Novel Three-Dimensional Organoid Model for Evaluation of the Interaction of Uropathogenic Escherichia coli with Terminally Differentiated Human Urothelial Cells

Abstract

Human bladder 5637 cells cultivated under microgravity conditions formed organoids that displayed characteristics of in vivo tissue-specific differentiation. Uropathogenic Escherichia coli (UPEC) strain CP9 colonized and penetrated the organoids and induced ␣-hemolysin-mediated exfoliation of uroepithelial cells. We propose these uro-organoids as models that simulate the interactions between UPEC and terminally differentiated human urothelium.

Inactivation of Host Akt/Protein Kinase B Signaling by Bacterial Pore-forming Toxins

“…For example, the HlyA-positive UPEC isolate CP9 was recently shown to induce rapid activation of proapoptotic executioner caspases in neutrophils much more efficiently than a HlyA-negative mutant (Russo et al, 2005). Relative to an isogenic hlyA mutant, the wild-type CP9 strain also induces more robust exfoliation of bladder epithelial cells (Smith et al, 2006), a phenomenon that seems to involve activation of apoptotic pathways (Mulvey et al, 1998;Schaeffer et al, 2004;Klumpp et al, 2006).…”

“…At high concentrations, HlyA is able to lyse erythrocytes and nucleated host cells, a process that may enable extraintestinal pathogens like UPEC to better cross mucosal barriers, damage effector immune cells, and gain enhanced access to host nutrients and iron stores (Cavalieri et al, 1984;Johnson, 1991). At sublytic concentrations, HlyA can induce the apoptosis of target host cells (like neutrophils, T lymphocytes, and renal cells) and promote the exfoliation of bladder epithelial cells, although the signaling pathways triggered by HlyA during such events have not been elucidated (Jonas et al, 1993;Weinrauch and Zychlinsky, 1999;Russo et al, 2005;Chen et al, 2006;Smith et al, 2006).…”

Culturing and Applications of Rotating Wall Vessel Bioreactor Derived 3D Epithelial Cell Models

“…microvilli), and cellular polarity. The majority of the data and examples presented here are using vaginal epithelial cells grown in the RWV system, however the RWV system also has been used to culture other epithelial cell types including small and large intestine, lung, bladder, liver, and tonsil epithelial cells to form 3-D organotypic models 1,[7][8][9][10][11][12][13]18 . Additionally, this system has been used to culture cell types other than epithelial cells, and currently the development of complex, multicellular organotypic culture models are underway 2,18 .…”

“…The progression from a monolayer of epithelial cells to a fully differentiated 3-D aggregate varies based on cell type 1,[7][8][9][10][11][12][13] . Periodic sampling from the bioreactor allows for monitoring of epithelial aggregate formation, cellular differentiation markers and viability ( Figure 1D).…”

“…The microenvironment provided by the bioreactor allows the cells to form three-dimensional (3-D) aggregates displaying in vivo-like characteristics often not observed under standard 2-D culture conditions ( Figure 1D). These characteristics include tight junctions, mucus production, apical/basal orientation, in vivo protein localization, and additional epithelial celltype specific properties.The progression from a monolayer of epithelial cells to a fully differentiated 3-D aggregate varies based on cell type 1,[7][8][9][10][11][12][13] . Periodic sampling from the bioreactor allows for monitoring of epithelial aggregate formation, cellular differentiation markers and viability ( Figure 1D).…”