Analyzing RNA posttranscriptional modifications to decipher the epitranscriptomic code.

The discovery of RNA silencing has revealed that non-protein-coding sequences (ncRNAs) can cover essential roles in regulatory networks and their malfunction may result in severe consequences on human health. These findings have prompted a general reassessment of the significance of RNA as a key player in cellular processes. This reassessment, however, will not be complete without a greater understanding of the distribution and function of the over 170 variants of the canonical ribonucleotides, which contribute to the breathtaking structural diversity of natural RNA. This review surveys the analytical approaches employed for the identification, characterization, and detection of RNA posttranscriptional modifications (rPTMs). The merits of analyzing individual units after exhaustive hydrolysis of the initial biopolymer are outlined together with those of identifying their position in the sequence of parent strands. Approaches based on next generation sequencing and mass spectrometry technologies are covered in depth to provide a comprehensive view of their respective merits. Deciphering the epitranscriptomic code will require not only mapping the location of rPTMs in the various classes of RNAs, but also assessing the variations of expression levels under different experimental conditions. The fact that no individual platform is currently capable of meeting all such demands implies that it will be essential to capitalize on complementary approaches to obtain the desired information. For this reason, the review strived to cover the broadest possible range of techniques to provide readers with the fundamental elements necessary to make informed choices and design the most effective possible strategy to accomplish the task at hand.

Prevalence of undiagnosed atrial fibrillation in elderly individuals and potential cost-effectiveness of non-invasive ambulatory electrocardiographic screening: The ASSERT-III study.

BACKGROUND: In approximately 10% of patients with implanted pacemakers or defibrillators, previously unrecognized atrial fibrillation (AF) is detected within 3 months. It is unknown whether elderly patients without implanted devices have a similar prevalence of undiagnosed AF using non-invasive ECG monitoring, and if this approach to screening in this population is cost-effective. METHODS: Individuals ≥80 years old attending outpatient clinics without a history of AF and with hypertension and one additional risk factor underwent 30 days of continuous ECG monitoring with an option for an additional 30 days of monitoring if no AF was detected. The primary outcome was AF ≥ 6 min. Cost-effectiveness to prevent stroke was estimated using a Markov model based on observed AF detection rates and data from the published literature. RESULTS: Among 129 patients enrolled, 100 initiated monitoring for an average duration of 36 ±â€¯21 days. The proportion of patients that completed at least 30 days of monitoring was 59%. Average age was 84 ±â€¯3 years and mean CHA2DS2-VASc score was 4.5 ±â€¯1.2. AF ≥ 6 min was documented in 14%, ≥6 h in 8%, and ≥24 h in 3%. One week of monitoring costed $50,000 per quality-adjusted life-year-gained, 30 days and 60 days of monitoring costed $70,000 and $84,000, respectively. CONCLUSIONS: Continuous non-invasive ECG monitoring is feasible in elderly patients. Undiagnosed AF is present in many elderly individuals, with 1 in 7 having episodes lasting ≥6 min. One week of monitoring may be cost-effective for stroke prevention in this population.

Design and rationale of the atrial fibrillation occurring transiently with stress (AFOTS) incidence study.

BACKGROUND: Atrial fibrillation (AF) is often detected for the first time in patients hospitalized for medical illness or non-cardiovascular surgery. AF occurring transiently with stress (AFOTS) describes this manifestation of AF, which may either be the result of a non-cardiac stressor, or existing paroxysmal AF that was not previously detected. Current estimates of AFOTS incidence are imprecise: ranging from 1 to 44%, owing to the marked heterogeneity in patient populations, identification and methods used to detect AFOTS. METHODS: The prospective, two-centre epidemiological AFOTS Incidence study will enroll 250 consecutive participants without a history of AF but with at increased risk of AF (Age ≥ 65 or >50 with one risk factor for AF) admitted to intensive care units (ICUs) for medical illness or non-cardiac surgery. Upon admission, participants will wear an ECG patch monitor that will remain in place for 14 days, or until discharge from hospital. Patients' consent to participation is deferred for up to 72 h after admission. The primary endpoint is the incidence of AF lasting ≥30 s. The study is powered to detect an AF incidence of 17% ±â€¯5%. RESULTS: We conducted a vanguard feasibility study, and 55 participants have completed participation. The median duration of monitoring was seven days. AF was detected by the clinical team in 8 participants (14%; 95% Confidence Interval 7-26%). CONCLUSIONS: The AFOTS Incidence study will employ a systematic and highly sensitive protocol for detecting AFOTS in medical illness and non-cardiac surgery ICU patients. This study is feasible and will provide a reliable estimate of the true incidence of AFOTS in this population.

Interactive effects of insomnia symptoms and trauma exposure on PTSD: Examination of symptom specificity.

OBJECTIVE: Although symptoms of posttraumatic stress disorder (PTSD) are common following exposure to a traumatic event, most people who experience trauma do not develop PTSD. Thus, the identification of risk factors that may interact with trauma exposure to confer vulnerability for the development of PTSD may highlight important targets for prevention and treatment. Recent research suggests that sleep disturbance amplifies the effect of maladaptive emotional processes on PTSD symptom severity. However, no study to date has examined the impact of sleep disturbance, such as insomnia symptoms, on the relationship between trauma exposure severity and specific PTSD symptom clusters. METHOD: The present study examined insomnia symptoms as a potential moderator of the relationship between trauma exposure severity and specific PTSD symptom clusters among combat exposed veterans (N = 72). RESULTS: Results revealed large associations between insomnia symptoms and PTSD symptom clusters, small to moderate associations between combat trauma severity and PTSD symptom clusters, and a significant interaction between insomnia symptoms and combat exposure to predict reexperiencing, but not avoidance or arousal, symptoms of PTSD. CONCLUSION: These findings suggest that poor sleep may be one risk context in which trauma exposure results in the development of reexperiencing symptoms of PTSD. The implications of these findings for the development and maintenance of reexperiencing symptoms of PTSD are discussed. (PsycINFO Database Record

Limited dorsiflexion predisposes to injuries of the ankle in children.

Injuries to the ankle are common in children. We investigated whether decreased dorsiflexion predisposes to such fractures and sprains. Passive dorsiflexion in children with ankle injuries was compared with that in a control group of patients with a normal ankle. The uninjured side was examined to determine flexibility in those patients with ankle injuries. In 82, the mean dorsiflexion was 5.7 degrees with the knee extended and 11.2 degrees with the knee flexed. In 85 controls, the mean dorsiflexion was 12.8 degrees with the knee extended and 21.5 degrees with the knee flexed (p < 0.001, Student's t-test). There was a strong association between decreased ankle dorsiflexion and injury in children. A flexible triceps surae appeared to absorb energy and protect the bone and ligaments, while stiffness predisposed to injury. We suggest that children with tight calf muscles should undergo a regimen of stretching exercises to improve their flexibility.

Bone marrow granulomas possibly associated with amiodarone.

Amiodarone is a class III antiarrhythmic agent that is effective in treating different types of cardiac dysrhythmias. It was approved only for treatment of life-threatening ventricular dysrhythmias refractory to other therapy; however, its use for atrial dysrhythmias such as atrial fibrillation is well accepted. Adverse effects associated with amiodarone include pulmonary, hepatic, thyroid, ocular, and neurologic toxicities. Our patient experienced intermittent fever, night sweats, and fatigue while taking the drug for treatment of atrial fibrillation. Bone marrow biopsy showed granuloma formation after 17 months of therapy with amiodarone. Amiodarone was discontinued due to significant hypotension and shortness of breath. To our knowledge, this is the third case report of granuloma formation in bone marrow possibly associated with this agent.

Monitoring of renal function in patients with spinal cord injury.

OBJECTIVE: To assess the sensitivity of serum creatinine level in detecting clinically important and early deterioration of renal function in patients with spinal cord injury (SCI), and to evaluate the optimal method of determining creatinine clearance in these patients. PATIENTS AND METHODS: The serum creatinine level of 36 patients (25 paraplegics and 11 quadriplegics) was evaluated and compared with the corresponding measured creatinine clearance rate. Correlations were also assessed between the creatinine clearance measured by 24-h endogenous clearance, single-shot 99mTc-labelled diethylenetriamine pentaacetic acid (99mTc-DTPA) clearance technique, and the Cockcroft-Gault formula, to test their validity. RESULTS: Of the 36 patients 11 (31%) had a measured creatinine clearance of < 100 mL/min (mean 84.8) and a corresponding normal serum creatinine level. Creatinine clearance calculated by the Cockcroft-Gault formula did not correlate well with that measured by the 24-h endogenous clearance (r = 0.426) and 99mTc-DTPA clearance (r = 0. 366), overestimating creatinine clearance in all but three patients. The mean (SD) difference between the creatinine clearance measured by the 24-h and DTPA clearance technique was 17.7 (16.5)% and the correlation between these techniques was good (r = 0.71). CONCLUSION: Serum creatinine level is not sensitive in detecting early deterioration of renal function in patients with SCI. The Cockcroft-Gault formula generally significantly overestimates the true creatinine clearance and is not recommended. The 24-h endogenous creatinine clearance measured on appropriately collected urine samples is an acceptable accurate and practical method of determining glomerular filtration rate in patients with SCI.

Assessment of a new transurethral balloon dilation catheter in the treatment of urethral stricture disease.

OBJECTIVES: To assess a newly designed balloon dilation catheter for the treatment of urethral stricture disease. The dilating capability of the catheter, the tolerability and safety of the procedure, and its short-term efficacy were evaluated. METHODS: Fifty-one patients with urethral strictures underwent dilation with the UrethraMax or a coude tip balloon dilation catheter. Efficacy parameters included measurement of the American Urological Association symptom score and maximum urinary flow rate 3, 6, and 12 months after treatment. The adequacy of dilation and the degree of mucosal trauma and hematuria were assessed endoscopically, and patient pain was measured using a visual analog scale. RESULTS: Forty-three patients (84.3%) were successfully dilated, achieving a urethral caliber of 20F or greater. Dilation resulted in statistically significant improvements in both the mean American Urological Association symptom score and mean maximum urinary flow rate at 3 and 6 months. Mucosal trauma was mild in all but 4 cases, and no patient developed significant hematuria. The mean visual analog pain score was 3.9 (range 0.1 to 9.4). CONCLUSIONS: Balloon dilation is a safe, well-tolerated, office-based procedure that theoretically offers several advantages over sequential rigid dilation and internal urethrotomy. It is associated with minimal complications, and its short-term efficacy is acceptable. We regard this as the dilation procedure of choice and first-line therapy for most strictures.

A randomized double-blind study assessing 4 versus 8 mg. doxazosin for benign prostatic hyperplasia.

PURPOSE: We compare the efficacy of 4 versus 8 mg. doxazosin for benign prostatic hyperplasia. MATERIALS AND METHODS: A total of 82 patients with benign prostatic hyperplasia successfully treated with 4 mg. doxazosin were randomized in a double-blind fashion to take 4 or 8 mg. Patients were evaluated with American Urological Association (AUA) symptom score, Boyarsky score, uroflowmetry and side effect profile before, and 1 and 3 months following randomization. RESULTS: Of the patients 42 and 40 were randomized to receive 4 and 8 mg. doxazosin, respectively. Both groups were similar with respect to patient age, baseline Boyarsky and AUA symptom scores, and baseline maximum urinary flow rate. At 3 months mean improvement from baseline plus or minus standard deviation in Boyarsky score was 0.6 +/- 6.5 and 4.9 +/- 6.6 in the 4 and 8 mg. groups (p <0.05), respectively, mean improvement in AUA symptom score was 1.6 +/- 5.3 and 5.3 +/- 8.0 (p <0.05), and mean maximum flow rate difference was -0.6 +/- 6.4 and +1.4 +/- 7.9 (p >0.05). Of the patients 7 and 8 in the 4 and 8 mg. groups dropped out of the study, and there were no statistical differences in side effects between dosages. CONCLUSIONS: A dose of 8 mg. doxazosin was more efficacious than 4 mg. and the side effects associated with both dosages appeared to be similar. The 8 mg. dose should be tried in patients who have not achieved an adequate therapeutic response to 4 mg. and are tolerating the medication. Consideration should be given to increasing the dosage to 8 mg. in patients who are clinically improved at lower dosages.

Prolongation of cardiac graft survival with anti-CD4Ig plus hCTLA4Ig in primates.

BACKGROUND: The aim of this study was to determine whether the use of combined immunotherapy with a brief course of humanized anti-CD4Ig and hCTLA4Ig would prolong heterotopic cardiac allograft survival in primates (rhesus monkeys). This model was based on work in "high responder" rats where a brief course of depletive anti-CD4mAb plus hCTLA4Ig was successful in inducing transplantation tolerance. METHODS: Heterotopic cardiac transplants were performed in rhesus recipients. Donor/recipient pairs between groups were confirmed to be reactive prior to transplantation by MLR matching. Humanized anti-CD4Ig, a recently developed anti-CD4mAb, was given at a dose of 20 mg/kg i.v. on days -3, -2, -1, and 0. hCTLA4Ig was administered at 6 mg/kg/dose i.v. on days 0 and 2 for the first recipient and days 0, 2, 4, and 6 for the second recipient. No further immunosuppression was administered. The treated (n = 2) or untreated (n = 5) recipients were followed for graft function by daily palpitation. RESULTS: Treatment with anti-CD4Ig plus hCTLA4Ig resulted in a significant prolongation of heart graft survival (42 days for the first recipient and 52 days for the second recipient) compared to untreated recipients (7 days x 4, 11 days x 1). FACS analysis demonstrated CD4 depletion of anti-CD4 treated animals to <2% on posttransplant day 1. The CD4+ T cells gradually repopulated to 50-70% pretransplant levels just prior to rejection. No adverse responses (fever, tachypnea, tachycardia, infections) were observed. CONCLUSIONS: These are the first results demonstrating that a brief course of combined specific induction immunotherapy with humanized anti-CD4Ig plus hCTLA4Ig, in the absence of adjuvant posttransplant immunosuppression, was well tolerated and resulted in marked prolongation of cardiac allograft survival in primates.

An analysis of open reduction of irreducible supracondylar fractures of the humerus in children.

OBJECTIVES: To review experience with irreducible supracondylar fractures requiring open reduction in children, and to propose guidelines for an open approach to supracondylar fractures. DESIGN: A chart review. SETTING: The Children's Hospital of Eastern Ontario (CHEO), a pediatric centre with a large referral base. PATIENTS: Forty-one children (18 boys 23 girls, average age 7 years), who had open reduction of irreducible supracondylar fractures at the CHEO over a 10-year period (1985 to 1995). Of these 41 children, 7 were lost to direct follow-up. INTERVENTIONS: After closed reduction of displaced supracondylar fractures of the humerus failed, all patients underwent open reduction and percutaneous fixation in the operating room. Before operation, 6 had no radial pulse, 5 lost their pulse with flexion after reduction and 4 had unstable fracture patterns. MAIN OUTCOME MEASURES: Assessment of elbow range of motion and carrying angle, distal neurovascular status and radiographic measurement of the Baumann angle and the humerocapitellar angle. RESULTS: In 25 children, the humerus was found to have "buttonholed" through the brachialis muscle; 1 had entrapment of the common flexor muscle at its origin and 1 had entrapment of the triceps. In 15 children there was entrapment or tethering of the median nerve and radial nerve or brachial artery, or both, but this was not predictive of preoperative neurovascular deficit, which was recorded in 21 patients (fully recovered). At follow-up, the Baumann angle and the humerocapitellar angle differed by an average of 2 degrees and 5.3 degrees respectively compared with the unaffected arm. Range of motion was satisfactory in 94% of patients, and there was no significant cubitus varus. CONCLUSION: Open reduction of supracondylar fractures is a safe and effective procedure, for which orthopedists should should lower their threshold, given certain appropriate indicators.

Cervicothoracic vascular injuries.

Arterial and venous trauma of the cervicothoracic region continues to present challenging problems for the surgeon, despite advances in vascular diagnostics and surgical technique. Whether due to penetrating or blunt mechanisms, overall incidence of these injuries is low, whereas morbidity and mortality remain high. Despite collective experience from busy trauma centers, there still remain controversies regarding diagnostic evaluation, operative approach, and surgical treatment of these potentially devastating injuries. Therefore, this article compares and contrasts recent literature and controversies surrounding the treatment of cervicothoracic trauma. Pros and cons of duplex ultrasonography and angiography in the diagnosis of carotid and vertebral artery injury are highlighted, and selective versus mandatory neck exploration for zone II penetrating injuries are discussed. Increasing awareness of blunt carotid artery injury is emphasized, including management dilemmas that frequently accompany this type of injury. In addition, we review interventional radiological techniques for the management of vertebral artery injury and surgical approaches for aortic arch branch vessel or major cervicothoracic vein injury.

Fractures of the capitellum in adolescents.

Fractures of the capitellum are rare in children. The treatment of these injuries has been controversial. At a major pediatric trauma center, seven capitellar fractures were seen in children between 1988 and 1994. The average age of the children was 14.7 years (range, 11-17). Six of these fractures were type I injuries, with large anterosuperior fragments that required operative reduction and internal fixation in five cases. Internal fixation methods used were K wires in three patients, Herbert screws in one patient, and cannulated screws in one patient. The remaining type I fracture was treated with a closed reduction. The seventh fracture was a type II fracture, treated nonoperatively. Five children did well with their respective treatments, but one required reoperation to remove an exostosis block to flexion. Accurate open reduction and internal fixation for the displaced capitellar fracture in children is an effective treatment to restore normal elbow function.

Randomized comparison of G-CSF + GM-CSF vs G-CSF alone for mobilization of peripheral blood stem cells: effects on hematopoietic recovery after high-dose chemotherapy.

Fifty patients with either lymphoid or selected solid tumor malignancies were apheresed an identical number of times for PBSC collection after being randomized to receive either G-CSF 10 microg/kg/day alone (arm I), or G-CSF at the same dose with GM-CSF 5 microg/kg/day (arm II). Growth factor(s) was/were given as the stem cell mobilizing agent for 5 days before the start of PBSC collection, and were continued throughout the 4 days of apheresis. Aspiration and cryopreservation of autologous bone marrow occurred on day 3 or 4 of growth factor(s). Thirty-one of 50 patients received one cycle only at time of evaluation, and 19 patients received two cycles of HDCT, each supported with PBSC with or without autologous bone marrow. No patients received growth factors post-autologous stem cell transplant, unless the absolute neutrophils count (ANC) failed to recover to > or = 100/microl by day +18 post-transplant. The median number of days to recovery of ANC to 100/microl, 500/microl and 1000/microl, and of platelet counts to 20000/microl, 50000/microl and 100000/microl after either cycle 1 or cycle 2 of HDCT and the number of febrile days and platelet and PRBC transfusion requirements was not significantly different between the two arms of the study. The duration of hospitalization was similar between study arms for cycle 1 of HDCT, but was 3.5 days less with arm II compared to arm I (P = 0.0248) for cycle 2 of HDCT. The bone marrow buffy coat and PBSC product mononuclear cell count (x 10(8)/kg) and CD34+ cell count (x 10(6)/kg) collected by each method of stem cell mobilization was not significantly different. There is questionable clinical benefit with PBSC products mobilized with the combination of G-CSF and GM-CSF vs G-CSF alone. Perhaps different dosages, schedules, or other growth factor combinations with G-CSF might enhance these differences.

Antifungal effects of yeast-derived rhu-GM-CSF in patients receiving high-dose chemotherapy given with or without autologous stem cell transplantation: a retrospective analysis.

Systemic fungal infections (SFI) in patients receiving high-dose chemotherapy (HDC) are a frequent cause of morbidity and mortality. Preclinical studies have reported augmented antifungal activity of monocytes, macrophage cells, and neutrophils exposed to certain colony-stimulating factors (CSF), including GM-CSF. We conducted a retrospective descriptive epidemiologic study to examine the characteristics of 145 consecutive patients receiving HDC administered with or without autologous stem cell transplantation (ASCT) and who subsequently received either GM-CSF and G-CSF, G-CSF alone, GM-CSF +/- IL-3 or no CSF. The analysis of this patient population sought to define the incidence of SFI and its relationship to therapy with monocyte/macrophage-stimulating (MMS group) cytokines (GM-CSF and G-CSF; GM-CSF +/- IL-3) or to cytokines which do not result in monocyte/macrophage stimulation (NMMS group, G-CSF alone or no CSF). Risk factors for the development of SFI were balanced between the MMS (n = 70) and NMMS (n = 75) groups. Two patients (2.9%) in the MMS and nine patients (12%) in the NMMS groups developed SFI. The risk ratio for developing SFI in the NMMS group compared to the MMS group was 4.20 (P = 0.023). This relationship was confounded, however, by the diagnosis of hematologic tumor or solid tumor (RR = 3.15, P = 0.082). SFI was the primary cause or major contributing factor in five of the 10 total deaths in our study population. Four SFI-related deaths occurred in the NMMS group and one SFI-related death occurred in the MMS group. Our data suggest a protective role for GM-CSF, IL-3 or other MMS cytokines in preventing SFI in patients receiving HDC. This should be further investigated as a potential complementary approach to conventional strategies in antifungal prophylaxis for patients receiving HDC.

Randomized study of growth factors post-peripheral-blood stem-cell transplant: neutrophil recovery is improved with modest clinical benefit.

PURPOSE: To evaluate the clinical value of growth factors (GFs) with peripheral-blood stem cells (PBSC) collected following mobilization with GFs, we randomized patients to receive or not to receive GFs following transplant. PATIENTS AND METHODS: Thirty-seven patients were apheresed after receiving the combination of granulocyte colony-stimulating factor (G-CSF) with granulocyte-macrophage colony-stimulating factor (GM-CSF) at doses of 10 micrograms/kg/d and 5 micrograms/kg/d, respectively, for 6 days before apheresis and during a median of 4 days of collections. One day after the infusion of autologous marrow and PBSC, patients were randomly assigned to receive no GFs or a combination of G-CSF (7.5 micrograms/kg/d) and GM-CSF (2.5 micrograms/kg/d), both as a 2-hour intravenous (i.v.) infusion twice per day until the neutrophil count was greater than 1,500/microL. RESULTS: The median days to recovery to an absolute neutrophil count (ANC) of 100/microL (9 v 11.5, P = .0005), 500/microL (10 v 16, P = .0004), or 1,000/microL (12 v 21, P = .0008) was shortened with the use of GFs, post-PBSC infusion. In addition, the duration of hospitalization was shorter (19 v 21 days, P = .0112) in the arm receiving GFs post-PBSC infusion. There was no significant difference between the two study arms in the duration of fever, documented septic episodes, or RBC or platelet transfusion requirements. CONCLUSION: Despite faster neutrophil recovery and shortened duration of hospitalization with GFs administered after PBSC transplantation, the measured clinical variables of febrile days, septic episodes, and transfusion requirements were similar between the study arms. The use of GFs post-PBSC transfusion is associated with a modest clinical benefit.

Effects of interferon-alpha monotherapy on hepatic drug metabolism in cancer patients.

1. The influence of interferon-alpha (IFN alpha) on the clearances of theophylline (TH), antipyrine (AP) and hexobarbitone (HB) was studied in seven cancer patients given IFN alpha as their only treatment. In addition, IFN alpha effects on drug clearance were correlated with changes in serum inflammatory cytokines and acute phase proteins. 2. A 'baseline' study was performed by administering an oral drug 'cocktail' of TH (150 mg), AP (250 mg) and HB (250 mg) with saline injected simultaneously and again 24 h later. One week later, an 'acute' study was performed at the initiation of IFN alpha therapy, 3 x 10(6) units injected with the drug cocktail and again 24 h later. After 2 weeks of IFN alpha treatment three times per week, a 'chronic' study was performed with IFN alpha injected the day prior to, simultaneously with, as well as 24 h after the drug cocktail. 3. Plasma samples were collected over 48 h and the clearances of TH, AP and HB were estimated. Serum samples were collected at various times for the measurement of tumor necrosis factor (TNF), interleukin-1 (IL-1), interleukin-6 (IL-6), C-reactive protein (C-RP) and alpha 1-acid glycoprotein (AGP). 4. IFN alpha caused a 33% decrease in the oral clearance of TH during the chronic study compared with baseline (P < or = 0.05). Although IFN alpha inhibited TH clearance by 16% during the acute study and AP clearance by 20-21% during both acute and chronic studies, these changes did not reach statistical significance. IFN alpha caused minimal changes in HB clearance. There were no chronic effects of IFN alpha on serum cytokines or acute phase proteins. 5. The findings confirm that the most commonly used dose of IFN alpha inhibits the hepatic clearance in humans of some but not all drugs and that this inhibition persists during IFN alpha therapy. Because inhibition was not associated with increases in serum cytokines or acute phase proteins, the mechanism by which IFN alpha inhibits cytochrome P450 activities in vivo does not appear to involve inflammatory mediators such as TNF. IL-1 or IL-6.