von Garnier/Blank Group
Our research team focuses on interactions between specifically manufactured biomedical nanoparticles and antigen presenting cells of the respiratory tract. While we are routinely using flow cytometry as an ideal approach to quantify particle-cell interactions as well as phenotypic and functional changes in immune cells upon particle exposure, there is a constant and growing need for novel approaches to detect and track nanoparticles in situ inside the tissue or even inside single cells in order to determine the fate of promising nanocarriers or to clarify possible pathways of immunomodulation.
Assessment of immunomodulatory effects in COPD following exposure with multi-walled carbon nanotubes in an occupational setting
Chronic obstructive pulmonary disease (COPD) is a major cause of morbidity and mortality worldwide. Carbon nanotubes (CNTs) represent a promising engineered nanomaterial but may represent a significant additional health risk, specifically in individuals with COPD. We isolate primary human nasal epithelial cells and human bronchial cells from healthy subjects or subjects suffering from COPD and the cells grow in vitro at the air-liquid interface (ALI). We expose the cells to multi-walled CNTs (MWCNTs) directly nebulise at the air-liquid surface of cultures. We check differentiation, integrity, pro-inflammatory status and viability of epithelial cultures using light microscopy, LDH analysis and ELISA. The ciliary beating frequency measure before and after exposure to MWCNTs.
Pulmonary immune responses to bio-mimetic antigen carriers for novel therapeutic approaches to treating allergic asthma
Asthma represents a considerable burden for health care systems globally, with rates increasing at an alarming pace in many countries. Despite an allergic component present in more than half of the asthmatic individuals, current therapy provides an anti-inflammatory, but no causal treatment. The respiratory tract with its ease of access, vast surface area and dense DC network represents an attractive target organ for a direct immune-modulatory approach with inhaled antigen carriers that are endowed with appropriate adjuvant properties.
Therefore we formulate virosomes and liposomes in our lab and surface-label them with ovalbumin (OVA), our protein of interest and a fluorescent dye (Atto647N) for detection by flow cytometry (FACS) and laser scanning microscopy (LSM). Virosomes furthermore contain hemagglutinin (HA) from the A/Brisbane strain. Both virosomes and liposomes are characterized for particle size, purity as well as for HA and OVA content.
In an In Vitro approach we isolate murine bone marrow and differentiate DCs (BMDCs) which are exposed to virosomes and liposomes overnight. Phenotype, particle uptake and antigen processing are determined by FACS and LSM.
Further In Vivo approaches (mouse) will show the fate of inhaled bio-mimetic antigen carriers in different respiratory tract compartments. Uptake by antigen-presenting cells and trafficking to regional draining lymph nodes with subsequent T cell activation will be monitored. Furthermore a mouse model of allergic asthmatic airway disease will be used to assess the potential of virosomes and liposomes as novel therapeutic agent for immune-modulation. Data will be analysed by FACS and LSM mostly.
The lung microbiome in COPD
Chronic Obstructive Pulmonary Disease (COPD) is one of the top 5 causes of death worldwide and its incidence is on the rise. Recent literature suggests that the microbiome in the airways of COPD patients is significantly altered and may influence the course of disease. Also, one of the key cell types involved in the pathogenesis, and possibly control of COPD, is the macrophage. Macrophages have a remarkable ability to respond to environment cues and differentiate into a range of phenotypes that can vary from anti-inflammatory or inflammatory to wound repair. In our research we aim to understand how the lung microbiome is altered in COPD patients, compared to non-COPD patients, and how it differs between distinct regions of the lung. Further, we rein the process to analyze how this microbiota influences local macrophage differentiation and effector function to gain insight into the role of the human microbiome in health and disease.
Specifically modified Gold Nanoparticles as immune modulators
The respiratory tract represents an ideal target organ for inhaled immuno-modulatory therapeutic approaches, given the vast surface where interactions between resident antigen-presenting cells (APCs), such as dendritic cells (DCs) and macrophages, and novel nanoparticle (NP)-based carriers may occur. However, our knowledge regarding immune responses to such nanocarriers remains poorly understood. NH2-PVA and COOH-PVA functionalized gold NPs (AuNPs) or their corresponding polymers were instilled intranasally in BALB/c mice, and their uptake by APC populations in broncho-alveolar lavage fluid (BAL), trachea, lung parenchyma and lung draining lymph nodes (DLNs) was assessed after 24h by flow cytometry. OVA-specific CD4+ T cell proliferation in non-DLNs and lung DLNs was furthermore examined following exposure to particles. Although uptake also occurred in DCs, the majority of both NH2-PVA and COOH-PVA AuNPs were found inside macrophages. In BAL and lung parenchyma, macrophages and DC subpopulations preferentially took up NH2-PVA AuNPs and NH2-PVA polymers compared to their COOH counterparts. Uptake of both types of particles or polymers generally increased the expression of CD40 and CD86 in analyzed APC populations. We further observed that NH2-PVA AuNPs induced enhanced proliferation of OVA-specific CD4+ T cell proliferation in lung DLNs compared to PBS or COOH-PVA AuNPs.