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Cytochrome P450 (CYP450) is a group of enzymes that play an essential role in Phase I metabolism, with 57 functional genes classified into 18 families in the human genome, of which the CYP1, CYP2, and CYP3 families are prominent. Beyond drug metabolism, CYP enzymes metabolize endogenous compounds such as lipids, proteins, and hormones to maintain physiological homeostasis. Thus, dysregulation of CYP450 enzymes can lead to different endocrine disorders. Moreover, CYP450 enzymes significantly contribute to fatty acid metabolism, cholesterol synthesis, and bile acid biosynthesis, impacting cellular physiology and disease pathogenesis. Their diverse functions emphasize their therapeutic potential in managing hypercholesterolemia and neurodegenerative diseases. Additionally, CYP450 enzymes are implicated in the onset and development of illnesses such as cancer, influencing chemotherapy outcomes. Assessment of CYP450 enzyme expression and activity aids in evaluating liver health state and differentiating between liver diseases, guiding therapeutic decisions, and optimizing drug efficacy. Understanding the roles of CYP450 enzymes and the clinical effect of their genetic polymorphisms is crucial for developing personalized therapeutic strategies and enhancing drug responses in diverse patient populations.

Neuroendocrine (NE) prostate cancer (NEPC) is a lethal subtype of castration-resistant prostate cancer (PCa) arising either de novo or from transdifferentiated prostate adenocarcinoma following androgen deprivation therapy (ADT). Extensive computational analysis has identified a high degree of association between the long noncoding RNA (lncRNA) H19 and NEPC, with the longest isoform highly expressed in NEPC. H19 regulates PCa lineage plasticity by driving a bidirectional cell identity of NE phenotype (H19 overexpression) or luminal phenotype (H19 knockdown). It contributes to treatment resistance, with the knockdown of H19 re-sensitizing PCa to ADT. It is also essential for the proliferation and invasion of NEPC. H19 levels are negatively regulated by androgen signaling via androgen receptor (AR). When androgen is absent SOX2 levels increase, driving H19 transcription and facilitating transdifferentiation. H19 facilitates the PRC2 complex in regulating methylation changes at H3K27me3/H3K4me3 histone sites of AR-driven and NEPC-related genes. Additionally, this lncRNA induces alterations in genome-wide DNA methylation on CpG sites, further regulating genes associated with the NEPC phenotype. Our clinical data identify H19 as a candidate diagnostic marker and predictive marker of NEPC with elevated H19 levels associated with an increased probability of biochemical recurrence and metastatic disease in patients receiving ADT. Here we report H19 as an early upstream regulator of cell fate, plasticity, and treatment resistance in NEPC that can reverse/transform cells to a treatable form of PCa once therapeutically deactivated., (© 2021. The Author(s).)

Background: Telemedicine for preanesthesia evaluation can decrease access disparities by minimizing commuting, time off work, and lifestyle disruptions from frequent medical visits. We report our experience with the first 120 patients undergoing telemedicine preanesthesia evaluation at Moffitt Cancer Center., Methods: This is a retrospective analysis of 120 patients seen via telemedicine for preanesthesia evaluation compared with an in-person cohort meeting telemedicine criteria had it been available. Telemedicine was conducted from our clinic to a patient's remote location using video conferencing. Clinic criteria were revised to create a tier of eligible patients based on published guidelines and anesthesiologist consensus., Results: Day-of-surgery cancellation rate was 1.67% in the telemedicine versus 0% in the in-person cohort. The two telemedicine group cancellations were unrelated to medical workup, and cancellation rate between the groups was not statistically significant ( P = .49). Median round trip distance and time saved by the telemedicine group was 80 miles [range 4; 1180] and 121 minutes [range 16; 1034]. Using the federal mileage rate, the median cost savings was $46 [range $2.30; 678.50] per patient. Patients were similar in gender and race in both groups ( P = .23 and .75, respectively), but the in-person cohort was older and had higher American Society of Anesthesiologists physical status classification ( P = .0003)., Conclusions: Telemedicine preanesthesia evaluation results in time, distance, and financial savings without increased day-of-surgery cancellations. This is useful in cancer patients who travel significant distances to specialty centers and have a high frequency of health care visits. American Society of Anesthesiologists Physical Status classification and age differences between cohorts indicate possible patient or provider selection bias. Randomized controlled trials will aid in further exploring this technology.

Introduction: Carpal Tunnel Questionnaire (CTQ) is widely used for assessing condition-specific impairments in individuals with carpal tunnel syndrome (CTS) or for assessing outcomes after carpal tunnel surgery (carpal tunnel release [CTR]). A systematic review of its measurement properties can greatly facilitate its evidence-based use in clinical practice. The purpose of this study was to systematically locate, appraise, and synthesize the evidence concerning the reliability, responsiveness, validity, minimal detectable change (MDC), and minimal clinically important difference (MCID) for the CTQ and its scales., Study Design: This is a systematic review of measurement properties., Methods: Using predefined keywords, PubMed, CINAHL, PsychInfo, and ProQuest were searched to locate primary studies that assessed measurement properties of the CTQ. The methodological quality of the included studies was assessed using a standardized tool. Data concerning the measurement properties were extracted and synthesized. The pooled estimates for the indices of test-retest reliability, standard error of measurement, responsiveness, MDC, and MCID were calculated from the included studies., Results: A total of 34 articles were deemed eligible and included in this review. The methodological quality of these 34 studies was generally good. Most studies suggested that the CTQ and its scales had good test-retest reliability and internal consistency. However, few studies found that the Symptom Severity Scale had more than one factor. The responsiveness of the CTQ and its scales was excellent across the studies. The pooled estimates for the MDC 90 and MCID for Symptom Severity Scale/Functional Status Scale were 0.72/0.79 and 1.05/1.13, respectively., Discussion: The results of this review support the use of CTQ and its scales in assessing conditions-specific impairments in individuals with CTS or after CTR. However, an effort should be made to review and modify the content of the symptom severity scale due to multiple reports challenging its unidimensional structure., Conclusions: The totality of evidence emerging from this systematic review suggests that the CTQ and its scales provide reliable and valid estimate of impairments resulting from CTS or after CTR., (Copyright © 2020 Hanley & Belfus. Published by Elsevier Inc. All rights reserved.)

Gynecologic brachytherapy procedures require targeted procedural and anesthetic needs including optimization of intra- and post-procedure analgesia, low rate of complications, and appropriate and timely transitions of care. It is uncertain whether neuraxial or general anesthesia is superior for these and other anesthetic outcomes. After a targeted search of the recent literature for anesthesia and analgesia studies for gynecologic brachytherapy, twenty studies were identified and appraised for potential review. Meta-analysis showed a decreased frequency in rescue analgesic administration in patients who underwent neuraxial anesthesia compared with general anesthesia for the procedure and literature review showed a comparable rate of anesthesia-related complications. Neuraxial anesthesia may be considered for gynecologic brachytherapy because of improved pain control, decreased opioid consumption, and similar rate of anesthesia complications., (Copyright © 2020 American Brachytherapy Society. Published by Elsevier Inc. All rights reserved.)

After traumatic brain injury, decompressive craniectomy (DC) is a second-tier, late therapy for refractory intracranial hypertension. We hypothesize that early DC, based on CT evidence of intracranial hypertension, improves intracranial pressure (ICP) and cerebral perfusion pressure (CPP). From September 2008 to January 2015, 286 traumatic brain injury patients requiring invasive ICP monitoring at a single Level I trauma center were reviewed. DC and non-DC patients were propensity score matched 1:1, based on demographics, hemodynamics, injury severity score (ISS), Glasgow Coma Scale (GCS), transfusion requirements, and need for vasopressor therapy. Data are presented as M ± SD or median (IQR) and compared at P ≤ 0.05. The study population was 42 ± 17 years, 84 per cent male, ISS = 29 ± 11, GCS = 6(5), length of stay (LOS) = 32(40) days, and 28 per cent mortality. There were 116/286 (41%) DC, of which 105/116 (91%) were performed at the time of ICP placement. For 50 DC propensity matched to 50 non-DC patients, the midline shift was 7(11) versus 0(5) mm (P < 0.001), abnormal ICP (hours > 20 mm Hg) was 1(10) versus 8(16) (P = 0.017), abnormal CPP (hours < 60 mm Hg) was 0(6) versus 4(9) (P = 0.008), daily minimum CPP (mm Hg) was 67(13) versus 62(17) (P = 0.010), and daily maximum ICP (mm Hg) was 18(9) versus 22(11) (P < 0.001). However, LOS [33(37) versus 25(34) days], mortality (24 versus 30%), and Glasgow Outcome Score Extended [3.0(3.0) versus 3.0(4.0)] did not improve significantly. Early DC for CT evidence of intracranial hypertension decreased abnormal ICP and CPP time and improved ICP and CPP thresholds, but had no obvious effect on the outcome.

Arginine vasopressin (AVP) is often used as an alternative pressor to catecholamines (CATs). However, unlike CATs, AVP is a powerful antidiuretic that could promote edema. We tested the hypothesis that AVP promoted cerebral edema and/or increased requirements for osmotherapy, relative to those who received CATs, for cerebral perfusion pressure (CPP) management after traumatic brain injury (TBI). This is a retrospective review of 286 consecutive TBI patients with intracranial pressure monitoring at a single institution from September 2008 to January 2015. Cerebral edema was quantitated using CT attenuation in prespecified areas of gray and white matter., Results: To maintain CPP >60 mm Hg, 205 patients required no vasopressors, 41 received a single CAT, 12 received AVP, and 28 required both. Those who required no pressors were generally less injured; required less hyperosmolar therapy and less total fluid; and had lower plasma Na, lower intracranial pressure, less edema, and lower mortality (all P < 0.05). Edema; daily mean, minimum, and maximum Na levels; and mortality were similar with AVP versus CATs, but the daily requirement of mannitol and 3 per cent NaCl were reduced by 45 and 35 per cent (both P < 0.05). In patients with TBI who required CPP therapy, AVP reduced the requirements for hyperosmolar therapy and did not delay resolution or increase cerebral edema compared with CATs.

Aim: Development of a sensitive method for DNA methylation profiling and associated mutation detection in clinical samples., Materials & Methods: Formalin-fixed and paraffin-embedded tumors received by clinical laboratories often contain insufficient DNA for analysis with bisulfite or methylation sensitive restriction enzymes-based methods. To increase sensitivity, methyl-CpG DNA capture and Coupled Abscription PCR Signaling detection were combined in a new assay, MethylMeter(®). Gliomas were analyzed for MGMT methylation, glioma CpG island methylator phenotype and IDH1 R132H., Results: MethylMeter had 100% assay success rate measuring all five biomarkers in formalin-fixed and paraffin-embedded tissue. MGMT methylation results were supported by survival and mRNA expression data., Conclusion: MethylMeter is a sensitive and quantitative method for multitarget DNA methylation profiling and associated mutation detection. The MethylMeter-based GliomaSTRAT assay measures methylation of four targets and one mutation to simultaneously grade gliomas and predict their response to temozolomide. This information is clinically valuable in management of gliomas., Competing Interests: Financial & competing interests disclosure This work was supported in part by funding from the NIH through NCI Small Business Innovative Grant 1R43CA132851 and NCI Contract HHSN261200900047C to RiboMed. D McCarthy and MM Hanna are employees of Ribomed Biotechnologies, Inc. and own Ribomed stock. OR Diago, DJ Hogan, and D Ostertag are employees of Tocagen Inc and own Tocagen stock. The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed. No writing assistance was utilized in the production of this manuscript.

U.S. Army Forward Surgical Teams (FSTs) are elite, multidisciplinary units that are highly mobile, and rapidly deployable. The mission of the FST is to provide resuscitative and damage control surgery for stabilization of life-threatening injuries in austere environments. The Army Trauma Training Center began in 2001 at the University of Miami Ryder Trauma Center under the direction of COL T. E. Knuth, MC USA (Ret.), as a multimodality combination of lectures, laboratory exercises, and clinical experiences that provided the only predeployment mass casualty and clinical trauma training center for all FSTs. Each of the subsequent five directors has restructured the training based on dynamic feedback from trainees, current military needs, and on the rapid advances in combat casualty care. We have highlighted these evolutionary changes at the Army Trauma Training Center in previous reviews. Under the current director, LTC J. M. Seery, MC USA, there are new team-building exercises, mobile learning modules and simulators, and other alternative methods in the mass casualty exercise. This report summarizes the latest updates to the state of the art training since the last review., (Reprint & Copyright © 2016 Association of Military Surgeons of the U.S.)

Background: This study assessed hemostatic function in cancer patients at high risk for venous thromboembolism., Methods: Thirty-eight female patients (age, 53 ± 9 years) undergoing immediate postmastectomy reconstruction were prospectively studied with informed consent. Blood was sampled preoperatively, on postoperative day 1, and at 1 week follow-up. Rotational thromboelastography clotting time, α-angle (clot kinetics), clot formation time, and maximum clot firmness were studied with three different activating agents: intrinsically activated test using ellagic acid, extrinsically activated test with tissue factor, and fibrin-based extrinsically activated test with tissue factor and the platelet inhibitor cytochalasin D. Thromboprophylaxis was unfractionated heparin plus sequential compression devices if not contraindicated. Hypercoagulability was defined by one or more parameters outside the reference range., Results: Preoperatively, 29 percent of patients were hypercoagulable, increasing to 67 percent by week 1 (p = 0.017). Clotting time, clot formation time, and α-angle remained relatively constant over time, but maximum clot formation increased in intrinsically activated test using ellagic acid, extrinsically activated test with tissue factor, and fibrin-based extrinsically activated test with tissue factor and the platelet inhibitor cytochalasin D (all p < 0.05). Body mass index was 28 ± 5 kg/m, 23 percent received preoperative chemotherapy, and 15 percent had a history of tobacco use, but there was no association between these risk factors and hypercoagulability., Conclusions: Despite perioperative thromboprophylaxis, two-thirds of patients undergoing combined tumor resection and reconstructive surgery for breast cancer were hypercoagulable 1 week after surgery. Hypercoagulability was associated with increased clot strength mediated by changes in platelet and fibrin function., Clinical Question/level of Evidence: Therapeutic, IV.

Background: Delayed gastric emptying (DGE) remains an unsolved complication after pancreaticoduodenectomy (PD) with conflicting reports of its cause. We aimed to compare the effect of surgical techniques involving the stomach in PD in lowering the risk of postoperative DGE., Methods: Online search and review of key bibliographies in PubMed, Medline, Embase, Scopus, Cochrane, and Google Scholar was performed. Studies comparing PD surgical techniques were identified. Primary outcome was postoperative DGE. Methodological quality was assessed using Strengthening the Reporting of Observational Studies in Epidemiology and Consolidated Standards of Reporting Trials. Calculated pooled relative risk and odds ratios (ORs) with the corresponding 95% confidence interval (CI) were used in the meta-analyses., Results: Overall, 376 studies were reviewed, of which 22 studies were selected including a total of 5172 patients. The incidence of DGE was lower in antecolic compared with retrocolic gastrojejunostomy (risk ratio [RR], 0.260; CI, 0.157-0.431; P < 0.001; n = 1067 patients) and in subtotal stomach preserving PD compared with pylorus preserving PD (RR, 0.527; CI, 0.363-0.763; P < 0.001; n = 663 patients). There was no significant difference between classic PD versus pylorus preserving PD (OR, 0.64; CI, 0.40-1.00; P = 0.05; n = 1209 patients), pancreaticogastrostomy versus pancreaticojejunostomy (RR, 1.02; CI, 0.62-1.68; P = 0.94; n = 961 patients), Roux-en-Y versus Billroth II gastrojejunostomy (RR, 0.946; CI, 0.788-1.136; P = 0.5513; n = 470 patients), or minimally invasive PD versus open PD (OR, 0.99; CI, 0.62-1.56; P = 0.96; n = 802)., Conclusions: In PD, surgical techniques using antecolic reconstruction route and subtotal stomach preserving PD seem to be associated with a lower risk of DGE. Further randomized controlled trials are necessary to evaluate these results taking other causes into consideration., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Background: The best gastrointestinal reconstruction route after pylorus preserving pancreaticoduodenectomy remains debatable. We aimed to evaluate the incidence of delayed gastric emptying (DGE) after antecolic (AC) and retrocolic (RC) duodenojejunostomy in these patients., Data Sources: Studies comparing AC to RC reconstruction after pylorus preserving pancreaticoduodenectomy were identified from literature databases (PubMed, MEDLINE, EMBASE, SCOPUS, and Cochrane). The meta-analysis included 10 studies with a total of 1,067 patients, where 504 patients underwent AC and 563 patients underwent RC reconstruction. The incidence of DGE was significantly lower with AC reconstruction in both randomized controlled trials (risk ratio = .44, confidence interval = .24 to.77, P = .005) and retrospective studies (risk ratio .21, confidence interval .14 to .30, P < .001) with less output and days of nasogastric tube use. AC reconstruction was associated with a decreased length of stay. There was no difference in operative time, blood loss, pancreatic fistula, and abdominal abscess/collections., Conclusions: AC reconstruction seems to be associated with less DGE, with no association with pancreatic fistula or abscess formation., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Background: A massive transfusion protocol (MTP) presents a logistical challenge for most blood banks and trauma centers. We compare the ratio of packed red blood cells (PRBC) and plasma transfused over serial time points in those requiring MTP (10-30 U PRBC/24 h) to those requiring "super" MTP (S-MTP; >30 U PRBC/24 h) and test the hypothesis that changes in allocation of blood products with use of readily transfusable liquid plasma (LP) improves the ratio of PRBC and plasma during S-MTP., Materials and Methods: All transfused trauma patients (n = 1305) from January 01, 2009-April, 03, 2015 were reviewed. PRBC:plasma ratio was compared for MTP (n = 277) and S-MTP (n = 61) patients, before and after the availability of LP at our institution. Data are reported as mean ± standard deviation or median (interquartile range)., Results: Age was 41 ± 19 y, 52% blunt mechanism, injury severity score 32 ± 16, and 46.3% mortality. In 24 h, requirements were 17 (14) U PRBC and 10 (11) U plasma, with a PRBC:plasma of 1.6 (0.8). Within the first hour, PRBC:plasma for S-MTP versus MTP was 2.1:1 versus 1.7:1 (P = 0.017). With LP, S-MTP patients received significantly lower PRBC:plasma at the first hour (P < 0.001). Before institutional changes, PRBC:plasma positively correlated with PRBC transfused at hour 1 (r = 0.410, R(2) = 0.168, P < 0.001); after institutional changes and the advent of LP, there was no correlation (r = 0.177, R(2) = 0.031, P = 0.219)., Conclusions: Within the first hour of transfusion, units of PRBC transfused positively correlated with PRBC:plasma, and patients receiving S-MTP had higher PRBC:plasma than those receiving MTP. Changes in our institution's MTP protocol to include LP improved the early PRBC:plasma transfused in patients requiring S-MTP., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Background: As the population ages, mortality from falls will soon exceed that from all other forms of injury. Tremendous resources are focused on this problem, but how these patients die is unclear. To fill this gap, we tested the hypothesis that falls among the elderly are related to patient, rather than to injury factors when compared with falls among younger adults., Methods: From January 2002 to December 2012, 7,293 fall admissions were reviewed. Data are reported as mean ± SD if normally distributed or median (interquartile range) if not., Results: In 2002 to 2007, 25% of all falls were in elderly patients (≥65 years), but in 2008 to 2012, this proportion increased to 30% (p < 0.001). When comparing adult (n = 5,216) with elderly (n = 2,077) admissions, characteristics were as follow: Injury Severity Score (ISS) of 8 (4-13) versus 9 (5-17), length of stay (in days) of 3 (1-7) versus 6 (2-11), and mortality of 3.8% versus 13.7% (all p < 0.001). After controlling for variables associated with mortality using multiple logistic regression, elderly age was the strongest independent predictor of mortality (odds ratio, 8.18; confidence interval, 4.88-13.71). When comparing adult (n = 198) with elderly (n = 285) fatalities, ground-level falls occurred in 31% versus 91%, ISS was 27 (25-41) versus 25 (16-36), and length of stay (in days) was 2 (0-6) versus 4 (1-11) (all p < 0.001). Death occurred directly from fall in 82% versus 63%, from complications in 10% versus 20%, and from a fatal event preceding the fall in 8% vs. 17% (all p < 0.001)., Conclusion: The proportion of fall admissions in the elderly is growing in this trauma system. Elderly age is the strongest independent predictor of mortality following a fall. In those who die, death is less likely a direct effect of the fall., Level of Evidence: Epidemiologic study, level III.

Background: Minimally invasive spleen-preserving distal pancreatectomy (SPDP) can be performed with either splenic vessel preservation (SVP) or resection [Warshaw procedure (WP)]. The aim of this study was to evaluate the postoperative clinical outcomes of patients undergoing both methods., Data Sources: Database search of PubMed, Embase, Scopus, Cochrane, and Google Scholar was performed (2000-2014); key bibliographies were reviewed. Qualified studies comparing patients undergoing SPDP with either SVP or WP, and assessing postoperative complications were included. Calculated pooled risk ratio (RR) with the corresponding 95% confidence interval (CI) by random effects methods were used in the meta-analyses., Results: The search yielded 215 studies, of which only 14 observational studies met our selection criteria. The studies included 943 patients in total; 652 (69%) underwent SVP and 291 (31%) underwent WP. Overall, there was a lower incidence of splenic infarction (RR=0.17; 95% CI: 0.09-0.33; P<0.001), gastric varices (RR=0.16; 95% CI: 0.05-0.51; P=0.002), and intra/postoperative splenectomy (RR=0.20; 95% CI: 0.08-0.49; P<0.001) in the SVP group. There was no difference in incidence of pancreatic fistula (WP vs SVP, 23.6% vs 22.9%; P=0.37), length of hospital stay, operative time or blood loss. There was moderate cross-study heterogeneity., Conclusions: SVP is a safe, efficient and feasible technique that may be used to preserve the spleen. WP may be more suitable for large tumors close to the splenic hilum or those associated with splenomegaly. Randomized clinical trials are justified to examine the long-term benefits of SVP-SPDP.

Background: Delayed gastric emptying (DGE) is one of the main complications after pancreaticoduodenectomy (PD). Literature review and meta-analysis were used to evaluate whether subtotal stomach-preserving pancreaticoduodenectomy (SSPPD) may have less incidence than pylorus-preserving pancreaticoduodenectomy (PPPD)., Methods: Online search for studies comparing PPPD to SSPPD was done. Primary outcome was DGE. Quality of included studies was evaluated and heterogeneity was assessed. Relative risk (RR) and 95% confidence intervals (CI) were calculated from pooled data in RCTs and retrospective studies., Results: Eight studies met our selection criteria, with a total of 663 patients undergoing pancreaticoduodenectomy; 309 underwent PPPD and 354 underwent SSPPD. Median age was 66 years. Average male/female ratio was 57 vs. 43%, respectively. There was lower incidence of DGE with SSPPD (RR 0.527; 95% CI 0.363-0.763; p < 0.001) and less nasogastric tube days with SSPPD (RR -0.544; 95% CI -876 to -0.008; p = 0.047). Operative blood loss was more in SSPPD (RR 0.285; 95% CI 0.071-0.499; p = 0.009). There was no statistical difference between the two groups regarding length of hospital stay, incidence of pancreatic fistula, abscesses, overall morbidity, or postoperative mortality., Conclusion: SSPPD was associated with less DGE than PPPD. Larger prospective randomized studies are needed to investigate the association of this result with other complications in more depth.

We hypothesize there are coagulation profile changes associated both with initiation of thromboporphylaxis (TPX) and with change in platelet levels in trauma patients at high-risk for venous thromboembolism (VTE). A total of 1203 trauma intensive care unit patients were screened with a VTE risk assessment profile. In all, 302 high-risk patients (risk assessment profile score ≥ 10) were consented for weekly thromboelastography. TPX was initiated between initial and follow-up thromboelastography. Seventy-four patients were analyzed. Upon admission, 87 per cent were hypercoagulable, and 81 per cent remained hypercoagulable by Day 7 (P = 0.504). TPX was initiated 3.4 ± 1.4 days after admission; 68 per cent received unfractionated heparin and 32 per cent received low-molecular-weight heparin. The VTE rate was 18 per cent, length of stay 38 (25-37) days, and mortality of 17.6 per cent. In all, 76 per cent had a rapid clotting time at admission versus 39 per cent at Day 7 (P < 0.001); correcting from 7.75 (6.45-8.90) minutes to 10.45 (7.90-15.25) minutes (P < 0.001). At admission, 41 per cent had an elevated maximum clot formation (MCF) and 85 per cent had at Day 7 (P < 0.001); increasing from 61(55-65) mm to 75(69-80) mm (P < 0.001). Platelets positively correlated with MCF at admission (r = 0.308, R(2) = 0.095, P = 0.008) and at Day 7 (r = 0.516, R(2) = 0.266, P < 0.001). Change in platelet levels correlated with change in MCF (r = 0.332, R(2) = 0.110, P = 0.005). In conclusion, hypercoagulability persists despite the use of TPX. Although clotting time normalizes, MCF increases in correlation with platelet levels. As platelet function is a dominant contributor to sustained trauma-evoked hypercoagulability, antiplatelet therapy may be indicated in the management of severely injured trauma patients.

Four series of condensed pyrrolo[1,2-c]pyrimidines 6a-d, 8a-d, 10a,b and 12a-e designed as PI3Kα inhibitors were synthesized and evaluated for inhibitory activity and selectivity toward different PI3K isoforms. The tested compounds displayed PI3Kα kinase inhibitory activity at either low micromolar or nanomolar level. In particular, the morpholino-pyrimidopyrrolopyrimidinones 8a-d and morpholino-pyridopyrrolopyrimidine-2-carbonitriles 12a-e proved to be highly potent and selective PI3Kα inhibitors (IC50 = 0.1-7.7 nM). Moreover, the target compounds exhibited considerable cytotoxic activity against cervical cancer cell line HeLa that over-expresses p110α (0.21-1.99 μM). Molecular modeling simulation revealed that, the designed compounds docked well into p110α active site and their complexes are stabilized by a key H-bonding with the backbone amide of Val851 as well as other favorable hydrophobic and H-bond interactions with different amino acids within the enzyme active site., (Copyright © 2015 Elsevier Masson SAS. All rights reserved.)

Four series of sulfonamides incorporating chromone moieties were synthesized and assessed for their cytotoxic activity against MCF-7 and A-549 cell lines, considering the fact that some of these tumors overexpress isoforms of carbonic anhydrase (CA, EC 4.2.1.1) which is inhibited by sulfonamides. Most new sulfonamides showed weak inhibitory activity against the offtarget, cytosolic isoforms hCA I, II but effectively inhibited the tumor-associated hCA IX and XII. The most active compounds featured a primary SO2NH2 group and were active in the low micromolar range against MCF-7 and A-549 cell lines. Compound 4a showed IC50 of 0.72 and 0.50 μM against MCF-7 and A-549 cell lines, respectively, and was further evaluated for its proapoptotic activity which proved enhanced in both tumor types., (Copyright © 2015 Elsevier Masson SAS. All rights reserved.)

Neutropenia frequently occurs in patients with Human immunodeficiency virus (HIV) infection. Causes for neutropenia during HIV infection are multifactoral, including the viral toxicity to hematopoietic tissue, the use of myelotoxic agents for treatment, complication with secondary infections and malignancies, as well as the patient's association with confounding factors which impair myelopoiesis. An increased prevalence and severity of neutropenia is commonly seen in advanced stages of HIV disease. Decline of neutrophil phagocytic defense in combination with the failure of adaptive immunity renders the host highly susceptible to developing fatal secondary infections. Neutropenia and myelosuppression also restrict the use of many antimicrobial agents for treatment of infections caused by HIV and opportunistic pathogens. In recent years, HIV infection has increasingly become a chronic disease because of progress in antiretroviral therapy (ART). Prevention and treatment of severe neutropenia becomes critical for improving the survival of HIV-infected patients.

New pyrimido[5,4-e]pyrrolo[1,2-c]pyrimidines were synthesized. A series of ylidene carbohydrazides 14a-i, and hydrazonate 15, were obtained from the prepared 3-carbohydrazide derivative 13. Pyrazole derivatives 12, 16a,b, 18, 19, 20, were also prepared through different reactions. The anti-inflammatory and analgesic activities of all new compounds were evaluated and most of them exerted comparable activity to indomethacin and celecoxib. Ulcer indexes for the most active compounds were calculated and most of them showed less ulcerogenic effect than the reference drugs. The most potent anti-inflammatory compound 12 showed an IC(50) of 6.00 μmol/kg and low ulcer index. COX-1/COX-2 activity ratio of compounds 12 and 16b showed almost equal inhibitory effect on both isoenzymes. 2D-QSAR studies revealed good predictive and statistically significant QSAR models., (Copyright © 2012 Elsevier Masson SAS. All rights reserved.)

2-(5-Arylidene-4-oxo-3-phenyl-thiazolidin-2-ylidene)-3-oxo-propionitriles 4a-j were prepared via condensation of aromatic aldehydes with 4-thiazolidinones 3a,b. The latter was obtained via electrophilic attack of phenylisothiocyanate on 3-oxo-propionitriles 1a,b followed by reaction with chloroacetyl chloride under basic condition. Additionally, 2-(5-heteroalicyclic methylene) analogues 5a-h were prepared via Mannich reaction of the appropriate secondary amines and formaldehyde with 4-thiazolidinones 3a,b. Many of the synthesized compounds exhibited promising antitumor properties against colon HCT116 and breast T47D cell lines. 3D-Pharmacophore modeling and quantitative structure-activity relationship (QSAR) analysis were combined to explain the observed antitumor properties.

Objective. To identify whether a standardised Echinacea formulation is effective in the prevention of respiratory and other symptoms associated with long-haul flights. Methods. 175 adults participated in a randomised, double-blind placebo-controlled trial travelling back from Australia to America, Europe, or Africa for a period of 1-5 weeks on commercial flights via economy class. Participants took Echinacea (root extract, standardised to 4.4 mg alkylamides) or placebo tablets. Participants were surveyed before, immediately after travel, and at 4 weeks after travel regarding upper respiratory symptoms and travel-related quality of life. Results. Respiratory symptoms for both groups increased significantly during travel (P < 0.0005). However, the Echinacea group had borderline significantly lower respiratory symptom scores compared to placebo (P = 0.05) during travel. Conclusions. Supplementation with standardised Echinacea tablets, if taken before and during travel, may have preventive effects against the development of respiratory symptoms during travel involving long-haul flights.

The starting material, 4-(1-indol-2-yl)phenol 1 was obtained via Fischer synthesis. Vilsmeir Haack(')s formylation of 1 gave the carboxaldehyde derivative 2 which was subjected to different reactions affording the 3-substituted compounds 3-10. Compound 1 reacted with halo esters to give 11 and 12a,b. The reaction of 12a with various amino derivatives gave compounds 13-16. The hydrazide derivative 15a reacted with 1,3-diketones, ethyl acetoacetate and aromatic carboxylic acid derivatives to give 17a,b, 18 and 19a-e, respectively. Antitumor activity of target compounds were tested against breast cancer cell lines (MCF-7) and (MDA-MB-231). The most potent compound was 3e with IC(50) = 1.60 nM against (MCF-7). Docking was performed on colchicine binding site of tubulin to study the binding mode of the designed compounds., (Copyright © 2011 Elsevier Masson SAS. All rights reserved.)

Measurements of calcaneus have been shown to be sexually dimorphic in American whites and blacks, Italians and South African whites and blacks. Since the validity of discriminant function equation in sex determination is population specific, the aim of the present study was to derive similar equations for the calcanei of Egyptians. Lateral radiographs of the ankles of 204 Egyptians aged 20-70 were analyzed with regards to sexual dimorphism, consisting of 104 males and 100 females. X-ray films were obtained from Minia Forensic Department. Six measurements were taken for every calcaneal X-ray film, three linear and three angular measurements. Data were analyzed by SPSS version 17, An independent samples t-test and discriminant function analysis were done. All linear measurements, but not angular, showed significant sexual differences. Maximum length was found to be the most sexually dimorphic (90.2%). Combination of maximum length with minimum height gave the same result of all linear measurements (92.6%) which was more than given by individual variables (81.4-90.2%)., (Copyright © 2010 Elsevier Ltd and Faculty of Forensic and Legal Medicine. All rights reserved.)

Novel series of bicyclic pyrrolo[1,2-c]pyrimidines 3a-g, 5, 6a, b, and 7a, b, tricyclic pyrimido[5,4-e]pyrrolo[1,2-c]pyrimidines 8a-c, 9a-g, 13a-c, 17, 18a, b, 19, 20a,b and 21 and tetracyclic condensed pyrimidines 14, 22 and 23 were synthesized through different chemical reactions. Structures of all synthesized pyrimidine derivatives were supported by spectral and elemental analyses. Analgesic activity evaluation was carried out using acetic acid-induced writhing assay, and all compounds exerted comparable activity to indomethacin. The anti-inflammatory activity evaluation was performed using carrageenan-induced paw edema in rats, the potency of the bicyclic derivatives 3a-f and 7b revealed comparable activity to indomethacin without gastric ulceration. The tricyclic derivatives 13a and 20a exerted good activity, however, they induced gastric ulcers while 13b and 13c showed moderate activity without ulceration. In case of tetracyclic derivatives, compound 14 exhibited the highest potency and safety profile.

A variety of tropane derivatives 14a-g were prepared via the reaction of the alcohol analogs 12a and 12b with substituted fluorobenzenes 13a-f. The prepared compounds were tested for their activity and selectivity toward the norepinephrine transporter (NET) and serotonin transporter (SERT) using yohimbine-induced mortality and 5-hydroxytryptophan-induced neurotoxicity in mice, respectively. All the tested compounds were found to be NE and 5-HT reuptake inhibitors except 14d which exhibited selective 5-HT reuptake inhibition activity.

The effects of haloperidol and olanzapine on polysomnographic measures made in bipolar patients during manic episodes were compared. Twelve DSM-IV mania patients were randomly assigned to receive either haloperidol (mean +/- SD final dosage: 5.8 +/- 3.8 mg) or olanzapine (mean +/- SD final dosage: 13.6 +/- 6.9 mg) in a 6-week, double-blind, randomized, controlled clinical trial. One-night polysomnographic evaluation was performed before and after the haloperidol or olanzapine treatment. Psychopathology and illness severity were rated respectively with the Young Mania Rating Scale (YMRS) and the Clinical Global Impressions - Bipolar version (CGI-BP). There was a significant improvement in the YMRS and CGI-BP scores at the end of the study for both groups. Mixed ANOVA used to compare the polysomnographic measures of both drugs demonstrated significant improvement in sleep measures with olanzapine. In the olanzapine group, statistically significant time-drug interaction effects on sleep continuity measures were observed: sleep efficiency (mean +/- SEM pre-treatment value: 6.7 +/- 20.3%; after-treatment: 85.7 +/- 10.9%), total wake time (pre-treatment: 140.0 +/- 92.5 min; after-treatment: 55.2 +/- 44.2 min), and wake time after sleep onset (pre-treatment: 109.7 +/- 70.8 min; after-treatment: 32.2 +/- 20.7 min). Conversely, improvement of polysomnographic measures was not observed for the haloperidol group (P > 0.05). These results suggest that olanzapine is more effective than haloperidol in terms of sleep-promoting effects, although olanzapine is comparatively as effective as haloperidol in treating mania. Polysomnography records should provide useful information on how manic states can be affected by psychopharmacological agents.

Vancomycin susceptibility was checked in isolates from initial and final blood samples obtained from 22 patients with persistent or recurrent methicillin-resistant Staphylococcus aureus bacteremia. The minimum inhibitory concentration of vancomycin was determined using Etest and found to have increased in 2 pairs of isolates, and results of screening in 4 mu g vancomycin and a modified population analysis profile suggested heteroresistance in 3 isolates (13.6%). Heteroresistance is not a common cause of persistent or recurrent bacteremia.

A new photocrosslinking purine analog was synthesized and evaluated as a transcription substrate for Escherichia coli RNA polymerase. This analog, 8-[(4-azidophenacyl)thio]adenosine 5'-triphosphate (8-APAS-ATP) contains an aryl azide photocrosslinking group that is attached to the ATP base via a sulfur-linked arm on the 8 position of the purine ring. This position is not involved in the normal Watson-Crick base pairing needed for specific hybridization. Although 8-APAS-ATP could not replace ATP as a substrate for transcription initiation, once stable elongation complexes were formed, 8-APAS-AMP could be site-specifically incorporated into the RNA, and this transcript could be further elongated, placing the photoreactive analog at internal positions in the RNA. Irradiation of transcription elongation complexes in which the RNA contained the analog exclusively at the 3' end of an RNA 22mer, or a 23mer with the analog 1 nt from the 3' end, produced RNA crosslinks to the RNA polymerase subunits that form the RNA 3' end binding site (beta, beta'). Both 8-APAS-AMP and the related 8-azido-AMP were subjected to conformational modeling as nucleoside monophosphates and in DNA-RNA hybrids. Surprisingly, the lowest energy conformation for 8-APAS-AMP was found to be syn, while that of 8-azido-AMP was anti, suggesting that the conformational properties and transcription substrate properties of 8-azido-ATP should be re-evaluated. Although the azide and linker together are larger in 8-APAS-ATP than in 8-N(3)-ATP, the flexibility of the linker itself allows this analog to adopt several different energetically favorable conformations, making it a good substrate for the RNA polymerase.

Labeled nucleic acid probes are used as diagnostic tools by detecting changes in gene expression upon hybridization to target RNAs or DNAs that are related to specific disease genes. 5-[S-(2, 4-Dinitrophenyl)-thio]-2'-deoxyuridine analog represents an excellent nucleic acid label, containing the DNP group which functions both as a probe and as a precursor for the introduction of a variety of fluorescent groups. This study describes thermal denaturation hybridization experiments with oligonucleotides containing the 5-[S-(2,4-dinitrophenyl)-thio]-2'-deoxyuridine analog. Using molecular modeling techniques, the effects of this analog on the hybrid structure and stability were examined, including (i) analog conformation, (ii) hydrogen bonding, (iii) stacking interactions and (iv) hybrid helical geometry. This analog does not prohibitively affect the hybrid thermal stability and incorporation of the analog does not compromise the structural integrity of the double helix. In particular, the sequence-dependence of the analog effects and the dependence on the modification site relative to the end(s) of the helix were investigated. Findings described here should provide guidelines in the rational design of nucleic acid probes., (Copyright 1999 John Wiley & Sons, Ltd.)

We report the synthesis and characterization of 5-thioacetamidofluorescein-uridine 5'-triphosphate (5-SF-UTP), and its application to the characterization of the environment of the nascent RNA during trans-cription. This analog specifically replaced UTP as a transcription substrate for Escherichia coli and T7 RNA polymerases, and yeast RNA polymerase III. Escherichia coli transcription complexes containing analog incorporated at only position +21 of the RNA were prepared. The RNA was then elongated in the absence of analog, moving the fluorescent group further away from the enzyme active site, and the fluorescence polarization was measured. Analog positioned near the 3' end of the transcript exhibited significantly increased polarization relative to that of free probe, consistent with the constrained environment of the RNA in the DNA-RNA hybrid. Analog positioned 14 nucleotides from the 3' end exhibited significantly decreased polarization relative to that at the 3' end of the RNA, but only slightly above that of free RNA, suggesting that the probe was on the solvent-exposed surface of the polymerase. Molecular modeling of these analog-substituted RNAs produced structures consistent with the experimental data. The excellent substrate properties of this analog make it useful for the characterization of the environment of RNA not only during transcription and translation, but in any type of ribonucleoprotein complex.

Proteins in a partially fractionated Escherichia coli extract that interact with the nascent RNA in active transcription complexes from several promoters were detected using the photocrosslinking ribonucleotide analogs 5-(azidophenacyl)thio-UTP or 5-(azidophenacyl)thio-CTP as transcription substrates. Upon irradiation of ternary transcription complexes, several extract proteins were crosslinked to the RNA. Most notably, a small protein was crosslinked to the RNA in complexes on seven of nine templates tested. This protein was purified and sequenced and found to match a hypothetical protein, MsmC/CspE, recently shown to be involved in chromatin partitioning. CspE has 69% amino acid sequence identity with the major cold shock protein in E. coli, CspA, which has been shown to bind to a DNA sequence designated the Y box, with the sequence 5'-ATTGG. Of the nine templates tested, CspE was found to be most heavily crosslinked to RNA from the lambda PR' promoter, which is modified by the Q antiterminator protein. CspE was very heavily crosslinked to RNA only ten nucleotides long in initial ternary complexes on this promoter, but not to this same RNA after it had been released from the transcription complex. However, even when present from the start of transcription, CspE did not crosslink to the RNA 82 nucleotides long in elongation complexes from this same promoter. Despite the loss of interaction with the RNA after polymerase had left the promoter, CspE inhibited Q-mediated transcriptional antitermination from PR' in vitro almost 200 nucleotides downstream from the promoter, presumably by interaction with the Y box DNA upstream from PR', which overlaps with the binding site for the Q. A potential role for CspE and transcription in chromosome condensation and nucleoid structure is discussed., (Copyright 1998 Academic Press.)

A new nucleotide analogue was developed for site-specific incorporation of a reactive thiol group into DNA. This creates a unique site for the post-synthetic modification of that nucleotide with a variety of molecular tags, such as photo-cross-linkers and fluorescent or spin-label moieties. 5'-O-(4,4'-Dimethoxytrityl)-5-[S-(2,4-dinitrophenyl)thio]-2'-deoxyuridin e 3'-O-(2-cyanoethyl N,N'-diisopropylphosphoramidite) was synthesized and incorporated at internal positions in several oligonucleotides using automated DNA synthesis and standard phosphoramidite chemistry. The coupling yield of the analogue was comparable to the coupling yield for a standard phosphoramidite, and no significant differences were observed in the overall yields of the dinitrophenyl-labeled oligonucleotides compared to the corresponding unmodified oligonucleotides. Characterization of the dinitrophenyl-modified oligonucleotides included enzymatic degradation, HPLC chromatography, and gel electrophoresis. Deprotection of the mercaptan group with beta-mercaptoethanol yielded an oligonucleotide containing 5-mercaptodeoxyuridine which was then selectively modified, without purification, by reaction with 5-(iodoacetamido)fluorescein. Incorporation of the dinitrophenyl-modified oligonucleotide into double-stranded DNA was achieved using the polymerase chain reaction. CHaracterization of the dinitrophenyl-labeled product by immunodetection with anti-dinitrophenyl antibodies confirmed the stability of the protecting group to the thermocycling and thus established the use of this thiol-protected mercaptodeoxyuridine phosphoramidite for preparation of site-specifically modified DNA.

The alpha subunit (alpha) of RNA polymerase (RNAP) is critical for assembly of polymerase and positive control of transcription initiation in Escherichia coli. Here, we report that alpha also plays a role in transcription elongation, and this involves a direct interaction between alpha and NusA factor. During in vitro transcription without NusA, alpha interacts with the nascent RNA, as revealed by photocrosslinking. When NusA is present, RNA crosslinks to NusA rather than to alpha. We show that this NusA-RNA interaction is diminished during transcription with an RNAP mutant that lacks the C-terminus of alpha beyond amino acid 235, including the so-called alpha CTD. The absence of alpha CTD also affects NusA's ability to enhance transcription pausing, termination at intrinsic terminators and anti-termination by the phage lambda Q anti-terminator, but not anti-termination by the lambda N anti-terminator. NusA functions are not recovered even when transcription with mutant RNAP is done with excess NusA, a condition which does restore NusA-RNA crosslinking. By affinity chromatography, we show that NusA interacts directly with alpha, and also with beta and beta', but not with mutant alpha. Hence, alpha-NusA interaction is vital for the control of transcript elongation and termination.

The Escherichia coli transcription factor NusA and the bacteriophage lambda antiterminator Q proteins were expressed as inducible glutathione S-transferase (GST) fusion proteins. The fusion proteins were purified under nondenaturing conditions by affinity chromatography on glutathione agarose. Thrombin cleavage of the glutathione agarose-bound fusion proteins yielded homogeneously pure NusAN+15 (5 mg/g cells) and almost homogeneously pure QN+13 protein (0.7 mg/g cells), where N+x indicates the presence of x additional amino acids at the N-terminus of the protein. The purified NusAN+15 exhibited the same activities as wildtype NusA in enhancement of transcriptional pausing, enhancement of termination at Rho-independent terminators, and enhancement of Q-mediated antitermination in vitro. The QN+13 protein exhibited both anti-pausing and antitermination activities in Q-mediated transcription antitermination. However, the antitermination activity of QN+13 was lost gradually during storage if the thrombin used for cleavage of the GST fusion protein was not removed. This was due to cleavage by thrombin after Arg22 within the Q protein itself, at a noncanonical thrombin cleavage site, so the truncated protein (QN+22) lacked the first 22 amino acids at the N-terminus of Q. The expression vectors described here can be used to rapidly produce large quantities of these proteins, and the truncated Q protein can be used to evaluate the requirement for the N-terminus of Q in antitermination, anti-pausing, interactions with the DNA template (qut site), and interaction with RNA polymerase itself.

The effects of NusA on the RNA polymerase contacts made by nucleotides at internal positions in the nascent RNA in Escherichia coli transcription complexes were analyzed by using the photocrosslinking nucleotide analog 5-[(4-azidophenacyl) thio]-UMP. It was placed at nucleotides between +6 and +15 in RNA transcribed from the phage lambda PR' promoter. Crosslinks of analog in these positions in RNAs which contained either 15, 28, 29, or 49 nt were examined. Contacts between the nascent RNA and proteins in the transcription complex were analyzed as the RNA was elongated, by placing the crosslinker nearest the 5' end of the RNA 10, 23, 24, or 44 nt away from the 3' end. The beta or beta' subunit of polymerase, and NusA when added, were contacted by RNA from 15 to 49 nt long. When the upstream crosslinker was 24 nt from the 3" end of the RNA (29-nt RNA), alpha was also contacted in the absence of NusA. The addition of NusA prevented RNA crosslinking to alpha. When the crosslinker was 44 nt from the 3' end (49-nt RNA), alpha crosslinks were still observed, but crosslinks to beta or beta' and NusA were greatly diminished. RNA crosslinking to alpha, and loss of this crosslink when NusA was added, was observed in the presence of NusB, NusE, and NusG and when transcription was carried out in the presence of an E. coli S100 cell extract. Peptide mapping localized the RNA interactions to the C-terminal domain of alpha.

We report a case of Grzybowski's generalized eruptive keratoacanthoma which demonstrates the characteristic features of this rare condition. The recurring nature of the eruption each summer supports the suggestion that UV irradiation may act as a precipitating factor in eruptive keratoacanthoma.

We report a modified synthesis for 5-mercapto-UTP (5-SH-UTP) and its use for analysis of protein-RNA interactions utilizing Escherichia coli and T7 RNA polymerases and yeast RNA polymerases I and III. 5-SH-UTP did not affect transcriptional pausing, Rho-independent termination or recognition of the E. coli transcription complex by NusA. RNA containing 5-SH-UMP did not crosslink to polymerase when irradiation was 302 or 337 nm. Transcription complexes containing RNA substituted with 5-SH-UMP were post-transcriptionally modified to attach a photocross-linking group to thiol-tagged nucleotides in the RNA on the surface of the polymerase of free in solution. The pKa for 5-SH-UTP was determined to be 5.6, so modification of the thiol groups in the RNA with p-azidophenacyl bromide could be carried out at pH 7. Addition of the transcription termination factor Rho, a RNA binding protein, to E. coli transcription complexes resulted in RNA crosslinking to Rho and to the beta and beta' subunits of polymerase. Using 5-SH-UTP, one can distinguish RNA binding domains on the surface of RNA polymerases or other RNA binding proteins from those buried within the protein.

We have examined the interaction between NusA and the nascent RNA in Escherichia coli transcription complexes on four different templates. Photocrosslinking CTP and UTP analogs were incorporated internally and at the 3' end of the RNA. Identical templates with and without boxA sequences were compared. We found that NusA did not contact the ten nucleotides nearest to the 3' end of the RNA in complexes containing RNA up to 20 nucleotides long. Longer RNA did crosslink to NusA with all four templates examined, however. We reported that RNA 80 nucleotides long from the bacteriophage T7 A1 promoter substituted in two RNA stem-loops with photocrosslinking UMP analogs did not crosslink to NusA, even though interaction between NusA and the transcription complex were demonstrated. Here, we report that when this same RNA is substituted at CMP residues, it does crosslink to NusA. Templates containing the E. coli ribosomal RNA promoter rrnG P2, with and without a boxA sequence downstream, were compared. Long RNAs from both crosslinked to NusA, and thus boxA RNA sequences are not required for interaction with NusA. NusA did not interact with the free RNA containing boxA once released from the transcription complex, nor did it interact with RNA in a binary complex containing only RNA polymerase and RNA, without the DNA template.

Background: A multi-dimensional approach was used to examine coping in chronic pain. The following hypotheses were tested: (a) patients who cope maladaptively also cope generally in a similar way; (b) patients' maladaptive coping is associated with childhood adversity., Method: Cross-sectional and retrospective data were collected from 68 consecutive patients (aged 18-70) at a pain clinic where their disease was non-systemic and the pain had lasted for at least three months. Sixty-one patients were interviewed using the Structured Clinical Interview for DSM-III-R, and the Measure of Parental Care in Childhood. All patients completed questionnaires on their pain and personality., Results: Two coping styles emerged from factor analysis. One was associated with chronicity, psychiatric morbidity, harm avoidance, immature defence style and reporting parental indifference., Conclusion: Patients may be predisposed to cope maladaptively after the experience of parental indifference in early life. Such coping is likely to reflect more general patterns.

A conformational change in Escherichia coli RNA polymerase induced by NusA was detected by utilizing photocrosslinking. A change in the binding site for the 3' end of the RNA occurred, and NusA increased interactions of the RNA with the beta subunit of the polymerase. NusA was not contacted by the 3' end of the RNA.

Some new substituted thiazolidinones, thioimidazolidinones and thiazolines have been synthesized from N1-substituted N2-(4-hydroxy-7-methyl-5H-furo[3,2-g] [1]benzopyran-5-on-9-yl)thioureas and monochloroacetic acid or alpha-halocarbonyl compounds. Some representative examples were tested for their anticonvulsant and antimicrobial activities.

A new photocrosslinking CTP analog that functioned as a substrate during transcription was synthesized and used to photoaffinity label E. coli and bacteriophage T7 RNA polymerases. This analog, 5-((4-azidophenacyl)thio) cytidine-5'-triphosphate (5-APAS-CTP) contains an aryl azide group approximately 10 A from the nucleotide base and specifically replaced CTP during synthesis of RNA by both polymerases. Analog was placed at the 3' end or internally within RNA. Both polymerases inefficiently incorporated two 5-APAS-CMP molecules sequentially, as was found for the related 5-APAS-UMP. Analog was placed at the 3' end of RNA in transcription complexes paused at the site of Q-modification of E. coli RNA polymerase, downstream of the lambda PR' promoter (+16), a pause that requires specific DNA sequences but no apparent RNA hairpin. Crosslinking was examined in the presence and absence of the NusA protein, which enhances the transcriptional pause at this site and is required for Q modification of the polymerase. Crosslinking of the 3' end of the RNA to NusA was not observed, consistent with our earlier results involving a NusA-enhanced pause site downstream from an RNA hairpin.

Regulation of transcription involves numerous specific protein-nucleic acid interactions. We have utilized photochemical crosslinking to identify interactions between Escherichia coli transcription proteins and the nascent RNA in several transcription complexes, including initiation, elongation, and antitermination complexes. We have developed new nucleotide analogs, 5-APAS-UTP and 5-APAS-CTP, which are tagged with photocrosslinking groups on base positions that do not interfere with normal Watson-Crick base-pairing. These analogs are incorporated at internal positions in RNA by E. coli RNA polymerase without disrupting RNA secondary structures. We have also used 8-azido-ATP, which can be incorporated uniquely into the 3' end of the RNA, to analyze interactions at the enzyme active site. Interactions between the RNA and the polymerase subunits, and the effect of various transcription factors, including NusA, NusB, NusE, and NusG, have been examined in complexes containing RNAs from 4 to approximately 80 nucleotides. At almost every RNA position examined, both the beta and beta' subunits are contacted, but never the alpha subunit or NusA. An effect of NusA on the core labeling has been observed in some complexes, however. Sigma is contacted by nucleotides within three nucleotides of the +1 position on the DNA.

We have used photocrosslinking to analyze the contacts between the 3' end of the RNA and Escherichia coli RNA polymerase during the early steps of RNA synthesis using the nucleotide analog 8-azido-ATP (8-N3-ATP). The crosslinking group on 8-N3-ATP contacts the beta, beta' and sigma subunits when the analog is bound to the holoenzyme. We show here that 8-N3-ATP is a substrate for E. coli RNA polymerase and acts as an RNA chain terminator when incorporated into the 3' end of nascent RNA. 8-N3-AMP was incorporated uniquely at the 3' end of tri-, tetra- and pentanucleotides synthesized from a poly[d(A-T)] template and at the 3' end of pentanucleotides from two promoters (lambda PR' and E. coli rrnB P1). The oligonucleotides were covalently attached to the RNA polymerase by irradiation of transcription complexes with ultraviolet light. All RNAs labeled the beta and beta' subunits, but sigma was contacted only by the trinucleotide and tetranucleotide on poly[d(A-T)]. Sigma is still present in transcription complexes containing the pentanucleotide on poly[d(A-T)], despite the lack of labeling. Neither pentanucleotide from the authentic promoters contacted sigma. We conclude that as holoenzyme moves downstream, either two separate conformational changes occur, after synthesis of the trinucleotide and tetranucleotide, which result in movement of sigma away from the nucleotide binding site or, alternatively, sigma remains fixed relative to the DNA while the domain on core polymerase forming the nucleotide binding site moves downstream.