Further, the functional double layer is composed of an upper mucus layer
and a lower semi-permeable polyamide membrane and has been conceived to potentially serve multiple objectives: i) to provide a mucosal area which can be colonized by the gut bacteria; ii) to allow the bilateral transport of low molecular weight metabolites; iii) to allow the transport of oxygen from the lower to the upper side of the mucosal layer in order to create microaerophilic MK0683 conditions at the bottom HSP inhibitor clinical trial of the growing biofilm; and iv) to protect the host’s cells from direct exposure to a complex microbial community and its toxic effects. In this study the HMI module has been used in i) short-term experiments to characterize different technical parameters and ii) in a long-term experiment, coupled to a SHIME system (as described in the related paragraph), to
assess the possibility to follow up the host’s response to a specific treatment up to 48 h. Figure 1 Scheme of the HMI module for long-term studies of the host-microbiota interaction in the GIT. A polyamide semipermeable membrane and a mucus layer form a double functional layer that separates the luminal compartment (upper one) from the lower compartment containing enterocyte cell lines. The HMI module allows to study the bacterial adhesion under relevant shear forces and microaerophilic conditions. It allows the reciprocal exchange of signals GSK1904529A in vitro and metabolites between compartments and it allows the exposure of cell lines to a complex microbial community, representative for the human colon, for up to 48 h. Characterization
of the technical parameters (shear stress, mucus thickness and oxygen diffusion) Urease In the first part of the work the newly developed model has been characterized with respect to a number of technological parameters in order to validate it with in vivo data. For these experiments the HMI module has been used as a separate unit (i.e. not coupled with a SHIME). The optimal shape of the HMI module was designed to provide a homogeneous fluid shear distribution on the surface of the mucus layer under different shear forces relevant for the GIT (Additional file 1: Figure S1). Analysis by Confocal Laser Scanning Microscopy (CLSM) of the mucus layer on a vertical section and the evaluation of the mucus thickness showed that 95% (i.e. residual thickness) of the original mucus layer (200 μm) was still present after 5 hours at medium shear stress (10 dynes/cm2) and 45% after high shear stress (20 dynes/cm2) (data not shown). Shear forces in the gut are a key factor in shaping the adhering community, in affecting bacterial gene expression and physiology, and can alternatively favor or disfavor the adhesion of specific strains [30–32]. Physiological levels of shear stress found in the intestinal epithelium during peristalsis may range between 35 and 0.02 dynes/cm2[25, 33, 34].