Standardized generation and differentiation of neural precursor cells from human pluripotent stem cells.

Precise, robust and scalable directed differentiation of pluripotent stem cells is an important goal with respect to disease modeling or future therapies. Using the AggreWell™400 system we have standardized the differentiation of human embryonic and induced pluripotent stem cells to a neuronal fate using defined conditions. This allows reproducibility in replicate experiments and facilitates the direct comparison of cell lines. Since the starting point for EB formation is a single cell suspension, this protocol is suitable for standard and novel methods of pluripotent stem cell culture. Moreover, an intermediate population of neural precursor cells, which are routinely >95% NCAM(pos) and Tra-1-60(neg) by FACS analysis, may be expanded and frozen prior to differentiation allowing a convenient starting point for downstream experiments.

A common variant in the 3'UTR of the GRIK4 glutamate receptor gene affects transcript abundance and protects against bipolar disorder.

Underactivity of the glutamatergic system is an attractive model for the pathophysiology of several major mental illnesses. We previously described a chromosome abnormality disrupting the kainate class ionotropic glutamate receptor gene, GRIK4/KA1, in an individual with schizophrenia and learning disability (mental retardation). We also demonstrated in a case-control study that two physically separated haplotypes within this gene were significantly associated with increased risk of schizophrenia and decreased risk of bipolar disorder, respectively. The latter protective haplotype was located at the 3' end of the gene. We now report the identification from carriers of the protective haplotype of a deletion variant within the 3' untranslated region of the gene. The deletion allele also was found to be negatively associated with bipolar disorder in both initial (P = 0.00000019) and replication (P = 0.0107) case-control studies. Expression studies indicated that deletion-carrying mRNA transcripts were relatively more abundant. We postulate that this may be a direct consequence of the differences in the RNA secondary structures predicted for the insertion and deletion alleles. These data suggest a mechanism whereby the genetic protective effect is mediated through increased kainate receptor expression.

A Lagrangian model of the evolution of the particulate size distribution of vehicular emissions.

The emission inventory for London indicates that nearly 80% of the particulate emissions derive from vehicular sources. Most of this mass is in the form of ultrafine submicrometer particles which are of concern because of their influence on lung function. The prediction of their dispersion in the atmosphere coupled to the physical and chemical transformations which affect their size distribution and concentration are of great importance. This paper reports the first results from a new meso-scale Lagrangian model which follows the particulate emissions and the evolution of their size distribution across the city. The vehicular emissions are based on the published inventory, corrected to time of day, while other emissions are assumed steady. The initial size distributions of background and emitted particles are represented by the sum of three lognormal distributions. Meteorological data are derived from Meteorological Office reports and are preprocessed to obtain the hourly values of boundary layer depth, Monin-Obukov (MO) length, friction velocity, etc., needed for the computation of the vertical dispersion process via eddy diffusivities and the aerodynamic component of the dry deposition process. In the vertical direction, three layers are assumed-surface layer (typically 50 m), canopy layer and one further layer up to the prevailing boundary layer depth. Currently, the model includes wet and dry deposition and coagulation but not chemical reaction, nucleation or deliquescence. Trajectories are evolved for several hours across the city and the number size distributions and mass concentrations (PM10, PM2.5, PM1 and PM0.1) output at each step. This enables the vehicular contributions over and above the background concentration in each size range to be studied in detail. Data from the model have been compared with experimental data for one of the London background sites where particle number size distribution up to 450 nm (SMPS), plus PM10 and PM2.5 (TEOM) data are available.

Carbon monoxide exposure from cooking in snow caves at high altitude.

OBJECTIVE: To determine the physiological consequences of acute CO exposure from cooking in snow caves at 3,200 m. We hypothesized that ambient CO and serum carboxyhemoglobin (COHb) levels would increase and that even low levels of COHb would be associated with symptoms of CO poisoning at high altitude. METHOD: This was a prospective observational study. Twenty-two healthy volunteers age 18 years or older were recruited during a winter camping trip at 3,200 m. Subjects filled out symptom questionnaires, and heart rate (HR), oxygen saturation (SaO2), serum COHb, and ambient CO were all measured before and after cooking inside snow caves. RESULTS: Median age of subjects was 32 years, and 87% were male. The median ambient CO level increased by 17 ppm (IQR, 2-27 ppm), P = .005. Mean serum COHb level rose from 0.3% (IQR, 0.2%-0.4%) to 1.2% (IQR, 0.7%-2.6%) after cooking, for a difference of 1% (IQR, 0.4%-2.3%), P < .001. There were no differences in symptom scores before and after cooking, and there was no significant effect on HR or SaO2. CONCLUSION: A single exposure to CO at 3,200 m increases ambient CO and COHb but not to clinically important levels. Further studies are needed to examine the risks of longer exposures at higher altitudes.

Effect of surgical cricothyrotomy on the unstable cervical spine in a cadaver model of intubation.

Cricothyrotomy is indicated for patients who require an immediate airway and in whom orotracheal or nasotracheal intubation is unsuccessful or contraindicated. Cricothyrotomy is considered safe with cervical spine (c-spine) injury; however, the amount of c-spine movement that occurs during the procedure has not been determined. In this experimental study, an established cadaver model of c-spine injury was used to quantify movement during cricothyrotomy. A complete C5--6 transection was performed by using an osteotome on 13 fresh-frozen cadavers. Standard open cricothyrotomy was performed on each cadaver, with c-spine images recorded in real time on fluoroscopy, then transferred to video and Kodachrome still images. Outcome measures included movement across the C5--6 site with regard to angulation expressed in degrees of rotation and linear measures of axial distraction and anterior-posterior (AP) displacement expressed as a proportion of C5 body width. Data were analyzed by using descriptive statistics to determine mean change from baseline in each of three planes of movement. Significance was assumed if 95% confidence intervals did not include zero. A significant amount of movement was observed with regard to AP displacement (6.3% of C5 width) and axial distraction (-4.5% of C5 width, indicating narrowing of the intervertebral space). These correspond to 1--2 mm AP displacement and less than 1 mm axial compression. No significant angular displacement was observed. In conclusion, cricothyrotomy results in a small but significant amount of movement across an unstable c-spine injury in a cadaver model. This degree of movement is less than the threshold for clinical significance.

Effects of cervical spine immobilization technique and laryngoscope blade selection on an unstable cervical spine in a cadaver model of intubation.

STUDY OBJECTIVE: Orotracheal intubation (OTI) is commonly used to establish a definitive airway in major trauma victims, with several different cervical spine immobilization techniques and laryngoscope blade types used. This experimental, randomized, crossover trial evaluated the effects of manual in-line stabilization and cervical collar immobilization and 3 different laryngoscope blades on cervical spine movement during OTI in a cadaver model of cervical spine injury. METHODS: A complete C5-C6 transection was performed by using an osteotome on 14 fresh-frozen cadavers. OTI was performed in a randomized crossover fashion by using both immobilization techniques and each of 3 laryngoscope blades: the Miller straight blade, the Macintosh curved blade, and the Corazelli-London-McCoy hinged blade. Intubations were recorded in real time on fluoroscopy and then transferred to video and color still images. Outcome measures included movement across C5-C6 with regard to angulation expressed in degrees of rotation and axial distraction and anteroposterior displacement with values expressed as a proportion of C5 body width. Cormack-Lehane visualization grades were also recorded as a secondary outcome measure. Data were analyzed by using multivariate analysis of variance to test for differences between immobilization techniques and between laryngoscope blades and to detect for interactions. Significance was assumed for P values of less than.05. RESULTS: Manual in-line stabilization resulted in significantly less movement than cervical collar immobilization during OTI with regard to anteroposterior displacement. Use of the Miller straight blade resulted in significantly less movement than each of the other 2 blades with regard to axial distraction. The Cormack-Lehane grade was significantly better with manual in-line stabilization versus cervical collar immobilization; no differences were observed between blades. CONCLUSION: Manual in-line stabilization results in less cervical subluxation and allows better vocal cord visualization during OTI in a cadaver model of cervical spine injury. The Miller laryngoscope blade allowed less axial distraction than the Macintosh or Corzelli-London-McCoy blades. The clinical significance of this degree of movement is unclear.

Safety and efficacy of the Rapid Four-Step Technique for cricothyrotomy using a Bair Claw.

The Rapid Four-Step Technique (RFST) has been demonstrated to be faster than standard open crico thyrotomy technique, but may have a higher incidence of cricoid injury with tracheal hook traction applied caudad. The "Bair Claw" is a novel device that may help eliminate these complications. This randomized, experimental trial used a fresh-frozen cadaver model of cricothyrotomy to compare speed and safety between RFST using a Bair Claw and standard open technique. Outcome measures included time to definitive airway, size of largest endotracheal (ET) tube able to be passed, and incidence of complications. We observed that RFST using a Bair Claw was significantly faster than standard open technique. There was no significant difference with regard to size of ET tube able to be passed with RFST using a Bair Claw versus standard open technique, and there was no damage to trachea or larynx observed with either technique. We concluded that RFST using a Bair Claw is faster and appears to be equally safe when compared to standard open technique in a fresh-frozen cadaver model of cricothyrotomy. The two techniques were equal with regard to maximal ET tube size.

Comparison of antibody titers determined by hemagglutination inhibition and enzyme immunoassay for JC virus and BK virus.

A comparison of antibody titers to JC virus (JCV) or BK virus (BKV) was made by hemagglutination inhibition (HI) and enzyme immunoassay (EIA) with 114 human plasma samples. Antibody titers to JCV or BKV determined by HI were lower than those determined by EIA. Nevertheless, as HI titers increased so did EIA titers. When antibody data were compared by the Spearman rank correlation test, highly significant correlations were found between HI and EIA titers. Results obtained by plotting EIA antibody titers for JCV against those for BKV generally showed a reciprocal relationship, i.e., samples with high antibody titers to JCV had lower antibody titers to BKV and vice versa. Some samples, however, had antibody titers to both viruses. Of the samples tested, 25.4% (25 of 114) had HI and EIA antibody titers to JCV and BKV which were identical or closely related. This is not the scenario one would expect for cross-reactive epitopes shared by the two viruses, but one suggesting that these samples were from individuals who had experienced infections by both viruses. Adsorption with concentrated JCV or BKV antigen of sera with high antibody titers to both JCV and BKV and testing by JCV and BKV EIA gave results which support this conclusion. Although 52.6% (51 of 97) of the samples from the Japanese population tested had very high antibody titers (>/=40,960) to either JCV or BKV, none of the samples were found by a dot blot immunoassay to have antibodies which cross-reacted with simian virus 40. The results from this study, in agreement with those of others, suggest that humans infected by JCV or BKV produce antibodies to species-specific epitopes on their VP1 capsid protein, which is associated with hemagglutination and cellular binding.

Comparison of wire-guided cricothyrotomy versus standard surgical cricothyrotomy technique.

We compared a wire-guided cricothyrotomy technique vs. standard surgical cricothyrotomy in terms of accuracy in placement, complications, performance time, incision length, and user preference. We conducted a randomized, crossover controlled trial in which Emergency Medicine (EM) attendings and residents performed cricothyrotomies by both standard and wire-guided techniques (using a commercially available kit) on human cadavers after a 15-min training session. Procedure time, incision length, and physician preference were recorded. Cadavers were inspected for accuracy of placement and complications. Airway placement was accurate in 13 of 15 cases for the standard technique (86.7%), and 14 of 15 cases for the wire-guided technique (93.3%). When comparing wire-guided vs. standard techniques, there were no differences in complication rates or performance times. The wire-guided technique resulted in a significantly smaller mean incision length than the standard technique (0.53 vs. 2.53 cm, respectively, p<0.0001). Overall, 14 of 15 physicians stated that they preferred the wire-guided to the standard technique. Our data suggest that this wire-guided cricothyrotomy technique is as accurate and timely to use as the standard technique and is preferred by our physician operators. In addition, the technique results in a smaller incision on human cadaver models.

Endodontic-orthodontic relationships: a review of integrated treatment planning challenges.

Literature review There is a paucity of information on the concise relationship between endodontics and orthodontics during treatment planning decisions. This relationship ranges from effects on the pulp from orthodontic treatment and the potential for resorption during tooth movement, to the clinical management of teeth requiring integrated endodontic and orthodontic treatment. This paper reviews the literature based on the definition of endodontics and the scope of endodontic practice as they relate to common orthodontic-endodontic treatment planning challenges. Literature data bases were accessed with a focus on orthodontic tooth movement and its impact on the viability of the dental pulp; its impact on root resorption in teeth with vital pulps and teeth with previous root canal treatment; the ability to move orthodontically teeth that were endodontically treated versus nonendodontically treated; the role of previous tooth trauma; the ability to move teeth orthodontically that have been subjected to endodontic surgery; the role of orthodontic treatment in the provision for and prognosis of endodontic treatment; and, the integrated role of orthodontics and endodontics in treatment planning tooth retention. Orthodontic tooth movement can cause degenerative and/or inflammatory responses in the dental pulp of teeth with completed apical formation. The impact of the tooth movement on the pulp is focused primarily on the neurovascular system, in which the release of specific neurotransmitters (neuropeptides) can influence both blood flow and cellular metabolism. The responses induced in these pulps may impact on the initiation and perpetuation of apical root remodelling or resorption during tooth movement. The incidence and severity of these changes may be influenced by previous or ongoing insults to the dental pulp, such as trauma or caries. Pulps in teeth with incomplete apical foramen, whilst not immune to adverse sequelae during tooth movement, have a reduced risk for these responses. Teeth with previous root canal treatment exhibit less propensity for apical root resorption during orthodontic tooth movement. Minimal resorptive/remodelling changes occur apically in teeth that are being moved orthodontically and that are well cleaned, shaped, and three-dimensionally obturated. This outcome would depend on the absence of coronal leakage or other avenues for bacterial ingress. A traumatized tooth can be moved orthodontically with minimal risk of resorption, provided the pulp has not been severely compromised (infected or necrotic). If there is evidence of pulpal demise, appropriate endodontic management is necessary prior to orthodontic treatment. If a previously traumatized tooth exhibits resorption, there is a greater chance that orthodontic tooth movement will enhance the resorptive process. If a tooth has been severely traumatized (intrusive luxation/avulsion) there may be a greater incidence of resorption with tooth movement. This can occur with or without previous endodontic treatment. Very little is known about the ability to move successfully teeth that have undergone periradicular surgical procedures. Likewise, little is known about the potential risks or sequelae involved in moving teeth that have had previous surgical intervention. Especially absent is the long-term prognosis of this type of treatment. During orthodontic tooth movement, the provision of endodontic treatment may be influenced by a number of factors, including but not limited to radiographic interpretation, accuracy of pulp testing, patient signs and symptoms, tooth isolation, access to the root canal, working length determination, and apical position of the canal obturation. Adjunctive orthodontic root extrusion and root separation are essential clinical procedures that will enhance the integrated treatment planning process of tooth retention in endodontic-orthodontic related cases.

Trends in hydrocarbon fleet emissions at four UK highway sites.

The on-road hydrocarbon emissions of vehicles at four sites in the UK have been monitored using remote sensing equipment. The single lane highway sites in London (Haringey and Southwark), Middlesborough and Leicester were monitored between May 1994 and October 1995. Analysis of the results shows that there is both a large majority of low emitting vehicles which contribute little to fleet hydrocarbon emissions and a small minority of high emitting vehicles which contribute significant proportions to fleet hydrocarbon emissions at all sites. This also results in a skewing of the data set so that a pattern of high mean values and lower median values is consistently observed. Analysis of model year data suggests a low association between vehicle age and mean hydrocarbon emissions for vehicles produced prior to 1983 but the relationship improves after 1983 with regression analyses giving r2 values as high as 0.96. Relatively new high polluting vehicles are the greatest contributors to fleet emissions with, on average, 52% of hydrocarbon fleet emissions being produced by these vehicles from model years 1985-1991. Therefore, fleet emissions could be significantly reduced if new highly polluting vehicles were subject to regular emissions testing followed by appropriate remedial action or were removed from the highway by the withdrawal of their vehicle registration. Older vehicles play a minor role in fleet emissions with, on average, only 13% of hydrocarbon fleet emissions being produced by vehicles registered prior to 1983.

Social awareness of air quality information.

In the UK, air quality information is made available through a range of media. However, limited attention has been paid to ensuring that the information is provided to the public in a format that is understandable and relevant to their needs. This research has begun the task of determining the nature and extent of public air quality requirements by performing a social survey (using a postal questionnaire) to provide a basic snapshot of the public's views and by determining the views of information providers and interested professionals. The paper identifies the main shortcomings in current public air quality information provision. The social survey and workshop results demonstrate that current information provision and dissemination does not match public requirements; the depth and breadth of local information needs to be enhanced. Local authorities need to improve their co-ordination and collaboration, the role of the mass-media needs to be considered carefully, air quality needs to be better described and its implications for individuals spelled out and certain public groups need special consideration. In addition, local authorities need more guidance on communicating air quality, possibly through a best practice guide. Further research is required to identify the best descriptors for air quality, to improve the effectiveness of public advice during episodes of poor air quality and to use public air quality information to effect behavioral changes.

Monitoring children's personal exposure to airborne particulate matter in London, UK--method development and study design.

The paper outlines the methodology selected for identifying the personal exposure of children to airborne particulate matter in a UK urban environment.

Reversal of midazolam-induced laryngospasm with flumazenil.

Midazolam is a commonly used benzodiazepine that is ideal for conscious sedation during a variety of procedures. Laryngospasm is listed by the manufacturer as a rare side effect of midazolam. Flumazenil is a competitive inhibitor of the benzodiazepines that is available for reversal at the end of such a procedure or when too much sedation has been achieved. We present a case of a 61-year-old man who was to undergo conscious sedation before cardioversion. Shortly after receiving midazolam, he developed laryngospasm and resultant respiratory distress; the laryngospasm was reversed by a small dose of flumazenil. The symptoms recurred after approximately 25 minutes and were again reversed with flumazenil. Neither midazolam-induced laryngospasm nor its reversal with flumazenil have previously been reported. A brief discussion regarding laryngospasm and the use of flumazenil are included.

Ethylene glycol poisoning: case report of a record-high level and a review.

Ethylene glycol is commonly found in automobile antifreeze and a variety of other commercial products. Ingestion of ethylene glycol, either accidentally or in a suicide attempt, is characterized by severe acidosis, calcium oxalate crystal formation and deposition, and a wide variety of end organ effects that may be fatal. We present a case of a patient who ingested a massive amount of ethylene glycol in a suicide attempt and yet survived with minimal sequelae. A comprehensive review of the literature on the pathology and pathophysiology of ethylene glycol toxicity on each organ system is provided, along with information on diagnosis and current treatment recommendations.

The efficacy and comfort of full-body vacuum splints for cervical-spine immobilization.

We performed a prospective crossover study to determine the cervical spine immobilization and comfort level of healthy subjects on a full-body vacuum splint in comparison with a standard backboard, with and without cervical spine collars. Twenty-six healthy volunteers were immobilized on a backboard (BB) and a full-body vacuum splint (VS), both with and without a cervical collar (CC). Pre- and post-immobilization cervical spine range-of-motion measurements were made using an electronic digital inclinometer and a standard handheld goniometer. Subjects were also asked to subjectively grade their immobilization and discomfort both overall and in seven specific body regions. No statistically significant difference was found between the VS+CC and the BB+CC for flexion and rotation, although the VS+CC combination provided significantly superior immobilization to the BB+CC for extension and lateral bending. The VS alone, in all cases except extension, provided superior immobilization to the BB alone. A statistically significant difference in subjective perception of immobilization was noted, with the BB being less effective than the other three alternatives and the VS+CC providing the best immobilization. A significant difference in overall comfort and occipital region comfort, favoring the vacuum splint, was found. In conclusion, the vacuum splint is an effective and more comfortable alternative to the background for cervical spine immobilization.

RNase inhibition of human immunodeficiency virus infection of H9 cells.

Onconase and bovine seminal RNase, two members of the RNase A superfamily, inhibit human immunodeficiency virus type 1 replication in H9 leukemia cells 90-99.9% over a 4-day incubation at concentrations not toxic to uninfected H9 cells. Two other members of the same protein family, bovine pancreatic RNase A and human eosinophil-derived neurotoxin, have no detectable antiviral activity, demonstrating a strikingly selective antiviral activity among homologous ribonucleases. The antiviral RNases do not appear to affect viral particles directly but inhibit replication in host cell cultures. Onconase, already in clinical trials for cancer therapy, and bovine seminal RNase have potential as antiviral therapeutics.

In situ hybridization detection of HTLV-I RNA in peripheral blood mononuclear cells of TSP/HAM patients and their spouses.

This is the first report of the direct detection of HTLV-I RNA in uncultured peripheral blood mononuclear cells (PBMNC's) of patients with tropical spastic paraparesis and HTLV-I-associated myelopathy (TSP/HAM) and their spouses, using the technique of in situ hybridization. Twenty-one Colombian patients were tested, all of whom had antibodies to HTLV-I; the presence of HTLV-I proviral DNA in their PBMNC's was confirmed by the polymerase chain reaction technique. Of the 21 patients 15 had a clinical diagnosis of tropical spastic paraparesis (TSP/HAM), 5 were asymptomatic relatives, and 1 patient had leukemia. In situ hybridization was positive in samples from 5 patients; 2 of these were TSP/HAM patients and the other 3 were healthy wives of TSP/HAM patients. This study demonstrates for the first time that viral RNA is expressed in uncultured PBMNC's of some patients with TSP/HAM in whom proviral DNA is also present; furthermore, the detection of HTLV-I RNA in the blood of female partners of TSP/HAM patients clearly illustrates the high likelihood of HTLV-I transmission through sexual contact.