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INTRODUCTION: To improve situational awareness in the operating room (OR), a virtual online operating room of hazards (ROH) with deliberately placed risks was created. We hypothesized that subjects first participating in the virtual online ROH would identify more hazards during an in-person ROH exercise in a physical OR than those in the control group who only received didactic training. METHODS: We conducted a randomized controlled trial at a major academic medical center, enrolling 48 pre-clinical medical students with no previous OR exposure during their classes. Control and experimental group subjects participated in a brief, online didactic orientation session conducted live over Zoom (Zoom Video Communications, Inc., San Jose, CA) to learn about latent hazards in the OR. Experimental group subjects further interacted with a virtual online operating ROH in which latent hazards were present. The fraction of deliberately created latent hazards placed in a physical, in-person OR identified by subjects was calculated. RESULTS: Experimental group subjects identified a significantly larger fraction of the created hazards (41.3%) than the control group (difference = 16.4%, 95% CI: 11.3% to 21.4%, P < 0.0001). There was no difference in the number of non-hazards misidentified as hazards between the groups. CONCLUSIONS: Participation in the virtual online environment resulted in greater recognition of latent operating room hazards during a simulation conducted in a physical, in-person OR than in a didactic experience alone. Because creating an in-room experience to teach the identification of latent hazards in an OR is resource-intensive and requires removing the OR from clinical use, we recommend the virtual online approach described for training purposes. Adding items most misidentified as hazards is suggested for future implementation.

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The glial scar is comprised of a heterogeneous population of reactive astrocytes. NG2 glial cells (also known as oligodendrocyte progenitor cells or polydendrocytes) may contribute to this heterogeneity by differentiating into astrocytes in the injured CNS, but there have been conflicting reports about whether astrocytes comprise a significant portion of the NG2 cell lineage. By using genetic fate mapping after spinal cord injury (SCI) and experimental autoimmune encephalomyelitis (EAE) in mice, the goal of this study was to confirm and extend upon previous findings, which have shown that NG2 cell plasticity varies across CNS injuries. We generated mice that express tdTomato in NG2 lineage cells and express GFP under the Aldh1l1 or Glt1 promoter so that NG2 glia-derived astrocytes can be detected by their expression of GFAP and/or GFP. We found that astrocytes comprise approximately 25% of the total NG2 cell lineage in the glial scar by 4 weeks after mid-thoracic contusive SCI, but only 9% by the peak of functional deficit after EAE. Interestingly, a subpopulation of astrocytes expressed only GFP without co-expression of GFAP, uncovering their heterogeneity and the possibility of an underestimation of NG2 glia-derived astrocytes in previous studies. Additionally, we used high performance liquid chromatography to measure the level of tamoxifen and its metabolites in the spinal cord and show that genetic labeling of NG2 glia-derived astrocytes is not an artifact of residual tamoxifen. Overall, our data demonstrate that a heterogeneous population of astrocytes are derived from NG2 glia in an injury type-dependent manner.

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In the version of this article originally published, the values for time shown on the x axis of Figure 5c were incorrect. The error has been corrected in all versions of the paper.

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Direct visualization and light control of several cellular processes is a challenge, owing to the spectral overlap of available genetically encoded probes. Here we report the most red-shifted monomeric near-infrared (NIR) fluorescent protein, miRFP720, and the fully NIR Förster resonance energy transfer (FRET) pair miRFP670-miRFP720, which together enabled design of biosensors compatible with CFP-YFP imaging and blue-green optogenetic tools. We developed a NIR biosensor for Rac1 GTPase and demonstrated its use in multiplexed imaging and light control of Rho GTPase signaling pathways. Specifically, we combined the Rac1 biosensor with CFP-YFP FRET biosensors for RhoA and for Rac1-GDI binding, and concurrently used the LOV-TRAP tool for upstream Rac1 activation. We directly observed and quantified antagonism between RhoA and Rac1 dependent on the RhoA-downstream effector ROCK; showed that Rac1 activity and GDI binding closely depend on the spatiotemporal coordination between these two molecules; and simultaneously observed Rac1 activity during optogenetic manipulation of Rac1.

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