Vicarious traumatization and coping in medical students: a pilot study.

OBJECTIVE: This study explored the impact of traumatic experiences on medical students during their clerkships. METHODS: Medical students completed an anonymous online survey inquiring about traumatic experiences on required clerkships during their third year of medical school, including any symptoms they may have experienced as well as coping strategies they may have used. RESULTS: Twenty-six percent of students reported experiencing vicarious traumatization (VT) during their third year of medical school. CONCLUSIONS: The experience of VT in medical students is relevant to medical educators, given that the resulting symptoms may impact student performance and learning as well as ongoing well-being. Fifty percent of the students who experienced VT in this study did so on the psychiatry clerkship. It is important for psychiatrists to recognize that this is a potential risk for students in order to increase the likelihood that appropriate supports are provided.

Investigating the role of the actin regulating complex ARP2/3 in rapid ischemic tolerance induced neuro-protection.

Neuronal morphology is highly sensitive to ischemia, although some re-organization may promote neuroprotection. In this study we investigate the role of actin regulating proteins (ARP2, ARP3 and WAVE-1) and their role in rapid ischemic tolerance. Using an established in vitro model of rapid ischemic tolerance, we show that WAVE-1 protein levels are stabilized following brief tolerance inducing ischemia (preconditioning). The stabilization appears to be due to a reduction in the ubiquitination of WAVE-1. Levels of ARP2, ARP3 and N-WASP were not affected by ischemic preconditioning. Immunocytochemical studies show a relocalization of ARP2 and ARP3 proteins in neurons following preconditioning ischemia, as well as a re-organization of actin. Blocking the protein kinase CK2 using emodin blocks ischemic tolerance, and our data suggests CK2 binds to WAVE-1 in neurons. We observe an increase in binding of the ARP2 subunit with WAVE-1. The neuroprotection observed following preconditioning is inhibited when cells are transduced with an N-WASP CA domain that blocks the activation of ARP2/3. Together these data show that ischemia affects actin regulating enzymes, and that the ARP2/3 pathway plays a role in rapid ischemic tolerance induced neuroprotection.

Identification of a novel Bcl-2-interacting mediator of cell death (Bim) E3 ligase, tripartite motif-containing protein 2 (TRIM2), and its role in rapid ischemic tolerance-induced neuroprotection.

We have previously shown that the cell death-promoting protein Bcl-2-interacting mediator of cell death (Bim) is ubiquitinated and degraded following a neuroprotection-conferring episode of brief ischemia (preconditioning). Here, we identify the E3 ligase that ubiquitinates Bim in this model, using a proteomics approach. Using phosphorylated GST-Bim as bait, we precipitated and identified by mass spectrometry tripartite motif protein 2 (TRIM2), a RING (really interesting new gene) domain-containing protein. The reaction between TRIM2 and Bim was confirmed using co-immunoprecipitation followed by immunoblotting. We show that TRIM2 binds to Bim when it is phosphorylated by p42/p44 MAPK but does not interact with a nonphosphorylatable Bim mutant (3ABim). 12-O-tetradecanoylphorbol-13-acetate activation of p42/p44 MAPK drives Bim ubiquitination in mouse embryonic fibroblast cells and is associated with an increased interaction between TRIM2 and Bim. One hour following preconditioning ischemia, the binding of Bim to TRIM2 increased, consistent with the time window of enhanced Bim degradation. Blocking p42/p44 MAPK activation following preconditioning ischemia with U0126 or using the nonphosphorylatable 3ABim reduced the binding between Bim and TRIM2. Immunodepletion of TRIM2 from cell lysates prepared from preconditioned cells reduced Bim ubiquitination. Finally, suppression of TRIM2 expression, using lentivirus transduction of shRNAmir, stabilized Bim protein levels and blocked neuroprotection observed in rapid ischemic tolerance. Taken together, these data support a role for TRIM2 in mediating the p42/p44 MAPK-dependent ubiquitination of Bim in rapid ischemic tolerance.

Rapid ischemic tolerance induced by adenosine preconditioning results in Bcl-2 interacting mediator of cell death (Bim) degradation by the proteasome.

Rapid ischemic tolerance, induced one hour following ischemic preconditioning, is mediated via the ubiq-uitin-proteasome system and the degradation of the pro-apoptotic bcl-2 family protein Bim. Previous studies implicate adenosine A1 receptors in mediating rapid ischemic tolerance. Since the A1 adenosine receptor antagonist DPCPX (10µM) blocked rapid ischemic tolerance in our model, we investigated whether adenosine-mediated preconditioning induces rapid ischemic tolerance via the proteasomal degradation of Bim. Cultured rat cortical neurons were incubated for 60 minutes with either adenosine (1µM) or (-)-N(6)-(2-Phenyl-isopropyl) adenosine (RPIA (1µM)), prior to a harmful dose of ischemia (120min oxygen and glucose deprivation). Preconditioned cells had significantly lower levels of cell death following harmful ischemia when compared to non-preconditioned cells. The proteasome inhibitor MG132 (0.1µM) blocked the protective effect of adenosine pre-conditioning. Immunoblot analysis revealed a decrease in Bim protein levels in adenosine and RPIA preconditioned neurons. Adenosine preconditioning induced neuroprotection and Bim degradation was blocked by the MEK inhibitor UO126 (10µM). Our data suggests that pharmacological preconditioning with adenosine results in proteasomal Bim degradation mediated by p42/44 MAPK. Therefore, pharmacological approaches may be able to induce rapid ischemic tolerance via similar molecular mechanisms as ischemic preconditioning.

Sumo-2/3-ylation following in vitro modeled ischemia is reduced in delayed ischemic tolerance.

Several recent studies suggest that sumo-2/3 modification of proteins occurs following harmful ischemia, however, sumo-2/3-ylation may also be associated with hibernation-mediated neuroprotection. Here we investigate the sumoylation of proteins following ischemia and ischemic tolerance using our established in vitro model of ischemia (oxygen and glucose deprivation; OGD). Following harmful ischemia (120 min OGD), we observed a significant increase in the sumo-2/3-ylation of high molecular weight proteins (>85 kDa), but not sumo-1-ylation of proteins. Sumo-2/3-ylation following 120 min OGD was reduced when cultures were preconditioned with non-harmful 30 min OGD 24 h earlier (delayed ischemic tolerance). However, we observed no change in sumo-2/3-ylation in a model of rapid ischemic tolerance. The effects of preconditioning on sumo-2/3-ylation following harmful ischemia were blocked by the protein synthesis inhibitor cycloheximide (1.0 muM), a known inhibitor of delayed ischemic tolerance. In addition, we observed a reduction in sumo-2/3-ylation using hypothermia (4 degrees C 30 min) as the preconditioning stimuli to induce delayed ischemic tolerance. Further studies show that sumo-2/3-ylation occurs during the ischemic insult and that preconditioning does not change expression of the sumo E1- and E2-ligases (UBA2 and Ubc9) or the sumo specific isopeptidases (SenP1-3). While sumo-2/3-ylation is enhanced under conditions of cell stress, it is not yet clear whether this is a cause or consequence of harmful ischemia-induced cell damage.

Ubiquitin proteasome-mediated synaptic reorganization: a novel mechanism underlying rapid ischemic tolerance.

Ischemic tolerance is an endogenous neuroprotective mechanism in brain and other organs, whereby prior exposure to brief ischemia produces resilience to subsequent normally injurious ischemia. Although many molecular mechanisms mediate delayed (gene-mediated) ischemic tolerance, the mechanisms underlying rapid (protein synthesis-independent) ischemic tolerance are relatively unknown. Here we describe a novel mechanism for the induction of rapid ischemic tolerance mediated by the ubiquitin-proteasome system. Rapid ischemic tolerance is blocked by multiple proteasome inhibitors [carbobenzoxy-L-leucyl-L-leucyl-L-leucinal (MG132), MG115 (carbobenzoxy-L-leucyl-L-leucyl-L-norvalinal), and clasto-lactacystin-beta-lactone]. A proteomics strategy was used to identify ubiquitinated proteins after preconditioning ischemia. We focused our studies on two actin-binding proteins of the postsynaptic density that were ubiquitinated after rapid preconditioning: myristoylated, alanine-rich C-kinase substrate (MARCKS) and fascin. Immunoblots confirm the degradation of MARCKS and fascin after preconditioning ischemia. The loss of actin-binding proteins promoted actin reorganization in the postsynaptic density and transient retraction of dendritic spines. This rapid and reversible synaptic remodeling reduced NMDA-mediated electrophysiological responses and renders the cells refractory to NMDA receptor-mediated toxicity. The dendritic spine retraction and NMDA neuroprotection after preconditioning ischemia are blocked by actin stabilization with jasplakinolide, as well as proteasome inhibition with MG132. Together these data suggest that rapid tolerance results from changes to the postsynaptic density mediated by the ubiquitin-proteasome system, rendering neurons resistant to excitotoxicity.