From animal models to gut-on-chip: the challenging journey to capture inter-individual variability in chronic digestive disorders.

Chronic digestive disorders are of increasing incidence worldwide with expensive treatments and no available cure. Available therapeutic schemes mainly rely on symptom relief, with large degrees of variability in patients' response to such treatments, underlining the need for new therapeutic strategies. There are strong indications that the gut microbiota's contribution seems to be a key modulator of disease activity and patients' treatment responses. Hence, efforts have been devoted to understanding host-microbe interactions and the mechanisms underpinning such variability. Animal models, being the gold standard, provide valuable mechanistic insights into host-microbe interactions. However, they are not exempt from limitations prompting the development of alternative methods. Emerging microfluidic technologies and gut-on-chip models were shown to mirror the main features of gut physiology and disease state, reflect microbiota modification, and include functional readouts for studying host responses. In this commentary, we discuss the relevance of animal models in understanding host-microbe interactions and how gut-on-chip technology holds promises for addressing patient variability in responses to chronic digestive disease treatment.

Lacticaseibacillus casei AMBR2 modulates the epithelial barrier function and immune response in a donor-derived nasal microbiota manner.

Live biotherapeutic products (LBP) are emerging as alternative treatment strategies for chronic rhinosinusitis. The selection of interesting candidate LBPs often involves model systems that do not include the polymicrobial background (i.e. the host microbiota) in which they will be introduced. Here, we performed a screening in a simplified model system of upper respiratory epithelium to assess the effect of nasal microbiota composition on the ability to attach and grow of a potential LBP, Lacticaseibacillus casei AMBR2, in this polymicrobial background. After selecting the most permissive and least permissive donor, L. casei AMBR2 colonisation in their respective polymicrobial backgrounds was assessed in more physiologically relevant model systems. We examined cytotoxicity, epithelial barrier function, and cytokine secretion, as well as bacterial cell density and phenotypic diversity in differentiated airway epithelium based models, with or without macrophage-like cells. L. casei AMBR2 could colonize in the presence of both selected donor microbiota and increased epithelial barrier resistance in presence of donor-derived nasal bacteria, as well as anti-inflammatory cytokine secretion in the presence of macrophage-like cells. This study highlights the potential of L. casei AMBR2 as LBP and the necessity to employ physiologically relevant model systems to investigate host-microbe interaction in LBP research.

Lactobacilli Have a Niche in the Human Nose.

Although an increasing number of beneficial microbiome members are characterized for the human gut and vagina, beneficial microbes are underexplored for the human upper respiratory tract (URT). In this study, we demonstrate that taxa from the beneficial Lactobacillus genus complex are more prevalent in the healthy URT than in patients with chronic rhinosinusitis (CRS). Several URT-specific isolates are cultured, characterized, and further explored for their genetic and functional properties related to adaptation to the URT. Catalase genes are found in the identified lactobacilli, which is a unique feature within this mostly facultative anaerobic genus. Moreover, one of our isolated strains, Lactobacillus casei AMBR2, contains fimbriae that enable strong adherence to URT epithelium, inhibit the growth and virulence of several URT pathogens, and successfully colonize nasal epithelium of healthy volunteers. This study thus demonstrates that specific lactobacilli are adapted to the URT and could have a beneficial keystone function in this habitat.

Dual and Triple Epithelial Coculture Model Systems with Donor-Derived Microbiota and THP-1 Macrophages To Mimic Host-Microbe Interactions in the Human Sinonasal Cavities.

The epithelium of the human sinonasal cavities is colonized by a diverse microbial community, modulating epithelial development and immune priming and playing a role in respiratory disease. Here, we present a novel in vitro approach enabling a 3-day coculture of differentiated Calu-3 respiratory epithelial cells with a donor-derived bacterial community, a commensal species (Lactobacillus sakei), or a pathobiont (Staphylococcus aureus). We also assessed how the incorporation of macrophage-like cells could have a steering effect on both epithelial cells and the microbial community. Inoculation of donor-derived microbiota in our experimental setup did not pose cytotoxic stress on the epithelial cell layers, as demonstrated by unaltered cytokine and lactate dehydrogenase release compared to a sterile control. Epithelial integrity of the differentiated Calu-3 cells was maintained as well, with no differences in transepithelial electrical resistance observed between coculture with donor-derived microbiota and a sterile control. Transition of nasal microbiota from in vivo to in vitro conditions maintained phylogenetic richness, and yet a decrease in phylogenetic and phenotypic diversity was noted. Additional inclusion and coculture of THP-1-derived macrophages did not alter phylogenetic diversity, and yet donor-independent shifts toward higher Moraxella and Mycoplasma abundance were observed, while phenotypic diversity was also increased. Our results demonstrate that coculture of differentiated airway epithelial cells with a healthy donor-derived nasal community is a viable strategy to mimic host-microbe interactions in the human upper respiratory tract. Importantly, including an immune component allowed us to study host-microbe interactions in the upper respiratory tract more in depth.IMPORTANCE Despite the relevance of the resident microbiota in sinonasal health and disease and the need for cross talk between immune and epithelial cells in the upper respiratory tract, these parameters have not been combined in a single in vitro model system. We have developed a coculture system of differentiated respiratory epithelium and natural nasal microbiota and incorporated an immune component. As indicated by absence of cytotoxicity and stable cytokine profiles and epithelial integrity, nasal microbiota from human origin appeared to be well tolerated by host cells, while microbial community composition remained representative for that of the human (sino)nasal cavity. Importantly, the introduction of macrophage-like cells enabled us to obtain a differential readout from the epithelial cells dependent on the donor microbial background to which the cells were exposed. We conclude that both model systems offer the means to investigate host-microbe interactions in the upper respiratory tract in a more representative way.

Modelling upper respiratory tract diseases: getting grips on host-microbe interactions in chronic rhinosinusitis using in vitro technologies.

Chronic rhinosinusitis (CRS) is a chronic inflammation of the mucosa of the nose and paranasal sinuses affecting approximately 11% of the adult population in Europe. Inadequate immune responses, as well as a dysbiosis of the sinonasal microbiota, have been put forward as aetiological factors of the disease. However, despite the prevalence of this disease, there is no consensus on the aetiology and mechanisms of pathogenesis of CRS. Further research requires in vitro models mimicking the healthy and diseased host environment along with the sinonasal microbiota. This review aims to provide an overview of CRS model systems and proposes in vitro modelling strategies to conduct mechanistic research in an ecological framework on the sinonasal microbiota and its interactions with the host in health and CRS.