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Schmid Lab

Pulmonary Aerosol Delivery

Our objective is to translate new concepts of aerosol research, biotechnology and physics into technological advances facilitating the development of novel inhaled drugs for the treatment of lung diseases.

Our objective is to translate new concepts of aerosol research, biotechnology and physics into technological advances facilitating the development of novel inhaled drugs for the treatment of lung diseases.

Our Key Questions

  • How can we improve aerosolized drug delivery to the lung in both preclinical (cells, animal models) and clinical (human) settings?
  • How can we determine both dose and bioactivity of the pulmonary drug dose in real-time?
  • Can physiologically more realistic cell-based drug screening methods improve the predictive power of these tests for the efficacy of novel drugs in clinical settings?
  • How can inhaled drugs be targeted specifically to the site of interest (e.g. bronchial or alveolar region or specific cell types depending on disease type)? 

Our Main Projects

We aim to design and validate systems for aerosolized drug delivery to pulmonary cells (in vitro) and to mouse models of disease (in vivo). Furthermore, methods for (real-time) in vivo monitoring of both delivered and bioactive pulmonary drug dose are developed. Currently, mainly optical imaging methods based on bioluminescence and fluorescence are examined.

Recently, the ALICE-CLOUD technology was introduced which combines ease-of-handling with rapid, efficient and accurate delivery of aerosolized drugs to cells cultured at air-liquid interface conditions. This technology may pave the way for screening of inhalable drugs under physiologically more relevant and thus clinically more predictive conditions than the currently used submerged cell culture systems.

Scientists at Schmid Lab

Dr. Ali Doryab

Acting project coordinator and head of the subproject of KLIMA

Anna Fuchs

Biologisch Technische Assistentin

Dr. Pramod Kumar

Postdoctoral Fellow

Vitus Prokosch

BSc Student

Andreas Schröppel

IT, Engineer

Penghang Chen

PhD Student

Publications

2022, Scientific Article in NanoImpact

An inter-laboratory effort to harmonize the cell-delivered in vitro dose of aerosolized materials.

Air-liquid interface (ALI) lung cell models cultured on permeable transwell inserts are increasingly used for respiratory hazard assessment requiring controlled aerosolization and deposition of any material on ALI cells. The approach presented herein aimed to assess the transwell insert-delivered dose of aerosolized materials using the VITROCELL® Cloud12 system, a commercially available aerosol-cell exposure system. An inter-laboratory comparison study was conducted with seven European partners having different levels of experience with the VITROCELL® Cloud12. A standard operating procedure (SOP) was developed and applied by all partners for aerosolized delivery of materials, i.e., a water-soluble molecular substance (fluorescence-spiked salt) and two poorly soluble particles, crystalline silica quartz (DQ12) and titanium dioxide nanoparticles (TiO2 NM-105). The material dose delivered to transwell inserts was quantified with spectrofluorometry (fluorescein) and with the quartz crystal microbalance (QCM) integrated in the VITROCELL® Cloud12 system. The shape and agglomeration state of the deposited particles were confirmed with transmission electron microscopy (TEM). Inter-laboratory comparison of the device-specific performance was conducted in two steps, first for molecular substances (fluorescein-spiked salt), and then for particles. Device- and/or handling-specific differences in aerosol deposition of VITROCELL® Cloud12 systems were characterized in terms of the so-called deposition factor (DF), which allows for prediction of the transwell insert-deposited particle dose from the particle concentration in the aerosolized suspension. Albeit DF varied between the different labs from 0.39 to 0.87 (mean (coefficient of variation (CV)): 0.64 (28%)), the QCM of each VITROCELL® Cloud 12 system accurately measured the respective transwell insert-deposited dose. Aerosolized delivery of DQ12 and TiO2 NM-105 particles showed good linearity (R2 > 0.95) between particle concentration of the aerosolized suspension and QCM-determined insert-delivered particle dose. The VITROCELL® Cloud 12 performance for DQ12 particles was identical to that for fluorescein-spiked salt, i.e., the ratio of measured and salt-predicted dose was 1.0 (29%). On the other hand, a ca. 2-fold reduced dose was observed for TiO2 NM-105 (0.54 (41%)), which was likely due to partial retention of TiO2 NM-105 agglomerates in the vibrating mesh nebulizer of the VITROCELL® Cloud12. This inter-laboratory comparison demonstrates that the QCM integrated in the VITROCELL® Cloud 12 is a reliable tool for dosimetry, which accounts for potential variations of the transwell insert-delivered dose due to device-, handling- and/or material-specific effects. With the detailed protocol presented herein, all seven partner laboratories were able to demonstrate dose-controlled aerosolization of material suspensions using the VITROCELL® Cloud12 exposure system at dose levels relevant for observing in vitro hazard responses. This is an important step towards regulatory approved implementation of ALI lung cell cultures for in vitro hazard assessment of aerosolized materials.

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2022, Scientific Article in European Journal of Pharmaceutical Sciences

Towards a gold standard functional readout to characterize In Vitro lung barriers.

The development of biomimetic in vitro lung models as an alternative to animal studies is urgent to improve the predictability of the pharmacokinetics of potential new drugs. For pharmacokinetics studies, advanced in vitro lung models such as lung-chips should mimic a functional air-blood barrier. Unlike in vivo conditions, stem/primary cells and cell lines do not necessarily form a functional and tight barrier when cultured in vitro. Here, we explore the two gold standard techniques for monitoring barrier integrity: transepithelial electrical resistance (TEER) and permeability. We discuss the advantages and limitations of these methods, provide recommendations for methodological improvements, and we elude on possible future directions.

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2022, Scientific Article in Biomedicines

Bioactive cell-derived ECM scaffold forms a unique cellular microenvironment for lung tissue engineering.

Chronic lung diseases are one of the leading causes of death worldwide. Lung transplantation is currently the only causal therapeutic for lung diseases, which is restricted to end-stage disease and limited by low access to donor lungs. Lung tissue engineering (LTE) is a promising approach to regenerating a replacement for at least a part of the damaged lung tissue. Currently, lung regeneration is limited to a simplified local level (e.g., alveolar-capillary barrier) due to the sophisticated and complex structure and physiology of the lung. Here, we introduce an extracellular matrix (ECM)-integrated scaffold using a cellularization-decellularization-recellularization technique. This ECM-integrated scaffold was developed on our artificial co-polymeric BETA (biphasic elastic thin for air-liquid interface cell culture conditions) scaffold, which were initially populated with human lung fibroblasts (IMR90 cell line), as the main generator of ECM proteins. Due to the interconnected porous structure of the thin (<5 µm) BETA scaffold, the cells can grow on and infiltrate into the scaffold and deposit their own ECM. After a mild decellularization procedure, the ECM proteins remained on the scaffold, which now closely mimicked the cellular microenvironment of pulmonary cells more realistically than the plain artificial scaffolds. We assessed several decellularization methods and found that 20 mM NH4OH and 0.1% Triton X100 with subsequent DNase treatment completely removed the fibroblasts (from the first cellularization) and maintains collagen I and IV as the key ECM proteins on the scaffold. We also showed the repopulation of the primary fibroblast from human (without chronic lung disease (non-CLD) donors) and human bronchial epithelial (16HBE14o-) cells on the ECM-integrated BETA scaffold. With this technique, we developed a biomimetic scaffold that can mimic both the physico-mechanical properties and the native microenvironment of the lung ECM. The results indicate the potential of the presented bioactive scaffold for LTE application.

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2022, Review in NanoImpact

Analytical and toxicological aspects of nanomaterials in different product groups: Challenges and opportunities.

The widespread integration of engineered nanomaterials into consumer and industrial products creates new challenges and requires innovative approaches in terms of design, testing, reliability, and safety of nanotechnology. The aim of this review article is to give an overview of different product groups in which nanomaterials are present and outline their safety aspects for consumers. Here, release of nanomaterials and related analytical challenges and solutions as well as toxicological considerations, such as dose-metrics, are discussed. Additionally, the utilization of engineered nanomaterials as pharmaceuticals or nutraceuticals to deliver and release cargo molecules is covered. Furthermore, critical pathways for human exposure to nanomaterials, namely inhalation and ingestion, are discussed in the context of risk assessment. Analysis of NMs in food, innovative medicine or food contact materials is discussed. Specific focus is on the presence and release of nanomaterials, including whether nanomaterials can migrate from polymer nanocomposites used in food contact materials. With regard to the toxicology and toxicokinetics of nanomaterials, aspects of dose metrics of inhalation toxicity as well as ingestion toxicology and comparison between in vitro and in vivo conclusions are considered. The definition of dose descriptors to be applied in toxicological testing is emphasized. In relation to potential exposure from different products, opportunities arising from the use of advanced analytical techniques in more unique scenarios such as release of nanomaterials from medical devices such as orthopedic implants are addressed. Alongside higher product performance and complexity, further challenges regarding material characterization and safety, as well as acceptance by the general public are expected.

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2022, Scientific Article in Advanced materials

Real-time measurement of cell mechanics as a clinically relevant readout of an in vitro lung fibrosis model established on a bioinspired basement membrane.

Lung fibrosis, as one of the major post-COVID complications, is a progressive and ultimately fatal disease without a cure. Here, we introduce an organ- and disease-specific in vitro mini-lung fibrosis model equipped with non-invasive real-time monitoring of cell mechanics as a functional readout. To establish an intricate multi-culture model under physiologic conditions, we developed a biomimetic ultrathin basement (BETA) membrane (<1 μm) with unique properties, including biocompatibility, permeability, and high elasticity (<10 kPa) for cell culturing under air-liquid interface (ALI) and cyclic mechanical stretch conditions. The human-based triple co-culture fibrosis model, which includes epithelial and endothelial cell lines combined with primary fibroblasts from idiopathic pulmonary fibrosis (IPF) patients established on the BETA membrane, is integrated into a millifluidic bioreactor system (CIVIC) with dose-controlled aerosolized drug delivery, mimicking inhalation therapy. We show the real-time measurement of cell/tissue stiffness (and compliance) as a clinical biomarker of the progression/attenuation of fibrosis upon drug treatment, which was confirmed for inhaled Nintedanib -an FDA-approved anti-fibrosis drug. The mini-lung fibrosis model allows the combined longitudinal testing of pharmacodynamics and pharmacokinetics of drugs, which is expected to enhance the predictive capacity of preclinical models and hence facilitate the development of approved therapies for lung fibrosis. This article is protected by copyright. All rights reserved.

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2022, Scientific Article in Nanomaterials

Aerosol–cell exposure system applied to semi-adherent cells for aerosolization of lung surfactant and nanoparticles followed by high quality RNA extraction.

Nanoparticle toxicity assessments have moved closer to physiological conditions while trying to avoid the use of animal models. An example of new in vitro exposure techniques developed is the exposure of cultured cells at the air–liquid interface (ALI), particularly in the case of respiratory airways. While the commercially available VITROCELL® Cloud System has been applied for the delivery of aerosolized substances to adherent cells under ALI conditions, it has not yet been tested on lung surfactant and semi-adherent cells such as alveolar macrophages, which are playing a pivotal role in the nanoparticle-induced immune response. Objectives: In this work, we developed a comprehensive methodology for coating semi-adherent lung cells cultured at the ALI with aerosolized surfactant and subsequent dose-controlled exposure to nanoparticles (NPs). This protocol is optimized for subsequent transcriptomic studies. Methods: Semi-adherent rat alveolar macrophages NR8383 were grown at the ALI and coated with lung surfactant through nebulization using the VITROCELL® Cloud 6 System before being exposed to TiO2 NM105 NPs. After NP exposures, RNA was extracted and its quantity and quality were measured. Results: The VITROCELL® Cloud system allowed for uniform and ultrathin coating of cells with aerosolized surfactant mimicking physiological conditions in the lung. While nebulization of 57 µL of 30 mg/mL TiO2 and 114 µL of 15 mg/mL TiO2 nanoparticles yielded identical cell delivered dose, the reproducibility of dose as well as the quality of RNA extracted were better for 114 µL.

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Contact

Dr. Otmar Schmid

Team Leader