Childhood adversities and risk of posttraumatic stress disorder and major depression following a motor vehicle collision in adulthood.

AIMS: Childhood adversities (CAs) predict heightened risks of posttraumatic stress disorder (PTSD) and major depressive episode (MDE) among people exposed to adult traumatic events. Identifying which CAs put individuals at greatest risk for these adverse posttraumatic neuropsychiatric sequelae (APNS) is important for targeting prevention interventions. METHODS: Data came from n = 999 patients ages 18-75 presenting to 29 U.S. emergency departments after a motor vehicle collision (MVC) and followed for 3 months, the amount of time traditionally used to define chronic PTSD, in the Advancing Understanding of Recovery After Trauma (AURORA) study. Six CA types were self-reported at baseline: physical abuse, sexual abuse, emotional abuse, physical neglect, emotional neglect and bullying. Both dichotomous measures of ever experiencing each CA type and numeric measures of exposure frequency were included in the analysis. Risk ratios (RRs) of these CA measures as well as complex interactions among these measures were examined as predictors of APNS 3 months post-MVC. APNS was defined as meeting self-reported criteria for either PTSD based on the PTSD Checklist for DSM-5 and/or MDE based on the PROMIS Depression Short-Form 8b. We controlled for pre-MVC lifetime histories of PTSD and MDE. We also examined mediating effects through peritraumatic symptoms assessed in the emergency department and PTSD and MDE assessed in 2-week and 8-week follow-up surveys. Analyses were carried out with robust Poisson regression models. RESULTS: Most participants (90.9%) reported at least rarely having experienced some CA. Ever experiencing each CA other than emotional neglect was univariably associated with 3-month APNS (RRs = 1.31-1.60). Each CA frequency was also univariably associated with 3-month APNS (RRs = 1.65-2.45). In multivariable models, joint associations of CAs with 3-month APNS were additive, with frequency of emotional abuse (RR = 2.03; 95% CI = 1.43-2.87) and bullying (RR = 1.44; 95% CI = 0.99-2.10) being the strongest predictors. Control variable analyses found that these associations were largely explained by pre-MVC histories of PTSD and MDE. CONCLUSIONS: Although individuals who experience frequent emotional abuse and bullying in childhood have a heightened risk of experiencing APNS after an adult MVC, these associations are largely mediated by prior histories of PTSD and MDE.

Prior differences in previous trauma exposure primarily drive the observed racial/ethnic differences in posttrauma depression and anxiety following a recent trauma.

BACKGROUND: Racial and ethnic groups in the USA differ in the prevalence of posttraumatic stress disorder (PTSD). Recent research however has not observed consistent racial/ethnic differences in posttraumatic stress in the early aftermath of trauma, suggesting that such differences in chronic PTSD rates may be related to differences in recovery over time. METHODS: As part of the multisite, longitudinal AURORA study, we investigated racial/ethnic differences in PTSD and related outcomes within 3 months after trauma. Participants (n = 930) were recruited from emergency departments across the USA and provided periodic (2 weeks, 8 weeks, and 3 months after trauma) self-report assessments of PTSD, depression, dissociation, anxiety, and resilience. Linear models were completed to investigate racial/ethnic differences in posttraumatic dysfunction with subsequent follow-up models assessing potential effects of prior life stressors. RESULTS: Racial/ethnic groups did not differ in symptoms over time; however, Black participants showed reduced posttraumatic depression and anxiety symptoms overall compared to Hispanic participants and White participants. Racial/ethnic differences were not attenuated after accounting for differences in sociodemographic factors. However, racial/ethnic differences in depression and anxiety were no longer significant after accounting for greater prior trauma exposure and childhood emotional abuse in White participants. CONCLUSIONS: The present findings suggest prior differences in previous trauma exposure partially mediate the observed racial/ethnic differences in posttraumatic depression and anxiety symptoms following a recent trauma. Our findings further demonstrate that racial/ethnic groups show similar rates of symptom recovery over time. Future work utilizing longer time-scale data is needed to elucidate potential racial/ethnic differences in long-term symptom trajectories.

Clinical and design factors influence the survivorship of custom flange acetabular components.

AIMS: Custom flange acetabular components (CFACs) are a patient-specific option for addressing large acetabular defects at revision total hip arthroplasty (THA), but patient and implant characteristics that affect survivorship remain unknown. This study aimed to identify patient and design factors related to survivorship. PATIENTS AND METHODS: A retrospective review of 91 patients who underwent revision THA using 96 CFACs was undertaken, comparing features between radiologically failed and successful cases. Patient characteristics (demographic, clinical, and radiological) and implant features (design characteristics and intraoperative features) were collected. There were 74 women and 22 men; their mean age was 62 years (31 to 85). The mean follow-up was 24.9 months (sd 27.6; 0 to 116). Two sets of statistical analyses were performed: 1) univariate analyses (Pearson's chi-squared and independent-samples Student's t-tests) for each feature; and 2) bivariable logistic regressions using features identified from a random forest analysis. RESULTS: Radiological failure and revision rates were 23% and 12.5%, respectively. Revisions were undertaken at a mean of 25.1 months (sd 26.4) postoperatively. Patients with radiological failure were younger at the time of the initial procedure, were less likely to have a diagnosis of primary osteoarthritis (OA), were more likely to have had ischial screws in previous surgery, had fewer ischial screw holes in their CFAC design, and had more proximal ischial fixation. Random forest analysis identified the age of the patient and the number of locking and non-locking screws used for inclusion in subsequent bivariable logistic regression, but only age (odds ratio 0.93 per year) was found to be significant. CONCLUSION: We identified both patient and design features predictive of CFAC survivorship. We found a higher rate of failure in younger patients, those whose primary diagnosis was not OA, and those with more proximal ischial fixation or fewer ischial fixation options. Cite this article: Bone Joint J 2019;101-B(6 Supple B):68-76.

The use of dual-mobility bearings in patients at high risk of dislocation.

AIMS: Instability continues to be a troublesome complication after total hip arthroplasty (THA). Patient-related risk factors associated with a higher dislocation risk include the preoperative diagnosis, an age of 75 years or older, high body mass index (BMI), a history of alcohol abuse, and neurodegenerative diseases. The goal of this study was to assess the dislocation rate, radiographic outcomes, and complications of patients stratified as high-risk for dislocation who received a dual mobility (DM) bearing in a primary THA at a minimum follow-up of two years. MATERIALS AND METHODS: We performed a retrospective review of a consecutive series of DM THA performed between 2010 and 2014 at our institution (Hospital for Special Surgery, New York, New York) by a single, high-volume orthopaedic surgeon employing a single prosthesis design (Anatomic Dual Mobility (ADM) Stryker, Mahwah, New Jersey). Patient medical records and radiographs were reviewed to confirm the type of implant used, to identify any preoperative risk factors for dislocation, and any complications. Radiographic analysis was performed to assess for signs of osteolysis or remodelling of the acetabulum. RESULTS: There were 151 patients who met the classification of high-risk according to the inclusion criteria and received DM THA during the study period. Mean age was 82 years old (73 to 95) and 114 patients (77.5%) were female. Mean follow-up was 3.6 years (1.9 to 6.1), with five patients lost to follow-up and one patient who died (for a reason unrelated to the index procedure). One patient (0.66%) sustained an intraprosthetic dislocation; there were no other dislocations. CONCLUSION: At mid-term follow-up, the use of a DM bearing for primary THA in patients at high risk of dislocation provided a stable reconstruction option with excellent radiographic results. Longer follow-up is needed to confirm the durability of these reconstructions.

Medial ball and socket total knee arthroplasty: five-year clinical results.

AIMS: Between 15% and 20% of patients remain dissatisfied following total knee arthroplasty (TKA). The SAIPH knee system (MatOrtho, Surrey, United Kingdom) is a medial ball and socket TKA that has been designed to replicate native knee kinematics in order to maximize the range of movement, stability, and function. This system is being progressively introduced in a stepwise fashion, with this study reporting the mid-term clinical and radiological outcomes. PATIENTS AND METHODS: A retrospective review was undertaken of the first 100 consecutive patients with five-year follow-up following SAIPH TKA performed by the senior authors. The data that were collected included the demographics of the patients, clinical findings, the rate of intraoperative ligamentous release, patient-reported outcome measures (PROMS), radiological assessment, complications, and all-cause revision. Revision data were cross-checked with a national registry. RESULTS: A total of 100 TKAs in 92 patients were included. Three patients died (three TKAs) and a further two TKAs were revised. Of the remaining 95 TKAs, five-year follow-up data were available for 81 TKAs (85%) in 87 patients. There were significant improvements in all PROMs and high satisfaction. The mean ROM at final follow-up was from 0° (full extension) to 124° flexion. There were seven major complications (7%): one infection, two deep vein thromboses, one cerebrovascular event, and two patients with stiffness requiring a manipulation under anaesthesia. Two patients required a lateral retinacular release to optimize patellar tracking in valgus knees; no additional ligament releases were performed in any patient. Radiological analysis demonstrated no evidence of implant-related complications. CONCLUSION: These results demonstrate satisfactory clinical and radiological outcomes at five years following a medial ball and socket TKA. The complication and revision rates are consistent with those previously reported for patients undergoing TKA. These results demonstrate the safety and efficacy of the SAIPH Knee TKA system and support its wider use.

Clinical, functional and radiological outcomes of extracorporeal irradiation in limb salvage surgery for bone tumours.

AIMS: We present a retrospective review of patients treated with extracorporeally irradiated allografts for primary and secondary bone tumours with the mid- and long-term survivorship and the functional and radiographic outcomes. PATIENTS AND METHODS: A total of 113 of 116 (97.4%) patients who were treated with extracorporeally irradiated allografts between 1996 and 2014 were followed up. Forms of treatment included reconstructions, prostheses and composite reconstructions, both with and without vascularised grafts. Survivorship was determined by the Kaplan-Meier method. Clinical outcomes were assessed using the Musculoskeletal Tumor Society (MSTS) scoring system, the Toronto Extremity Salvage Score (TESS) and Quality of Life-C30 (QLQ-30) measures. Radiographic outcomes were assessed using the International Society of Limb Salvage (ISOLS) radiographic scoring system. RESULTS: There were 61 (54%) men with a mean age of 22 years (6 to 70) and 52 (46%) women with a mean age of 26 years (3 to 85). There were 23 deaths. The five-year patient survivorship was 82.3% and the ten-year patient survivorship was 79.6%. The mean follow-up of the 90 surviving patients was 80.3 months (2 to 207). At the last follow-up, 105 allografts (92.9%) were still in place or had been at the time of death; eight (7%) had failed due to infection, local recurrence or fracture. Outcome scores were comparable with or superior to those in previous studies. The mean outcome scores were: MSTS 79% (sd 8); TESS 83% (sd 19); QLQ 82% (sd 16); ISOLS 80.5% (sd 19). Pearson correlation analysis showed a strong relationship between the MSTS and ISOLS scores (r = 0.71, p < 0.001). CONCLUSION: This study shows that extracorporeal irradiation is a versatile reconstructive technique for dealing with large defects after the resection of bone tumours with good functional and radiographic outcomes. Functional outcomes as measured by MSTS, TESS and QLQ-30 were strongly correlated to radiographic outcomes. Cite this article: Bone Joint J 2017;99-B:1681-8.

Matrix-induced autologous chondrocyte implantation in sheep: objective assessments including confocal arthroscopy.

The assessment of cartilage repair has largely been limited to macroscopic observation, magnetic resonance imaging (MRI), or destructive biopsy. The aims of this study were to establish an ovine model of articular cartilage injury repair and to examine the efficacy of nondestructive techniques for assessing cartilage regeneration by matrix-induced autologous chondrocyte implantation (MACI). The development of nondestructive assessment techniques facilitates the monitoring of repair treatments in both experimental animal models and human clinical subjects. Defects (Ø 6 mm) were created on the trochlea and medial femoral condyle of 21 sheep randomized into untreated controls or one of two treatment arms: MACI or collagen-only membrane. Each group was divided into 8-, 10-, and 12-week time points. Repair outcomes were examined using laser scanning confocal arthroscopy (LSCA), MRI, histology, macroscopic ICRS grading, and biomechanical compression analysis. Interobserver analysis of the randomized blinded scoring of LSCA images validated our scoring protocol. Pearson correlation analysis demonstrated the correlation between LSCA, MRI, and ICRS grading. Testing of overall treatment effect independent of time point revealed significant differences between MACI and control groups for all sites and assessment modalities (Asym Sig < 0.05), except condyle histology. Biomechanical analysis suggests that while MACI tissue may resemble native tissue histologically in the early stages of remodeling, the biomechanical properties remain inferior at least in the short term. This study demonstrates the potential of a multisite sheep model of articular cartilage defect repair and its assessment via nondestructive methods.

Laser scanning confocal arthroscopy of a fresh cadaveric knee joint.

OBJECTIVE: Osteoarthritis (OA) inflicts an enormous burden upon sufferers and healthcare systems worldwide. Continuing efforts to elucidate the aetiology of OA have indicated the need for non-destructive methods of in vivo microstructural assessment of articular cartilage (AC). In this study, we describe the first use of a recently developed laser scanning confocal arthroscope (LSCA) to image the cartilage of a fresh frozen cadaveric knee from a patient with OA. DESIGN: Using an adaptation of the International Cartilage Repair Society (ICRS) joint mapping protocol, the joint was divided into three discrete regions (femoral condyle, patella and tibial plateau) for grading according to the ICRS (Outerbridge) system. The LSCA was used to generate images from each area within the three regions. Following imaging, the joint was sectioned and histology was performed on the corresponding sites with histological grading (modified-Mankin). RESULTS: Quantitative results of ICRS, LSCA and histological OA assessment were compared using intraclass correlation (ICC) and Pearson correlation analysis. The LSCA enabled visualisation of chondrocyte morphology and cell density, with classical OA changes such as chondrocyte clustering, surface fibrillation and fissure formation evident. Obvious qualitative similarities between LSCA images and histology were observed, with fair to moderate agreement (P<0.05) demonstrated between modalities. CONCLUSIONS: In this study, we have shown the viability of the LSCA for non-destructive imaging of the microstructure of OA knee cartilage. LSCA technology is potentially a valuable research and clinical tool for the non-destructive assessment of AC microstructure in early to late OA.

Fiber depolymerization.

Depolymerization is, by definition, a crucial process in the reversible assembly of various biopolymers. It may also be an important factor in the pathology of sickle cell disease. If sickle hemoglobin fibers fail to depolymerize fully during passage through the lungs then they will reintroduce aggregates into the systemic circulation and eliminate or shorten the protective delay (nucleation) time for the subsequent growth of fibers. We study how depolymerization depends on the rates of end- and side-depolymerization, k(end) and k(side), which are, respectively, the rates at which fiber length is lost at each end and the rate at which new breaks appear per unit fiber length. We present both an analytic mean field theory and supporting simulations showing that the characteristic fiber depolymerization time tau= square root 1/k(end)k(side) depends on both rates, but not on the fiber length L, in a large intermediate regime 1 << k(side)L(2)/k(end) << (L/d)(2), with d the fiber diameter. We present new experimental data which confirms that both mechanisms are important and shows how the rate of side depolymerization depends strongly on the concentration of CO, acting as a proxy for oxygen. Our theory remains rather general and could be applied to the depolymerization of an entire class of linear aggregates, not just sickle hemoglobin fibers.

Confocal laser scanning microscopy in orthopaedic research.

Confocal laser scanning microscopy (CLSM) is a type of high-resolution fluorescence microscopy that overcomes the limitations of conventional widefield microscopy and facilitates the generation of high-resolution 3D images from relatively thick sections of tissue. As a comparatively non-destructive imaging technique, CLSM facilitates the in situ characterization of tissue microstructure. Images generated by CLSM have been utilized for the study of articular cartilage, bone, muscle, tendon, ligament and menisci by the foremost research groups in the field of orthopaedics including those teams headed by Bush, Errington, Guilak, Hall, Hunziker, Knight, Mow, Poole, Ratcliffe and White. Recent evolutions in techniques and technologies have facilitated a relatively widespread adoption of this imaging modality, with increased "user friendliness" and flexibility. Applications of CLSM also exist in the rapidly advancing field of orthopaedic implants and in the investigation of joint lubrication.

Gestational diabetes and its impact on the neonate.

Gestational diabetes mellitus is a relatively common medical condition that was described as early as the nineteenth century. This article discusses the maternal and fetal pathophysiology and the impact of the maternal condition on the neonate. Fetal macrosomia and infant respiratory distress syndrome, cardiomyopathy, hypoglycemia, hypocalcemia, hypomagnesemia, polycythemia, and hyperviscosity all can occur as a result of maternal hyperglycemia and are discussed in detail. Therapeutic approaches and treatment options for the mother, manifestations and diagnosis of the infant, and current research related to this condition are also included.

Characterisation of a binding-protein-dependent, active transport system for short-chain amides and urea in the methylotrophic bacterium Methylophilus methylotrophus.

Three genes (fmdCAB) encoding an outer-membrane porin for short-chain amides and urea, formamidase, and a putative regulatory protein in Methylophilus methylotrophus have previously been cloned and characterised. Three genes have now been identified downstream of fmdB, viz fmdD encoding a hydrophilic protein containing an N-terminal signal sequence, and fmdEF encoding hydrophobic transmembrane proteins. The derived amino acid sequence of mature FmdD (predicted molecular mass 41,870 Da) was similar to the cytoplasmic, amide-binding protein (AmiC) from Pseudomonas aeruginosa and to several periplasmic, solute-binding proteins from other bacteria. Mature FmdD was purified and shown to be a monomer (40-45 kDa) with the predicted N-terminal amino acid sequence (ADYPTA-). Equilibrium dialysis showed that the purified protein bound short-chain amides and urea with high affinity (Kd 7.2 microM for [14C]urea). SDS/PAGE and western blotting using antiserum to mature FmdD showed it was induced by short-chain amides and urea, and repressed by excess ammonia. The derived amino acid sequences of FmdE (32,822 Da) and FmdF (incomplete; >25,435 Da) were similar to the transmembrane proteins BraD/LivH and BraE/LivM, respectively, in various leucine/isoleucine/valine transport systems. Uptake of [14C]urea by washed cells was inhibited by the uncoupling agent carbonyl cyanide p-trifluoromethoxyphenylhydrazone and unlabelled formamide. It is concluded that FmdDEF comprise part of a high-affinity, binding-protein-dependent active-transport system for short-chain amides and urea in M. methylotrophus.

Poly-3-hydroxybutyrate production by washed cells of Alcaligenes eutrophus; purification, characterisation and potential regulatory role of citrate synthase.

Washed cells prepared from carbon-limited continuous cultures of Alcaligenes eutrophus synthesised poly-3-hydroxybutyrate (PHB) rapidly when supplied with glucose, DL-lactate or L-lactate. Unlike growing cultures, washed cells excreted significant amounts of pyruvate. The combined rates of PHB production (qPHB) and pyruvate excretion (qPyr) were linearly related to the rate of carbon substrate utilisation (qS), showing that washed cells behaved similarly to growing cultures when corrected for the absence of non-PHB biomass production. The addition of formate (as a potential source of NADH and/or ATP) significantly stimulated both qPHB and qPyr, but slightly decreased qS and substantially decreased the flux of carbon through the tricarboxylic acid cycle (qTCA). Citrate synthase activity of broken cells was inhibited by physiological concentrations of NADH, but not of ATP, in a manner that was not reversible by AMP. Citrate synthase was purified and shown to be a "large" form of the enzyme (Mr 227,000), comprising a single type of subunit (Mr 47,000) as found in several other gram-negative aerobes. The potential role of citrate synthase in the regulation of PHB production via its ability to control carbon flux into the tricarboxylic acid cycle is discussed.

A post-marketing surveillance study of Voltarol 75 mg SR in the primary care setting.

A total of 7438 patients suffering from a wide variety of painful conditions was included in the final analysis of a post-marketing surveillance (PMS) study monitoring the use of Voltarol 75 mg SR in a primary care setting. Follow-up data were collected at visits conducted one, four and 12 months after the initial consultation. Improvement of symptoms was the most common reason for discontinuation of treatment (47% of patients who discontinued). Adverse events led to the withdrawal of 18% of patients overall. The rate of serious gastrointestinal complications was low (0.4%) and deleterious hepatic or renal effects were not apparent. There were significantly more events experienced by female patients and there was a significant effect of age on severe gastrointestinal events. In this PMS study Voltarol 75 mg SR was used successfully once or twice daily without any unexpected adverse effects in a manner consistent with current recommendations for the use of non-steroidal anti-inflammatory drugs.

Molecular characterisation of formamidase from Methylophilus methylotrophus.

A 3.2-kbp PstI fragment of DNA encoding formamidase from the methylotrophic bacterium Methylophilus methylotrophus which had previously been cloned (pNW3) [Wyborn, N.R., Scherr, D.J. & Jones, C.W. (1994) Microbiology 140, 191-195], was subcloned as a 2.3 kbp HindIII fragment (pNW323). Nucleotide sequencing showed that the subclone contained two genes which encoded formamidase (fmdA) and a possible regulatory protein (fmdB). Predicted molecular masses for FmdA and FmdB were 44438 Da (compared with approximately 44500 Da by electrospray mass spectrometry and 51000 Da by SDS/PAGE of the purified enzyme) and 12306 Da, respectively. The derived amino acid sequence of formamidase was supported by N-terminal amino acid sequencing of the enzyme and of proteolytic fragments prepared from it using V8 endoproteinase and was 57% similar to that of the acetamidase from Mycobacterium smegmatis. The structural similarities between these two enzymes, and their existence as a separate class of bacterial amidase, were confirmed by immunological investigations.

Agrobacterium radiobacter and related organisms take up fructose via a binding-protein-dependent active-transport system.

Washed cells of Agrobacterium radiobacter prepared from a fructose-limited continuous culture (D 0.045 h-1) transported D(-)[U-14C]fructose in a linear manner for up to 4 min at a rate several-fold higher than the rate of fructose utilization by the growing culture. D(-)[U-14C]Fructose transport exhibited a high affinity for fructose (KT < 1 microM) and was inhibited to varying extents by osmotic shock, by the uncoupling agent carbonyl cyanide p-trifluoromethoxyphenylhydrazone, and by unlabelled sugars (D-fructose/D-mannose > D-ribose > D-sorbose > D-glucose/D-galactose/D-xylose; no inhibition by D-arabinose). Prolonged growth of A. radiobacter in fructose-limited continuous culture led to the selection of a novel strain (AR100) which overproduced a fructose-binding protein (FBP) and showed an increased rate of fructose transport. FBP was purified from osmotic-shock fluid using anion-exchange fast protein liquid chromatography (FPLC). The monomeric protein (M(r) 34,200 by SDS-PAGE and 37,700 by gel-filtration FPLC) bound D-[U-14C]-fructose stoichiometrically (1.17 nmol nmol FBP-1) and with high affinity (KD 0.49 microM) as shown by equilibrium dialysis. Binding of D-[U-14C]fructose by FBP was variably inhibited by unlabelled sugars (D-fructose/D-mannose > D-ribose > D-sorbose; no inhibition by D-glucose, D-galactose or D-arabinose). The N-terminal amino acid sequence of FBP (ADTSVCLI-) was similar to that of several sugar-binding proteins from other species of bacteria. Fructose transport and FBP were variably induced in batch cultures of A. radiobacter by growth on different carbon sources (D-fructose > D-ribose/D-mannose > D-glucose; no induction by succinate). An immunologically similar protein to FBP was produced by Agrobacterium tumefaciens and various species of Rhizobium following growth on fructose. It is concluded that fructose is transported into A. radiobacter and related organisms via a periplasmic fructose/mannose-binding-protein-dependent active-transport system, in contrast to the phosphotransferase system used by many other species of bacteria.

The effect of nutrient limitation on glycerol uptake and metabolism in continuous cultures of Pseudomonas aeruginosa.

Pseudomonas aeruginosa NM48, a non-mucoid derivative of an alginate-producing strain isolated from a cystic fibrosis patient, was grown in batch culture with glycerol, glucose or succinate as carbon source, and in continuous culture (D 0.05 h-1) under glycerol or glucose limitation. Glycerol uptake, glycerol kinase and glycerol-3-phosphate dehydrogenase were induced by glycerol, but not by glucose or succinate. Linear uptake of [14C]glycerol by washed cells (Km < or = 2 microM) was inhibited by unlabelled glycerol and glyceraldehyde, but not by cyanide or the uncoupling agent carbonyl cyanide p-trifluoromethoxyphenylhydrazone (FCCP), and was accompanied by substantial intracellular accumulation of glycerol-3-phosphate and/or dihydroxyacetone phosphate but not glycerol. Prolonged growth under glycerol limitation led to substantial increases in the activities and/or concentrations of the enzymes catalysing glycerol uptake and metabolism, together with a 48,000 M(r) outer-membrane protein which was also over-expressed following prolonged growth under glucose limitation. The N-terminal amino acid sequence (AEAFSPN-) and electrophoretic properties of this protein were the same as those of the previously characterized glucose porin (OprB) from P. aeruginosa, indicating that this porin is active with both glucose and glycerol. It is concluded that during growth under glycerol limitation, glycerol is transported into P. aeruginosa NM48 via OprB and a high-affinity, binding-protein-independent facilitated-diffusion system.

Interspecific comparisons of the unusually long 5' leader of the Drosophila ecdysone-inducible gene E74A.

The Drosophila melanogaster E74A gene is expressed in response to the steroid hormone ecdysone. Its product is a site-specific DNA-binding protein that is believed to play an important role in the normal development and metamorphosis of the fly. In addition to being under the transcriptional control of ecdysone and its receptor, there is evidence for translational regulation of the E74A mRNA. The E74A mRNA 5' leader is unusually long (nearly 1.9 kb) and contains 17 AUGs, suggesting that translation of the mRNA would be rather inefficient. To identify features and sequences that might be important in regulating E74A translation, we determined the nucleotide sequences of the 5' leaders from the E74A-homologous genes of D. pseudoobscura and D. virilis, comparing them to D. melanogaster. Several conserved characteristics and specific sequences were identified. In addition to conservation of the relative distances separating the three E74A gene exons encoding the 5' leader, all three species have maintained the extensive length of the leader as well as multiple AUGs. Within the 5' leader are numerous conserved sequences, several of which are found at the two ends of the leader and at splice site junctions, where sequence conservation might be expected. A 53-nucleotide sequence element and a 30-nucleotide element in exon 2 are highly conserved and are proposed as candidates for an internal ribosome entry site (IRES) of the E74A mRNA.

The reliability of a solid-phase extraction system for the analysis of benzoylecgonine in urine.

Gas chromatographic-mass spectrometric (GC-MS) analysis for benzoylecgonine (BE), a metabolite of cocaine, requires an initial extraction from urine. Although liquid-liquid extraction methods are frequently used, solid-phase extraction (SPE) may be preferable for obtaining reliable results and clean chromatograms. We describe a 12-month study that evaluates the accuracy, precision, variability between analysts, variability between column lots, and cleanliness of BE extracts using SPE columns followed by GC-MS analysis. The overall mean for a control urine sample prepared at 150 ng/mL is 151 ng/mL (N = 293) with a standard deviation of 8.59 and a coefficient of variation (CV) of 5.7%. Within-run precision (measured as CV) at 75, 150, and 2000 ng/mL is 4.0, 1.8, and 0.8%, respectively. Mean results from 10 different analysts vary a maximum of 4.6% from the overall mean of 151 ng/mL, and the CV for 9 out of 10 analysts is 7.0% or less. The CV for the remaining analyst is 10.4%. Quantitative results from nine different lots of SPE columns fluctuate 3.3% from the overall mean of 151 ng/mL, and the CV varies from 3.5 to 6.2%. GC-MS chromatograms following SPE are significantly cleaner (i.e., reduced baseline signal and no interfering peaks) than those from two types of liquid-liquid extractions.

Chemical compatibility of regional anesthetic drug combinations.

OBJECTIVE: To define the physical and chemical compatibilities of several classes of drugs that may be used in combination for peridural anesthesia. DESIGN: Morphine, fentanyl, bupivacaine, lidocaine, tetracaine, ketamine, and clonidine were admixed for one hour in five groups of three-drug combinations, plus one group of all seven drugs. The combinations were inspected macroscopically and microscopically to determine physical compatibility. The admixtures were evaluated by gas chromatography/mass spectroscopy (GC/MS) and compared with known standards to determine chemical compatibility. RESULTS: The admixtures showed no physical incompatibility on microscopic or macroscopic evaluation. Chemical compatibility of all mixtures was confirmed by GC/MS. Ion chromatograms of the drugs in admixtures were identical to previously established standards. CONCLUSIONS: The agents evaluated demonstrated physical and chemical compatibility under conditions that would be observed during the administration of peridural anesthesia. Combinations of these drugs therefore could be safely admixed for use in anesthesia.