Effects of epigenetic pathway inhibitors on corticotroph tumour AtT20 cells.

Medical treatments for corticotrophinomas are limited, and we therefore investigated the effects of epigenetic modulators, a new class of anti-tumour drugs, on the murine adrenocorticotropic hormone (ACTH)-secreting corticotrophinoma cell line AtT20. We found that AtT20 cells express members of the bromo and extra-terminal (BET) protein family, which bind acetylated histones, and therefore, studied the anti-proliferative and pro-apoptotic effects of two BET inhibitors, referred to as (+)-JQ1 (JQ1) and PFI-1, using CellTiter Blue and Caspase Glo assays, respectively. JQ1 and PFI-1 significantly decreased proliferation by 95% (P < 0.0005) and 43% (P < 0.0005), respectively, but only JQ1 significantly increased apoptosis by >50-fold (P < 0.0005), when compared to untreated control cells. The anti-proliferative effects of JQ1 and PFI-1 remained for 96 h after removal of the respective compound. JQ1, but not PFI-1, affected the cell cycle, as assessed by propidium iodide staining and flow cytometry, and resulted in a higher number of AtT20 cells in the sub G1 phase. RNA-sequence analysis, which was confirmed by qRT-PCR and Western blot analyses, revealed that JQ1 treatment significantly altered expression of genes involved in apoptosis, such as NFκB, and the somatostatin receptor 2 (SSTR2) anti-proliferative signalling pathway, including SSTR2. JQ1 treatment also significantly reduced transcription and protein expression of the ACTH precursor pro-opiomelanocortin (POMC) and ACTH secretion by AtT20 cells. Thus, JQ1 treatment has anti-proliferative and pro-apoptotic effects on AtT20 cells and reduces ACTH secretion, thereby indicating that BET inhibition may provide a novel approach for treatment of corticotrophinomas.

Epigenetic pathway inhibitors represent potential drugs for treating pancreatic and bronchial neuroendocrine tumors.

Cancer is associated with alterations in epigenetic mechanisms such as histone modifications and methylation of DNA, and inhibitors targeting epigenetic mechanisms represent a novel class of anti-cancer drugs. Neuroendocrine tumors (NETs) of the pancreas (PNETs) and bronchus (BNETs), which may have 5-year survivals of <50% and as low as 5%, respectively, represent targets for such drugs, as >40% of PNETs and ~35% of BNETs have mutations of the multiple endocrine neoplasia type 1 (MEN1) gene, which encodes menin that modifies histones by interacting with histone methyltransferases. We assessed 9 inhibitors of epigenetic pathways, for their effects on proliferation, by CellTiter Blue assay, and apoptosis, by CaspaseGlo assay, using 1 PNET and 2 BNET cell lines. Two inhibitors, referred to as (+)-JQ1 (JQ1) and PFI-1, targeting the bromo and extra terminal (BET) protein family which bind acetylated histone residues, were most effective in decreasing proliferation (by 40-85%, P<0.001) and increasing apoptosis (by 2-3.6 fold, P<0.001) in all 3 NET cell lines. The anti-proliferative effects of JQ1 and PFI-1 remained present for at least 48 hours after removal of the compound. JQ1, but not PFI-1, had cell cycle effects, assessed by propidium iodide staining and flow cytometry, resulting in increased and decreased proportions of NET cells in G1, and S and G2 phases, respectively. RNA Sequencing analysis revealed that these JQ1 effects were associated with increased histone 2B expression, and likely mediated through altered activity of bromodomain-containing (Brd) proteins. Assessment of JQ1 in vivo, using a pancreatic beta cell-specific conditional Men1 knockout mouse model that develops PNETs, revealed that JQ1 significantly reduced proliferation (by ~50%, P<0.0005), assessed by bromodeoxyuridine incorporation, and increased apoptosis (by ~3 fold, P<0.0005), assessed by terminal deoxynucleotidyl transferase dUTP nick end labelling, of PNETs. Thus, our studies demonstrate that BET protein inhibitors may provide new treatments for NETs.

Clustering of lifestyle risk behaviours among residents of forty deprived neighbourhoods in London: lessons for targeting public health interventions.

BACKGROUND: Clustering of lifestyle risk behaviours is very important in predicting premature mortality. Understanding the extent to which risk behaviours are clustered in deprived communities is vital to most effectively target public health interventions. METHODS: We examined co-occurrence and associations between risk behaviours (smoking, alcohol consumption, poor diet, low physical activity and high sedentary time) reported by adults living in deprived London neighbourhoods. Associations between sociodemographic characteristics and clustered risk behaviours were examined. Latent class analysis was used to identify underlying clustering of behaviours. RESULTS: Over 90% of respondents reported at least one risk behaviour. Reporting specific risk behaviours predicted reporting of further risk behaviours. Latent class analyses revealed four underlying classes. Membership of a maximal risk behaviour class was more likely for young, white males who were unable to work. CONCLUSIONS: Compared with recent national level analysis, there was a weaker relationship between education and clustering of behaviours and a very high prevalence of clustering of risk behaviours in those unable to work. Young, white men who report difficulty managing on income were at high risk of reporting multiple risk behaviours. These groups may be an important target for interventions to reduce premature mortality caused by multiple risk behaviours.

Impact of preoperative evaluation of tumour grade by core needle biopsy on clinical risk assessment and patient selection for adjuvant systemic treatment in breast cancer.

BACKGROUND: Histological characteristics are important when making a decision on adjuvant systemic treatment in breast cancer. Preoperative assessments of core needle biopsy (CNB) specimens are becoming increasingly relevant as novel minimally invasive ablative techniques are introduced, because a surgical specimen is no longer obtained with these methods. The clinical impact of potential underestimation of tumour grade on preoperative CNB on clinical decision-making was evaluated. METHODS: Histological tumour grade was reassessed on CNB and resection specimens from consecutive invasive ductal carcinomas diagnosed between 2010 and 2013. For each patient, the indication for systemic therapy was assessed, based on either CNB or surgical excision, in combination with clinical characteristics and imaging findings. The clinical impact of discordance between tumour grade on CNB versus the resection specimen was assessed. RESULTS: The analysis included 213 invasive ductal carcinomas in 199 patients. Discordance in tumour grade between CNB and the resection specimen was observed in 64 (30.0 per cent) of 213 tumours (κ = 0.53, 95 per cent c.i. 0.43 to 0.63). A decision on adjuvant treatment based on CNB would have resulted in overtreatment in seven (3.5 per cent) and undertreatment in three (1.5 per cent) of 199 patients. In the undertreated patients, incorrect omission of adjuvant systemic treatment would have increased the predicted 10-year mortality rate by 2.6-5.2 per cent and 10-year recurrence rate by 8.2-15.3 per cent based on the online risk assessment tool Adjuvant! CONCLUSION: The substantial discordance in tumour grading between CNB and resection specimens from breast cancer affects the indication for adjuvant therapy in only a small minority of patients with invasive ductal carcinoma. Assessment of tumour grade by CNB is feasible and accurate for the planning of postoperative treatment.

TGF-ß/NF1/Smad4-mediated suppression of ANT2 contributes to oxidative stress in cellular senescence.

Oxidative stress and persistent activation of DNA damage response (DDR) are causally involved in the development of cellular senescence, a phenomenon implicated in fundamental (patho)physiological processes such as aging, fetal development and tumorigenesis. Here, we report that adenine nucleotide translocase-2 (ANT2) is consistently down-regulated in all three major forms of cellular senescence: replicative, oncogene-induced and drug-induced, in both normal and cancerous human cells. We previously reported formation of novel NF1/Smad transcription repressor complexes in growth-arrested fibroblasts. Here we show that such complexes form in senescent cells. Mechanistically, binding of the NF1/Smad complexes to the NF1-dependent repressor elements in the ANT2 gene promoter repressed ANT2 expression. Etoposide-induced formation of these complexes and repression of ANT2 were relatively late events co-incident with production and secretion of, and dependent on, TGF-ß. siRNA-mediated knock-down of ANT2 in proliferating cells resulted in increased levels of reactive oxygen species (ROS) and activation of the DDR. Knock-down of ANT2, together with etoposide treatment, further intensified ROS production and DNA damage signaling, leading to enhanced apoptosis. Together, our data show that TGF-ß-mediated suppression of ANT2 through NF1/Smad4 complexes contributes to oxidative stress and DNA damage during induction of cellular senescence.

Influence of body mass index on mortality after surgery for perforated peptic ulcer.

BACKGROUND: Body mass index (BMI) is a strong predictor of mortality in the general population. In spite of the medical hazards of obesity, a protective effect on mortality has been suggested in surgical patients: the obesity paradox. The aim of the present nationwide cohort study was to examine the association between BMI and mortality in patients treated surgically for perforated peptic ulcer (PPU). METHODS: This was a national prospective cohort study of all Danish patients treated surgically for PPU between 1 February 2003 and 31 August 2009, for whom BMI was registered. Non-surgically treated patients and those with malignant ulcers were excluded. The primary outcome measure was 90-day mortality. The association between BMI and mortality was calculated as crude and adjusted relative risks (RRs) with 95 per cent confidence intervals (c.i.). RESULTS: Of 2668 patients who underwent surgical treatment for PPU, 1699 (63.7 per cent) had BMI recorded. Median age was 69.4 (range 17.6-100.9) years and 53.7 per cent of the patients were women. Some 1126 patients (66.3 per cent) had at least one of six co-morbid diseases; 728 (42.8 per cent) had an American Society of Anesthesiologists grade of III or more. A total of 471 patients (27.7 per cent) died within 90 days of surgery. Being underweight was associated with a more than twofold increased risk of death following surgery for PPU (adjusted RR 2.26, 95 per cent c.i. 1.37 to 3.71). No statistically significant association was found between obesity and mortality. CONCLUSION: Being underweight was associated with increased mortality in patients with PPU, whereas being overweight or obese was neither protective nor an adverse prognostic factor.

Immune cell subtyping in the cerebrospinal fluid of patients with neurological diseases.

The analysis of cerebrospinal fluid (CSF) with the assessment of CSF cell counts and proteins is an important method in the diagnostic workup of neurological diseases. As an addition to this standard approach, we here present data on the distribution of CSF immune cell subsets in common neurological diseases, and provide reference values along with cases of rare neurological diseases. CD4+ and CD8+ T cells, the CD4/CD8 ratio, B cells, plasmablasts, monocytes and NK cells in the CSF of 319 patients with inflammatory or non-inflammatory neurological diseases were analysed by seven-color flow cytometry. Diagnoses included headache, idiopathic intracranial hypertension, Guillain-Barré syndrome, multiple sclerosis, Lyme neuroborreliosis, bacterial and viral meningitis, human immunodeficiency virus (HIV) infection, stroke, and CNS malignancies, among others. T cells were the predominant population in the CSF with CD4+ T cells being more prevalent than CD8+ T cells. Mostly in HIV patients, and under other conditions of immunosuppression, CD4+ and CD8+ T cells were significantly altered and the CD4/CD8 ratio reduced. B cells and plasmablasts could hardly be detected in non-inflammatory diseases but were consistently elevated in inflammatory diseases. Monocytes were reduced in neuroinflammation and showed a negative correlation with B cells. NK cells were slightly elevated in neuroinflammation. Both monocytes and NK cells were slightly elevated in CNS malignancies. The analysis of immune cell subsets in the CSF adds valuable information to clinicians and is a promising tool for the differential diagnosis of neurological diseases.

Clinical relevance of cranial nerve injury following carotid endarterectomy.

OBJECTIVES: The benefit of carotid endarterectomy (CEA) may be diminished by cranial nerve injury (CNI). Using a quality improvement registry, we aimed to identify the nerves affected, duration of symptoms (transient vs. persistent), and clinical predictors of CNI. METHODS: We identified all patients undergoing CEA in the Vascular Study Group of New England (VSGNE) between 2003 and 2011. Surgeon-observed CNI rate was determined at discharge (postoperative CNI) and at follow-up to determine persistent CNI (CNIs that persisted at routine follow-up visit). Hierarchical multivariable model controlling for surgeon and hospital was used to assess independent predictors for postoperative CNI. RESULTS: A total of 6,878 patients (33.8% symptomatic) were included for analyses. CNI rate at discharge was 5.6% (n = 382). Sixty patients (0.7%) had more than one nerve affected. The hypoglossal nerve was most frequently involved (n = 185, 2.7%), followed by the facial (n = 128, 1.9%), the vagus (n = 49, 0.7%), and the glossopharyngeal (n = 33, 0.5%) nerve. The vast majority of these CNIs were transient; only 47 patients (0.7%) had a persistent CNI at their follow-up visit (median 10.0 months, range 0.3-15.6 months). Patients with perioperative stroke (0.9%, n = 64) had significantly higher risk of CNI (n = 15, CNI risk 23.4%, p < .01). Predictors for CNI were urgent procedures (OR 1.6, 95% CI 1.2-2.1, p < .01), immediate re-exploration after closure under the same anesthetic (OR 2.0, 95% CI 1.3-3.0, p < .01), and return to the operating room for a neurologic event or bleeding (OR 2.3, 95% CI 1.4-3.8, p < .01), but not redo CEA (OR 1.0, 95% CI 0.5-1.9, p = .90) or prior cervical radiation (OR 0.9, 95% CI 0.3-2.5, p = .80). CONCLUSIONS: As patients are currently selected in the VSGNE, persistent CNI after CEA is rare. While conditions of urgency and (sub)acute reintervention carried increased risk for postoperative CNI, a history of prior ipsilateral CEA or cervical radiation was not associated with increased CNI rate.

Surgical delay is a critical determinant of survival in perforated peptic ulcer.

BACKGROUND: Morbidity and mortality following perforated peptic ulcer (PPU) remain substantial. Surgical delay is a well established negative prognostic factor, but evidence derives from studies with a high risk of bias. The aim of the present nationwide cohort study was to evaluate the adjusted effect of hourly surgical delay on survival after PPU. METHODS: This was a cohort study including all Danish patients treated surgically for PPU between 1 February 2003 and 31 August 2009. Medically treated patients and those with a malignant ulcer were excluded. The associations between surgical delay and 30-day survival are presented as crude and adjusted relative risks (RRs) with 95 per cent confidence intervals (c.i.). RESULTS: A total of 2668 patients were included. Their median age was 70·9 (range 16·2-104·2) years and 55·4 per cent (1478 of 2668) were female. Some 67·5 per cent of the patients (1800 of 2668) had at least one of six co-morbid diseases and 45·6 per cent had an American Society of Anesthesiologists fitness grade of III or more. A total of 708 patients (26·5 per cent) died within 30 days of surgery. Every hour of delay from admission to surgery was associated with an adjusted 2·4 per cent decreased probability of survival compared with the previous hour (adjusted RR 1·024, 95 per cent c.i. 1·011 to 1·037). CONCLUSION: Limiting surgical delay in patients with PPU seems of paramount importance.

Presenting features and early management of childhood intermittent exotropia in the UK: inception cohort study.

AIM: To investigate factors associated with early management of intermittent exotropia (X(T)) in hospital eye departments in the UK in a prospective cohort study. METHODS: An inception cohort of 460 children aged <12 years with previously untreated X(T) (mean age 3.6 years, 55.9% girls) was recruited from 26 UK hospital children's eye clinics and orthoptic departments. Participants received a standard ophthalmic examination at recruitment and orthoptic assessment at three-monthly intervals thereafter. The influence of severity of exotropia (control measured by Newcastle Control Score (NCS), and angle of strabismus, visual acuity and stereoacuity) and age on the type of management was investigated. RESULTS: Within the first 12 months following recruitment, 297 (64.6%) children received no treatment, either for impaired visual acuity or for strabismus. Ninety-six (21%) children had treatment for impaired visual acuity. Eighty-nine (19.4%) received treatment for strabismus (22 of whom also received treatment for defective visual acuity); in 54 (11.7%) treatment was non-surgical and in 35 (7.6%) eye muscle surgery was performed. Children with poor (score 7-9) control of strabismus at recruitment were more likely to have surgery than children with good (score 1-3) control (p<0.001). Children who had no treatment were younger (mean age 3.38 years) than those who were treated (mean 4.07 years) (p<0.001). Stereoacuity and size of the angle of strabismus did not influence the type of management received. CONCLUSIONS: X(T) can be a presenting sign of reduced visual acuity. Most children with well controlled X(T) receive no treatment within 12 months following presentation.

Distance stereoacuity in intermittent exotropia.

BACKGROUND: Studies of distance stereoacuity in intermittent exotropia suggest that normal stereoacuity corresponds to good control of the deviation and that reduced or negative stereoacuity signifies poorer control. AIM: : To evaluate distance stereoacuity in intermittent exotropia using the Frisby Davis Distance stereo test (FD2). METHODS: Children with intermittent exotropia where the near angle was less than or equal to distance were eligible for recruitment. Standardised prospective data collection included FD2 distance stereoacuity. This was a longitudinal study in which outcomes are reported for baseline, last follow-up (> or =6 months before any surgery) or preoperative and last postoperative visits for those undergoing surgery. RESULTS: 110 children with intermittent exotropia had FD2 stereoacuity tested at baseline: 70 comprehended the test. Mean (standard deviation (SD)) age was 4.6 (1.7) years (range 2-10 years). 41/70 (59%) showed positive responses: mean (SD) stereoacuity 30 (12) s of arc. The mean follow-up period before any surgery was 13 months (range 6-27 months). At follow-up, mean (SD) stereoacuity was 24 (11) s of arc. Preoperative and postoperative stereoacuity were not significantly different from those not undergoing surgery. CONCLUSION: This study was the first to report distance stereoacuity in intermittent exotropia using the FD2 stereo test: patients with intermittent exotropia can achieve normal levels of distance stereoacuity, but a considerable proportion, despite comprehending, showed a negative response. This suggests that using the FD2, distance stereoacuity in intermittent exotropia is either absent or normal rather than reduced. Possible reasons for this and its implications are discussed.

Analysis of the C/T(-1) single nucleotide polymorphism in the CD40 gene in multiple sclerosis.

The costimulatory CD40-CD40L pathway plays a critical role in the generation and maintenance of adaptive immune responses. Genetic interference of CD40-CD40L interactions strongly influences the onset and course in many autoimmune disease models including experimental autoimmune encephalomyelitis. We analysed the association of a single nucleotide polymorphism of the CD40 gene (C/T(-1)) in 287 patients with multiple sclerosis (MS) and 184 matched controls. No significant differences were found in the frequency of the C/T(-1) polymorphism between the patients with MS and the controls (53% vs 49%) or among different MS subtypes. Cell surface expression of CD40 did not differ within the different genotypes, but carriers of the T allele showed a trend for a lower stimulatory index compared with individuals with the CC genotype. Although these subtle differences indicate functional consequences in the immune stimulatory capabilities related to the CD40 C/T(-1) polymorphism, our population-based study found no association with disease susceptibility or disease course in MS.

Potential and limitations of the automatic detection of fiducial markers using an amorphous silicon flat-panel imager.

Amorphous silicon electronic portal imaging devices (a-Si EPIDs) allow fast acquisition of high resolution portal images (PI). A visualization of organ movement for adaptive image-guided radiotherapy (IGRT) can be reached by implantation and automatic detection of fiducial markers. A method of automatic detection has been developed for fiducial spherical tungsten markers on PIs, acquired with an a-Si flat-panel imager. The detection method consists of a 2D Mexican hat filter (MHF), whose parameters are tuned to the particular marker signal. The high selectivity of this filter allows a reliable and precise detection of tungsten markers down to a diameter of 1.5 mm. The presented method allows fast, automatic and unsupervised detection of markers. Inevitably, the detection is hampered by image background (bone structures, etc) and noise. A detection success rate higher than 95% was reached, analysing PIs of patients with markers fixed on their skin. Furthermore, this approach to automatic marker detection can also be generalized to elliptic MHFs for the detection of cylindrical markers. The accuracy and detection probability of this method may allow accurate and fast online localization of the considered organ.

Smoking and quitting with the aid of nicotine replacement therapies in the English adult population. Results from the Health Education Monitoring Survey 1995.

BACKGROUND: Many clinical trials have shown the usefulness of nicotine replacement therapies, family physician advice and other cessation aids in enhancing smoking cessation. However, many trials have taken place amongst selected patients and doctors. This paper assesses cessation amongst a representative general adult population in England in order to see whether such aids are as successful amongst more typical smokers. METHODS: Descriptive information on the pattern of smoking cessation aids use in the English population is presented for the first time. This is followed by multivariate probit analysis of the 'Health in England 1995' survey data set in order to predict successful smoking cessation as a function of several variables including use of nicotine gum and patches, willpower, advice from family physicians, social class, education, gender, marital status and family composition. RESULTS: The results tend to confirm that lower social class and lower educational achievement are linked to continuing smoking status. However, users of nicotine gum and patches were found to be less likely to have given up smoking than non-users. This may be explained by the fact that only those who find it difficult to give up will go to the time and expense of considering such aids. These findings could also be explained by the fact that gum and patches have only been available recently in England. The pattern of results also suggests that 'willpower' is a marker for success in cessation as opposed to a determinant of giving up in its own right. CONCLUSION: This research confirms a lot of other research on the determinants of smoking cessation and has for the first time looked at the use and impact of nicotine replacement therapy on giving up in a representative English adult population. More research is needed on observational data sets in order to tease out the true impact of nicotine replacement on cessation rates in representative smoking populations.

Rapid production of recombinant allergens in Nicotiana benthamiana and their impact on diagnosis and therapy.

BACKGROUND: Type I allergies are immunological disorders that afflict a quarter of the world's population. Recombinant allergens have improved the diagnosis of allergic diseases and allow the formulation of new therapeutic approaches. Over 50% of all allergens are of plant origin. OBJECTIVE: We have applied a novel method of overexpressing plant allergens in the tobacco-related species Nicotiana benthamiana. METHOD: This method is based on the use of a chimeric tobacco mosaic virus that harbors a foreign gene sequence and directs its transcription after the infection of the host plant. RESULTS: We have expressed the model allergen Bet v 1, the major birch pollen allergen, and two Hevea brasiliensis latex allergens, the spina-bifida-associated allergens Hev b 1 and Hev b 3, in N. benthamiana using such a viral vector. Bet v 1, Hev b 1 and Hev b 3 produced by this method were recognized by patients' IgE suggesting that the plant-produced allergens were properly folded. Nonpurified Bet v 1 expressed in N. benthamiana leaves had the same immunogenicity as purified Bet v 1 expressed in Escherichia coli or natural Bet v 1 when tested in a murine model of type I allergy. CONCLUSION: We conclude that this plant expression system offers a viable alternative to fermentation-based production of allergens in bacteria or yeasts.

Recombinant human acidic fibroblast growth factor and fibrin carrier regenerates bone.

Bone regeneration promoted by acidic recombinant human fibroblast growth factor (rhFGF-1), rabbit demineralized bone matrix (rDBM), and a fibrin (f) delivery system was measured in critical-sized defects in rabbits' radii. A unilateral segmental defect 20 mm in length was prepared in radii of 48 skeletally mature New Zealand White rabbits divided equally between 4- and 8-week cohorts. The temporal cohorts were divided equally among four treatment groups: rDBM, rDBM/f, rDBM/rhFGF-1/f, and rhFGF-1/f. Data for the fifth group, untreated critical-sized defects, were exploited from previous published reports from this laboratory. In response to experimental treatments, radiomorphometric and histomorphometric methods were used to derive quantitative outcome data that were tested by analysis of variance and post hoc multiple comparison tests (significance p

Direct isolation of human central nervous system stem cells.

Stem cells, which are clonogenic cells with self-renewal and multilineage differentiation properties, have the potential to replace or repair damaged tissue. We have directly isolated clonogenic human central nervous system stem cells (hCNS-SC) from fresh human fetal brain tissue, using antibodies to cell surface markers and fluorescence-activated cell sorting. These hCNS-SC are phenotypically 5F3 (CD133)(+), 5E12(+), CD34(-), CD45(-), and CD24(-/lo). Single CD133(+) CD34(-) CD45(-) sorted cells initiated neurosphere cultures, and the progeny of clonogenic cells could differentiate into both neurons and glial cells. Single cells from neurosphere cultures initiated from CD133(+) CD34(-) CD45(-) cells were again replated as single cells and were able to reestablish neurosphere cultures, demonstrating the self-renewal potential of this highly enriched population. Upon transplantation into brains of immunodeficient neonatal mice, the sorted/expanded hCNS-SC showed potent engraftment, proliferation, migration, and neural differentiation.

BDCA-2, BDCA-3, and BDCA-4: three markers for distinct subsets of dendritic cells in human peripheral blood.

We have generated a panel of mAbs that identify three presumably novel human dendritic cell Ags: BDCA-2, BDCA-3, and BDCA-4. In blood, BDCA-2 and BDCA-4 are expressed on CD11c(-) CD123(bright) plasmacytoid dendritic cells, whereas BDCA-3 is expressed on small population of CD11c(+) CD123(-) dendritic cells. All three Ags are not detectable on a third blood dendritic cell population, which is CD1c(+) CD11c(bright) CD123(dim), or on any other cells in blood. BDCA-4 is also expressed on monocyte-derived and CD34(+) cell-derived dendritic cells. Expression of all three Ags dramatically changes once blood dendritic cells undergo in vitro maturation. BDCA-2 is completely down-regulated on plasmacytoid CD11c(-) CD123(bright) dendritic cells, expression of BDCA-3 is up-regulated on both plasmacytoid CD11c(-) CD123(bright) dendritic cells and CD1c(+) CD11c(bright) CD123(dim) dendritic cells, and expression of BDCA-4 is up-regulated on CD1c(+) CD11c(bright) CD123(dim) dendritic cells. BDCA-2 is rapidly internalized at 37 degrees C after mAb labeling. The three presumably novel Ags serve as specific markers for the respective subpopulations of blood dendritic cells in fresh blood and will be of great value for their further analysis and to evaluate their therapeutic potential.

Ventricular shunts and the prevalence of sensitization and clinically relevant allergy to latex in patients with spina bifida.

Patients with spina bifida represent the highest-risk group for developing hypersensitivity to latex. Recognized risk factors for these patients are repeated surgery and an atopic disposition. Our aim was to study the influence of a ventricular shunt as an independent risk factor. One hundred and sixty-one patients with spina bifida (median age 10 years) were investigated for the presence of a shunting device and the number, type and date of previous surgical interventions. Additionally, skin-prick tests and provocation tests were performed in order to classify sensitized and symptomatic latex-allergic individuals. Eighty-eight patients (54.7%) were sensitized; 55 (34.2%) were provocation-positive. Patients with a shunt system had undergone a significantly higher number of surgical procedures (p < 0.0001) and showed significantly higher levels of immunoglobulin E (IgE)-antibodies to latex (p < 0.0001) than patients without a shunting device. The total number of operations correlated significantly with the level of IgE-antibodies to latex in serum (p < 0.0001), whereas the number of shunt operations in patients with a ventricular shunt did not significantly correlate with the degree of sensitization. In conclusion, patients with spina bifida who have a ventricular shunt are at particularly high risk because they undergo a significantly higher number of operations than patients without a ventricular shunt. However, the ventricular shunt does not seem to be an independent risk factor. Furthermore, this study emphasizes that individuals at high risk, such as patients with spina bifida, should be handled latex-free from the very beginning of life.

Nicotine administration in rabbits using Habitrol nicotine patches and nicotine nasal spray.

1. Several methods are used to provide predictable and effective nicotine to experimental animals in scientific studies. Due to the expense and technical challenges of these methods, we sought suitable alternatives. Consequently, the purpose of the present study was to develop a reliable experimental nicotine protocol in rabbits that included either Habitrol nicotine patches (Novartis Consumer Health Inc., Summit, NJ, USA) or nicotine nasal spray. 2. Administration of one of three doses of nicotine (2.5, 5, or 10 mg) was accomplished daily on 13 rabbits divided between either the patch or spray groups. Systemic nicotine and cotinine levels at 0 h, 15 min and 8 and 24 h were assayed. Data were analysed by a Fisher's protected least significant difference test at P = 0.05. 3. Rabbits treated with Habitrol patches exhibited consistent and predictable systemic nicotine levels. The nicotine nasal spray produced an immediate dose-dependent response with no measurable nicotine serum levels at 8 h. 4. For nicotine administration in rabbits, nicotine patches are easy to administer and provide a nicotine serum level between 5 and 25 ng/mL, which is consistent with the average daily level found in a patient who smokes cigarettes.