Role of a natural microbiome in the course of cystic fibrosis-like lung disease in mice

Principle Investigator

Scientific interest within the context of the graduate college:

The microbiota, i.e. the entirety of microbes in an organism, plays a crucial role in the maintenance of health and the development of diseases. A diverse microbiota not only produces essential metabolites and supports the uptake of nutrients, but also is important for the development of a functional immune system.^1,2 Mice that are born and kept under laboratory, specific pathogen-free (SPF) conditions have only a reduced microbiome and thus, possess an immature immune system, resembling more likely a newborn than an adult human.³ This may have contributed to failure in translating results from previous preclinical studies into the clinics. However, mice are an invaluable tool in preclinical research, as their genome is well studied and various genetic models exist that are necessary to understand molecular mechanisms of disease pathogenesis. To improve translatability, Wildling mice have been developed that harbor a natural microbiome, comparable to mice in the wild, and consequently, have an immune system being more similar to that of an adult human.⁴ As part of the “Wildling Mice in Health and Disease (HeaD)”-Consortium at Charité we will validate the Wildling model in our preclinical mouse model of muco-obstructive lung disease, called βENaC-transgenic (βENaC-Tg) mice.⁵ These mice overexpress the β-subunit of the epithelial Na⁺ channel (ENaC) specifically in the airways to produce airway surface dehydration, resulting in the development of airway mucus plugging, dysbiosis, chronic inflammation and structural lung damage, i.e. key features of patients with cystic fibrosis (CF) and other muco-obstructive lung diseases.^5-7 Although this mouse model mimics airway surface dehydration as a key disease mechanism in CF lung disease, important clinical aspects like spontaneous Pseudomonas aeruginosa infections are not recapitulated in SPF mice. Hence, we hypothesize that a natural airway microbiome will critically affect the infection status in βENaC-Tg mice including the development of chronic Pseudomonas infection that is a key determinant of lung disease severity and lung function decline. Such a novel model of chronic Pseudomonas infection would enable in vivo testing of novel therapeutic strategies to eradicate Pseudomonas from the airways and thus, target a high unmet need in patients with CF.

Project description:

Introduction: The overall aim of this research project is to evaluate whether βENaC-Tg mice with CF-like lung disease that harbour a “wild”, natural airway microbiome will develop chronic Pseudomonas infections, which is currently the missing human phenotype in SPF βENaC-Tg mice, and consequently, will be of major advance for preclinical testing of novel anti-infective strategies. For this purpose, we will generate Wildling βENaC-Tg mice and the development of the airway microbiota throughout distinct developmental stages (neonatal, juvenile, adult mice) will be carefully followed by culture-dependent and -independent (16S rRNA gene sequencing) techniques of bronchoalveolar lavage (BAL) and lung homogenates (Aim 1). Further, distinct lung immune cell populations will be characterized (Aim 2), concomitant with the assessment of inflammation markers and airway mucus obstruction to study thoroughly the impact of a natural microbiota on the progression of chronic lung disease (Aim 3). Finally, transcriptomic analyses will be conducted to obtain insights into molecular mechanisms underlying the pathogenesis of muco-obstructive lung disease in presence of a natural airway microbiota. Through a direct comparison with SPF animals, these analyses will comprehensively evaluate whether Wildling βENaC-Tg mice will mimic more closely disease mechanisms in patients with muco-obstructive lung diseases, including the development of chronic Pseudomonas infection, which is a key determinant of lung disease outcomes in patients with CF, and would thus authorize in vivo testing of novel anti-infective therapeutic strategies. In addition to basic molecular biology laboratory work, this experimental MD thesis also applies state-of-the-art multi-color flow cytometry and sequencing technologies.

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