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Katherine S Monroe, Lindsey Amerson, William Bundy, Yulanda Greene, Gregory Jarvis, Jennifer Stever, Gaurav P Patel Department of Anesthesiology, Anesthesiologist Assistant Program, Emory University School of Medicine, Atlanta, GA, USACorrespondence: Gaurav P Patel, Department of Anesthesiology, Anesthesiologist Assistant Program, Emory University School of Medicine, 57 Executive Park South, Suite 300, Atlanta, GA, 30329, USA, Email gppatel@emory.eduBackground: The Anesthesiologist Assistant (AA) profession is gaining popularity, and as such, the number of applicants and the number of AA programs are also increasing. There is no published data on what makes a good AA student – multiple studies exist in this regard for anesthesiology residents and medical students. The Medical College Admissions Test (MCAT) and the Graduate Record Examination (GRE) have shown to have some correlation with student success in other health professions. There is no data to suggest that they can indicate potential success or academic problems during AA school. As such, this study evaluated the impact of these standardized test scores and student success in both the preclinical and clinical curricula at the Emory AA program.Methods: A retrospective analysis of MCAT scores and GRE scores was performed. 2-tailed Pearson coefficients were calculated to see if there was a relationship between higher MCAT and GRE scores and performance in our program.Results: 2- tailed Pearson coefficients showed a moderately strong correlation between MCAT scores and our AA program’s science curriculum. There was a weak correlation as well between GRE scores here. The correlation dropped slightly for the MCAT and performance during our clinical curriculum, and a similar drop was noted for the GRE.Discussion: Our study shows that the MCAT appears to more strongly correlated with better performance in both the science curriculum of our program and also with our clinical program. The study likely implies that the MCAT appears to be a better gauge as to how a student will do in an AA program in comparison to the GRE. Further studies are needed to understand how to appropriately admit students who will successfully complete the rigorous AA program curriculum both at Emory and beyond.Keywords: admission metrics, certified anesthesiologist assistant, anesthesia care team, student success, standardized test scores

Katherine S Monroe, William Bundy, Yulanda Greene, Gregory Jarvis, Jennifer Stever, Gaurav P Patel Department of Anesthesiology, Emory University School of Medicine, Atlanta, GA, USACorrespondence: Gaurav P Patel, Emory University, 57 Executive Park South, Suite 300, Atlanta, GA, 30329, USA, Email gppatel@emory.eduPurpose: The Anesthesiologist Assistant career is gaining significant popularity in the health professions in the United States. Given that this medical occupation is relatively young, there is limited information regarding student success in this demanding graduate-level program. Assessing if pre-admission metrics influence how students perform during the curriculum is essential to recruiting the appropriate candidates. Grade point averages have been shown to correlate with student success in medical education programs for both medical students and physician assistant students, but there is currently no information regarding anesthesiologist assistant students.Methods: Pre-matriculation science and cumulative grade point averages were accessed in a deidentified manner for Emory University Anesthesiologist Assistant Students, and 2-tailed Pearson coefficients were calculated to see if there was a correlation with performance during the science/didactic curriculum of our program and with the clinical curriculum of the program.Results: The 2-tailed Pearson coefficients showed a moderately strong positive correlation between pre-admission science and cumulative grade point averages and performance during the science curriculum of the Emory program (r=0.522). Data also suggested a moderate correlation with grade point averages at graduation from our program (r=0.484). Similar results were found with cumulative grade point averages as well.Conclusion: Given the limited information, we have regarding pre-admission metrics and performance in an Anesthesiologist Assistant program, our study shows that pre-admission science scores and grades in general in undergraduate studies does in fact mimic the information found from studies of other health profession students. Further studies are needed to elucidate how to choose the most appropriate candidates for admission to anesthesiologist assistant programs.Keywords: admission metrics, certified anesthesiologist assistant, anesthesia care team, student success

Up to 50% of those diagnosed with HIV in the U.S. are not retained in medical care. Care retention provides opportunity to monitor benefits of HIV therapy and enable viral suppression. To increase retention, there is a need to prioritize best practices appropriate for translation and dissemination for real-world implementation. Eighteen interventions from CDC's Compendium of Evidence-Based Interventions were scored using the RE-AIM framework to determine those most suitable for dissemination. A CDC Division of HIV Prevention workgroup developed a RE-AIM scale with emphasis on the Implementation and Maintenance dimensions and less emphasis on the Efficacy dimension since all 18 interventions were already identified as evidence-based or evidence-informed. Raters referenced primary efficacy publications and scores were averaged for a ranked RE-AIM score for interventions. Of 18 interventions, four included care linkage and 7 included viral suppression outcomes. Interventions received between 20.6 and 35.3 points (45 maximum). Scores were converted into a percentage of the total possible with ranges between 45.8 and 78.4%. Top four interventions were ARTAS (78.4%); Routine Screening for HIV (RUSH) (73.2%); Optn4Life (67.4%) and Virology Fast Track (65.9%). All four scored high on Implementation and Maintenance dimensions. Select items within the scale were applicable to health equity, covering topics such as reaching under-served focus populations and acceptability to that population. Navigation-enhanced Case Management (NAV) scored highest on the health equity subscale. RE-AIM prioritization scores will inform dissemination and translation efforts, help clinical staff select feasible interventions for implementation, and support sustainability for those interventions., (© 2022. This is a U.S. Government work and not under copyright protection in the US; foreign copyright protection may apply.)

On September 30, 2022, after >3 years with no confirmed cholera cases (1), the Directorate of Epidemiology, Laboratories and Research (DELR) of the Haitian Ministry of Public Health and Population (Ministère de la Santé Publique et de la Population [MSPP]) was notified of two patients with acute, watery diarrhea in the metropolitan area of Port-au-Prince. Within 2 days, Haiti's National Public Health Laboratory confirmed the bacterium Vibrio cholerae O1 in specimens from the two patients with suspected cholera infection, and an outbreak investigation began immediately. As of January 3, 2023, >20,000 suspected cholera cases had been reported throughout the country, and 79% of patients have been hospitalized. The moving 14-day case fatality ratio (CFR) was 3.0%. Cholera, which is transmitted through ingestion of water or food contaminated with fecal matter, can cause acute, severe, watery diarrhea that can rapidly lead to dehydration, shock, and death if not treated promptly (2). Haiti is currently facing ongoing worsening of gang violence, population displacement, social unrest, and insecurity, particularly in the metropolitan area of Port-au-Prince, including Belair, Bas-Delmas, Centre-Ville, Martissant, Cité Soleil, Croix-des Bouquets, and Tabarre, creating an environment that has facilitated the current resurgence of cholera (3). This report describes the initial investigation, ongoing outbreak, and public health response to cholera in Haiti. Cholera outbreak responses require a multipronged, multisectoral approach including surveillance; case management; access to safe water, sanitation, and hygiene (WASH) services; targeted oral cholera vaccine (OCV) campaigns; risk communication; and community engagement. This activity was reviewed by CDC and was conducted consistent with applicable federal law and CDC policy., Competing Interests: All authors have completed and submitted the International Committee of Medical Journal Editors form for disclosure of potential conflicts of interest. No potential conflicts of interest were disclosed.

Since 2003, the US President's Emergency Plan for AIDS Relief (PEPFAR) has supported implementation and maintenance of health information systems for HIV/AIDS and related diseases, such as tuberculosis, in numerous countries. As the COVID-19 pandemic emerged, several countries conducted rapid assessments and enhanced existing PEPFAR-funded HIV and national health information systems to support COVID-19 surveillance data collection, analysis, visualization, and reporting needs. We describe efforts at the US Centers for Disease Control and Prevention (CDC) headquarters in Atlanta, Georgia, USA, and CDC country offices that enhanced existing health information systems in support COVID-19 pandemic response. We describe CDC activities in Haiti as an illustration of efforts in PEPFAR countries. We also describe how investments used to establish and maintain standards-based health information systems in resource-constrained settings can have positive effects on health systems beyond their original scope.

Background: Haiti's first COVID-19 cases were confirmed on March 18, 2020, and subsequently spread throughout the country. The objective of this study was to describe clinical manifestations of COVID-19 in Haitian outpatients and to identify risk factors for severity of clinical manifestations., Methods: We conducted a retrospective study of COVID-19 outpatients diagnosed from March 18-August 4, 2020, using demographic, epidemiological, and clinical data reported to the Ministry of Health (MoH). We used univariate and multivariate analysis, including multivariable logistic regression, to explore the risk factors and specific symptoms related to persons with symptomatic COVID-19 and the severity of symptomatic COVID-19 disease., Results: Of 5,389 cases reported to MOH during the study period, 1,754 (32.5%) were asymptomatic. Amongst symptomatic persons 2,747 (75.6%) had mild COVID-19 and 888 (24.4%) had moderate-to-severe disease; the most common symptoms were fever (69.6%), cough (51.9%), and myalgia (45.8%). The odds of having moderate-to-severe disease were highest among persons with hypertension (aOR = 1.72, 95% Confidence Interval [CI] (1.34, 2.20), chronic pulmonary disease (aOR = 3.93, 95% CI (1.93, 8.17)) and tuberculosis (aOR = 3.44, 95% CI (1.35, 9.14)) compared to persons without those conditions. The odds of having moderate-to-severe disease increased with age but was also seen among children aged 0-4 years (OR: 1.73, 95% CI (0.93, 3.08)), when using 30-39 years old as the reference group. All of the older age groups, 50-64 years, 65-74 years, 75-84 years, and 85+ years, had significantly higher odds of having moderate-to-severe COVID-19 compared with ages 30-39 years. Diabetes was associated with elevated odds of moderate-to-severe disease in bivariate analysis (OR = 2.17, 95% CI (1.58,2.98) but, this association did not hold in multivariable analyses (aOR = 1.22,95%CI (0.86,1.72))., Conclusion: These findings from a resource-constrained country highlight the importance of surveillance systems to track emerging infections and their risk factors. In addition to co-morbidities described elsewhere, tuberculosis was a risk factor for moderate-to-severe COVID-19 disease., Competing Interests: The authors have declared that no competing interests exist.

Background: Haiti introduced a monovalent human group A rotavirus (RVA) vaccine (Rotarix) into its routine infant immunization program in April 2014. The goal of the surveillance program was to characterize RVA strains circulating in Haiti before and after RVA vaccine introduction., Methods: Stool samples were collected from children <5 years old presenting with acute gastroenteritis at 16 hospitals in Haiti. RVA antigen enzyme immunoassay (EIA) testing was performed, and G and P genotypes were determined for positive specimens. In this study, genotype data for samples collected from May 2012 through April 2014 (the pre-vaccine introduction era) and May 2014 through July 2019 (post-vaccine introduction era) were analyzed., Results: A total of 809 specimens were tested by the Centers for Disease Control and Prevention. During the pre-vaccine introduction era (May 2012 through April 2014), G12P[8] was the predominant genotype, detected in 88-94% of specimens. There was a high prevalence of the equine-like G3P[8] genotype among Haitian children with RVA after vaccine introduction., Conclusions: The predominance of equine-like G3P[8] in three of five RVA seasons post-vaccine introduction suggests possible vaccine-specific selection pressure in Haiti. These temporal variations in RVA genotype predominance will require continued monitoring in Haiti as the vaccination program continues., (© 2022 Published by Elsevier Ltd on behalf of International Society for Infectious Diseases.)

Rotavirus is responsible for 26% of diarrheal deaths in Latin America and the Caribbean. Haiti introduced the monovalent rotavirus vaccine in April 2014. The objective of this analysis is to describe the impact of the rotavirus vaccine on hospitalizations among Haitian children younger than 5 years old during the first 5 years after introduction. This analysis includes all children with diarrhea who were enrolled as part of a sentinel surveillance system at two hospitals from May 2013 to April 2019. We compare the proportion of rotavirus-positive specimens in each post-vaccine introduction year to the pre-vaccine period. To account for the potential dilution of the proportion of rotavirus-positive specimens from a waning cholera outbreak, we also analyzed annual trends in the absolute number of positive stools, fit a two-component finite-mixture model to the negative specimens, and fit a negative binomial time series model to the pre-vaccine rotavirus-positive specimens to predict the number of rotavirus diarrhea hospital admissions in the absence of rotavirus vaccination. The overall percentage of rotavirus-positive specimens declined by 22% the first year after introduction, increased by 17% the second year, and declined by 33% to 50% the subsequent 3 years. All sensitivity analyses confirmed an overall decline. We observed a clear annual rotavirus seasonality before and after vaccine introduction, with the greatest activity in December through April, and a biennial pattern, with high sharp peaks and flatter longer periods of increased rotavirus activity in alternating years, consistent with suboptimal vaccination coverage. Overall, our study shows evidence that the introduction of the rotavirus vaccine reduced the burden of severe rotavirus diarrhea.

Background: Rotavirus vaccines are effective in preventing severe rotavirus. Haiti introduced 2-dose monovalent (G1P[8]) rotavirus vaccine recommended for infants at 6 and 10 weeks of age in 2014. We calculated the effectiveness of rotavirus vaccine against hospitalization for acute gastroenteritis in Haiti., Methods: We enrolled children 6-59 months old admitted May 2014-September 2019 for acute watery diarrhea at any sentinel surveillance hospital. Stool was tested for rotavirus using enzyme immunoassay (EIA) and genotyped with multiplex one-step RT-PCR assay and Sanger sequencing for stratification by genotype. We used a case-negative design where cases were children positive for rotavirus and controls were negative for rotavirus. Only children eligible for vaccination were included and a child was considered vaccinated if vaccine was given ≥ 14 days before enrollment. We used unconditional logistic regression to calculate odds ratios and calculated 2-dose and 1-dose vaccine effectiveness (VE) as (1 - odds ratio) * 100., Results: We included 129 (19%) positive cases and 543 (81%) negative controls. Among cases, 77 (60%) were positive for equine-like G3P[8]. Two doses of rotavirus vaccine were 66% (95% CI: 44, 80) effective against hospitalizations due to any strain of rotavirus and 64% (95% CI: 33, 81) effective against hospitalizations due to the equine-like G3P[8] genotype., Conclusions: These findings are comparable to other countries in the Americas region. To the best of our knowledge, this is the first VE estimate both against the equine-like G3P[8] genotype and from a Caribbean country. Overall, these results support rotavirus vaccine use and demonstrate the importance of complete vaccination., Competing Interests: Declaration of Competing Interest At the time of data collection and manuscript preparation Negar Aliabadi was an employee of CDC, but have since taken on employment at Pfizer, Inc. Eyal Leshem received personal consulting fees from Sanofi Pasteur outside the area of work in this study. Mark Katz received consulting fees from Roche in 2018 and 2019 for participation on an advisory board related to Baloxivir. Mark Katz have worked with the Clalit Research Institute, in Tel Aviv, Israel, which has received a grant from Pfizer for work on pneumococcal disease., (Published by Elsevier Ltd.)

Background: The aim of this review was to summarize the data on HIV/AIDS epidemiology and affected populations in Ghana and to describe the United States President's Emergency Plan for Emergency Relief's (PEPFAR) response to the epidemic., Design: We conducted a literature review focusing on PEPFAR's contribution to the HIV response in Ghana. Additionally, we summarized the epidemiology of HIV. We searched both peer-reviewed and grey literature., Setting: Ghana., Results: Overall, HIV prevalence in Ghana is 1.6% with regional variation. Key populations (KPs) are disproportionately affected by HIV in the country. FSW and their clients, and MSM, account for 28% of all new infections. PEPFAR provides technical assistance (TA) to Ghana to maximize the quality, coverage and impact of the national HIV/AIDS response. To ensure adequate supply of antiretrovirals (ARVs), in 2016-2017, PEPFAR invested $23.7 million as a onetime supplemental funding to support Ghana's ARV treatment program. In addition, the National AIDS Control Programme in collaboration with PEPFAR is implementing a scale up of viral load testing. PEPFAR is also implementing a comprehensive package of prevention services in five regions to help reach MSM and FSW and to expand HIV testing services for KPs., Conclusions: Ghana is making changes at both policy and program level in the fight against HIV/AIDS and is working towards achieving the UNAIDS' 90-90-90 targets. PEPFAR is providing TA to ensure these goals can be achieved., Funding: This manuscript has been supported by the U.S. President's Emergency Plan for AIDS Relief (PEPFAR) through the Centers for Disease Control and Prevention (CDC)., Competing Interests: Conflict of interest: None declared

OBJECTIVE To determine the source of a healthcare-associated outbreak of Pantoea agglomerans bloodstream infections. DESIGN Epidemiologic investigation of the outbreak. SETTING Oncology clinic (clinic A). METHODS Cases were defined as Pantoea isolation from blood or catheter tip cultures of clinic A patients during July 2012-May 2013. Clinic A medical charts and laboratory records were reviewed; infection prevention practices and the facility's water system were evaluated. Environmental samples were collected for culture. Clinical and environmental P. agglomerans isolates were compared using pulsed-field gel electrophoresis. RESULTS Twelve cases were identified; median (range) age was 65 (41-78) years. All patients had malignant tumors and had received infusions at clinic A. Deficiencies in parenteral medication preparation and handling were identified (eg, placing infusates near sinks with potential for splash-back contamination). Facility inspection revealed substantial dead-end water piping and inadequate chlorine residual in tap water from multiple sinks, including the pharmacy clean room sink. P. agglomerans was isolated from composite surface swabs of 7 sinks and an ice machine; the pharmacy clean room sink isolate was indistinguishable by pulsed-field gel electrophoresis from 7 of 9 available patient isolates. CONCLUSIONS Exposure of locally prepared infusates to a contaminated pharmacy sink caused the outbreak. Improvements in parenteral medication preparation, including moving chemotherapy preparation offsite, along with terminal sink cleaning and water system remediation ended the outbreak. Greater awareness of recommended medication preparation and handling practices as well as further efforts to better define the contribution of contaminated sinks and plumbing deficiencies to healthcare-associated infections are needed. Infect Control Hosp Epidemiol 2017;38:314-319.

Purpose: The lack of an ongoing surveillance system for hemoglobinopathies in the United States impedes the ability of public health organizations to identify individuals with these conditions, monitor their health-care utilization and clinical outcomes, and understand the effect these conditions have on the health-care system. This article describes the results of a pilot program that supported the development of the infrastructure and data collection methods for a state-based surveillance system for selected hemoglobinopathies., Methods: The system was designed to identify and gather information on all people living with a hemoglobinopathy diagnosis (sickle cell diseases or thalassemias) in the participating states during 2004-2008. Novel, three-level case definitions were developed, and multiple data sets were used to collect information., Results: In total, 31,144 individuals who had a hemoglobinopathy diagnosis during the study period were identified in California; 39,633 in Florida; 20,815 in Georgia; 12,680 in Michigan; 34,853 in New York, and 8,696 in North Carolina., Conclusion: This approach provides a possible model for the development of state-based hemoglobinopathy surveillance systems.

Fungal endophthalmitis is a rare but serious infection. In March 2012, several cases of probable and laboratory-confirmed fungal endophthalmitis occurring after invasive ocular procedures were reported nationwide. We identified 47 cases in 9 states: 21 patients had been exposed to the intraocular dye Brilliant Blue G (BBG) during retinal surgery, and the other 26 had received an intravitreal injection containing triamcinolone acetonide. Both drugs were produced by Franck's Compounding Lab (Ocala, FL, USA). Fusarium incarnatum-equiseti species complex mold was identified in specimens from BBG-exposed case-patients and an unopened BBG vial. Bipolaris hawaiiensis mold was identified in specimens from triamcinolone-exposed case-patients. Exposure to either product was the only factor associated with case status. Of 40 case-patients for whom data were available, 39 (98%) lost vision. These concurrent outbreaks, associated with 1 compounding pharmacy, resulted in a product recall. Ensuring safety and integrity of compounded medications is critical for preventing further outbreaks associated with compounded products.

The effects of transcranial magnetic stimulation (TMS) on hallucination severity and neurocognition were studied in 9 medication-resistant hallucinating patients. A statistically significant improvement was observed on a hallucination scale after 10 days of TMS at the left auditory cortex.

A residents' morning report, adapted from the traditional internal medicine model, was introduced into a psychiatry residency curriculum to supplement bedside teaching for junior residents during inpatient service rotations by focusing on case presentations, clinical reasoning, and evidence-based decision-making skills. This paper describes the design of the report and findings from surveys of residents who participated in the report over a 3-year period. The surveys demonstrate high levels of satisfaction with the exercise and offer insight into factors that may optimize or impede learning in the morning report setting in psychiatry.

Cognitive dysfunction occurs in up to 65% of patients with multiple sclerosis (MS), but there is no effective treatment for the symptoms. The authors conducted a 12-week, open-pilot study to assess the efficacy and tolerability of donepezil HCl administered in patients with MS and cognitive impairment. Seventeen patients at a long-term care facility with Mini-Mental State Examination scores of < or = 25 received 5 mg of donepezil HCl for a 4-week period, followed by 8 weeks of 10 mg of donepezil HCl. Cognitive, neurologic, functional, and behavioral assessments were conducted at baseline and at 4 and 12 weeks. Statistically significant improvement was observed in several cognitive domains including attention, memory, and executive functioning, as well as different aspects of behavior. These data suggest that donepezil HCl merits further study as a potentially viable treatment option for patients with cognitive impairment associated with MS.

Camphor, alpha-pinene (the major component of turpentine), and thujone (a constituent in the liqueur called absinthe) produced an increase in porphyrin production in primary cultures of chick embryo liver cells. In the presence of desferrioxamine (an iron chelator which inhibits heme synthesis and thereby mimics the effect of the block associated with acute porphyria), the terpenes enhanced porphyrin accumulation 5- to 20-fold. They also induced synthesis of the rate-controlling enzyme for the pathway, 5-aminolevulinic acid synthase, which was monitored both spectrophotometrically and immunochemically. These effects are shared by well-known porphyrogenic chemicals such as phenobarbital and glutethimide. Camphor and glutethimide alone led to the accumulation of mostly uro- and heptacarboxylporphyrins, whereas alpha-pinene and thujone resulted in lesser accumulations of porphyrins which were predominantly copro- and protoporphyrins. In the presence of desferrioxamine, plus any of the three terpenes, the major product that accumulated was protoporphyrin. The present results indicate that the terpenes tested are porphyrogenic and hazardous to patients with underlying defects in hepatic heme synthesis. There are also implications for the illness of Vincent van Gogh and the once popular, but now banned liqueur, called absinthe.

Relationships between activities of delta-aminolevulinate synthase and heme oxygenase, respectively the rate-limiting enzymes of heme biosynthesis and degradation, have been studied in chick embryo liver cell cultures following exposure of the cultures to glutethimide and iron, a combination known to produce a synergistic induction of both enzymes. In time-course experiments, synergistic induction of heme oxygenase activity by glutethimide and iron preceded that of delta-aminolevulinate synthase by 4 h. Effects of selective inhibitors of both heme synthesis and degradation have also been studied with respect to effects on delta-aminolevulinate synthase and heme oxygenase activities. The synergistic induction of heme oxygenase by glutethimide and iron appears to be dependent upon cellular heme synthesis because addition of inhibitors of heme biosynthesis, 4,6-dioxoheptanoic acid or N-methyl-mesoporphyrin abolishes this synergistic induction. Exposure of cultures to tin-mesoporphyrin, a potent inhibitor of heme oxygenase, prevented the synergistic induction of delta-aminolevulinate synthase produced by glutethimide and iron, or, when added after induction was already established, promptly halted any further induction. These results suggest that the level of activity of heme oxygenase can reciprocally modulate intracellular heme levels and thus activity of delta-aminolevulinate synthase.

Monospecific polyclonal rabbit antibodies to a purified form of haem oxygenase of chick liver, showing sequence similarity to mammalian haem oxygenase-1, were raised and used to study characteristics of the oxygenase. The antibodies inhibited activity of the purified oxygenase, but not other enzyme components (NADPH:cytochrome reductase and biliverdin reductase) of the standard assay mixture of haem oxygenase. In addition, the antibodies inhibited activity of haem oxygenase in microsomes (microsomal fractions) from Cd(2+)-treated chick liver, spleen, testis and brain. Western (immuno-) blots of microsomal proteins of selected organs from chick, rat and man, and homogenates of chick-embryo liver-cell cultures, probed with the antibodies, showed a major protein with a molecular mass of 33-34 kDa and a lower-molecular-mass protein (28-29 kDa) of variable intensity. Studies with trypsin and selected proteinase inhibitors established that the smaller peptide was a proteolytic product of the larger. Treatment of chick-embryo liver-cell cultures with CdCl2, a potent inducer of haem oxygenase, increased the degree of proteinase-mediated cleavage of the 33 kDa protein to the lower-molecular-mass form. These results indicate that, under at least some conditions, such cultures should be homogenized in the presence of trypsin inhibitor to prevent proteolytic degradation of the enzyme and allow maximal expression of haem oxygenase activity. The antibodies also reacted with haem oxygenase from spleen, testis and brain of both chicks and rats, and the spleen of humans. A method for quantifying the amount of haem oxygenase protein was developed with use of slot-blots and laser densitometry; linearity was observed from 0 to 5 ng of haem oxygenase protein per slot, and the method was applied to sonicated cultured chick-embryo liver cells treated with Cd2+ (0.3 mM) or iron plus glutethimide. In both cases, increases in enzyme activity were of similar magnitude to increases in amounts of enzyme protein. Approximate amounts of haem oxygenase protein in microsomes of several organs from intact animals could also be estimated by the use of slot-blot-laser densitometry, and the amounts measured were increased by the addition of purified haem oxygenase to the microsomal preparations. Results of these studies indicated that haem oxygenase-1 could be detected in microsomes from all chick or rat organs studied, including testis and brain.(ABSTRACT TRUNCATED AT 400 WORDS)

A cDNA from a chick liver library that encodes for haem oxygenase has been cloned and sequenced. Positive clones were identified with monospecific antibodies to the purified enzyme from chick liver and a cDNA of rat haem oxygenase-1. The length of the cDNA is 1258 bases. An open reading frame of 888 bases was identified by comparison of nucleotide and amino acid sequences with those previously identified for haem oxygenase of mammalian or avian origin. The protein corresponding to this fragment of DNA is composed of 296 amino acid residues and has a molecular mass of 33,509 Da, which is similar to that previously estimated for haem oxygenase purified from chick liver. Unequivocal identification of this clone as that complementary to haem oxygenase was provided by (a) comparison of amino acid compositions and partial sequences with those previously established for the purified enzyme, (b) comparison with nucleotide and amino acid sequences for haem oxygenase from rat and human sources and (c) expression in Escherichia coli with production of high levels of mRNA, protein and haem oxygenase activity after exposure of the transfected bacteria to isopropyl beta-D-thiogalactopyranoside (IPTG). Overall, the similarity of chick haem oxygenase to rat and human haem oxygenase (nucleotides 66% and amino acids 62%) is moderately high. The region between proline-129 and alanine-157 is identical in all three enzymes, including histidine-135, which is proposed to play a key role in binding the substrate haem at the active centre of the enzyme. Northern blots also show that treatment of chicks with CdCl2, a potent inducer of haem oxygenase, results in increases in 1.65-1.70 kb mRNA, which hybridizes selectively to the full-length cDNA or to a synthetic 24-base oligonucleotide with sequence identical to that of a portion of the haem oxygenase cDNA. These results suggest that Cd-dependent induction of haem oxygenase is due to increased transcription of the gene or stabilization of its message.

Heme oxygenase, the rate controlling enzyme for heme catabolism, is inducible by a variety of treatments, some of which induce by a heme-dependent mechanism and others by a heme-independent mechanism. This work shows that, in cultured chick embryo liver cells, synergistic induction of heme oxygenase by iron, added with the phenobarbital-like drug, glutethimide was heme-dependent. Addition of an inhibitor of heme biosynthesis abolished the synergistic induction of heme oxygenase providing evidence for the heme-dependent mechanism of induction. Glutethimide and iron appeared to induce at the transcriptional level since both heme oxygenase mRNA and protein levels correlate with changes in heme oxygenase activity.

This study measured the incidence of cause-specific trauma in the Cleveland and Lorain-Elyria Standard Metropolitan Statistical Areas (SMSAs), population 2.2 million, as reported to hospital emergency departments (ED). Cases were selected according to a stratified probability sampling plan (N = 9268). The participating hospitals accounted for 97.6 per cent of 903,346 ED visits in 1977; 52 per cent of these visits were for trauma (ICDA-8 E800-E999). The trauma incidence rate was 197 per 1,000 population. The six leading causes of injury were: falls, 24.4 per cent; cut/piercing injury, 14.2 per cent; striking or struck by object, 13.8 per cent; motor vehicle collisions (MVC), 11.6 per cent; overexertion/strain, 8.2 per cent; and assault, 4.3 per cent. Only falls, MVCs, and assaults were leading causes of both injury and death. The injury incidence rates for vehicular crashes and assault were 1.4 and 3.8 times higher, respectively, than the official incidence rates for these SMSAs. These differences point to a significant underreporting of data needed for public health decision making. Because data were not collected on cases treated outside the participating hospitals, the incidence rates reported here represent a conservative estimate of the magnitude of the problem.

Guinea pigs fed a normal diet show the expected diurnal variation in 3-hydroxy-3-methylglutaryl coenzyme A reductase activity. Vitamin C deficiency, however, suppresses the diurnal peak activity of reductase, due to a decrease in active (unphosphorylated) enzyme. Inhibition of reductase is paralleled by both a fall in hepatic cholesterol synthesis and a rise in serum cholesterol. Incubation of normal guinea pig hepatic microsomes with physiologic and supraphysiologic concentrations of sodium ascorbate also leads to a concentration-dependent inhibition of reductase activity. Thus, dietary extremes of vitamin C may exert similar effects on reductase activity and cholesterolgenesis. Moreover, the changes in enzyme activity induced by ascorbic acid appear to be due in part to a direct effect of the vitamin on the microsomally bound enzyme.

3-Hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase activity in microsomes isolated from cultured lymphoid (IM-9) cells or freshly isolated human leukocytes was markedly decreased by either ascorbic acid or its oxidized derivative, dehydroascorbate. Inhibition of IM-9 leukocyte HMG-CoA reductase activity was log linear between 0.01 and 10 mM ascorbic acid (25 and 81% inhibition, respectively) and 0.1 and 10 mM dehydroascorbate (5 and 75% inhibition, respectively). Inhibition was noncompetitive with respect to HMG-CoA (Km = 10.2 microM (RS); ascorbic acid, Ki = 6.4 mM; dehydroascorbate, Ki = 15 mM) and competitive with respect to NADPH (Km = 16.3 microM; acetic acid, Ki = 6.3 mM; dehydroascorbate, Ki = 3.1 mM). Ascorbic acid and dehydroascorbate are interconverted through the free radical intermediate monodehydroascorbate. Reducing agents are required to convert dehydroascorbate to monodehydroascorbate, but prevent formation of the free radical from ascorbate. In microsomes from IM-9 cells, the reducing agent, dithiothreitol, abolished HMG-CoA reductase inhibition by ascorbate but enhanced inhibition by dehydroascorbate. In addition, the concentration of monodehydroascorbate present in ascorbate solutions was directly proportional to the degree of HMG-CoA reductase inhibition by 1.0 mM ascorbate. Fifty per cent inhibition of enzyme activity occurred at a monodehydroascorbate concentration of 14 microM. These data indicate that monodehydroascorbate mediates inhibition of HMG-CoA reductase by both ascorbate and dehydroascorbate. This effect does not appear to be due to free radical-induced membrane lipid modification, however, since both ascorbate and dehydroascorbate inhibited the protease-solubilized, partially purified human liver enzyme. Since inhibition of HMG-CoA reductase occurs at physiological concentrations of ascorbic acid in the human leukocyte (0.2-1.72 mM), this vitamin may be important in the regulation of endogenous cholesterol synthesis in man.

Methods were developed for determination of human mononuclear leukocyte HMG-CoA reductase protein concentration by a noncompetitive, solid phase, bridged biotin-avidin enzyme immunoassay procedure. Leukocyte microsomal HMG-CoA reductase, first immobilized onto a nitrocellulose filter, is sequentially reacted with 1) monospecific, polyclonal rabbit anti-rat liver HMG-CoA reductase antiserum, which crossreacts with the human liver and leukocyte enzymes; 2) biotinylated donkey anti-rabbit immunoglobulin; 3) a streptavidin-horseradish peroxidase conjugate; and 4) 4-chloro-1-naphthol and H2O2 to visualize the quantity of horseradish peroxidase bound to the immunocomplex. Color development was proportional to the quantity of either purified liver or leukocyte microsomal HMG-CoA reductase applied to the nitrocellulose. Color development was not observed, however, when HMG-CoA reductase was omitted from the nitrocellulose, when one of the reactant species was omitted from the incubation reactions, or when anti-rat liver HMG-CoA reductase antiserum was pre-absorbed with either rat liver or human leukocyte HMG-CoA reductase. Immunoreactivity of microsomal HMG-CoA reductase was independent of the phosphorylation state of the enzyme, but was inversely related to the concentration of thiol-reducing agents present in the microsomal preparation up to 4 mM. Further increases in thiol-reductant failed to produce changes in immunoreactivity. Freshly isolated mononuclear leukocyte microsomal HMG-CoA reductase protein concentration in leukocytes from 31 healthy, normocholesterolemic subjects was a linear function of HMG-CoA reductase activity (R = 0.65; P less than 0.001). The catalytic efficiency of the freshly isolated mononuclear leukocyte enzyme was 313 +/- 34 pmol of mevalonate formed per min of incubation at 37 degrees C per mg immunoreactive protein. This methodology, in conjunction with that recently developed to measure human leukocyte HMG-CoA reductase activity (1984. J. Lipid Res. 25: 967-978), should prove useful in discriminating between HMG-CoA reductase regulatory mechanisms involving changes in enzyme protein concentration and those resulting from changes in enzyme catalytic efficiency.