined and compared with omeprazole. Phytosomes with spherical smooth particles with size range 0.106-0.217 µm and good encapsulation efficiencies (range = 67.61-72.8%) were obtained. Drug release increased with time irrespective of phospholipid concentration or dissolution medium. The extract possessed antiulcer activity (23.33%) which was increased to 33.33, 43.33 and 56.67% by formulating it as phytosomal formulations containing extract:P90G ratios of 1:1, 1:3, 1:5, respectively. However, omeprazole and its formulations gave significantly (p<0.05) greater antiulcer activity when compared with both the ethanol extract and phytosomes. Pentaclethra macrophylla stem-bark possessed antiulcer activity, which was improved via phytosomal formulation. This would serve as potential safer and cheaper alternative therapeutics for ulcer given the side-effects associated with omeprazole.