operating-biocontainment-procedures-facility-for-laboratory-mice-with

Operating and Biocontainment Procedures of a Facility for Laboratory Mice with a Natural Microbiome: Immunophenotyping Procedure

The use of laboratory mice with a natural microbiome, such as &#34;Wildling mice&#34;, offers a promising research tool for both basic and applied science ...

University Hospital Erlangen, Friedrich-Alexander-Universität Erlangen-Nürnberg (FAU)

Medical Center - University of Freiburg

Synucleinopathies are a group of neurodegenerative disorders, classically characterized by the accumulation of aggregated alpha synuclein (aSyn) in the central nervous system. Parkinson's disease (PD) and multiple system atrophy (MSA) are the two prominent members of this family. Current treatment options mainly focus on the motor symptoms of these diseases. However, non-motor symptoms, including gastrointestinal (GI) symptoms, have recently gained particular attention, as they are frequently associated with synucleinopathies and often arise before motor symptoms. The gut-origin hypothesis has been proposed based on evidence of an ascending spreading pattern of aggregated aSyn from the gut to the brain, as well as the comorbidity of inflammatory bowel disease and synucleinopathies. Recent advances have shed light on the mechanisms underlying the progression of synucleinopathies along the gut-brain axis. Given the rapidly expanding pace of research in the field, this review presents a summary of the latest findings on the gut-to-brain spreading of pathology and potential pathology-reinforcing mediators in synucleinopathies. Here, we focus on 1) gut-to-brain communication pathways, including neuronal pathways and blood circulation, and 2) potential molecular signalling mediators, including bacterial amyloid proteins, microbiota dysbiosis-induced alterations in gut metabolites, as well as host-derived effectors, including gut-derived peptides and hormones. We highlight the clinical relevance and implications of these molecular mediators and their possible mechanisms in synucleinopathies. Moreover, we discuss their potential as diagnostic markers in distinguishing the subtypes of synucleinopathies and other neurodegenerative diseases, as well as for developing novel individualized therapeutic options for synucleinopathies.

Gut-to-brain spreading of pathology in synucleinopathies: A focus on molecular signalling mediators.

Allergic disorders are caused by a combination of hereditary and environmental factors. The hygiene hypothesis postulates that early-life microbial exposures impede the development of subsequent allergic disease. Recently developed "wildling" mice are genetically identical to standard laboratory specific pathogen-free (SPF) mice but are housed under seminatural conditions and have rich microbial exposures from birth. Thus, by comparing conventional SPF mice with wildlings, we can uncouple the impact of lifelong microbial exposures from genetic factors on the allergic immune response. We found that wildlings developed larger populations of antigen-experienced T cells than conventional SPF mice, which included interleukin-10-producing CD4 T cells specific for commensal  strains and allergy-promoting T helper 2 (T2) cells. In models of airway exposure to house dust mite (HDM), recombinant interleukin-33, or , wildlings developed strong allergic inflammation, characterized by eosinophil recruitment, goblet cell metaplasia, and antigen-specific immunoglobulin G1 (IgG1) and IgE responses. Wildlings developed robust de novo T2 cell responses to incoming allergens, whereas preexisting T2 cells could also be recruited into the allergic immune response in a cytokine-driven and TCR-independent fashion. Thus, wildling mice, which experience diverse and lifelong microbial exposures, were not protected from developing pathological allergic immune responses. Instead, wildlings mounted robust allergic responses to incoming allergens, shedding new light on the hygiene hypothesis.

Laboratory mice with a wild microbiota generate strong allergic immune responses.

Natural killer (NK) cells are present in the circulation and can also be found residing in tissues, and these populations exhibit distinct developmental requirements and are thought to differ in terms of ontogeny. Here, we investigate whether circulating conventional NK (cNK) cells can develop into long-lived tissue-resident NK (trNK) cells following acute infections. We found that viral and bacterial infections of the skin triggered the recruitment of cNK cells and their differentiation into Tcf1CD69 trNK cells that share transcriptional similarity with CD56TCF1 NK cells in human tissues. Skin trNK cells arose from interferon (IFN)-γ-producing effector cells and required restricted expression of the transcriptional regulator Blimp1 to optimize Tcf1-dependent trNK cell formation. Upon secondary infection, trNK cells rapidly gained effector function and mediated an accelerated NK cell response. Thus, cNK cells redistribute and permanently position at sites of previous infection via a mechanism promoting tissue residency that is distinct from Hobit-dependent developmental paths of NK cells and ILC1 seeding tissues during ontogeny.

Circulating NK cells establish tissue residency upon acute infection of skin and mediate accelerated effector responses to secondary infection.

Mice with a natural microbiome are a promising research model for basic and applied science because of their closer resemblance to the human superorganism compared to mice born and raised under stringent hygiene conditions. Consequently, biomedical therapies developed and tested in “Wildling mice” hold great potential for successful translation into clinical applications. Over the past four years, scientists, veterinarians and institutional officials at Charité – Universitätsmedizin Berlin, supported by the University Hospital Erlangen, have designed a facility for Wildling mice and developed a conceptual framework for safe and ethical preclinical research involving mice with a natural microbiome.

A facility for laboratory mice with a natural microbiome at Charité - Universitätsmedizin Berlin.

Primary hemophagocytic lymphohistiocytosis (HLH) is a life-threatening hyperinflammatory syndrome caused by inborn errors of cytotoxicity. Patients with biallelic PRF1 null mutations (encoding perforin) usually develop excessive immune cell activation, hypercytokinemia, and life-threatening immunopathology in the first 6 months of life, often without an apparent infectious trigger. In contrast, perforin-deficient (PKO) mice only develop HLH after systemic infection with lymphocytic choriomeningitis virus (LCMV). We hypothesized that restricted microbe-immune cell interactions due to specific pathogen-free (SPF) housing might explain the need for this specific viral trigger in PKO mice. To investigate the influence of a "wild" microbiome in PKO mice, we fostered PKO newborns with Wildling microbiota ('PKO-Wildlings') and monitored them for signs of HLH. PKO-Wildlings survived long-term without spontaneous disease. Also, systemic infection with vaccinia virus did not reach the threshold of immune activation required to trigger HLH in PKO-Wildlings. Interestingly, after infection with LCMV, PKO-Wildlings developed an altered HLH pattern. This included lower IFN-γ serum levels along with improved IFN-γ-driven anemia, but more elevated levels of IL-17 and increased liver inflammation compared with PKO-SPF mice. Thus, wild microbiota alone is not sufficient to trigger HLH in PKO mice, but host-microbe interactions shape inflammatory cytokine patterns, thereby influencing manifestations of HLH immunopathology.

Article	Year
Gut-to-brain spreading of pathology in synucleinopathies: A focus on molecular signalling mediators. Behavioural brain research\| PubMed ID: 37423320	2023
Laboratory mice with a wild microbiota generate strong allergic immune responses. Science immunology\| PubMed ID: 37774008	2023
Circulating NK cells establish tissue residency upon acute infection of skin and mediate accelerated effector responses to secondary infection. Immunity\| PubMed ID: 38157853	2024
A facility for laboratory mice with a natural microbiome at Charité - Universitätsmedizin Berlin. Lab animal\| PubMed ID: 39533119	2024
The Microbiome Modifies Manifestations of Hemophagocytic Lymphohistiocytosis in Perforin-Deficient Mice. European journal of immunology\| PubMed ID: 39548906	2024

Stephan P. Rosshart

Biography

JoVE Journal Publications

Other Publications