Comparison of mesencephalic free-floating tissue culture grafts and cell suspension grafts in the 6-hydroxydopamine-lesioned rat

Ventral mesencephalon (VM) of fetal rat and human origin grown as free-floating roller-tube (FFRT) cultures can survive subsequent grafting to the adult rat striatum. To further explore the functional efficacy of such grafts, embryonic day 13 ventral mesencephalic tissue was grafted either after 7 days in culture or directly as dissociated cell suspensions, and compared with regard to neuronal survival and ability to normalize rotational behavior in adult rats with unilateral 6-hydroxydopamine (6-OHDA) lesions. Other lesioned rats received injections of cell-free medium and served as controls. The amphetamine-induced rotational behavior of all 6-OHDA-lesioned animals was monitored at various time points from 18 days before transplantation and up to 80 days after transplantation. Tyrosine hydroxylase (TH) immunostaining of the histologically processed brains served to assess the long-term survival of grafted dopaminergic neurons and to correlate that with the behavioral effects. Additional cultures and acutely prepared explants were also fixed and stored for histological investigation in order to estimate the loss of dopaminergic neurons in culture and after transplantation. Similar behavioral improvements in terms of significant reductions in amphetamine-induced rotations were observed in rats grafted with FFRT cultures (127%) and rats grafted with cell suspensions (122%), while control animals showed no normalization of rotational behavior. At 84 days after transplantation, there were similar numbers of TH-immunoreactive (TH-ir) neurons in grafts of cultured tissue (775 +/- 98, mean +/- SEM) and grafts of fresh, dissociated cell suspension (806 +/- 105, mean +/- SEM). Cell counts in fresh explants, 7-day-old cultures, and grafted cultures revealed a 68.2% loss of TH-ir cells 7 days after explantation, with an additional 23.1% loss after grafting, leaving 8.7% of the original number of TH-ir cells in the intracerebral grafts. This is to be compared with a survival rate of 9.1% for the TH-ir cells in the cell-suspension grafts. Immunostaining for the calcium-binding proteins calretinin, calbindin, and parvalbumin showed no differences in the neuronal expression of these proteins between the two graft types. In conclusion, we found comparable dopaminergic cell survival and functional effects of tissue-culture grafts and cell-suspension grafts, which currently is the type of graft most commonly used for experimental and clinical grafting. In this sense the result is promising for the development of an effective in vitro storage of fetal nigral tissue, which at the same time would allow neuroprotective and neurotrophic treatment prior to intracerebral transplantation.

Septic shock resuscitation: what goals and how to achieve them?

The need to achieve adequate tissue oxygen delivery early in patients with septic shock is well established. However, it is less well recognized that tissue hypoperfusion can exist despite normalization of systemic hemodynamics. Efforts to resuscitate septic patients until adequate tissue perfusion has been achieved can potentially improve outcome. In a multicenter study, 130 patients with septic shock were resuscitated within 12 hours of diagnosis using a protocol including goals for mean arterial and pulmonary artery occluded pressures, urinary output, arterial pH, and hemoglobin goals. They were then randomly assigned to further resuscitation with either a cardiac index (>or= 3 l/minute per m2) or a gastric mucosal pH (>or= 7.32) target. The intensive care unit length of stay and 28-day mortality did not differ between groups, but more patients in the cardiac index group were in the target range, both at baseline and after resuscitation, as compared with the gastric mucosal pH group. In contrast to cardiac index, gastric mucosal pH at baseline and at 24 and 48 hours predicted mortality. Whether other targets for the chosen variables, or different and--in particular--earlier resuscitation efforts would have favored one group cannot be concluded from the data provided.

Intensive insulin treatment

Hyperglycaemia is common in acute illness and more severe hyperglycaemia is associated with worse outcomes in critically ill patients in general and after acute myocardial infarction, stroke, and trauma. Normalization of blood glucose by intensive insulin therapy has been shown to reduce morbidity and mortality in one study in surgical intensive care patients; a subsequent study in medical intensive care patients resulted in reduced morbidity but not a reduction in mortality. Multicentre studies and current meta-analyses in the critically ill have not demonstrated improved outcomes when normalization of blood glucose was targeted; furthermore all studies to date have detected an increased risk of hypoglycaemia in patients subjected to intensive insulin therapy. At present, universal treatment guidelines or recommendations to target strict normoglycaemia must be considered premature. Further data will be available after the completion of the NICE-SUGAR study which has recruited 6103 patients; the NICE SUGAR study will add significant power to future meta-analyses and may help define the role of intensive insulin therapy in critically ill patients.

Selecting control genes for RT-QPCR using public microarray data

BACKGROUND: Gene expression analysis has emerged as a major biological research area, with real-time quantitative reverse transcription PCR (RT-QPCR) being one of the most accurate and widely used techniques for expression profiling of selected genes. In order to obtain results that are comparable across assays, a stable normalization strategy is required. In general, the normalization of PCR measurements between different samples uses one to several control genes (e.g. housekeeping genes), from which a baseline reference level is constructed. Thus, the choice of the control genes is of utmost importance, yet there is not a generally accepted standard technique for screening a large number of candidates and identifying the best ones. RESULTS: We propose a novel approach for scoring and ranking candidate genes for their suitability as control genes. Our approach relies on publicly available microarray data and allows the combination of multiple data sets originating from different platforms and/or representing different pathologies. The use of microarray data allows the screening of tens of thousands of genes, producing very comprehensive lists of candidates. We also provide two lists of candidate control genes: one which is breast cancer-specific and one with more general applicability. Two genes from the breast cancer list which had not been previously used as control genes are identified and validated by RT-QPCR. Open source R functions are available at http://www.isrec.isb-sib.ch/~vpopovic/research/ CONCLUSION: We proposed a new method for identifying candidate control genes for RT-QPCR which was able to rank thousands of genes according to some predefined suitability criteria and we applied it to the case of breast cancer. We also empirically showed that translating the results from microarray to PCR platform was achievable.

[Cross-talk between heart and kidney--mechanisms and management of the cardiorenal syndrome from a nephrologists view]

Renal dysfunction represents a frequent comorbidity in patients with in chronic heart failure and is not only a strong predictor of mortality, but also causally linked to the development and progression of CHF. Mechanisms involved in the cross-talk between the kidney and the heart include the up-regulated sympathetic nerve system, activation of the renin-angiotensin-aldosterone system, vasopressin release and decreased activity of arterial baroreceptors and natriuretic peptides resulting in abnormal salt and water retention. The main therapeutic goals for patients with the so-called cardiorenal syndrome is the normalization of volume status while avoiding overdiuresis and renal dysfunction as well as the implementation of an evidence-based pharmacologic treatment to improve patient outcome. If these two goals are not achieved with conventional therapy, renal replacement therapy should be discussed in an interdisciplinary approach. All current renal replacement techniques have proved to be useful in controlling hypervolemia and ameliorating functional cardiac parameters and quality of life in patients with heart failure. Nevertheless, the influence of renal replacement therapy on long-term survival of affected patients has not been addressed in large controlled studies.

Quantifying the periodicty of Inter Movement Intervals of Leg Movements

ims: Periodic leg movements in sleep (PLMS) are a frequent finding in polysomnography. Most patients with restless legs syndrome (RLS) display PLMS. However, since PLMS are also often recorded in healthy elderly subjects, the clinical significance of PLMS is still discussed controversially. Leg movements are seen concurrently with arousals in obstructive sleep apnoea (OSA) may also appear periodically. Quantitative assessment of the periodicity of LM/PLM as measured by inter movement intervals (IMI) is difficult. This is mainly due to influencing factors like sleep architecture and sleep stage, medication, inter and intra patient variability, the arbitrary amplitude and sequence criteria which tend to broaden the IMI distributions or make them even multi-modal. Methods: Here a statistical method is presented that enables eliminating such effects from the raw data before analysing the statistics of IMI. Rather than studying the absolute size of IMI (measured in seconds) we focus on the shape of their distribution (suitably normalized IMI). To this end we employ methods developed in Random Matrix Theory (RMT). Patients: The periodicity of leg movements (LM) of four patient groups (10 to 15 each) showing LM without PLMS (group 1), OSA without PLMS (group 2), PLMS and OSA (group 3) as well as PLMS without OSA (group 4) are compared. Results: The IMI of patients without PLMS (groups 1 and 2) and with PLMS (groups 3 and 4) are statistically different. In patients without PLMS the distribution of normalized IMI resembles closely the one of random events. In contrary IMI of PLMS patients show features of periodic systems (e.g. a pendulum) when studied in normalized manner. Conclusions: For quantifying PLMS periodicity proper normalization of the IMI is crucial. Without this procedure important features are hidden when grouping LM/PLM over whole nights or across patients. The clinical significance of PLMS might be eluded when properly separating random LM from LM that show features of periodic systems.

Aromatase deficiency in a female who is compound heterozygote for two new point mutations in the P450arom gene: impact of estrogens on hypergonadotropic hypogonadism, multicystic ovaries, and bone densitometry in childhood

We report on a female who is compound heterozygote for two new point mutations in the CYP19 gene. The allele inherited from her mother presented a base pair deletion (C) occurring at P408 (CCC, exon 9), causing a frameshift that results in a nonsense codon 111 bp (37 aa) further down in the CYP19 gene. The allele inherited from her father showed a point mutation from G-->A at the splicing point (canonical GT to mutational AT) between exon and intron 3. This mutation ignores the splice site and a stop codon 3 bp downstream occurs. Aromatase deficiency was already suspected because of the marked virilization occurring prepartum in the mother, and the diagnosis was confirmed shortly after birth. Extremely low levels of serum estrogens were found in contrast to high levels of androgens. Ultrasonographic follow-up studies revealed persistently enlarged ovaries (19.5-22 mL) during early childhood (2 to 4 yr) which contained numerous large cysts up to 4.8 x 3.7 cm and normal-appearing large tertiary follicles already at the age of 2 yr. In addition, both basal and GnRH-induced FSH levels remained consistently strikingly elevated. Low-dose estradiol (E2) (0.4 mg/day) given for 50 days at the age of 3 6/12 yr resulted in normalization of serum gonadotropin levels, regression of ovarian size, and increase of whole body and lumbar spine (L1-L4) bone mineral density. The FSH concentration and ovarian size returned to pretreatment levels shortly (150 days) after cessation of E2 therapy. Therefore, we recommend that affected females be treated with low-dose E2 in amounts sufficient to result in physiological prepubertal E2 concentrations using an ultrasensitive estrogen assay. However, E2 replacement needs to be adjusted throughout childhood and puberty to ensure normal skeletal maturation and adequate adolescent growth spurt, normal accretion of bone mineral density, and, at the appropriate age, female secondary sex maturation.

Electrocardiographic changes during stress test in a patient with "Brugada syndrome".

We report a 28 year old man with the Brugada syndrome characterised by an electrocardiographic pattern of a right bundle branch block and an ST segment elevation in the right precordial leads as well as syncope. During an exercise test, we observed a normalization of the ST segment in V2+ while in the postexercise phase, the ST segment elevation in the right leads was established. This is the first case reported of the Brugada syndrome in Mexico, with spontaneous changes on the EKG masked during exercise and apparent during postexercise phase.

The risk of early mortality of polytrauma patients associated to ISS, NISS, APACHE II values and prothrombin time

BACKGROUND: The early hemodynamic normalization of polytrauma patients may lead to better survival outcomes. The aim of this study was to assess the diagnostic quality of trauma and physiological scores from widely used scoring systems in polytrauma patients. METHODS: In total, 770 patients with ISS > 16 who were admitted to a trauma center within the first 24 hours after injury were included in this retrospective study. The patients were subdivided into three groups: those who died on the day of admission, those who died within the first three days, and those who survived for longer than three days. ISS, NISS, APACHE II score, and prothrombin time were recorded at admission. RESULTS: The descriptive statistics for early death in polytrauma patients who died on the day of admission, 1--3 days after admission, and > 3 days after admission were: ISS of 41.0, 34.0, and 29.0, respectively; NISS of 50.0, 50.0, and 41.0, respectively; APACHE II score of 30.0, 25.0, and 15.0, respectively; and prothrombin time of 37.0%, 56.0%, and 84%, respectively. These data indicate that prothrombin time (AUC: 0.89) and APACHE II (AUC: 0.88) have the greatest prognostic utility for early death. CONCLUSION: The estimated densities of the scores may suggest a direction for resuscitative procedures in polytrauma patients.Trial registration: "Retrospektive Analysen in der Chirurgischen Intensivmedizin" StV01-2008.http://www.kek.zh.ch/internet/gesundheitsdirektion/kek/de/home.html.

Ureteroileal Strictures After Urinary Diversion with an Ileal Segment—Is There a Place for Endourological Treatment at All?

Purpose We compared the long-term results of minimally invasive endourological intervention and open surgical revision in patients with a nonmalignant ureteroileal stricture. Materials and Methods We retrospectively evaluated the records of 74 patients (85 renal units) treated for unilateral or bilateral nonmalignant ureteroileal strictures. Overall, 96 endourological and 35 open surgical procedures were performed. Balloon dilatation and Acucise® or Ho:YAG laser endoureterotomy were used as minimally invasive endourological interventions. Open surgical revision with stricture resection and open ureteroileal end-to-side-reanastomosis was the alternate therapy. Treatment success was defined as radiological normalization or improvement of upper urinary tract morphology combined with absent flank pain, infection, ureteral stents or percutaneous nephrostomies. Results Median followup was 29 months (range 2 to 177). The overall success rate was 26% (25 of 96 cases) for endourological intervention vs 91% (32 of 35) for open surgical revision (p <0.001). Subgroup analysis showed a significant difference in the success rate of minimally invasive endourological interventions vs open surgical revision for strictures greater than 1 cm (3 of 52 cases or 6% vs 19 of 22 or 86%, p <0.001). The success rate of endourological and open surgical procedures for strictures 1 cm or less was 50% (22 of 44 cases) and 100% (13 of 13), respectively. After adjusting for multiple preoperative stricture characteristics, only stricture length was strongly and inversely associated with a successful outcome (p <0.001). Conclusions Open surgical revision produces better results than minimally invasive endourological intervention for ureteroileal strictures, particularly those greater than 1 cm. The success rate of endourological intervention is acceptable only for ureteroileal strictures 1 cm or less. Therefore, ureteroileal strictures greater than 1 cm should be primarily managed by open surgical revision.

Robust Proximal Femur Segmentation in Conventional X-Ray Images via Random Forest Regression on Multi-resolution Gradient Features

In this paper, we propose a fully automatic, robust approach for segmenting proximal femur in conventional X-ray images. Our method is based on hierarchical landmark detection by random forest regression, where the detection results of 22 global landmarks are used to do the spatial normalization, and the detection results of the 59 local landmarks serve as the image cue for instantiation of a statistical shape model of the proximal femur. To detect landmarks in both levels, we use multi-resolution HoG (Histogram of Oriented Gradients) as features which can achieve better accuracy and robustness. The efficacy of the present method is demonstrated by experiments conducted on 150 clinical x-ray images. It was found that the present method could achieve an average point-to-curve error of 2.0 mm and that the present method was robust to low image contrast, noise and occlusions caused by implants.

High precision determination of the terrestrial 40K abundance

Recent improvements in the precision of mass spectrometric measurements have reduced the uncertainty of K-Ar and 39Ar-40Ar ages measured on geological materials. Now the major sources of uncertainty are the uncertainties on the 40K decay constant and the absolute abundance of 40K. In order to improve on this situation we determined the abundance of the 40K isotope in terrestrial standards. A ThermoFischer Triton+ thermal ionization mass spectrometer was used for K isotope ratio measurements of the NIST K standard reference materials SRM 918b and SRM 985. Ion beams were measured in Faraday cups with amplifiers equipped with 1E10, 1E11 and 1E12 Ω resistors. Three measurement protocols were used: (A) dynamic measurement with in-run fractionation correction by normalization to the IUPAC recommended isotope ratio 41K/39K = 0.0721677; (B) total evaporation; (C) a modified total evaporation with interblock baseline measurements. Different measurement protocols were combined with different loading procedures. The best results were obtained by loading samples on single tantalum filaments with 0.1M H3PO4. The total ion yields (ionization + transmission) were tested for the evaporation procedures (B) and (C) and ranged up to 48 %. The resulting best estimate for the 40K/39K ratio is 0.000 125 116 ± 57 (2σ), corresponding to 40K/K = (1.1668 ± 8; 2σ) x 10-4.

Frameless linac-based (Novalis TX) stereotactic treatment for vestibular schwannoma that extend distally into the iac (Internal Acoustic Canal): an analysis of the dose to the cochlea

Objectives: We compare the dose parameters between 3 different radiosurgery delivery techniques which may have an impact on cochlea function. Methods: Five patients with unilateral vestibular schwannoma (VS) were selected for this study. Planning procedure was carried out using the BrainLAB® iPlan planning system v. 4.5. For each patient three different planning techniques were used: dynamic arc (DA) with 5 arcs per plan, hybrid arc (HA) with 5 arcs per plan and IMRT with 8 fields per plan. For each technique, two plans were generated with different methods: with the first method (PTV coverage) it was the goal to fully cover the PTV with at least 12 Gy (normalization: 12 Gy covered 99% of the PTV) and with the second method (cochlea sparing) it was the goal to spare the cochlea (normalization: 12 Gy covers 50% of the PTV/V4Gy of cochlea lower than 1%). Plan evaluation was done considering target volume and coverage (conformity and homogeneity) and OAR constraints (mean (Dmean) and maximum dose (Dmax) to cochlea, Dmax to brainstem and cochlea). The total number of monitor units (MU) was analyzed. Results: The median tumor volume was 0.95 cm³ (range, 0.86-3 cm³). The median PTV was 1.44 cm³ (range, 1-3.5 cm³). The median distance between the tumor and the cochlea's modiulus was 2.7 mm (range, 1.8-6.3 mm). For the PTV coverage method, when we compared the cochlear dose in VS patients planned with DA, HA and IMRT, there were no significant differences in Dmax (p = 0.872) and in Dmean (p= 0.860). We found a significant correlation (p< 0.05) between the target volume and the cochlear Dmean for all plans with Pearson's coefficient correlation of 0.90, 0.92 and 0.94 for the DA, HA and IMRT techniques, respectively. For the cochlea sparing method, when we compared the cochlear dose in VS patients planned with DA, HA and IMRT, there were no significant differences in Dmax (p = 0.310) and in Dmean (p= 0.275). However, in this group the V4Gy of the ipsilateral cochlea represents less than 1%. When using the HA or IMRT technique, the homogeneity and conformity in the PTV, but also the number of MUs were increased in comparison to the DA technique. Conclusion: VS tumors that extend distally into the IAC had an equivalent sparing of cochlea with DA approach compared with the HA and IMRT techniques. Disclosure: No significant relationships.

Characterization of microcirculation in multiple sclerosis lesions by dynamic texture parameter analysis (DTPA)

OBJECTIVE Texture analysis is an alternative method to quantitatively assess MR-images. In this study, we introduce dynamic texture parameter analysis (DTPA), a novel technique to investigate the temporal evolution of texture parameters using dynamic susceptibility contrast enhanced (DSCE) imaging. Here, we aim to introduce the method and its application on enhancing lesions (EL), non-enhancing lesions (NEL) and normal appearing white matter (NAWM) in multiple sclerosis (MS). METHODS We investigated 18 patients with MS and clinical isolated syndrome (CIS), according to the 2010 McDonald's criteria using DSCE imaging at different field strengths (1.5 and 3 Tesla). Tissues of interest (TOIs) were defined within 27 EL, 29 NEL and 37 NAWM areas after normalization and eight histogram-based texture parameter maps (TPMs) were computed. TPMs quantify the heterogeneity of the TOI. For every TOI, the average, variance, skewness, kurtosis and variance-of-the-variance statistical parameters were calculated. These TOI parameters were further analyzed using one-way ANOVA followed by multiple Wilcoxon sum rank testing corrected for multiple comparisons. RESULTS Tissue- and time-dependent differences were observed in the dynamics of computed texture parameters. Sixteen parameters discriminated between EL, NEL and NAWM (pAVG = 0.0005). Significant differences in the DTPA texture maps were found during inflow (52 parameters), outflow (40 parameters) and reperfusion (62 parameters). The strongest discriminators among the TPMs were observed in the variance-related parameters, while skewness and kurtosis TPMs were in general less sensitive to detect differences between the tissues. CONCLUSION DTPA of DSCE image time series revealed characteristic time responses for ELs, NELs and NAWM. This may be further used for a refined quantitative grading of MS lesions during their evolution from acute to chronic state. DTPA discriminates lesions beyond features of enhancement or T2-hypersignal, on a numeric scale allowing for a more subtle grading of MS-lesions.

Transconvolution and the virtual positron emission tomograph--a new method for cross calibration in quantitative PET∕CT imaging

PURPOSE Positron emission tomography (PET)∕computed tomography (CT) measurements on small lesions are impaired by the partial volume effect, which is intrinsically tied to the point spread function of the actual imaging system, including the reconstruction algorithms. The variability resulting from different point spread functions hinders the assessment of quantitative measurements in clinical routine and especially degrades comparability within multicenter trials. To improve quantitative comparability there is a need for methods to match different PET∕CT systems through elimination of this systemic variability. Consequently, a new method was developed and tested that transforms the image of an object as produced by one tomograph to another image of the same object as it would have been seen by a different tomograph. The proposed new method, termed Transconvolution, compensates for differing imaging properties of different tomographs and particularly aims at quantitative comparability of PET∕CT in the context of multicenter trials. METHODS To solve the problem of image normalization, the theory of Transconvolution was mathematically established together with new methods to handle point spread functions of different PET∕CT systems. Knowing the point spread functions of two different imaging systems allows determining a Transconvolution function to convert one image into the other. This function is calculated by convolving one point spread function with the inverse of the other point spread function which, when adhering to certain boundary conditions such as the use of linear acquisition and image reconstruction methods, is a numerically accessible operation. For reliable measurement of such point spread functions characterizing different PET∕CT systems, a dedicated solid-state phantom incorporating (68)Ge∕(68)Ga filled spheres was developed. To iteratively determine and represent such point spread functions, exponential density functions in combination with a Gaussian distribution were introduced. Furthermore, simulation of a virtual PET system provided a standard imaging system with clearly defined properties to which the real PET systems were to be matched. A Hann window served as the modulation transfer function for the virtual PET. The Hann's apodization properties suppressed high spatial frequencies above a certain critical frequency, thereby fulfilling the above-mentioned boundary conditions. The determined point spread functions were subsequently used by the novel Transconvolution algorithm to match different PET∕CT systems onto the virtual PET system. Finally, the theoretically elaborated Transconvolution method was validated transforming phantom images acquired on two different PET systems to nearly identical data sets, as they would be imaged by the virtual PET system. RESULTS The proposed Transconvolution method matched different PET∕CT-systems for an improved and reproducible determination of a normalized activity concentration. The highest difference in measured activity concentration between the two different PET systems of 18.2% was found in spheres of 2 ml volume. Transconvolution reduced this difference down to 1.6%. In addition to reestablishing comparability the new method with its parameterization of point spread functions allowed a full characterization of imaging properties of the examined tomographs. CONCLUSIONS By matching different tomographs to a virtual standardized imaging system, Transconvolution opens a new comprehensive method for cross calibration in quantitative PET imaging. The use of a virtual PET system restores comparability between data sets from different PET systems by exerting a common, reproducible, and defined partial volume effect.