Introduction: 

Increasing multi-resistant bacteria and rising numbers of infections during hospitalization require new strategies to test inflammatory processes in complex tissues in vitro. Embryonic stem cells (ES cells) are a powerful model for the development of different cell types of the cellular immune response, e.g. leukocytes, granulocytes, macrophages and lymphocytes. In this study we analyzed the defense- and phagocytic capacity of Embryoid Bodies (EBs) using in vitro inflammatory conditions caused by Escherichia coli (E.coli).

Question: 

Differentiated murine ES cells of the cell line CGR8 exert an anti-inflammatory and anti-bacterial phagocytic capacity. Reactive oxygen species (ROS) play a role by mediating immune defense mechanisms in the EB system.

Methods: 

ES cell culture of the cell line CGR8, conventional- and real-time PCR, Western Blot, ELISA, confocal microscopy as well as physiological ROS measurements.

Results: 

Spontaneously differentiated 18-day-old EBs contain sub-populations of CD68⁺-, F4/80⁺-, CD45⁺- and CD20⁺-cells at certain percentages. Exposure of EBs to E.coli with a defined infection dose of bacterial colony forming units (CFUs) led to a significant time-dependent reduction of CFUs within 10 h, indicating an immune response exerted by EBs. Within the same time window an up-regulation of typical inflammatory cytokines i.e. IL-1beta and TNF-alpha occurred. This was paralleled by increased production of ROS in beating foci of differentiated cardiomyocytes during the inflammation process. Western Blot analysis of infected EBs indicated an up-regulation of CD14 antigen and cytochrome b-245 heavy chain (NOX2). Co-localization strategies with GFP-labeled E.coli revealed time-dependent phagocytosis of bacteria by ES cell-derived leukocytes within differentiated EBs.

Conclusion: 

A primitive immune response potentially co-mediated by ROS in combination with efficient phagocytosis was responsible for the observed antibacterial capacity of differentiated ES cells.