Use of Prescribed Psychotropic Medications in an Opioid Substitution Therapy Cohort.

Objective: Comorbid mental illness is extremely common in individuals receiving opioid substitution therapy. The use of common psychiatric medications is complex in this cohort with increased risks of drug-drug interaction, overdose, and diversion or abuse of prescribed medication. We have therefore investigated rates of co-prescribing and psychiatric comorbidity in a cohort of individuals receiving therapeutic methadone or buprenorphine. Methods: Comprehensive electronic medical records were accessed for a cohort of individuals (n = 698) receiving opioid substitution therapy at a single center in London, United Kingdom. The following was collected for each individual: demographic data, current prescribed medications (including opioid substitution therapy agents), duration of prescription, indication for each prescription, and psychiatric diagnoses. Results: A total of 610 individuals were included in the final analysis. High rates of psychotropic co-prescribing were observed, with 36.7% of individuals receiving a psychotropic medication in addition to their opioid substitution drug, including 35.4% receiving an antidepressant, 9.2% an antipsychotic, 8.6% a benzodiazepine, and 4.5% a gabapentinoid, rates that are far in excess of the local population prescription frequency; 75.5% of antipsychotic prescriptions and 47.7% of benzodiazepine prescriptions were for an unlicensed indication. Conclusions: This highlights the need for evidence-based treatment of comorbid mental illness for individuals receiving opioid substitution therapy.

Completion CT of Chest, Abdomen, and Pelvis after Acute Head and Cervical Spine Trauma: Incidence of Acute Traumatic Findings in the Setting of Low-Velocity Trauma.

PURPOSE: To determine the frequency of acute traumatic findings in computed tomographic (CT) chest abdomen pelvis (CAP) examinations in patients with acute traumatic head and/or cervical spine injury and no evidence suggesting bodily injury. MATERIALS AND METHODS: After institutional review board approval with a waiver of informed consent was obtained, a HIPAA-compliant retrospective study was performed. A review of the electronic medical records and dictated reports identified patients who met the following criteria: CT-documented acute head and/or cervical spine trauma, CT CAP performed at least 20 minutes after initial brain and/or cervical spine CT, and no evidence of bodily injury at physical examination or on initial plain radiographs. The types of head and/or cervical injury, as well as mechanisms of injury in these patients, were analyzed. The frequency of acute traumatic injury in the CT CAP examinations was also determined, and 95% confidence intervals were calculated. RESULTS: There were 115 patients who met the study criteria (average age, 67.3 years). Sixty-three (54.8%) patients were male. The average injury severity score was 9.3. No patients who met the criteria for this study were found to have an acute traumatic injury to the chest, abdomen, or pelvis. These 115 CT CAP examinations comprised 7.5% (115 of 1530) of all CT CAP examinations performed in the emergency department over the 15-month study period. CONCLUSION: CT CAP examinations rarely if ever reveal acute traumatic injury in patients who have experienced low-velocity trauma and have acute head and/or cervical spine trauma in the absence of evidence of bodily injury.

Benign colonic metaplasia at a previous stoma site in a patient without adenomatous polyposis.

There are few reported cases of cutaneous intestinal metaplasia or primary adenocarcinoma arising at the ileostomy site following panproctocolectomy. These complications have been seen almost exclusively in patients with familial adenomatous polyposis and inflammatory bowel disease (IBD). However, benign intraepidermal colonic mucosa at a reversed ileostomy site in a patient without familial adenomatous polyposis or IBD has not been documented. We report a case of a 51-year-old female with a history of colonic adenocarcinoma who presented with pruritic, erythematous, scaly plaques on the right lower abdomen, present since reversal of her ileostomy in 2007. Skin biopsy revealed benign foci of colonic epithelium with no evidence of adenomatous change. Benign intraepidermal colonic mucosa was diagnosed based on histopathologic findings and immunohistochemistry. To our knowledge, this is the first case of intraepidermal benign colonic metaplasia forming in a patient following ostomy reversal. The case emphasizes the importance of patient education and physical examination of the stoma or stoma remnants for detection of unusual or changing lesions due to the risk for malignant transformation. It also demonstrates that benign colonic mucosa should be considered in the differential diagnosis when evaluating lesions near ileostomy sites, regardless of whether the patient has a history of familial adenomatous polyposis or IBD.

Emergency physician focused cardiac ultrasound improves diagnosis of ascending aortic dissection.

STUDY OBJECTIVE: Ascending aortic dissection (AAD) is an uncommon, time-sensitive, and deadly diagnosis with a nonspecific presentation. Ascending aortic dissection is associated with aortic dilation, which can be determined by emergency physician focused cardiac ultrasound (EP FOCUS). We seek to determine if patients who receive EP FOCUS have reduced time to diagnosis for AAD. METHODS: We performed a retrospective review of patients treated at 1 of 3 affiliated emergency departments, March 1, 2013, to May 1, 2015, diagnosed as having AAD. All autopsies were reviewed for missed cases. Primary outcome measure was time to diagnosis. Secondary outcomes were time to disposition, misdiagnosis rate, and mortality. RESULTS: Of 386547 ED visits, targeted review of 123 medical records and 194 autopsy reports identified 32 patients for inclusion. Sixteen patients received EP FOCUS and 16 did not. Median time to diagnosis in the EP FOCUS group was 80 (interquartile range [IQR], 46-157) minutes vs 226 (IQR, 109-1449) minutes in the non-EP FOCUS group (P = .023). Misdiagnosis was 0% (0/16) in the EP FOCUS group vs 43.8% (7/16) in the non-EP FOCUS group (P = .028). Mortality, adjusted for do-not-resuscitate status, for EP FOCUS vs non-EP FOCUS was 15.4% vs 37.5% (P = .24). Median rooming time to disposition was 134 (IQR, 101-195) minutes for EP FOCUS vs 205 (IQR, 114-342) minutes for non-EP FOCUS (P = .27). CONCLUSIONS: Patients who receive EP FOCUS are diagnosed faster and misdiagnosed less compared with patients who do not receive EP FOCUS. We recommend assessment of the thoracic aorta be performed routinely during cardiac ultrasound in the emergency department.

Aperius interspinous device for degenerative lumbar spinal stenosis: a review.

With an aging population, degenerative lumbar spinal stenosis (DLSS) leading to neurogenic intermittent claudication (NIC) is a growing problem. For patients suffering from this condition, interspinous process distraction devices (IPDs) offer an effective and cheap alternative to conservative or decompressive surgery. Aperius is one such device that has been gaining popularity for its percutaneous insertion under local anesthetic, short operative time, and low risk of complications. The main objective of this review was to carry out a comprehensive search of the literature to evaluate the effectiveness and potential complications of Aperius. A database search, including PubMed, Clinical trials.gov, Cochrane (CENTRAL), MEDLINE, CINAHL, EMBASE, and Scopus, was carried out to identify relevant articles written in English reporting on complications with a minimum 12-month follow-up. The literature search resulted in six eligible studies; two nonrandomized comparative and four prospective case series were available. The analysis revealed that in total, 433 patients underwent treatment with Aperius, with all studies demonstrating an improvement in outcome measures. The average follow-up was 17 months with an overall complication rate of 10.62%. Overall, the quality of evidence is low, suggesting that currently, the evidence is not compelling and further prospective randomized trials including cost-effectiveness studies are required.

Reshaping the emergency radiology call experience after the merger of two programs: the Yale experience.

In the summer of 2013, 16 radiology residents from the Hospital of Saint Raphael (HSR) joined the 38 residents of Yale-New Haven Hospital (YNHH) to become a single 54-resident program. This posed a significant challenge given the number of residents and very different call structures of the two institutions. After evaluating the emergency radiology volume at both hospitals, it was determined that implementing YNHH's traditional call system at HSR would increase call by approximately 25 %. In order to negate this increase, the SRC rotation was created at HSR. This Monday-Friday rotation covered by R3s starts at 1 p.m. with afternoon conference. Residents then read cases on a subspecialty service from 2-5 p.m. and then cover the entire hospital until 10 p.m. with a single attending. Because of this rotation, call did not increase for the YNHH residents and third year residents were provided with increased responsibility. For programs not undergoing a merger, call rotations can also be extremely beneficial. These rotations allow third year residents to have more "call-free" weeks prior to the ABR core exam. Also, patient care can be improved, as the shift length for on-call residents is reduced, which has been shown to improve accuracy.

Chicago Ebola Response Network (CERN): A Citywide Cross-hospital Collaborative for Infectious Disease Preparedness.

The 2014-2015 Ebola virus disease (EVD) epidemic and international public health emergency has been referred to as a "black swan" event, or an event that is unlikely, hard to predict, and highly impactful once it occurs. The Chicago Ebola Response Network (CERN) was formed in response to EVD and is capable of receiving and managing new cases of EVD, while also laying the foundation for a public health network that can anticipate, manage, and prevent the next black swan public health event. By sharing expertise, risk, and resources among 4 major academic centers, Chicago created a sustainable network to respond to the latest in a series of public health emergencies. In this respect, CERN is a roadmap for how a region can prepare to respond to public health emergencies, thereby preventing negative impacts through planning and implementation.

Reported clinical outcomes of high-fidelity simulation versus classroom-based end-of-life care education.

BACKGROUND: Nursing students often experience increased anxiety, lack of clinical skills and poor coping mechanisms when dealing with death and dying for the first time. AIMS: To design, use and assess the effectiveness of high-fidelity simulation teaching versus classroom-based end-of-life care (EoLC) teaching for first-year nursing students on dealing with their first death experiences in clinical placements. METHODS: A qualitative phenomenography approach was taken. First-year nursing students (n=187) were randomly allocated to receive either classrtoom-based (n=139) or high-fidelity simulation-based EoLC teaching (n=48). When all participants had returned from their first clinical placement, 12 individual in-depth interviews were carried out. Data were analysed using framework analysis. RESULTS: Comparative data analysis revealed four key themes as clinical outcomes: recognising death and dying; knowledge into practice; preparedness for clinical eventualities; and emotional preparedness. CONCLUSIONS: Despite the fact that both strategies improved students' knowledge of EoLC, simulation-based EoLC education was perceived as better in terms of enhanced practical skills and improved emotional experience in their first clinical placement.

Disclosure of financial conflicts of interest: an evaluation of orthopaedic surgery patients' understanding.

BACKGROUND: Industry and orthopaedic surgeons often partner to develop new technology, which can lead to orthopaedic surgeons having financial conflicts of interest (FCOI). It is essential these FCOI be conveyed clearly to patients. It is unclear, however, whether and to what degree patients understand the ramifications of physician FCOI. QUESTIONS/PURPOSES: We evaluated (1) patients' concerns regarding their surgeon having FCOI or the presence of institutional FCOI, (2) the effect of surgeon FCOI on patients' willingness to have surgery, and (3) patients' understanding of FCOI. METHODS: We asked 101 patients (66% female) receiving total joint arthroplasty from the orthopaedic practices of two surgeons at an academic health center to complete a descriptive, correlational designed survey at their 6-week followup appointment. The data collected included patient demographics, knowledge of FCOI, and the influence of FCOI on patient attitudes toward surgery and their surgeon. RESULTS: A minority of patients (13%) reported discussing FCOI with prior physicians and only 55% agreed or strongly agreed a surgeon should disclose FCOI. Only 15% of patients believed such conflicts would make them less likely to have their surgeon operate on them. Level of education was weakly correlated (Spearman's rho = 0.29) with patient understanding of FCOI. CONCLUSIONS: Overall, patients had a poor understanding of FCOI. Both level of education and previous discussions of FCOI predicted better understanding. This study emphasizes communication of FCOI with patients needs to be enhanced.

A retrospective review of out of hospital cardiac arrest at Gillette Stadium: 10 years of experience at a large sports venue.

Introduction: Rates of out-of-hospital cardiac arrest (OHCA) at major sporting events are as high as 0.7 per 100,000 attendees. However, factors contributing to OHCA at mass gatherings have not been well-described. We describe our experience with ten years of medical oversight and OHCA care at a professional football stadium. Methods: We performed a retrospective review of OHCA events between August 2010 and January 2020 at a 65,878-seat football stadium, with a single transporting EMS agency and a single receiving hospital. We analyzed EMS incident reports and matched patients to hospital records for outcome data. Results: A total of 7,767,345 people attended 115 football games during the study period. There were 21 OHCAs (0.27 per 100,000 attendees). Ninety-five percent of OHCAs were witnessed and 71.4% had an initial shockable rhythm, with bystander AED use in 47.6%. Median EMS response time was 2 minutes (IQR 1-6). For 7 patients defibrillated by EMS, time to defibrillation was 4 minutes (IQR 4-11). Return of spontaneous circulation (ROSC) occurred in 71%, with 47% having good 30-day neurologic survival. All patients with an initial rhythm of asystole died. Conclusion: The ROSC rate at our stadium exceeded 70% with almost half surviving with good neurologic outcomes, substantially higher than that reported for the general public. We hope that our experience will provide valuable lessons to other similarly sized stadiums.

Should gabapentinoids be prescribed long-term for anxiety and other mental health conditions?

Prescription rates for gabapentinoids are rising in England. Pregabalin is currently recommended by NICE for the treatment of anxiety. Gabapentinoids have some overlap with the action of benzodiazepines, and have similar issues with tolerance, dependence, addiction and withdrawal. They were scheduled as class C controlled drugs in 2019 because of these risks. There were 244 deaths due to poisoning recorded by the ONS in 2019 involving pregabalin. Poisonings due to pregabalin usually involve concomitant use of opioids or other drugs. The rate of deaths involving pregabalin has been rising steeply for the last 10 years, and now exceeds those attributed to diazepam, fentanyl, the tricyclics as a group or SSRIs as a group. Evidence for the use of pregabalin in anxiety is derived from short-term trials, with marginal differences from placebo, which do not take into account the longer term effects of tolerance, dependence and withdrawal. We call on NICE to re-evaluate their support for use of pregabalin in anxiety in light of its known harms. The use of gabapentinoids off-label for other psychiatric conditions should also be re-considered. In general, psychotropic medications require longer term efficacy and safety studies before allowing widespread use.

Alcohol and cocaine use prior to suspected suicide: Insights from toxicology.

INTRODUCTION: This study investigates whether there is a relationship between alcohol and cocaine use in deaths where suicide by self-injury is the suspected cause of death. METHODS: Adults referred by coroners to the Imperial College London Toxicology Unit for toxicological analysis between 2012 and 2016 were reviewed for inclusion criteria. Those who died by self-injury reasoned to be deliberate were included in the analysis. Femoral blood alcohol concentration (BAC) and presence of cocaine or benzoylecognine (a metabolite of cocaine) in blood and/or urine were tabulated and odds ratios calculated. RESULTS: A total of 1722 decedents met inclusion criteria. BAC was ≥50 mg/dL in 29% of decedents. Cocaine was detected in 8.4% of cases. The likelihood of testing positive for cocaine increased with BAC and was most frequent between 100 and 199 mg/dL, consistent with moderate to severe intoxication (odds ratio 5.88, 95% confidence interval 3.80, 9.09; P ≤ 0.001) compared to those with BAC <10 mg/dL. DISCUSSION AND CONCLUSIONS: This study demonstrates a correlation between increasing BAC and likelihood of cocaine use prior to suspected suicide, up to a level consistent with severe intoxication. Cocaine use was found in a high proportion of cases relative to the general population reporting regular use. This pattern of drug and alcohol use has previously been given little attention in suicide prevention strategies and clinical prioritisation.

Expression and localization of rat aldo-keto reductases and induction of the 1B13 and 1D2 isoforms by phenolic antioxidants.

The aldo-keto reductase (AKR) phase I drug metabolism enzyme superfamily is implicated in detoxification or bioactivation of a wide variety of carbonyl-bearing compounds. In this study, we have used antibodies raised against purified recombinant rat AKR isoforms 1A3, 1B4, 1C9, 1D2, and 7A1 to characterize the expression profile of these superfamily members in the rat and define their localization by immunohistochemistry. Western blotting showed that AKR1A3, AKR1B4, and AKR1C9 are ubiquitously expressed, whereas AKR1D2 and AKR7A1 are present in liver, adrenal gland, and kidney, with the latter also present in testis, spleen, and stomach. Immunohistochemical analysis of the kidney demonstrated the localization of AKR1A3 in proximal convoluted tubules, AKR1B4 in the loop of Henle, and AKR1C9 in the pars recta S3 segment of proximal tubules. We also report localization of AKR1B4 in the adrenal gland (parenchymal cells of the zona reticularis) and testis (Sertoli cells and late spermatids), of AKR1D2 in the liver (hepatocyte nuclei), and of AKR7A1 in the pancreatic duct and bronchiolar epithelium. Previous studies have shown that expression of AKR7A1 is induced in response to dietary administration of the phenolic antioxidants butylated hydroxyanisole and ethoxyquin. Here we identify AKR1B13 and AKR1D2 as further inducible members of the rat AKR superfamily.

Role of Mdm2 acid domain interactions in recognition and ubiquitination of the transcription factor IRF-2.

Mdm2 (murine double minute 2)-mediated ubiquitination of the p53 tumour suppressor requires interaction of the ligase at two distinct binding sites that form general multiprotein-docking sites for the p53 protein. The first Mdm2-binding site resides in the transactivation domain of p53 and is an allosteric effector site for Mdm2-mediated p53 ubiquitination; the second site requires the acid domain of Mdm2 to recognize a 'ubiquitination signal' within p53's DNA-binding core. In order to expand on fundamental requirements for a protein to function as an Mdm2 substrate and the role of the acid domain in recognition, we have carried out a bioinformatics search for open reading frames that have homology with the Mdm2-docking sites in p53. IRF-2 [IFN (interferon) regulatory factor-2], an IFN-regulated transcription factor, has been identified as an Mdm2-binding protein and substrate requiring interactions with both the hydrophobic pocket and the acid domain of Mdm2. Mutation of either of the two Mdm2-binding sites on IRF-2 can attenuate substrate ubiquitination, confirming the requirement of a dual-site substrate interaction mechanism. Ligands that bind to the hydrophobic pocket are not sufficient to inhibit Mdm2 E3-ligase activity. Rather, acid domain-binding ligands act as E3-ligase inhibitors, lending additional support to the idea that the acid domain of Mdm2 is key to understanding its mechanism of action. The ability of Mdm2 and IRF-2 to form a complex in cells complements the biochemical assays and together establishes a novel substrate with which to develop insights into E3-ubiquitin ligase-substrate interactions in vitro and in cells.

Incidence and treatment costs of severe bacterial infections among people who inject heroin: A cohort study in South London, England.

BACKGROUND: People who inject drugs often get bacterial infections. Few longitudinal studies have reported the incidence and treatment costs of these infections. METHODS: For a cohort of 2335 people who inject heroin entering treatment for drug dependence between 2006 and 2017 in London, England, we reported the rates of hospitalisation or death with primary causes of cutaneous abscess, cellulitis, phlebitis, septicaemia, osteomyelitis, septic arthritis, endocarditis, or necrotising fasciitis. We compared these rates to the general population. We also used NHS reference costs to calculate the cost of admissions. RESULTS: During a median of 8.0 years of follow-up, 24 % of patients (570/2335) had a severe bacterial infection, most commonly presenting with cutaneous abscesses or cellulitis. Bacterial infections accounted for 13 % of all hospital admissions. The rate was 73 per 1000 person-years (95 % CI 69-77); 50 times the general population, and the rate remained high throughout follow-up. The rate of severe bacterial infections for women was 1.50 (95 % CI 1.32-1.69) times the rate for men. The mean cost per admission was £4980, and we estimate that the annual cost of hospital treatment for people who inject heroin in London is £4.5 million. CONCLUSIONS: People who inject heroin have extreme and long-term risk of severe bacterial infections.

Telephone delivered incentives for encouraging adherence to supervised methadone consumption (TIES): Study protocol for a feasibility study for an RCT of clinical and cost effectiveness.

The majority of people receiving treatment for their heroin addiction, are prescribed methadone; for which there is an extensive evidence base. When treatment starts, people take their daily dose of methadone under supervision at a community pharmacy. Supervision guarantees methadone is taken as directed by the individual for whom it has been prescribed, helps to ensure individuals take their correct dose every day, and safeguards against diversion and overdose. However, individuals often fail to attend the pharmacy to take their methadone. Each missed dose is of concern. If a patient misses their daily dose of methadone, they will start to experience opiate withdrawal and cravings and are more likely to use heroin. If they miss three days dose, there are concerns that they may lose tolerance to the drug and may be at risk of overdose when the next dose is taken. Hence there is an urgent need to develop effective interventions for medication adherence. Research suggests that incentive-based medication adherence interventions may be very effective, but there are few controlled trials and the provision of incentives requires time and organisational systems which can be challenging in pharmacies. The investigators have developed the technology to deliver incentives by mobile telephone. This cluster randomised trial will test the feasibility of conducting a future trial evaluating the clinical and cost effectiveness of using telephone delivered incentives (praise and modest financial rewards via text messaging) to encourage adherence with supervised consumption of methadone in community pharmacies. Three drug services (each with two or three community pharmacies supervising methadone consumption that will enrol 20 individuals, a total of 60 participants) will be recruited and randomly allocated to deliver either i) telephone delivered incentives, ii) telephone delivered reminders or iii) no telephone system. Acceptability, recruitment, follow-up, and suitable measures of clinical and cost effectiveness will be assessed. Findings from this feasibility study will be assessed against stated progression criteria and used to inform a future confirmatory trial of the clinical and cost effectiveness of telephone delivered incentives to encourage medication adherence. TRIAL REGISTRATION: ISRCTN58958179 (retrospectively registered).

Droplet Combinations: A Scalable Microfluidic Platform for Biochemical Assays.

Droplet-based microfluidics holds enormous potential for transforming high-throughput drug screening. Miniaturization through droplets in combination with automation contributes to reduce reagent use and analysis time as well as minimizing or eliminating labor-intensive steps leading to associated reductions in cost. In this paper, we demonstrate the potential of automated and cost-effective microfluidic droplet-generating technology in the context of an enzymatic activity assay for screening collagenase inhibitors. Experimental results show reproducible and accurate creation and mixing of droplet combinations resulting in biochemical data comparable to data produced by an industry standard instrument. This microfluidic platform that can generate and combine multiple droplets represents a promising tool for high-throughput drug screening.

1-Cyano-2,3-epithiopropane is a novel plant-derived chemopreventive agent which induces cytoprotective genes that afford resistance against the genotoxic alpha,beta-unsaturated aldehyde acrolein.

Epithionitriles represent a previously unrecognized class of cancer chemopreventive phytochemical generated from alkenyl glucosinolates in cruciferous vegetables. In rat liver RL-34 epithelial cells, 1-cyano-2,3-epithiopropane (CETP), 1-cyano-3,4-epithiobutane (CETB) and 1-cyano-4,5-epithiopentane (CETPent) were shown to induce cytoprotective enzymes including NAD(P)H:quinone oxidoreductase 1 (NQO1), glutathione (GSH) S-transferase A3 and the glutamate-cysteine ligase modifier subunit; CETP was more potent in this regard than were either CETB or CETPent, with 50 microM CETP eliciting a remarkable approximately 10-fold induction of NQO1. Furthermore, 50 microM CETP stimulated a 2.0-fold overproduction of GSH in RL-34 cells. Transfection experiments demonstrated that epithionitriles induced gene expression through an antioxidant response element (ARE) and that transactivation of an Nqo1-luciferase reporter plasmid was dependent on NF-E2 p45-related factor 2 (Nrf2), a cap'n'collar basic region leucine zipper transcription factor. Evidence is presented that CETP affected Nrf2-mediated induction of ARE-driven transcription by inhibiting Kelch-like ECH-associated protein 1 (Keap1), a ubiquitin ligase substrate adaptor that negatively regulates Nrf2. We found that Nqo1 was expressed constitutively at high levels in Keap1(-/-) mouse embryonic fibroblasts (MEFs) and it was not further induced by CETP. However, knock-in of mouse Keap1 or zebrafish Keap1a into Keap1(-/-) MEFs repressed Nqo1-luciferase reporter gene activity, but repression by the murine or zebrafish proteins was antagonized by CETP. Pre-treatment of Nrf2(+/+) MEFs, but not Nrf2(-/-) MEFs, with 15 microM CETP for 24 h conferred 2.4-fold resistance against subsequent exposure to the alpha,beta-unsaturated aldehyde acrolein, indicating that the phytochemical exerts chemopreventive properties against genotoxic xenobiotics.

Transcription factor Nrf2 mediates an adaptive response to sulforaphane that protects fibroblasts in vitro against the cytotoxic effects of electrophiles, peroxides and redox-cycling agents.

Sulforaphane can stimulate cellular adaptation to redox stressors through transcription factor Nrf2. Using mouse embryonic fibroblasts (MEFs) as a model, we show herein that the normal homeostatic level of glutathione in Nrf2(-/-) MEFs was only 20% of that in their wild-type counterparts. Furthermore, the rate of glutathione synthesis following its acute depletion upon treatment with 3 micromol/l sulforaphane was very substantially lower in Nrf2(-/-) MEFs than in wild-type cells, and the rebound leading to a approximately 1.9-fold increase in glutathione that occurred 12-24 h after Nrf2(+/+) MEFs were treated with sulforaphane was not observed in Nrf2(-/-) fibroblasts. Wild-type MEFs that had been pre-treated for 24 h with 3 micromol/l sulforaphane exhibited between 1.4- and 3.2-fold resistance against thiol-reactive electrophiles, including isothiocyanates, alpha,beta-unsaturated carbonyl compounds (e.g. acrolein), aryl halides and alkene epoxides. Pre-treatment of Nrf2(+/+) MEFs with sulforaphane also protected against hydroperoxides (e.g. cumene hydroperoxide, CuOOH), free radical-generating compounds (e.g. menadione), and genotoxic electrophiles (e.g. chlorambucil). By contrast, Nrf2(-/-) MEFs were typically approximately 50% less tolerant of these agents than wild-type fibroblasts, and sulforaphane pre-treatment did not protect the mutant cells against xenobiotics. To test whether Nrf2-mediated up-regulation of glutathione represents the major cytoprotective mechanism stimulated by sulforaphane, 5 micromol/l buthionine sulfoximine (BSO) was used to inhibit glutathione synthesis. In Nrf2(+/+) MEFs pre-treated with sulforaphane, BSO diminished intrinsic resistance and abolished inducible resistance to acrolein, CuOOH and chlorambucil, but not menadione. Thus Nrf2-dependent up-regulation of GSH is the principal mechanism by which sulforaphane pre-treatment induced resistance to acrolein, CuOOH and chlorambucil, but not menadione.