As stubborn as they are common, urinary tract infections (UTIs) have attracted intense scrutiny, but much about them remains hidden. Scientists know that they begin with intracellular bacterial communities (IBCs), and that these communities, which grow inside the cells lining the bladder’s wall, burst free, unleashing an infectious cascade that eventually reaches the bladder’s deeper layers, where quiescent intracellular reservoirs (QIRs) form. However, scientists haven’t been able to observe these events in detail. Consequently, scientists have little idea of how to prevent or eliminate QIRs, which are notorious for promoting antibiotic resistance and causing recurrent infections.

According to scientists at Ecole Polytechnique Fédérale de Lausanne (EPFL), UTI dynamics could be studied more closely if better models of UTI infection were used. Current models include tissue explant cultures and animal models. The EPFL scientists, however, suggest bladder organoids and bladder chips. In fact, the EPFL scientists have already developed these types of models, as reported in two recent papers.

The first paper, which appeared in eLife, is titled, “Dynamic persistence of UPEC intracellular bacterial communities in a human bladder-chip model of urinary tract infection.” The second paper, which appeared in Cell Reports, is titled, “Early invasion of the bladder wall by solitary bacteria protects UPEC from antibiotics and neutrophil swarms in an organoid model.”

“We develop a human bladder-chip model wherein umbrella cells and bladder microvascular endothelial cells are co-cultured under flow in urine and nutritive media respectively, and bladder filling and voiding mimicked mechanically by application and release of linear strain,” the EPFL researchers wrote in the first paper. “Using time-lapse microscopy, we show that rapid recruitment of neutrophils from the vascular channel to sites of infection leads to swarm and neutrophil extracellular trap formation but does not prevent IBC formation.”

Additional observations, the researchers noted, “reinforce a dynamic role for IBCs as harbors of bacterial persistence, with significant consequences for noncompliance with antibiotic regimens.”

In the second paper, the EPFL team describe how they tracked the formation of QIR-like bacteria in a bladder organoid model that recapitulates the stratified uroepithelium within a volume suitable for high-resolution live-cell imaging. The authors of this paper observed that “single bacteria penetrate deeper layers of the organoid wall, where they localize within or between uroepithelial cells.” Then these “solitary” bacteria, which are morphologically distinct from bacteria in IBCs, evade killing by antibiotics and neutrophils.

“We conclude,” the authors wrote, “that bacteria with QIR-like properties may arise at early stages of infection, independent of IBC formation and rupture.”

The lead author of the two studies, Kunal Sharma, a PhD student at EPFL, suggested why current UTI models are inadequate: “Infection dynamics are difficult to capture from static imaging of tissue explants at serial time points. Thus far, in vitro models have not recapitulated bladder architecture with sufficient fidelity to study the time course of these events.”

To overcome this limitation, the group of John McKinney, PhD, professor, at EPFL’s School of Life Sciences developed two complementary bladder models: first, bladder organoids that recreate the 3D stratified architecture of the bladder epithelium; second, a bladder-on-a-chip that incorporates physiological stimuli. (The chip simulates the mechanical effect of bladder filling and voiding, as well as the migration of immune cells through the vasculature to sites of infection.)

“By generating organoids from a mouse with a fluorescent label incorporated within cell membranes, we could use live-cell confocal imaging to identify specific bacterial niches within the organoid with a high spatial resolution,” said Sharma. “By imaging multiple organoids, we managed to identify heterogeneity and diverse outcomes of host-pathogen interactions. This proof-of-concept system has shown promising potential for follow-up studies on bacterial persistence to antibiotics and the dynamics of immune cell responses to infection.”

“Microphysiological models bridge the gap between simple cell culture systems and animal models,” said Vivek V. Thacker, PhD, a senior author on both studies and a scientist in McKinney’s group. “The two models complement each other well and are tailored to study specific aspects of the disease. We hope they will serve as a resource for the wider microbiology community and advance the synergies between the tissue engineering and infectious diseases communities.”

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