Identification of TLT2 as an engulfment receptor for apoptotic cells.

Clearance of apoptotic cells (efferocytosis) is critical to the homeostasis of the immune system by restraining inflammation and autoimmune response to intracellular Ags released from dying cells. TLRs-mediated innate immunity plays an important role in pathogen clearance and in regulation of the adaptive immune response. However, the regulation of efferocytosis by activation of TLRs has not been well characterized. In this study, we found that activation of TLR3 or TLR9, but not of TLR2, enhances engulfment of apoptotic cells by macrophages. We found that the activation of TLR3 upregulates the expression of triggering receptor expressed on myeloid cells (TREM)-like protein 2 (TLT2), a member of the TREM receptor family, on the surface of macrophages. Blocking TLT2 on the macrophage surface by either specific anti-TLT2 Ab or soluble TLT2 extracellular domain attenuated the enhanced ability of macrophages with TLR3 activation to engulf apoptotic cells. To the contrary, overexpression of TLT2 increased the phagocytosis of apoptotic cells. We found that TLT2 specifically binds to phosphatidylserine, a major "eat me" signal that is exposed on the surface of apoptotic cells. Furthermore, we found that TLT2 mediates phagocytosis of apoptotic cells in vivo. Thus, our studies identified TLT2 as an engulfment receptor for apoptotic cells. Our data also suggest a novel mechanism by which TREM receptors regulate inflammation and autoimmune response.

A Decade Later-Progress and Next Steps for Pediatric Simulation Research.

SUMMARY STATEMENT: A decade ago, at the time of formation of the International Network for Pediatric Simulation-based Innovation, Research, and Education, the group embarked on a consensus building exercise. The goal was to forecast the facilitators and barriers to growth and maturity of science in the field of pediatric simulation-based research. This exercise produced 6 domains critical to progress in the field: (1) prioritization, (2) research methodology and outcomes, (3) academic collaboration, (4) integration/implementation/sustainability, (5) technology, and (6) resources/support/advocacy. This article reflects on and summarizes a decade of progress in the field of pediatric simulation research and suggests next steps in each domain as we look forward, including lessons learned by our collaborative grass roots network that can be used to accelerate research efforts in other domains within healthcare simulation science.

Rapidly building surge capacity within a pandemic response using simulation-based clinical systems testing.

Introduction: As the SARS-CoV-2 virus spread across the globe, hospitals around the USA began preparing for its arrival. Building on previous experience with alternative care sites (ACS) during surge events, Texas Children's Hospital (TCH) opted to redeploy their mobile paediatric emergency response teams. Simulation-based clinical systems testing (SbCST) uses simulation to test preoccupancy spaces and new processes. We developed rapid SbCST with social distancing for our deployed ACS, with collaboration between emergency management, paediatric emergency medicine and the simulation team. Methods: A two-phased approach included an initial virtual tabletop activity followed by SbCST at each campus, conducted simultaneously in-person and virtually. These activities were completed while also respecting the need for social distancing amidst a pandemic response. Each activity's discussion was facilitated using Promoting Excellence and Reflective Learning in Simulation (PEARLS) for systems integration debriefing methodology and was followed by compilation of a failure mode and effects analysis (FMEA), which was then disseminated to campus leaders. Results: Within a 2-week period, participants from 20 different departments identified 109 latent safety threats (LSTs) across the four activities, with 71 identified as being very high or high priority items. Very high and high priority threats were prioritised in mitigation efforts by hospital leadership. Discussion: SbCST can be rapidly implemented to hone pandemic responses and identify LSTs. We used SbCST to allow for virtual participation and social distancing within a rapidly accelerated timeline. With prioritised FMEA reporting, leadership was able to mitigate concerns surrounding the four Ss of surge capacity: staff, stuff, structure and systems.

The receptor for urokinase regulates TLR2 mediated inflammatory responses in neutrophils.

The urokinase-type plasminogen activator receptor (uPAR), a glycosylphosphatidylinositol (GPI) anchored membrane protein, regulates urokinase (uPA) protease activity, chemotaxis, cell-cell interactions, and phagocytosis of apoptotic cells. uPAR expression is increased in cytokine or bacteria activated cell populations, including macrophages and monocytes. However, it is unclear if uPAR has direct involvement in the response of inflammatory cells, such as neutrophils and macrophages, to Toll like receptor (TLR) stimulation. In this study, we found that uPAR is required for optimal neutrophil activation after TLR2, but not TLR4 stimulation. We found that the expression of TNF-α and IL-6 induced by TLR2 engagement in uPAR-/- neutrophils was less than that in uPAR+/+ (WT) neutrophils. Pretreatment of neutrophils with PI-PLC, which cleaves GPI moieties, significantly decreased TLR2 induced expression of TNF-α in WT neutrophils, but demonstrated only marginal effects on TNF-α expression in PAM treated uPAR-/- neutrophils. IκB-α degradation and NF-κB activation were not different in uPAR-/- or WT neutrophils after TLR2 stimulation. However, uPAR is required for optimal p38 MAPK activation after TLR2 engagement. Consistent with the in vitro findings that uPAR modulates TLR2 engagement induced neutrophil activation, we found that pulmonary and systemic inflammation induced by TLR2, but not TLR4 stimulation is reduced in uPAR-/- mice compared to WT counterparts. Therefore, our data suggest that neutrophil associated uPAR could be a potential target for treating acute inflammation, sepsis, and organ injury related to severe bacterial and other microbial infections in which TLR2 engagement plays a major role.