Nanoparticle clearance from the airways : development and testing of a new in vitro model to investigate mucociliary clearance of aerosol particles

The current challenge in pulmonary drug delivery is to overcome and to control drug clearance from the lung. Here, the interaction with the mucociliary lung clearance plays a key role for any drug formulation technology. By today many open questions on mucociliary clearance (MC) have to be answered but suffer from limited experimental options. Considering these facts, the central part of this work was to develop a new in vitro test model to investigate the complex mechanism of MC more detailed. In a next step the new model was employed to study the influence of different factors on mucociliary particle clearance. From the overall results it can be concluded that: 1) A new embryonic chicken trachea-based in vitro model to study MC was successfully developed and characterized. 2) The model exhibits a stability and sensitivity for environmental as well as chemical influences reflecting the in vivo situation. 3) Size and ζ-potential (ZP) seem to be minor critical factors for mucociliary particle clearance, which could be shown for different polymeric particles covering a broad size and ZP range. 4) The chemical surface structure of nanoparticles was found to be a most relevant factor for MC, offering promising possibilities to achieve faster or slower particle transport. In summary, this model may help to clarify open questions on lung clearance, strongly limiting the benefit of pulmonary drug delivery, and to realize the \u273R\u27; concept in animal testing: Reduce, Refine, and Replace.Die kontrollierte Arzneistoffelimination aus der Lunge stellt für die Inhalative Therapie eine große Herausforderung dar. Von zentraler Bedeutung, doch bis heute mit vielen offenen Fragen verbunden, ist hierbei die mukoziliäre Clearance (MC). Ziel dieser Arbeit war daher die Entwicklung eines neuen in vitro Modells für die gezielte Untersuchung mukoziliärer Transportprozesse sowie die anschließende Testung verschiedener Einflussfaktoren auf die MC von Partikeln. Aufgrund der Ergebnisse dieser Arbeit können folgende Schlüsse gezogen werden: 1) Ein auf embryonaler Hühnertrachea basierendes in vitro Testsystem zur Untersuchung der MC wurde erfolgreich entwickelt und charakterisiert. 2) Das Modell weist gegenüber äußeren Einflüssen und Arzneistoffen eine mit der in vivo Situation vergleichbare Stabilität sowie Sensitivität auf. 3) Partikelgröße sowie ζ-Potenzial scheinen für die MC von untergeordneter Bedeutung zu sein, was für verschiedene Polymerpartikel unterschiedlicher Ladung und Größe gezeigt werden konnte. 4) Die chemische Struktur und Oberflächenbeschaffenheit nanoskaliger Partikel scheint ein wichtiger Parameter für die MC zu sein und stellt damit einen Erfolg versprechenden Ansatz für die kontrollierte Arzneistoffelimination aus der Lunge dar. Zusammenfassend besteht mit diesem neuen in vitro Modell künftig die Option, mukoziliäre Transportprozesse unter Beachtung des 3R-Konzepts für Tierexperimente (Reduce, Refine, and Replace) gezielt zu untersuchen und aufzuklären

Towards understanding crossover for Cartesian Genetic Programming

Unlike in traditional Genetic Programming, Cartesian Genetic Programming (CGP) does not commonly feature a recombination/crossover operator, although recombination plays an important role in other evolutionary techniques, including Genetic Programming from which CGP originates. Instead, CGP mainly depends on mutation and selection operators in their evolutionary search. To this day, it is still unclear as to why CGP’s performance does not generally improve with the addition of crossover. In this work, we argue that CGP’s positional bias might be a reason for this phenomenon. This bias describes a skewed distribution of active and inactive nodes, which might lead to destructive behaviour of standard recombination operators. We provide a first assessment with preliminary results. No final conclusion to this hypothesis can be drawn yet, as more thorough evaluations must be done first. However, our first results show promising trends and may lay the foundationf or future work

3.5 kW coherently combined ultrafast fiber laser

An ultrafast laser based on the coherent beam combination of four ytterbium-doped step-index fiber amplifiers is presented. The system delivers an average power of 3.5 kW and a pulse duration of 430 fs at an 80 MHz repetition rate. The beam quality is excellent (M2<1.24·1.10), and the relative intensity noise is as low as 1% in the frequency span from 1 Hz to 1 MHz. The system is turn-key operable, as it features an automated spatial and temporal alignment of the interferometric amplification channels

Enhancement of the Zero Phonon Line emission from a Single NV-Center in a Nanodiamond via Coupling to a Photonic Crystal Cavity

Using a nanomanipulation technique a nanodiamond with a single nitrogen vacancy center is placed directly on the surface of a gallium phosphide photonic crystal cavity. A Purcell-enhancement of the fluorescence emission at the zero phonon line (ZPL) by a factor of 12.1 is observed. The ZPL coupling is a first crucial step towards future diamond-based integrated quantum optical devices

Elemental Bioimaging of Sheep Bone and Articular Cartilage After Single Application of Gadolinium-Based Contrast Agents

BACKGROUND: Gadolinium-based contrast agents (GBCAs) are applied to enhance magnetic resonance imaging. Gadolinium (Gd), a rare earth metal, is used in a chelated form when administered as GBCA to patients. There is an ongoing scientific debate about the clinical significance of Gd retention in tissues after administration of GBCAs. It is known that bone serves as Gd reservoir, but only sparse information on localization of Gd in bone is available. PURPOSE: The aim of this study was to compare Gd tissue concentration and spatial distribution in femoral epiphysis and diaphysis 10 weeks after single-dose injection of linear and macrocyclic GBCAs in a large animal model. MATERIALS AND METHODS: In this prospective animal study, Swiss-Alpine sheep (n = 36; age range, 4-10 years) received a single injection (0.1 mmol/kg) of macrocyclic (gadobutrol, gadoteridol, and gadoterate meglumine), linear (gadodiamide and gadobenate dimeglumine) GBCAs, or saline. Ten weeks after injection, sheep were killed, and femur heads and shafts were harvested. Gadolinium spatial distribution was determined in 1 sample of each treatment group by laser ablation-inductively coupled plasma-mass spectrometry. All bone specimens were analyzed histopathologically. RESULTS: Injection of GBCAs in female Swiss-Alpine sheep (n = 36) resulted in Gd localization at the endosteal and periosteal surface and in a subset of GBCAs additionally at the cement lines and the bone cartilage junction. No histopathological alterations were observed in the investigated tissue specimens. CONCLUSIONS: Ten weeks after single injection of a clinically relevant dose in adult sheep, both linear species of GBCA resulted in considerably higher accumulation than macrocyclic GBCAs. Gadolinium deposits were restricted to distinct bone and cartilage compartments, such as in bone linings, cement lines, and bone cartilage junctions. Tissue histology remained unaffected

Differential Glucose Uptake in Quadriceps and Other Leg Muscles During One-Legged Dynamic Submaximal Knee-Extension Exercise

One-legged dynamic knee-extension exercise (DKE) is a widely used model to study the local cardiovascular and metabolic responses to exercise of the quadriceps muscles. In this study, we explored the extent to which different muscles of the quadriceps are activated during exercise using positron emission tomography (PET) determined uptake of [18F]-fluoro-deoxy-glucose (GU) during DKE. Five healthy male subjects performed DKE at 25 W for 35 min and both the contracting and contralateral resting leg were scanned with PET from mid-thigh and distally. On average, exercise GU was the highest in the vastus intermedius (VI) and lowest in the vastus lateralis (VL; VI vs VL, p < 0.05), whereas the coefficient of variation was highest in VL (VL vs VI, p < 0.05). Coefficient of variation between the mean values of the four quadriceps femoris (QF) muscles in the exercising leg was 35 ± 9%. Compared to mean GU in QF (=100%), GU was on average 73% in VL, 84% in rectus femoris, 115% in vastus medialis, and 142% in VI. Variable activation of hamstring muscles and muscles of the lower leg was also observed. These results show that GU of different muscles of quadriceps muscle group as well as between individuals vary greatly during DKE, and suggests that muscle activity is not equal between quadriceps muscles in this exercise model. Furthermore, posterior thigh muscles and lower leg muscles are more active than hitherto thought even during this moderate exercise intensity

Cryo-EM structural studies of the agonist complexed human TRPV4 ion-channel reveals novel structural rearrangements resulting in an open-conformation

The human transient receptor potential vanilloid 4 (hTRPV4) ion channel plays a critical role in a variety of biological processes. Whilst the activation of hTRPV4 gating properties has been reported for a broad spectrum of stimuli, including synthetic 4α-phorbols, the molecular basis of the activation is poorly understood. Here we report the novel cryo-EM structure of the hTRPV4 determined in the presence of the archetypical phorbol acid agonist, 4α-PDD. Complementary mutagenesis experiments support the EM-identified binding site as well as allowing rationalization of disruptive mutants located outside of the 4α-PDD binding site. This work represents the first structural information of hTRPV4 in a ligand-induced open conformation. Together, our data reveal the underlying molecular mechanisms resulting in the opening of the central pore and ion-channel activation and provide a structural template for designing inhibitors targeting the open-state conformation of hTRPV4

Patients with Acute Limb Ischemia Might Benefit from Endovascular Therapy—A 17-Year Retrospective Single-Center Series of 985 Patients

Acute lower limb ischemia (ALI) is a common vascular emergency, requiring urgent revascularization by open or endovascular means. The aim of this retrospective study was to evaluate patient demographics, treatment and periprocedural variables affecting the outcome in ALI patients in a consecutive cohort in a tertiary referral center. Primary outcome events (POE) were 30-day (safety) and 180-day (efficacy) combined mortality and major amputation rates, respectively. Secondary outcomes were perioperative medical and surgical leg-related complications and the 5-year combined mortality and major amputation rate. Statistical analysis used descriptive and uni- and multivariable Cox regression analysis. In 985 patients (71 ± 9 years, 56% men) from 2004 to 2020, the 30-day and 180-day combined mortality and major amputation rates were 15% and 27%. Upon multivariable analysis, older age (30 d: aHR 1.17; 180 d: 1.27) and advanced Rutherford ischemia stage significantly worsened the safety and efficacy POE (30 d: TASC IIa aHR 3.29, TASC IIb aHR 3.93, TASC III aHR 7.79; 180 d: TASC IIa aHR 1.97, TASC IIb aHR 2.43, TASC III aHR 4.2), while endovascular treatment was associated with significant improved POE after 30 days (aHR 0.35) and 180 days (aHR 0.39), respectively. Looking at five consecutive patient quintiles, a significant increase in endovascular procedures especially in the last quintile could be observed (17.5% to 39.5%, p < 0.001). Simultaneously, the re-occlusion rate as well as the number of patients with any previous revascularization increased. In conclusion, despite a slightly increasing early re-occlusion rate, endovascular treatment might, if possible, be favorable in ALI treatment

Prognostic Impact of Bcl-2 Depends on Tumor Histology and Expression of MALAT-1 lncRNA in Non–Small-Cell Lung Cancer

IntroductionApoptosis is a crucial pathway in tumor growth and metastatic development. Apoptotic proteins regulate the underlying molecular cascades and are thought to modulate the tumor response to chemotherapy and radiation. However, the prognostic value of the expression of apoptosis regulators in localized non–small-cell lung cancer (NSCLC) is still unclear.MethodsWe investigated the protein expression of apoptosis regulators Bcl-2, Bcl-xl, Mcl-1, and pp32/PHAPI, and the expression of the lncRNA MALAT-1 in tumor samples from 383 NSCLC patients (median age: 65.6 years; 77.5% male; paraffin-embedded tissue microarrays). For statistical analysis correlation tests, Log rank tests and Cox proportional hazard models were applied.ResultsTumor histology was significantly associated with the expression of Bcl-2, Bcl-xl and Mcl-1 (all p < 0.001). Among the tested apoptotic markers only Bcl-2 demonstrated prognostic impact (hazard ratio = 0.64, p = 0.012). For NSCLC patients with non-adenocarcinoma histology, Bcl-2 expression was associated with increased overall survival (p = 0.036). Besides tumor histology, prognostic impact of Bcl-2 was also found to depend on MALAT-1 lncRNA expression. Gene expression analysis of A549 adenocarcinoma cells with differential MALAT-1 lncRNA expression demonstrated an influence on the expression of Bcl-2 and its interacting proteins.ConclusionsBcl-2 expression was specifically associated with superior prognosis in localized NSCLC. An interaction of Bcl-2 with MALAT-1 lncRNA expression was revealed, which merits further investigation for risk prediction in resectable NSCLC patients