Intrathecal antibody synthesis is a well-documented phenomenon in infectious neurological diseases as well as in demyelinating diseases. Intrathecal antibody synthesis against HTLV-1 has been reported in HAM/TSP, but little is known about the role of B cells and humoral immune responses in the central nervous systems (CNS) of HAM/TSP patients. Here we demonstrate profiles of HTLV-1-specific antibodies in cerebrospinal fluid (CSF) of HAM/TSP patients. Of 36 HAM/TSP patients, antibody responses against Gag and Tax were detected in CSF of all the patients. CSF/Serum antibody ratio was elevated in anti-Gag (mean 1.20) more than in anti-Tax (mean 0.85), but importantly HAM/TSP patients with lesions or atrophy in spinal cord showed higher CSF/Serum anti-Gag antibody ratio. Antibody response against Env was detected in CSF of 94.4% of patients, but CSF/serum anti-Env ratio was significantly lower than those of anti-Gag and anti-Tax (mean 0.18). 19 patients were further studied for oligoclonal band (OCB) specificity to HTLV-1 antigens and all of them were found to have bands specific for at least one antigen studied. Significantly higher proportion of patients had Gag- or Env-specific OCBs than Tax-specific OCBs. Interestingly, CXCL13 (B cell attracting chemokine-1) was increased in CSF of HAM/TSP patients, which was associated with higher HTLV-1-specific antibody responses in CSF and was correlated with decrease of plasma blasts in peripheral blood. These results highlight the importance of the B cell compartment in HAM/TSP where production of HTLV-1-specific antibody may be required to control viral persistence and/or may be associated with HAM/TSP disease progression.


The resulting clinical impact of the combined use of G-CSF with chemotherapy as a chemosensitizing strategy for treatment of acute myeloid leukemia (AML) patients is still controversial. In this study, the effect of ex vivo treatment with G-CSF on AML primary blasts was studied.Peripheral blood mononuclear cells from AML patients were treated with G-CSF at increasing doses, alone or in co-culture with HS-5 stromal cells. Cell viability and surface phenotype was determined by flow cytometry 72 h after treatment. For clonogenicity assays, AML primary samples were treated for 18 h with G-CSF at increasing concentrations and cultured in methyl-cellulose for 14 days. Colonies were counted based on cellularity and morphology criteria.The presence of G-CSF reduced the overall viability of AML cells co-cultured with bone marrow stroma; whereas, in absence of stroma, a negligible effect was observed. Moreover, clonogenic capacity of AML cells was significantly reduced upon treatment with G-CSF. Interestingly, reduction in the AML clonogenic capacity correlated with the sensitivity to chemotherapy observed in vivo.These ex vivo results would provide a biological basis to data available from studies showing a clinical benefit with the use of G-CSF as a priming agent in patients with a chemosensitive AML and would support implementation of further studies exploring new strategies of chemotherapy priming in AML.The online version of this article (doi:10.1186/s12935-015-0272-3) contains supplementary material, which is available to authorized users. According to the hierarchic model of cancer, AML cells are maintained by a subset of cells, called leukemic stem cells (LSCs), which have the capacity of self-renewal and differentiation . Due to In clinics, the simultaneous administration of G-CSF and chemotherapy as a priming strategy has yielded conflicting results. Thus, some reports have described a favorable effect of G-CSF priming in favorable and the intermediate risk AML patients, without clinical benefit in patients with unfavorable cytogenetics , 7.In coG-CSF is the main cytokine that drives granulopoiesis exerting its function through the G-CSF receptor (G-CSFR). G-CSFR is a single transmembrane receptor that belongs to the cytokine receptor type I superfamily . The intHere, we analyzed the ex vivo effect of G-CSF on primary AML samples in order to elucidate the biological mechanisms underlying chemotherapy priming strategies with this agent –18. Our In order to study the effect of G-CSF on cell viability, bulk AML blast population was treated with increasing doses of G-CSF. In concordance with previous data , no signtranswell system  exclusion and cell count was obtained by volume in a FACSCanto II cytometer (BD). In co-culture experiments, AML cells were discriminated based on their CD45 expression (BD). For CXCR4 determination, cells were stained on the surface with an anti-CXCR4 antibody  and samples were acquired in a FACSCantoII (BD). Flow cytometry fcs files were analyzed using FlowJo software (TriStar) and statistics were calculated in GraphPad (Prism software). Each experimental point was normalized against the mean value corresponding to the vehicle treated control triplicates. A 2-tailed Mann–Whitney test was performed to determinate statistical significance.Five-hundred cells per ml were cultured in 96-well plates in complete IMDM medium (PAA). G-CSF (PrepoTech) was added at different concentrations. Co-culture experiments with stromal cells were performed seeding 30 × 10Membranes with 1 μm pore size  were used in a 24-wells format. A total of 100,000 HS-5 cells were added to the lower compartment. 300,000 primary AML patient samples were added to the upper compartment. Both compartments were filled with complete media as described above. Cells were incubated 24 h and analyzed as for viability assays.AML primary samples were treated for 18 h with G-CSF (PrepoTech) at different concentrations in complete IMDM medium and cultured in MethoCult H4034 (StemCell Technologies) for 14 days. Colonies were counted based on cellularity and morphology criteria.


A plasma stratum (cell free layer or CFL) generated by flowing blood interposed between the red blood cell (RBC) core and the endothelium affects generation, consumption, and transport of nitric oxide (NO) in the microcirculation. CFL width is a principal factor modulating NO diffusion and vessel wall shears stress development, thus significantly affecting NO bioavailability. Since the CFL is bounded by the surface formed by the chaotically moving RBCs and the stationary but spatially non-uniform endothelial surface, its width fluctuates randomly in time and space. We analyze how these stochastic fluctuations affect NO transport in the CFL and NO bioavailability. We show that effects due to random boundaries do not average to zero and lead to an increase of NO bioavailability. Since endothelial production of NO is significantly enhanced by temporal variability of wall shear stress, we posit that stochastic shear stress stimulation of the endothelium yields the baseline continual production of NO by the endothelium. The proposed stochastic formulation captures the natural continuous and microscopic variability, whose amplitude is measurable and is of the scale of cellular dimensions. It provides a realistic model of NO generation and regulation. Nitric oxide (NO) plays a critical role in the local control of smooth muscle tone and the regulation of blood flow at the microvascular level. Its distribution in the microcirculation is determined by the balance between NO production and consumption in the blood and tissue compartments. The local concentration of NO in blood results from the competition between NO diffusing from the endothelium and NO scavenging by hemoglobin in red blood cells (RBCs) or at times dissolved in plasma. Mathematical modeling was used  treat the interface between the RBC column and plasma as a smooth deterministic surface. In the present study, we adopt a more realistic approach by treating the surface between flowing RBCs and the CFL as a random field whose statistics are obtained from experimental studies , are modeled by treating it as a random function of both angular coordinate θ and time t, i.e., r1 = r1 with ω ∈ Ω indicating a realization (“coordinate”) in the probability space Ω. This renders the CFL width w = r2 − r1 random, i.e., w = w. Our goal is to capture the effects of stochastic fluctuations of the RBC-CFL interface r1 on distribution of NO concentration, CNO, in the Krogh tissue cylinder r, θ):0 ≤ r ≤ r4, 0 ≤ θ ≤ 2π}.Our analysis deals with NO transport in the geometrical configuration associated with the standard Krogh tissue cylinder model. We consider an arteriolar cross-section that consists of the RBC-rich core , the concentration CNO satisfies a reaction-diffusion equationIn each region of the computational domain, Di and ki are the diffusion coefficient and degradation (reaction) rate in the i-th region, respectively. These four equations are coupled by the continuity conditions at the interfaces ri ,where − and + indicate the left and right limits of the corresponding quantities at the i-th interface, Fn = Fi · ni is the normal component of Fick's flux Fi = −Di∇CNO at the i-th interface whose outward unit normal is ni, and r = ri. Since only endothelium cells are involved in NO production, r = r4, so thatHere the superscripts r = ri  propagates uncertainty (randomness) in the topology of the RBC-CFL interface r1 through the modeling process, leading to randomly varying NO concentration CNO throughout the Krogh tissue cylinder.The coupling of the reaction-diffusion Equations (1) at the interfaces ur) and angular (uθ) components of flow velocity ur = uθ ≡ 0, its longitudinal component is V = V(r), and NO concentration CNO = CNO(r). This results in advective flux of NO in the blood vessel, u · ∇CNO, that is identically zero. In the stochastic setting with randomly fluctuating CFL width, advective flux u·∇CNO is zero in the mean –(3) implicitly assumes that blood flow is laminar, fully-developed, and incompressible, vessel walls are impermeable to blood flow, NO concentration at the vessel inlet equals that at the vessel outlet, and all the reaction rates are spatially uniform. In the deterministic setting with a uniform CFL width, these assumptions imply that the radial (ki in the endothelium (i = 3) and tissue (i = 4) can be considered constant, the reaction rate in the RBC-rich core (k1) is related to hemoglobin levels. The latter depends on hematocrit H(r) and radial component of blood flow velocity V. Let ks denote a reference rate of NO scavenging by RBCs at a reference level of hematocrit Hs. Then, the NO scavenging rate k1 corresponding to a given hematocrit level Hc is given by Ong et al.  separating the two regions. This implies that flow velocity V is random as well, being given by a solution of corresponding flow equations in random domains ,The first is based on a lubrication approximation in which random geometry parameterizes an otherwise deterministic velocity profile  CFL width. Following Martini et al. (c = 0.1678Hc − 4.348 between a hematocrit level Hc and the viscosity of the RBC core μc. Specifying a (random) radial distribution of hematocrit, H = H, as a step functionwhere i et al.  and manyk1 by combining Equations (4)–(7). First, the system of Equations (5)–(7) was solved using Matlab function “solve” to compute Hc for μp = 1.2 cP and two values of Hs. Then k1 was obtained from Equation (4).enables one to compute the randomly fluctuating NO scavenging rate k1 treats blood as a single-phase fluid with a parabolic velocity profileThe second alternative for obtaining r1 with a radial distribution of hematocrit,This formulation replaces the CFL and the random RBC-CFL interface k1. This approach was used by Ong et al.  as constant.Substituting Equations (8) and (9) into Equations (4) and (5) yields an alternative expression for the NO scavenging rate g et al.  in the dw exerted on the endothelium walls by blood flow. Following Ong et al.  = r2 − r1.where Di in Equation (1) to be the same and equal to D. Its value and the values of the remaining parameters used in our are model are reported in Table In the numerical results reported below we assume the diffusion coefficients Equation  to be thWe represent spatio-temporal variations of the RBC-CFL interface,rt and rθ, respectively. A Reynolds decomposition is used to represent each of these fields, r′. Setting as the product of mutually uncorrelated temporal and angular fluctuations w is its standard deviation.where Since the random field n(ω) are complex zero-mean random variables, whose real and imaginary parts are mutually independent for all n. Each has zero mean and variance provides its natural representation. Here the eigenvalues νCθ isare coefficients of the Fourier cosine expansion of a 2π-periodic covariance function Cθ of the random field lθ. The decay of the Fourier cosine coefficients Cn determines the number of terms Nθ in the expansion in Equation (13) that is required to achieve a given truncation error. As the correlation length lθ decreases, Nθ increases.Then We represent the random field Ym(ω) (m ≥ 1) are independent random variables, and λm and fm(t) are, respectively, the eigenvalues and eigenfunctions of Fredholm equations,where lt > 0, the eigenvalue problems in Equation (17) admit an analytical solution  depends on the correlation length lt. The smaller the correlation length, the more terms Nt are necessary to represent the random field r′ with a given degree of accuracy.where γlθ and lt. Experimental data, such as those reported by Kim et al. , which maps the original stochastic domain B = {:−1 ≤ ξ1 ≤ 1, 0 ≤ ξ2 ≤ 2π}. A mapping We introduce a new coordinate system  is represented by the expansions described above.where The mapping defined by Equation (19) renders the transformation JacobianK = 2Nθ + Nt independent random variables Nt variables Y1, …, YNt coincide with those introduced in Equation (16) and the remaining 2Nθ variables represent their counterparts in Equation (13), such that YNt + 1 = νNθ−, …, YK = νNθ. Consequently, the deterministic reaction-diffusion Equations (1) are transformed into stochastic equations of the form (Appendix)and other related metrics stochastic, i.e., dependent on a set of A11, A12 = A21, and A22 are given by Equation (A2) in the Appendix.where the random coefficients B can be solved with a variety of well-established techniques, including perturbation-based moment equations –(3) are given in terms of their statistical moments. Ensemble means, e.g., mean NO concentration w = r2 − r1 in which the random RBC-CFL interface r1 is given by Equation (11). These data, which represent temporal fluctuations of w at a single spatial location , give rise to the histogram and auto-correlation reported in Figures r2) indicates that the random field w is non-Gaussian. We fit the histogram in Figure B = Γ(α + β)∕[Γ(α)Γ(β)] is the beta function, Γ(·) is the complete gamma function, 0 ≤ W ≤ r2, and α > 0 and β > 0 are shape parameters. Setting α = 4.358 and β = 32.9 provides the best data fit, resulting in the mean CFL width lt = 0.007 s.The CFL width measurements . In analogy with its temporal counterpart rt, we chose rθ to have the beta distribution with unit mean and variance lθ. In the formulation provided by Equation (12), the amplitude of spatio-temporal (in the angular coordinate θ and time t) fluctuations of both the RBC-CFL interface r1 and CFL width w = r2 − r1 increases with the variances lθ and lt. This behavior, which reflects chaotic motion of RBCs in the blood core, is demonstrated by two representative realizations of the random CFL width shown in Figure Experimental limitations preclude data acquisition at multiple azimuths θ, which requires us to postulate a probabilistic model for w in Equation (10) is inversely proportional to the wall shear stress (WSS) τw. Hence the random spatio-temporal fluctuations in w induce corresponding fluctuations in τw. In the computations of the WSS we use the values of the edge velocity Ve = 0.54 mm/s and the corresponding pressure gradient J = 2.15 × 104 computed by fitting the smooth-wall model to the experimentally observed peak NO concentration of 11.2 nM.The CFL width w.  that the rate of NO production by the endothelium, k1 in Equation (4) given by the constitutive model in Equations (8) and (9). We start by computing a (deterministic) reference NO concentration r1 and r2. It serves as an initial condition for transient stochastic simulations.Unless specified otherwise, the results reported below correspond to the hematocrit-dependent reaction rate r = r2 and r3) are the same, the NO scavenging rate in the RBC core (0 ≤ r ≤ r1) is higher than that in the muscle tissue (r > r3). That is why the peak NO concentration is at the endothelium surface facing the tissue (r = r3).The mean concentration profiles computed with these simulations, r1: the higher CVw and/or the smaller the correlation lengths lt and lθ, the rougher the interface is.Figure The simulation results reported in Table k1 given by the constitutive law in Equations (8), (9), which treats blood as a single-phase fluid. The alternative constitutive model for k1, which explicitly accounts for the CFL presence, is given by Equations (6) and (7). Our simulations demonstrate that the difference between the mean peak NO concentrations predicted with the two models is less than 1% . To match the decrease in the reported peak NO concentration from 11.2 to 9.5 nM, we recalculated the value of the pressure gradient J = 2.15 × 104 and 1.72 × 104.In the experiments reported by Ong et al. , infusioFigure We developed a computational framework to quantify the impact of spatio-temporal fluctuations in the CFL width on the production and transport of NO. This is accomplished by treating the RBC-CFL interface (and the corresponding CFL width) as a space-time correlated random field. This surface is represented via Karhunen-Loéve and Fourier expansions. The differential equations describing blood flow and NO production and transport, defined on random simulation domains, were solved by using a stochastic collocation method.Our analysis demonstrates that the two-phase nature of microcirculatory flow, partitioned between a central blood column and a peripheral cell free plasma layer, causes stochastic flow variability in the CFL that influences NO bioavailability. Our findings are qualitatively comparable to those obtained with the deterministic analysis conducted by Ong et al. . HoweverFlow variability has a homeostatic role in the microcirculation where it is a factor in the control of blood flow and inflammation through biochemical mechanotransduction modulation of the production of NO and prostaglandins. Large Reynolds numbers and flow variability in the central circulation can cause the up-regulation of virtually all atherogenic or pro-inflammatory genes ultimately promoting the development of atherosclerotic plaques  are similar throughout the circulation, as proposed by the “uniform shear stress hypothesis”  or plasma (possibly due to diabetes and hypertension).SP carried out numerical implementation of the proposed algorithms, participated in the design of the study and drafted the manuscript; MI formulated the physiological problem, participated in the design of the study, and helped draft the manuscript; DT designed and coordinated the study, and helped draft the manuscript. All authors gave final approval for publication.This research was supported in part by NIH under award numbers R01-HL064395 and R24-HL64395, Air Force Office of Scientific Research under award number DE-FG02-07ER25815, and by National Science Foundation under award number DMS-1522799.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Background: Literature review suggests that adherence to immunosuppressive drugs may be lower in recipients of living than of deceased donor kidney grafts, possibly because of profile differences.Objective: To compare the level of immunosuppressive adherence levels between patients with deceased and living  donor grafts in Switzerland.Methods: Using data from two similar cross-sectional studies at two transplant centers in Switzerland, the level of adherence between the two groups was compared. Medication adherence was assessed by self-report or electronic monitoring. Possible explanatory factors included age, beliefs regarding immunosuppressive drugs, depressive symptomatology, pre-emptive transplantation, and the number of transplants received, were also considered. Data were analyzed using logistic regression analysis. Results: Unadjusted non-adherence odds were 2 to 3 times higher in living-related than deceased donor transplantation . Adjustment for confounders showed that these differences were associated most with the younger age of living-related subjects and the belief that immunosuppressive drugs are less important for living-related donations. ie, health beliefs regarding their immunosuppressive needs), knowledge of which may enhance adherence if addressed.Conclusion: There is a lower immunosuppressive adherence in recipients of living-related donor kidneys, possibly owing to differences in patient profile ( The studies differed from each other in their measurement of the number of variables reflecting possible reasons for adherence differences, but were similar in their measurement of the key variables, The Zürich study Design, Sample and SettingThe first source of data was a study conducted at the nephrology outpatient clinic at University Hospital Zürich , which iProcedureClinical variables were retrieved from participants’ medical files, including graft donor type  and pre-emptive transplantation status (yes/no). Demographic and adherence data were assessed by interview. The item “how often have you not taken immunosuppressive medication in the past four weeks?” was selected as a measure of “taking adherence” from the 4-item Basel Assessment of Adherence Scale for Immunosuppressives (BAAS-IS) , 17. The Basel study Design, Sample and SettingThe second source of data was a study conducted at the nephrology outpatient clinic of the University Hospital Basel , a similProcedurePatients were recruited during their annual check-up visits to the hospital, when demographic and adherence data were collected. Clinical variables retrieved from the patients’ medical files included graft donor type , pre-emptive transplantation status (yes/no), number of HLA-mismatches, and number of transplants received. Depressive symptomatology was measured using the 21-item, 4-point Beck Depression Inventory , 19. Pat®-V TrackCap system  [® uses a medication bottle fitted with a cap containing a chip registering the time and date of every opening. For this analysis, taking adherence was defined as the ratio of registered openings to the number of prescribed doses [A subsample of patients agreed to electronic adherence monitoring by means of a MEMSx, Ltd.) . The MEMed doses . For anaData analysis 2 and Mann-Whitney U tests. Testing for differences in adherence levels between graft types was done by ordinal logistic regression analysis [The two studies were analyzed separately. Data were not pooled to allow separate corroboration of our main hypothesis in the two different settings. Moreover, the Zürich study did not have the extended set of confounding factors. Variables were described using percentages, means and standard deviations. Simple comparisons of demographic and clinical variables between graft types were performed using χanalysis , which cSample characteristics Sample characteristics are outlined in Differences in non-adherence among graft type groups In both studies living-unrelated graft recipients had adherence levels similar to those of deceased-donor recipients . SimilarA single confounder’s contribution to explaining adherence differences between deceased and living-related transplants was inferred by omitting one explanatory variable at a time from the model , revealiie, that the belief that an individual who receives a living-donor kidney needs less immunosuppressive medication than a deceased-donor kidney recipient contributed more than any other variable to the adherence difference. A sense of immunity to rejection in this group may indeed be a major factor promoting non-adherence [Our analysis confirmed the observation that adherence differences exist between recipients of different types of kidney graft, however, indicated living-related transplantations as the factor responsible for the lower adherence in the larger living donor transplantation group. The living-unrelated recipients showed adherence levels similar to those of deceased donor transplant recipients, which suggests that relatedness is at the core of the observed adherence differences, rather than the fact that the donor is alive and known to the recipient, as is true in most cases. This in turn agrees with our second analytic finding, dherence . While tdherence . The effPre-emptive transplantation ie, n=2). Further evidence that pre-emptive transplantation does not contribute to post-transplantation adherence levels arises from the fact that living-unrelated transplant recipients  show adherence levels similar to those of deceased donor transplants. The seemingly large effect of pre-emptive transplantation on adherence differences observed in the Zürich is thus very likely a statistical deviation invoked by the small sample size of dialysisless transplantations in the deceased donor reference category (A limitation of the study is that we did not have access to variables characterizing the relationship between living-(un)related donors and recipients, such as the perceived closeness or quality of the donor-recipient relationship. The emotional bond of the donor-recipient pair may be an important extra-confounding factor to be investigated.In conclusion, our analysis supported the hypothesis of lower adherence to immunosuppressive medication in living-related kidney transplants, an association we found to be a proxy of underlying risk factors, which the modifiable risk factor “patients beliefs” (rather than experience with chronic illness) is an example of. The finding may help identifying patients at risk for rejection, which is reportedly more frequent and severe in this patient population , and to


Torsion of the appendix testis is a rare cause of scrotal swelling in the neonatal period. We present a case of torsion of the appendix testis in a one-day-old male. We discuss the physical examination and radiologic studies used to make the diagnosis. Nonoperative therapy was recommended and the patient has done well. Recognition of this condition in the neonatal period can prevent surgical intervention and its associated risks. There are several causes of scrotal swelling in the neonate. The most common causes include testicular torsion, neoplasms, supernumerary testis, splenogonadal fusion, and adrenal rests , 2. TorsA one-day-old newborn was found on initial exam to have right sided scrotal swelling and urologic consultation was obtained. The patient was the product of a 39-week gestation born via an uncomplicated vaginal delivery. The pregnancy was notable only for well-controlled maternal diabetes. On examination, the patient was well appearing. His abdomen was soft and without masses. No swelling was appreciated in his groins. He had had a bluish discoloration of the right hemiscrotal skin. Scrotal ultrasonography demonstrated normal testicular morphology and good testicular blood flow. Large right hydrocele was identified but no testicular masses were noted . AdjacenThe appendix testis is a vestige of the Müllerian duct that remains as a nonfunctional remnant during male embryological development . The morEtiologies of scrotal swelling in the neonate are categorized as intratesticular or extratesticular. Intratesticular causes include testicular torsion, neoplasm, and supernumerary testis, whereas extratesticular causes include hematoma, hydrocele, inguinal hernia, and extension of systemic disease . TorsionTorsion of the appendix testis is an extremely rare cause of scrotal swelling in neonates. Ultrasonography is an important diagnostic tool in the diagnosis of a neonate with scrotal swelling. If one suspects appendix testis torsion and emergent causes of scrotal swelling are ruled out, conservative management can be considered.Torsion of the appendix testis is a rare cause of scrotal swelling in neonates. Accurate diagnosis is important as the treatment is nonoperative. Ultrasonography is an important diagnostic tool and if one is confident about the diagnosis, conservative management can be considered.


The processing of information through the spatiotemporal coordination of neuronal activity is still poorly understood . Here we


Objectives: The ortopantomography (OPG) can be a valuable way for an early detection of calcified atheroma plaques, thus contributing for a preliminary stroke risk evaluation. The study looks for the existence of calcified atheroma plates through the use of OPG, comparing the results with the stenosis percentage found through eco-doppler. It has been analyzed the correlation of the number of years as a smoker, arterial hypertension and body mass index, against the risk of having calcified atheroma plaques.Study Design: Observational, transversal and prospective study with 84 patients from the Dental Center of Hospital Particular de Lisboa. First the patients answered to an inquiry and them they were submitted to an OPG and an eco-doppler.Results and Conclusions: It is possible to detect calcified atheroma plaques in the carotid artery through an OPG and patients who have them have got a fifteen fold greater risk of suffering from carotid stenosis. In this study, it has been confirmed the increase in carotid stenosis for long term smokers . The study results show that hypertension patients have a probability 5,426 greater than normal of developing atheroma plaques . Amid analyzed patients, the correlation between obesity and the existence of carotid atheroma plaques was significant, although negative . OPG can help find patients with higher risk of isquemic stroke. Key words:Orthopantomography, Stroke, Carotid disease, Calcified atheroma. A CVA (cerebrovascular accident) is defined as a sudden loss of brain function due to ischemia or hemorrhaging in the central nervous system. In Spain, the mortality rate associated with CVAs has decreased in recent decades, but it is still the second-highest cause of death among men and the first in women . AccordiThere are various significant risk factors to consider regarding carotid atherosclerosis. Age is one such factor, due to both the cellular changes inherent in the aging process and great exposure to other risk factors, such as tobacco, hypertension, cholesterol and obesity . Also, oAdditionally, tobacco use is an important independent risk factor for CVAs (increasing the risk by up to six times) . ArteriaOrthopantomographs (OPGs) are frequently used in dentistry and offer a general view of patients’ upper and lower teeth; they also provide information about the temporomandibular joints and the cervical vertebrae ,7. In 19In a study of 11,854 patients carried out in Rotterdam by the Bots . Based on this, we aim to study the correlation between the presence of calcified plaques on the OPG and the degree of stenosis. From a cardiovascular point of view, we know that stenosis greater than 70% is considered to be high and is closely correlated with an increased risk of CVA .The carotid bifurcation shows hemodynamic changes and a reduction in nitric oxide production, which leads to an increase in vascular damage. The nitric oxide produced in the endothelium regulates different mechanisms such as vasodilatation, inhibition of activation, adhesion and plaque buildup, and also participates in the regulation of basal blood pressure . IndividTo detect the presence of calcified atherormatous plaques in the carotid artery using OPG and their correlation with stenosis found using eco-doppler.To relate the results obtained with age, number of years of tobacco use, obesity and hypertension.An observational, longitudinal and prospective study is carried out with patients of the Servicio de Medicina Dentária (Dentistry Unit) at the Hospital Particular de Lisboa in Lisbon, Portugal. The project has been approved by the Comité Ético de Investigación Clínica del Hospital Particular de Lisboa .After a pilot study for a 95% test power and in accordance with the relevant literature for calculating the sample sizes, the number 42 was arrived at for the patients in each group (42 people with atheromatous plaques in the OPGs and a control group of 42 patients without atheromatous plaques in the OPGs).Patients were selected upon their arrival to the dental medicine office by a dentist who evaluated the OPGs of 1,176 patients (from 2008 to 2012) until finding 42 patients with the presence of calcified atheromatous plaques in their OPGs.The OPGS of all the patients were taken using a Kodak 8000C Digital panoramic and cephalometric system machine .To carry out the study, a questionnaire was prepared . A form explaining the study was provided to them and their consent was requested.The first 42 patients who met the requirements  made up the control group.The duration of the radiographic examination was 14 seconds, with a 6mA current and an 85 kV voltage. An eco-doppler test was later done  on all 84 patients to evaluate carotid flow, determining the percentage of stenosis in those such cases. This examination was done in the Servicio de Cirugía Vascular (Vascular Surgery Unit) in the same hospital.The percentage of stenosis for each patient was entered into the data spreadsheet (using Microsoft Excel 2007), and then statistical analysis of the questionnaire responses and the results was performed using SPSS version 15.0.All OPGs were analyzed by the same dentist, looking for the presence of atheromas in the area of carotid bifurcation (C3-C4) and examining circular or vertical line radiopacities, to differentiate between the OPGs with and without atheromas. The dentist analyzed the x-rays using a computer, focusing on the bifurcation of the carotid arteries, 2 to 4 cm below the mandibular angle.For studying obesity, the universal body mass index formula was used, and patients with a value equal to or greater than 30 were considered obese.Of 1,142 orthopantomographs, 42 showed plaques; in other words, 3.68% of the x-rays presented images com-patible with calcified atheroma.p=0.049, phi coefficient=0.214 using chi-square). 61.9% of the participants who presented calcified atheromatous plaques in the OPGs had diagnosed hypertension (The correlation between having atheromatous plaques and hypertension was significant (rtension .p=0.047), but in a negative direction (phi=-0.218). In the group with atheromatous plaques, there are less patients with obesity (n=3) than in the group without atheromatous plaques (n=8) (The correlation between having plaques and obesity was significant (es (n=8) .p=0.156 and phi=0.309) , but in the group with plaques, the participants who smoked  had been doing so for more years. There is a significant correlation between having plaques and the number of years of smoking (i=0.309) .p=0.034 and OR=1.114) and hypertension (p=0.031 and OR=5.246)  .It was determined that there is a correlation between having atheromatous plaques visible on the OPG and the percentage of stenosis on the eco-doppler, with a confidence interval of 95%. The group with plaques on the OPG had a risk 15 times greater of stenosis greater than 70% on the eco-doppler . Of the Some authors indicate that the age 55 and over population is the one with the highest risk of atheromatous plaques . In thisHypertension is the most predictive factor for CVAs . In thisThe positive correlation between calcifications in the carotid artery and smoking is also identified ,17. In tWhile obesity is positively correlated with the presence of calcified atheroma , we haveThere are many studies that speak of obesity as a risk factor  for cardIt is crucial to always obtain a detailed clinical history to learn about patients’ harmful habits and any prior pathological, personal and family events.In conclusion, we have stated taht identifying calcified atheromatous plaques using panoramic x-rays may detect a specific risk of a CVA when used as a secondary instrument . TherefoDentists are in a unique position; given that they can perform a non-invasive examination such as the OPG, they can contribute to reducing mortality due to CVA.


Organ donation after circulatory death (DCD) has become an accepted strategy to reduce the shortage of organs for transplantation in many European countries. The use and number of DCD donors varies between countries. The purpose of this study was to evaluate the available protocols for DCD in Europe.We contacted national transplant societies and responsible transplant co-ordinators in the countries that perform DCD to obtain DCD protocols. We compared information on the protocols and additional data including: inclusion and exclusion criteria for donation, legislation, determination of death and preservation methods.In ten European countries DCD is performed, eight of which describe the methods in protocols. There are large differences in used DCD categories, legislation and the way death is determined. Protocols differ in the detail in which DCD procedures are described and the way methods are supported by additional consensus statements and ethical frameworks.Although DCD is an established strategy to enlarge the donor pool and to contribute to the reduction of the waiting list for transplantation, its potential has not been fully utilized yet. To further promote DCD transplantation, it is important to share expertise and obtain consensus, so that this can be translated into more uniform and solid protocols supported by the competent authorities, transplant and intensive care professionals, which may eventually result in a further promotion of DCD transplantation in Europe. Kidney transplantation increases life expectancy and quality of life in patients with end-stage renal disease compared with renal dialysis ,2. HowevOne way to expand the organ donor pool is the use of organs from donors who die after circulatory death (DCD). These organs are inevitably subjected to a period of warm ischemia, which adversely affects transplant outcome . DespiteNext to DCD kidney transplantation, which has been a generally accepted source of donor organs in many countries, DCD transplantation of the liver, lung and pancreas is possible and has been increasingly performed in the past decade. The results of DCD liver transplantation are comparable with DBD liver transplantation, with equivalent 1-year and 3-year graft survival and patient survival, although there is a higher risk of biliary strictures in DCD livers . The numIn 1995, at the First International Workshop on Non-heart Beating Donation in Maastricht, four categories of nonheart-beating donation were identified, which include controlled and uncontrolled donors , DCD activity is restricted to one or two hospitals. In the Netherlands and the UK, DCD donation is possible in almost every hospital. Ten other countries are planning to start a DCD program in the near future, including Cyprus, Estonia, Luxembourg, Norway, Poland, Portugal, Romania, Slovak Republic, Slovenia and Sweden. In seven countries  there is no planned DCD activity, mainly because of legal restriction. The most important reason for not performing DCD next to legislation includes organizational difficulties .Table Out of the 10 countries with DCD, seven countries (70%) described inclusion and exclusion criteria of controlled and/or uncontrolled donation. Seven countries actively performed controlled as well as uncontrolled DCD , of which four separated inclusion and exclusion criteria for controlled and uncontrolled donation in their protocols, which mainly included differences in donor age limits. Inclusion criteria for donation generally included positive registration as a donor or permission of the next of kin if a donor had not been registered with a donor registry, witnessed arrest (in uncontrolled DCD) and age limits. The most commonly used exclusion criteria were hypothermia, intoxication, malignancy, sepsis (untreated) and organ-specific contra-indications such as renal failure and liver cirrhosis. Next to the internationally used Maastricht DCD categories, Spain and Italy describe additional categories that include donation after sudden irreversible cardiac arrest in the ICU (Spain) and death during extracorporeal membrane oxygenation . In most countries DCD donors are predominantly Maastricht category III donors – that is, donors who die after withdrawal of treatment – which results in a high percentage of DCD donors in the UK, the Netherlands and Belgium Table . France In all countries with a donor protocol, there was an organ donation law. Although the actual donation act was often not included in the protocol, a reference was always available. Three out of 10 countries 30%) performing DCD in Europe have an opt-in registration system; this means that explicit consent for donation is necessary by donor card or registration in a national registry . In the Netherlands and Switzerland, registration as a donor is sufficient to proceed to organ procurement, and the relatives are informed. In the UK, the opt-in registration system requires additional consent by the next of kin. A registered objection to donation always excludes organ donation.in situ preservation (ISP) in uncontrolled DCD donors to preserve organs if a person has not objected to donation. ISP is a minimally invasive technique to preserve the organs after determination of death and a no-touch period. A special catheter with proximal and distal occlusion balloons is inserted through the femoral artery into the aorta, which enables selective flushing of the kidneys. ISP preserves organ viability and provides the opportunity to meet legal and logistical requirements before organ procurement. ISP also gives relatives time to decide about organ donation.All three protocols allow use of Seven countries use the system of presumed consent for donation. Six out of seven protocols require additional permission from the next of kin; Spain requires written permission. Austria does not specify informed consent of the relatives in its protocol. In clinical practice, in most countries with an opt-out system, the relatives are consulted to confirm the consent for donation. In Latvia, the next of kin has to submit their objection in writing if there is no information in the donor registry. In uncontrolled donors, most protocols allow ISP awaiting the relatives’ consent. In Spain, approval has to be requested from the local court. In most countries, it is allowed or otherwise not specifically mentioned in the protocol for controlled donors, to maintain or optimise the condition of the potential donor after the decision has been made to withdraw treatment if this does not harm the patient or cause discomfort. In Switzerland, (opt-in) consent of the next of kin is required before starting any medical treatment to improve the condition of the donor.lege artis), to more specific guidelines and tests that have to be performed  to detailed protocols with complementary consensus statements and ethical frameworks, as in the UK .Reasons for not performing DCD at all include conflicts with national law or difficulties with the infrastructure to initiate a DCD program . Also, rA notable difference between protocols is the way in which death of the patient is determined. When a deceased patient is an eligible organ donor, both the accurate determination of death and a fast transition from treating a patient to preserving a donor is essential to enable organ preservation with minimized ischemic injury to the organs. Definitions of the criteria to determine death and the no-touch period, to ensure irreversibility of cardiocirculatory arrest, are therefore crucial. Determination of death varies in the different protocols, but also between ICUs. There is a lack of uniformity -21. ThisDonor procurement strategies differ throughout Europe, and the implementation of protocols differs even more. Strategies to procure organs each have their advantages and disadvantages and depend on consent systems, legal opportunities, in-hospital and out-of-hospital institution and financial possibilities. Protocols can be helpful to obtain and maintain support for DCD programs and to help other institutes or countries to develop, implement and continue DCD. The protocols also provide answers to important ethical questions concerning end-of-life management and care .DCD is an established strategy to enlarge the donor pool and to contribute to the reduction of the waiting list for transplantation. The DCD potential in Europe is not yet fully utilized. DCD protocols in European countries and even in individual hospitals are heterogeneous. Restrictions by national laws, organizational problems or ethical struggles are important factors for this. Following these conclusions, we feel it is important that experiences with DCD are shared and consensus is obtained, so these can be translated into more uniform and solid protocols supported by the competent authorities, transplant and intensive care professionals, which may eventually result in a further promotion of DCD transplantation in Europe.• Transplantation of organs from donors after circulatory death (DCD) contributes to the reduction of the waiting list for transplantation, but the DCD potential in Europe is not yet fully utilized.• There is a large variability within European countries to perform DCD and existing DCD protocols are heterogeneous.• Protocols can be helpful to develop, implement and improve DCD performance and answer ethical questions.In situ preservation.DBD: Donation after brain death; DCD: Donation after circulatory death; ISP: The authors declare that they have no competing interests.JW conceived and designed the study, analyzed the data and wrote the manuscript. MF collected the data and revised the manuscript. TCvS collected the data and revised the manuscript. ELWvH designed the study and helped to draft the manuscript. All authors read and approved the final manuscript for publication.


Bacterial meningitis (BM) is characterized by an intense host inflammatory reaction, which contributes to the development of brain damage and neuronal sequelae. Activation of the kynurenine (KYN) pathway (KP) has been reported in various neurological diseases as a consequence of inflammation. Previously, the KP was shown to be activated in animal models of BM, and the association of the SNP AADAT + 401C/T (kynurenine aminotransferase II - KAT II) with the host immune response to BM has been described. The aim of this study was to investigate the involvement of the KP during BM in humans by assessing the concentrations of KYN metabolites in the cerebrospinal fluid (CSF) of BM patients and their relationship with the inflammatory response compared to aseptic meningitis (AM) and non-meningitis (NM) groups.The concentrations of tryptophan (TRP), KYN, kynurenic acid (KYNA) and anthranilic acid (AA) were assessed by HPLC from CSF samples of patients hospitalized in the Giselda Trigueiro Hospital in Natal . The KYN/TRP ratio was used as an index of indoleamine 2,3-dioxygenase (IDO) activity, and cytokines were measured using a multiplex cytokine assay. The KYNA level was also analyzed in relation to AADAT + 401C/T genotypes.In CSF from patients with BM, elevated levels of KYN, KYNA, AA, IDO activity and cytokines were observed. The cytokines INF-γ and IL-1Ra showed a positive correlation with IDO activity, and TNF-α and IL-10 were positively correlated with KYN and KYNA, respectively. Furthermore, the highest levels of KYNA were associated with the AADAT + 401 C/T variant allele.This study suggests a downward modulatory effect of the KP on CSF inflammation during BM. Despite the advances in antimicrobial and intensive care therapies against bacterial meningitis (BM), high mortality and morbidity rates have been observed . Neurolo+de novo synthesis [Bacterial invasion and proliferation within the cerebrospinal fluid (CSF) induce an intense inflammatory response that leads to the activation of several metabolic pathways. One such pathway is the kynurenine (KYN) pathway (KP), which has been shown to be activated in experimental pneumococcal meningitis ,6. The Kynthesis ,9.+ as a cofactor, and its high activity induces energetic depletion, leading to the cell death and neuronal injury typical of BM [+de novo synthesis and KYNA production, the KP is possibly a neuroprotective pathway in BM, despite its neurotoxic metabolites [In the inflamed brain, TRP is metabolized through the KP to form neuroactive metabolites such as QUIN, an agonist of excitotoxic NMDA receptors, showing a cytotoxic effect, and KYNA, a neuroprotector antagonist of these NMDA receptors ,11. Becaal of BM . Due to abolites .Important immunomodulatory properties, mainly related to the immunosuppressive effect of IDO, have also been attributed to this pathway ,9,17. A In a previous work, we identified the association of the SNP AADAT + 401C/T (kynurenine aminotransferase II -– KAT II) with the host immune response to BM, and our results suggested that this SNP may affect the host’s ability to recruit leukocytes to the infection site . This evThis is a prospective study involving 28 patients with the clinical suspicion of meningitis who were admitted to the Giselda Trigueiro Hospital in Natal , a reference center for infectious diseases. Ethical approval for this study was given by Committees on Medical Ethics of the Giselda Trigueiro Hospital and by the National Committee in Ethics (CONEP) with number 0052.1.051.000-05. Informed consent was obtained from each patient participating in this study. For child patients, informed consent was obtained from their parents or legal guardians.Lumbar puncture (LP) was performed to obtain CSF for diagnostic purposes. Twenty-eight CSF samples were collected at the time of admission. Immediately after sampling, the CSF was kept at 4°C before centrifugation . The supernatant were immediately frozen and stored at −80°C until assayed. The CSF samples were anonymized.3), with predominantly polymorphonuclear granulocytes (PMN). Aseptic meningitis (AM) was diagnosed in seven patients by acute onset, fever, meningeal irritation signs, mild increase in protein content and normal glucose levels and the absence of the detection of bacterial pathogens. Because LP is a very invasive procedure, CSF samples were obtained only from patients undergoing procedures for the diagnosis of meningitis. Eight patients had a negative diagnosis for CNS infection and were included as non-meningitis controls (NM) (Table Thirteen patients were diagnosed with BM by the following criteria: 1) positive CSF bacterial culture, (2) detection of the pathogen in the CSF by gram staining plus clinical signs  and/or (3) positive blood culture or Gram stain in the presence of clinical signs of meningitis, (4) positive bacterial antigen detection in the CSF or blood using the latex agglutination test, with clinical signs of meningitis, and (5) clinical signs plus CSF parameters of increased protein content (>40 mg/dL), reduced glucose levels (<40 mg/dL) and the presence of CSF pleocytosis  to quantify the levels of TRP, KYN, KYNA and AA. Briefly, CSF samples were mixed 4:1 with 6% perchloric acid (PCA) to precipitate proteins. The acidified samples were centrifuged 10,000 × g at 4°C for 10 minutes, and the supernatant was filtered through 0.2-μm nylon membranes. An 80-μL aliquot of the filtrate was applied onto a C18 reverse-phase HPLC column  with a guard column . TRP metabolites were eluted isocratically at a flow rate of 0.8 mL/minute with a mobile phase consisting of 100 mM zinc acetate and 3% acetonitrile (v/v), pH 6.2 ,18. TRP plex Kit  with Luminex Technology . The assay was performed according to the manufacturer’s instructions. Samples were diluted to fit within the dynamic range of the assay. The cytokine concentrations were calculated by the Bio-Plex Manager software using a five-parametric logistic standard curve derived from the recombinant cytokine standards provided in the kit.The concentration of six cytokines  was assessed in CSF samples using the human cytokine LincoP < 0.05 was considered to be statistically significant. Multiple comparisons with Tukey’s test were performed if the distribution was normal. Differences between two groups were analyzed with the Mann–Whitney test when the distribution was not normal and with Dunn’s post test when more than two groups were analyzed. The table and graph data are presented as the median ± interquartiles when not following a Gaussian distribution and the mean ± SEM when following a Gaussian distribution.Data were analyzed by analysis of variance (ANOVA), with the Kruskal-Wallis test if needed . A Streptococcus pneumoniae (n = 7), Neisseria meningitidis (n = 1) and Proteus mirabilis (n = 1); for 4 patients, the bacterial species could not be identified. Three patients died during hospitalization. Two belonged to the BM group and 1 to the AM group.Patients were divided into three groups, BM, AM and NM, according to the diagnostic criteria Table . The CSFP < 0.001) and AM (P < 0.05)    and observed a high frequency of the T allele and TT genotype in BM patients . As the CSF concentrations of KP metabolites have been demonstrated in a variety of neurological diseases -10. In tInflammation in the CNS results in the recruitment of activated T-cells and macrophages from the periphery into the neural parenchyma . Activatin vitro studies have attributed antioxidant activity to certain KP metabolites [Reports linking increased KYN metabolism with neuronal injury, particularly during neuroinflammatory disease, have emerged. This neuronal injury would be mainly attributed to the production of neurotoxic metabolites and oxidative molecules, such as superoxide anion and hydrogen peroxide, during the metabolism of KYN -10,25,26abolites -29.de novo NAD synthesis [Due to these roles, the KP has been investigated as a target for therapeutic intervention. The pharmacological modulation of KYN metabolism can affect ynthesis ,30. NAD ynthesis . In thisynthesis .During the course of bacterial meningitis, anti-inflammatory cytokines are produced to mitigate the induced inflammation ,31. Somekynurenine aminotransferase II gene (AADAT + 401C/T) was associated with reduced levels of TNF-α, IL-1β, IL-6, MIP-1 /CCL3 and MIP-1/CCL4 in BM patients; a reduction in cell count with a high correlation with cytokine and chemokine levels was also observed, suggesting a reduced leukocyte recruitment ability in these patients [et al., showing that KYNA improves the outcomes of heatstroke in rats. In this model, KYNA inhibited the expression of inflammatory molecules such as TNF-α and ICAM-1 and enhanced IL-10 levels [Recently, our group demonstrated that a polymorphism in the patients . This SNpatients . In agre AADAT + 01C/T was0 levels . Corrobo0 levels . KYNA wa0 levels .In addition to cytokines, other molecules can regulate the KP. There are known interactions between TRP depletion and nitric oxide (NO) production. Several findings have demonstrated that NO is able to inhibit the IDO enzyme by direct interaction or accelerating proteasomal degradation ,40. FurtAltogether, these findings suggest that IDO has prominent importance during bacterial meningitis due to the following: its capacity to maintain PARP-1 activity through NAD synthesis, protecting against cell death ,16; the In conclusion, our results demonstrate a specific upregulation of the KP in BM patients and suggest that KYN metabolites contribute directly to the disease. Despite the neurotoxic properties reported in the literature to date, we propose a positive and essential role of this metabolic pathway in BM.


The introduction of modern troponin assays has facilitated diagnosis of acute myocardial infarction due to improved sensitivity with corresponding loss of specificity. Atrial fibrillation (AF) is associated with elevated levels of troponin. The aim of the present study was to evaluate the diagnostic performance of troponin I in patients with suspected acute coronary syndrome and chronic AF.Contemporary sensitive troponin I was assayed in a derivation cohort of 90 patients with suspected acute coronary syndrome and chronic AF to establish diagnostic cut-offs. These thresholds were validated in an independent cohort of 314 patients with suspected myocardial infarction and AF upon presentation. Additionally, changes in troponin I concentration within 3 hours were used.In the derivation cohort, optimized thresholds with respect to a rule-out strategy with high sensitivity and a rule-in strategy with high specificity were established. In the validation cohort, application of the rule-out cut-off led to a negative predictive value of 97 %. The rule-in cut-off was associated with a positive predictive value of 88 % compared with 71 % if using the 99th percentile cut-off. In patients with troponin I levels above the specificity-optimized threshold, additional use of the 3-hour change in absolute/relative concentration resulted in a further improved positive predictive value of 96 %/100 %.Troponin I concentration and the 3-hour change in its concentration provide valid diagnostic information in patients with suspected myocardial infarction and chronic AF. With regard to AF-associated elevation of troponin levels, application of diagnostic cut-offs other than the 99th percentile might be beneficial.The online version of this article (doi:10.1186/s12916-015-0410-8) contains supplementary material, which is available to authorized users. Acute chest pain is one of the most common causes of admission to an emergency department . IntroduIn addition to the classical cascade of plaque rupture and subsequent thrombus formation followed by necrosis of dependent myocardium, other causes of myocardial infarction (MI), such as myocardial injury due to an imbalance of oxygen supply and demand, have been defined. According to the universal definition of myocardial infarction used worldwide , 7, whicAtrial fibrillation (AF) is the most common arrhythmia in the general population, and due to changing demographics, the number of patients with AF is continuously increasing , 10. A lThe primary aim of the present study was to evaluate the clinical performance of a contemporary sensitive troponin I assay in diagnosing type 1 MI in patients presenting with AF and clinical symptoms suggestive of an ACS in two large, distinct study cohorts. An additional objective was to define and validate an optimal diagnostic troponin I threshold in such a setting. Precise identification is of utmost importance in these individuals, as patients at risk with type 1 MI benefit from an early and aggressive MI-specific treatment  comparedderivation cohort to define optimal troponin I thresholds for identification of MI type 1 in patients with AF experiencing chest pain. Second, these specifically calculated diagnostic cut-offs were applied to a large, multicenter, real-world population of patients presenting with suspected ACS and AF as a validation cohort to test for the diagnostic performance compared with the standard 99th percentile troponin I cut-off. Only individuals of the two cohorts with available investigational troponin I measurements as well as available electrocardiogram on admission were used in the present post hoc analyses.The present study investigated the clinical application of a contemporary sensitive troponin I assay in patients presenting with concomitant AF and suspected ACS. To describe the optimal usage of troponin I determination in such a setting, two independent prospective cohorts were used. First, an ACS registry was used as a The Bad Nauheim ACS registry served as the derivation cohort. In this registry patients were enrolled consecutively from April 2003 until November 2006 and referred for early coronary angiography or primary percutaneous coronary intervention due to a potential ACS with an episode of chest pain within the prior 48 hours. Patients were either admitted directly by the emergency medical system or transferred from community hospitals. Medical history and data on the acute medical situation were assessed as described earlier [A large, multicenter, all-comers study was used as a validation cohort. Patients were enrolled consecutively who presented with pain suspected to be due to ACS at the chest pain unit of the Johannes Gutenberg-University Medical Centre in Mainz, the Federal Armed Forces Hospital in Koblenz, or the University Hospital Hamburg-Eppendorf in Hamburg between January 2007 and December 2008. An electrocardiogram was acquired directly upon admission, and blood was drawn at admission and after 3 hours. Data on the acute medical situation and the patients’ medical history were assessed as described earlier . ParticiThe final diagnosis of MI was adjudicated according to the universal definition of MI , 8 in boRoutine laboratory parameters, including creatinine, were measured immediately after blood withdrawal by standardized methods in both study cohorts. Additionally, venous blood samples were collected on admission and after 3 hours in the validation cohort, processed immediately, and stored at −80 °C until assay.In-house troponin, represented by cardiac troponin T in the derivation cohort and two study centers of the validation cohort, was measured in serum using a conventional commercial one-step electrochemiluminescence immunoassay . The lower detection limit of this assay is 0.01 ng/mL, the 99th percentile is <0.01 ng/mL, and the lowest concentration measurable with a coefficient of variation (CV) <10 % is 0.03 ng/mL, which was used as the diagnostic cut-off. At the third study center of the validation cohort troponin I was used as in-house troponin for adjudication of final diagnosis. A conventional troponin I assay was used  with lower detection limit of 0.040 ng/mL and measuring range of 0.04 to 40.0 ng/mL. The 99th percentile is 0.07 ng/mL, and the 10 % CV used as the diagnostic cut-off is 0.14 ng/mL.As investigational troponin a contemporary sensitive troponin I assay  was measured in both cohorts. For this assay, the level of detection is 0.01 ng/mL with a measuring range of 0.01–50.0 ng/mL, and the 99th percentile and the lowest concentration with CV of 10 % is 0.032 ng/mL . This inContinuous skewed variables are described as median and interquartile range and symmetric variables are presented as mean with standard deviation. Receiver operating characteristic (ROC) curves based on continuous troponin I levels were calculated in both cohorts.In the derivation cohort of 90 patients with AF, optimized thresholds were computed by determining the cut-offs that maximized i) the sum of specificity and sensitivity  and those that yielded ii) 90 % sensitivity and iii) 90 % specificity, respectively. In addition we have considered the 99th percentile of the assay as cut-off. The uncertainty of choices i) – iii) is reflected by 95 % confidence intervals that were obtained nonparametrically by taking the 2.5 % and 97.5 % percentiles from 2,000 bootstrap replications of these evaluations.These cut-offs have been applied to the validation cohort of 314 patients. Sensitivity, specificity, positive predictive values (PPVs), and negative predictive values (NPVs) for the individual patient groups were calculated by applying the different troponin I cut-off values and consecutively calculating the corresponding values from a two-by-two factorial design. Corresponding confidence intervals for all these proportions were calculated according to Clopper-Pearson.Relative and absolute changes in the concentration between admission and after 3 hours  in the validation cohort were regarded as new biomarkers that gave rise to analogously defined cut-offs. Empirical kernel density estimations of these absolute and relative changes have been plotted for both subgroups, MI and non-MI patients Fig. , where tP < 0.05 was considered significant. All analyses were carried out using R 2.15 and 3.1.1 .A one-sided The derivation cohort included a total of n=90 patients with AF out of n=1,574 patients presenting with suspected MI as published . Of thesThe validation cohort consisted of n=314 patients with documented AF in the ECG obtained at presentation out of a total of n=1,818 patients presenting consecutively with symptoms suggestive of an acute spontaneous MI . After dIn these 314 patients presenting with AF, data on previously known arrhythmias, based on information given by the patients, was available in 66 patients leading to a subcohort of 248 individuals with presumably new onset AF of whom 52 had the final diagnosis MI type 1.Baseline characteristics of the derivation and validation cohorts are provided in Table Use of a single contemporary sensitive troponin I determination upon presentation to the emergency department in patients with AF and symptoms suggestive of an ACS of the derivation cohort yielded an area under the curve (AUC) in the ROC analyses of 0.905  for identification of type 1 MI. Based on these ROC analyses and using an unweighted approach by maximizing the Youden index led to an optimized diagnostic threshold of 0.04 ng/mL  to identify type 1 MI. For a rule-out strategy the cut-off associated with a sensitivity of 90 % was calculated to be 0.019 ng/mL . Regarding valid rule-in of type 1 MI, the optimal cut-off associated with a specificity of 90 % was calculated at 0.09 ng/mL . Figure Application of the three different diagnostic thresholds that were calculated based on the derivation cohort with respect to overall performance (0.04 ng/mL), rule-out 0.019 ng/mL), and rule-in (0.09 ng/mL) as well as the proposed 99th percentile cut-off of the assay 0.032 ng/mL) in a large, real-world validation cohort of patients presenting with acute chest pain or equivalent symptoms and chronic AF led to the diagnostic performance presented in Table  ng/mL in9 ng/mL, P < 0.001) if comparing the 99th percentile cut-off and the specificity-optimized cut-off. Concerning rule-out, a slight increase in the NPV from 0.96 to 0.97 (P = 0.169) if comparing the 99th percentile cut-off and the sensitivity-optimized cut-off was observed.This approach of different diagnostic thresholds relating to different clinical strategies improved potential rule-in with an increase of the PPV from 0.71 to 0.88  for the absolute change and of 0.3 %  for the relative change was calculated via ROC analyses. Optimization for specificity (90 %) leads to a 0.023 ng/mL  absolute change and a 40 %  relative change threshold.The diagnostic performance of these different absolute and relative changes in troponin I concentration within 3 hours alone and in combination with troponin I determined directly upon admission is presented in Tables The availability of robust, sensitive cardiac troponin assays has been shown to substantially improve the timely diagnostic workup , 4 as werule-in. Measurement of troponin I on admission and application of the 99th percentile thresholds in 314 patients with chronic AF taken from a large, multicenter, real-world cohort of patients with suspected ACS led to the expected lower specificity with a positive predictive value of 71 % compared with 80.9 % [rule in patients, use of the calculated specificity-optimized threshold improved this positive predictive value to 88 %. Concerning rule-out of type 1 MI, troponin I determination 3 hours after admission was associated with a negative predictive value of 100 % if using the 99th percentile cut-off or an optimized threshold. Based on the assumption that myocardial necrosis is associated with a larger increase in cardiac troponin compared with troponin release due to ischemic imbalance, use of the change in troponin concentration should further facilitate diagnostic discrimination. This is supported by our data showing that patients with type 1 MI have larger changes in troponin I concentration within 3 hours after presentation compared with patients without type 1 MI   irrespecSeveral limitations should be considered. The final diagnosis of type 1 MI, aside from being based on clinical parameters and imaging data, is based on serial troponin measurements in line with the universal definition of MI; considering the influence of AF on troponin levels, this might introduce a bias. Further studies, such as those using magnetic resonance imaging to define the diagnosis of MI according to loss of myocardial tissue, might therefore further elucidate the value of troponin testing in patients with AF and suspected MI. Several patients in the derivation cohort have been transferred from a tertiary hospital leading to higher median time between onset of symptoms and presentation which might have an influence on troponin concentrations upon admission and therefore on threshold calculation. In contrast, the high percentage of coronary angiography-confirmed type 1 MI in the derivation cohort demonstrates the low percentage of other causes of troponin elevation by parameters influencing ischemic myocardial injury. These are important aspects which need to be reflected by interpreting the data.In addition, the proportion of MI in the validation cohort is comparable to those of other European all-comer studies but higher than those in non-European cohorts, which might limit the generalizability of the result.The use of troponin I assayed upon admission and after 3 hours to obtain data on troponin kinetics provides valid diagnostic information in patients with suspected acute spontaneous myocardial infarction and chronic atrial fibrillation. Considering the higher troponin I levels observed in patients with atrial fibrillation, application of diagnostic cut-offs other than the 99th percentile might be beneficial.


Simple surgical intervention advocated by the World Health Organization can alleviate trachomatous trichiasis (TT) and prevent subsequent blindness. A large backlog of TT cases remain unidentified and untreated. To increase identification and referral of TT cases, a novel approach using standard screening questions, a card, and simple training for Community Treatment Assistants (CTAs) to use during Mass Drug Administration (MDA) was developed and evaluated in Kongwa District, a trachoma-endemic area of central Tanzania.p < 0.05). While specificity was above 90% for both groups, the sensitivity for the novel screening tool was 31.2% compared to 5.6% for the usual care group (p < 0.05).A community randomized trial was conducted in 36 communities during MDA. CTAs in intervention villages received an additional half-day of training and a TT screening card in addition to the training received by CTAs in villages assigned to usual care. All MDA participants 15 years and older were screened for TT, and senior TT graders confirmed case status by evaluating all screened-positive cases. A random sample of those screened negative for TT and those who did not present at MDA were also evaluated by the master graders. Intervention CTAs identified 5.6 times as many cases (n = 50) as those assigned to usual care  is recommended by the WHO as a cost-effective strategy to mitigate blinding trachoma. However, a large surgical backlog exists and many individuals suffering with TT remain unknown to the health system. To identify TT cases, we designed a standard set of screening questions, a card, and simple training for Community Treatment Assistants (CTAs) to identify trichiasis during community-wide Mass Drug Administrations (MDA). To evaluate the sensitivity, specificity, and positive predictive value of this approach, we conducted a community randomized trial in 36 communities in trachoma-endemic Kongwa District, Tanzania. Additional training and the use of a TT screening card increased the sensitivity of TT identification and resulted in more cases identified compared to the usual training of CTAs. The positive predictive value was low, indicating a need for further verification of TT cases identified by the enhanced screening. MDA appears to be a good opportunity for TT screening by CTAs, but further training to improve screening sensitivity is suggested. Chlamydia trachomatis leading to the development of conjunctival scarring in a portion of individuals [Trachoma, the leading cause of infectious blindness, affects an estimated 84 million individuals worldwide . Althougividuals –5. As scividuals .Surgery to prevent blindness from TT, Antibiotics to reduce the infectious burden in children, and Facial cleanliness along with Environmental improvement to reduce transmission of C. trachomatis [To control trachoma and mitigate potentially blinding TT, the Alliance for Global Elimination of Trachoma by 2020 (GET 2020) and World Health Organization (WHO) advocate the multifaceted SAFE strategy, an approach consisting several components: chomatis . As partchomatis –10.Surgical management of TT involves a tarsal rotation procedure, which can be performed by a trained surgical technician within the community ,12. HoweThis community randomized trial sought to develop and test a novel community-based screening approach designed to enhance the ability of CTAs to recognize and report TT cases during MDA. We developed a set of standard screening questions, a TT screening and recognition card, along with additional training. In the randomly selected intervention villages, the approach was introduced immediately prior to MDA, during training of CTAs. Control communities had standard training only. To evaluate the new approach, we compared the number of cases found, as well as the sensitivity and specificity, between the communities with enhanced training group of CTAs and the communities with the control group of CTAs.A controlled community randomized trial design was implemented, using two parallel arms to compare the novel TT screening tool (intervention) to usual care (control) during mass drug administration. The trial was conducted in 36 geographically distinct communities in Kongwa district, Tanzania, between January and November of 2013. Communities were selected based on eligibility to receive antibiotic treatment through MDA.The community served as the unit of randomization, and enrolled villages were randomly allocated on a 1:1 basis to either intervention (n = 18) or usual care (n = 18) arms.This study was built on to the baseline study of another project, which enrolled 52 communities in Kongwa Tanzania, of whom 36 were to receive MDA. Communities were eligible for this study if they were eligible to receive MDA, and had the consent of village leadership. Within the 36 study villages, individual members of the community were eligible for TT screening if they were 15 years of age or older.In each community, 2–10 CTAs were chosen to distribute antibiotic treatment. CTAs were selected by local leadership based on standing in the community and basic reading skills. Supervisory staff conducted training sessions for CTAs prior to MDA, including how to measure heights of children for antibiotic dosage, to mix liquid doses, to observe treatment provided, to log and manage medication, and to ask about TT and refer self-identified cases of trichiasis for surgery at the local hospital.In communities assigned to the intervention arm, CTAs received an expanded training session on trichiasis case recognition in addition to the standard instruction on MDA protocol. This additional training lasted one afternoon and consisted of both didactic learning and role-play designed to teach a basic understanding of trichiasis and focusing on four key components:What is trichiasis and how does it cause blindness?What does trichiasis look like, and what are common symptoms?How do you look for trichiasis, using a torchWhat does the surgery involve, and what can one expect after surgery?How is an individual added to a referral form, and what will happen after he or she is added?In addition to this training, CTAs received a TT screening card containing simple questions on symptoms in Swahili, instructions on performing a TT exam, and images depicting proper TT examination and examples of TT cases for reference . InterviIn communities assigned to the usual care arm, CTAs received the normal half-day of training on MDA procedures and a half hour basic overview of trichiasis and TT case recognition. CTAs were directed to ask during MDA whether individuals self-identified as having TT, and to record suspected cases of TT on the treatment log for each person. CTAs in usual care communities were given Maglite Solitaire torches identical to those used by CTAs in intervention villages but no other instructions.Following completion of MDA in each village, a survey was conducted by an experienced trichiasis grader, the “gold standard.” He was masked to TT grade assigned during MDA screening. We also attempted to mask him to the village intervention assignment, but once in the village it was difficult to maintain that masking. The follow-up survey was designed to:Confirm suspected TT cases screened positive by CTAsIn a random sample of those screened negative for TT by CTAs, confirm they are truly negative in order to estimate missed TT cases during screeningIdentify TT in a random sample of those who did not attend MDA in order to estimate the number of TT cases missed due to lack of MDA attendanceFor the survey, all TT cases reported by CTAs were re-examined to confirm their status. In addition, treatment records were used to randomly select a sample of 100 adults per community initially screened as negative for TT to determine the rate of cases missed by the CTAs during MDA. A random sample of 50 adults per community who did not attend MDA was also selected to estimate the number of TT cases missed because of absence from the MDA.To conduct the follow-up surveys, invitations were sent to selected individuals following MDA. Invitations notified individuals of the time, date, and location within the community of the survey and requested their attendance. On the day of the survey, the master TT grader examined all presenting individuals for trichiasis. For those found to have confirmed TT, the option of surgical treatment was explained, and interested individuals were added to a surgical referral list for the upcoming surgical camps to be held after the MDA program had concluded. After the initial day of survey, the TT grader continued follow-up on an individual basis until at least 70% participation was achieved for those who had attended MDA.CTA screening results obtained during MDA were recorded on paper forms in the mass treatment books. These forms were entered into a customized Access database by staff with double data entry performed for fields of particular importance. All ocular examinations for TT were entered into Samsung Tab 2.0 tablets using a customized ODK electronic form. The ocular exam form included identification information, personal details, and TT status. All electronic data was uploaded into a customized Access database and transferred electronically to Johns Hopkins University for analysis.All statistical analysis was conducted using SAS, and Stata version 12 –19.To assess comparability between study arms, differences in demographic and socioeconomic characteristics at the community level were evaluated using Wilcoxson rank sum tests. Additionally, characteristics of participants who were re-examined were compared to those lost to follow up within each arm, using chi-square tests or Fisher’s exact test as appropriate.Sensitivity, specificity, positive predictive value and negative predictive value were estimated for the intervention and usual care arms using a 2 × 2 table comparing the extrapolated results from the samples verified from the Master Grader with those obtained by the CTAs during MDA, as depicted below in Values used in these calculations as well as in the estimation of disease prevalence were obtained using a simple extrapolation method applying the findings from the random samples in the survey to the study population obtained from the full census of the community. This extrapolation was performed as follows:A and B in Proportions of confirmed TT cases in groups initially screened positive by CTAs and successfully followed up with by the master grader were extrapolated to the total number of individuals initially screened positive by CTAs in each arm, included those lost to follow-up .A simple extrapolation was deemed appropriate after review of the samples’ representativeness of the respective study population based on age and gender indicators. This method of extrapolation allowed for the comparison of confirmed cases to the estimated total missed cases in both arms, accounting for the rescreening of all individuals initially screened positive and only a sample of individuals initially screened negative.Oral informed consent was obtained for eligible adults prior to screening. All study protocols and procedures were approved by the Johns Hopkins Institutional Review Board and the National Institute for Medical Research in Tanzania. This randomized trial is registered in the ClinicalTrials.gov database under identifier NCT01783743.A total of 27,473 individuals in the 36 villages holding MDA were eligible for this study, of whom 19,607 (71.4%) attended MDA and were screened for trichiasis. p = 0.56).Baseline characteristics of these villages were compared between arms , with noThe master trichiasis grader re-examined at follow up 3,233 (81.6%) of the 3,962 individuals who were initially screened during MDA and who were chosen for follow-up. Additionally, 1,695 individuals who did not attend MDA were randomly selected for follow-up to estimate the number of TT cases missed during MDA. Of these 1695 persons, 1,149 (67.8%) were examined. Loss to follow-up was attributed primarily to refusal or being away from the home, as many residents travel long distances to farmland during the planting and harvest seasons. n = 409, (4.3%)) than those assigned to usual care , p < 0.01) (p = 0.070). Among those that we could verify, CTAs in the intervention communities identified 5.9 times as many true cases as did those in the control arm (n = 50 (0.525%) and 9 (0.089%), respectively), but also had almost ten times as many false positives (n = 316 vs. 32). Within the random sample of individuals screened negative by CTAs, the proportions of missed TT cases were not significantly different between the two arms (p = 0.269), although fewer cases were missed in the intervention group (1.6%) compared to usual care (2.1%).After conclusion of MDA, CTAs in the intervention arm reported a significantly higher number of suspected cases ( < 0.01) . The prop < 0.05), indicating that the intervention significantly increased the ability of CTAs to identify TT cases. PPV of this screening method did not statistically differ from that of usual care, although both were low . The specificity in the usual care arm was higher (99.7%) than that of the intervention , although specificity for both groups was high. Based on case finding and the estimates derived from the random samples, the prevalence of TT in the study communities in those aged 15 and older was estimated to be 2.2% in both arms.The sensitivity, specificity, and positive predictive value (PPV) for both groups were calculated using extrapolated values from the follow-up survey . SensitiOf the random sample of individuals who did not present to MDA, 14 (1.2%) of the 1,149 individuals examined had TT.n = 5) answered the questions negatively again despite having visible TT. However, those who had not been asked questions during MDA (n = 5) and those who had been both interviewed and examined (n = 5) answered positively to at least one interview question.Six months after the follow-up survey, we re-contacted 19 of the 23 TT cases who were screened negative by the intervention CTAs to determine possible reasons for being incorrectly screened during MDA. Four persons were quite elderly and had no recollection of attending MDA or being screened despite mass treatment logs noting their attendance and completion of screening. Of the remaining 15 individuals, 5 (33%) stated that they had not been asked screening questions, 5 (33%) said they had been asked the questions but had not been examined, and 5 (33%) said they had received both questions and examination. When these 15 individuals were asked again the questions using the TT screening card, those who had initially been screened out based on the interview questions  in national onchocerciasis programs in Nigeria and South Sudan left after their first year due largely to a lack of incentives offered ,28. In oIn summary, though surgical management of trichiasis has been shown to be cost-effective, the identification of cases needing surgery remains challenging and often resource-intensive. Our study evaluated an innovative approach to TT screening that sought to take advantage of the efforts already being made by trachoma control programs—the use of MDA—as a stage for TT screening. Additional training and use of a TT screening card significantly increased the ability of community treatment assistants to recognize and refer TT cases during MDA; however, further efforts are needed to improve case detection and reduce the number of false positive cases.