Application of adjunct techniques in cytologic material in the diagnosis of rhabdomyosarcoma: case report and review of the literature

A 4(1/2)-yr-old female presented with right-sided pleural effusion and a retroperitoneal mass. Cytologic analysis of the pleural fluid yielded malignant small round blue cells, which were noncohesive, 3-4 times the size of lymphocytes. The malignant cells had hyperchromatic, pleomorphic nuclei with moderate amounts of vacuolated cytoplasm. A few fiber-shaped cells were also seen. Immunostains for desmin, muscle-specific actin were positive; ultrastructural findings of thick and thin actin-myosin filaments confirmed the diagnosis of embryonal rhabdomyosarcoma. This case illustrates the importance of performing appropriate immunohistochemical stains and ultrastructural studies on cytological material to arrive at a definitive diagnosis.     

Is the diaphragm motion probability density function normally distributed?

During radiotherapy treatment planning, the margins given to the clinical target volume to form the planning target volume accounts for internal motion and set-up error. Most margin formulas assume that the underlying distributions are independent and normal. Clinical data suggests that the set-up error probability density function (pdf) can be considered to have an approximately normal distribution. However, there is evidence that internal motion does not have a normal distribution. Thus, in general, a convolution of the two pdfs should be performed to determine the total geometric error. The goals of this article were to (1) determine if the internal motion pdf due to respiration can be characterized using a normal distribution, and (2) if not, determine if the total geometric uncertainty for combining internal motion and set-up error can be characterized by a normal distribution. Sixty fluoroscopy diaphragm motion data sets were obtained using three breathing training types: free breathing, audio instruction, and visual feedback. Diaphragm motion was used as a surrogate for liver and lung cancer motion. The data were analyzed with normality tests in the following groups: (1) single motion measurements, (2) combined motion measurements for each patient, and (3) combined motion measurements for all patients. Following this analysis, the diaphragm motion pdfs were convolved with a set-up error pdf, and the standard deviation of the set-up error pdf at which the total geometric error pdf became normal was determined. At set-up error standard deviation values of at least 0.27 and 0.1 cm for free breathing, 0.57 and 0.42 cm for audio instruction, and 0.55 and 0 cm for visual feedback, for single motion measurements and combined motion measurements for each patient, respectively, total geometric error pdfs became approximately normal. When the motion measurements for all the patients were combined, diaphragm motion pdfs were approximately normal for all feedback types. Therefore, for treatment planning purposes in the absence of individual patient measurements, the diaphragm motion pdf can be considered an approximately normal distribution. However, care should be taken when determining a margin based on individual patients measurements as the total geometric error will, in general, not be normally distributed.     

Comparative evaluation of commonly used laboratory tests for post-mortem diagnosis of rabies

Animal brain samples received at WHO Collaborating Centre laboratory at National Institute of Communicable Diseases (NICD) during the years 1991-2002 were tested by Seller's stain, Fluorescent Antibody Test (FAT) and Mouse Innoculation Test (MIT) as methods of rabies diagnosis. Negri bodies on Seller's staining could be detected in 52.5% of MIT positive brains, the concordance of this test with MIT was found to be 77.8%. FAT was positive in 91.5% of MIT positive brains, though it showed concordance of 95.7% with MIT results in the total samples. 12.2% of the samples were found positive by FAT of which 1/3rd also showed the presence of Negri bodies when MIT was negative i.e. showing that the virus is present in inactivated form. Thus emphasizing the need for timely and proper collection and transportation of specimens for testing. Seller's stain and FAT give reliable diagnosis of rabies in the brain samples in majority of the cases. MIT being time-intensive test, is of academic value only in decision making as regards initiation of Post Exposure Treatment (PET), it is recommended that in cases where Seller's stain and FAT have yielded negative results the decision to initiate PET should give due consideration to the nature and circumstances of the animal bite and other epidemiological features.     

The Clinical Health Economics System Simulation (CHESS): a teaching tool for systems- and practice-based learning.

Academic medical centers face barriers to training physicians in systems- and practice-based learning competencies needed to function in the changing health care environment. To address these problems, at the University of Virginia School of Medicine the authors developed the Clinical Health Economics System Simulation (CHESS), a computerized team-based quasi-competitive simulator to teach the principles and practical application of health economics. CHESS simulates treatment costs to patients and society as well as physician reimbursement. It is scenario based with residents grouped into three teams, each team playing CHESS using differing (fee-for-service or capitated) reimbursement models. Teams view scenarios and select from two or three treatment options that are medically justifiable yet have different potential cost implications. CHESS displays physician reimbursement and patient and societal costs for each scenario as well as costs and income summarized across all scenarios extrapolated to a physician's entire patient panel. The learners are asked to explain these findings and may change treatment options and other variables such as panel size and case mix to conduct sensitivity analyses in real time. Evaluations completed in 2003 by 68 (94%) CHESS resident and faculty participants at 19 U.S. residency programs preferred CHESS to a traditional lecture-and-discussion format to learn about medical decision making, physician reimbursement, patient costs, and societal costs. Ninety-eight percent reported increased knowledge of health economics after viewing the simulation. CHESS demonstrates the potential of computer simulation to teach health economics and other key elements of practice- and systems-based competencies.     

Microdissection genotyping of mixed glial and primitive neuroectodermal central nervous system neoplasm

A 22-year-old man with previous radiation treatment for childhood astrocytoma underwent resection of a right parietooccipital lesion. Histopathology revealed a malignant neoplasm with areas of astrocytic and primitive neuroectodermal components. To resolve the relationship and cellular origin, representative tissue was microdissected from several targets, obtaining a balanced mixture of each element. Nonneoplastic brain parenchyma was separately microdissected to determine polymorphic marker informativeness and to serve as an internal negative control. Despite the relatively small quantity of tissue removed for each microdissection target, sufficient material was available for reliable, balanced, polymerase chain reaction-format genotyping encompassing a panel of tumor suppressor genes and genetic loci associated with these forms of neoplasia. The findings revealed distinct discordant genotypic profiles for each of the neoplastic components. The efficacy of the approach used for molecular analysis of this complex neoplasm and the implication of the genotypic findings are discussed.     

Development and validation of a simple sensitive enzyme immunoassay (EIA) for GH determination in buffalo plasma

A simple and highly sensitive enzymeimmunoassay (EIA) for GH determination in buffalo plasma on microtitreplates using biotin-streptavidin amplification system and the second antibody coating was developed. Biotin was coupled to GH and used to bridge between streptavidin-peroxidase and immobilized antiserum in competitive assay. The EIA was carried out directly in 100 microL buffalo plasma. The GH standards ranging from 0.05 ng/well/100 microL to 12.8 ng/well/100 microL were prepared in hormone free plasma collected from an aged (> 15 years) senile buffalo. The sensitivity of the EIA procedure was 50 pg/well GH. which corresponded to 0.50 ng/mL plasma; the 50% relative binding sensitivity was seen at 800 pg/well/100 microL. Plasma volumes for the EIA, viz., 25, 50, and 100 microL did not influence the shape of standard curve, even though a slight drop in the OD450 was seen with higher plasma volumes. For the biological validation of the assay, 12 Murrah buffalo calves were used. Six of these were administered synthetic bovine growth hormone-releasing factor (10 microg/100 kg body weight, i.v., and the remaining six animals were administered sterile normal saline and kept as controls. Jugular blood samples were collected at -60, -45, -30, -15, -10, -5, 5, 10, 15, and 30 min and, thereafter, at an interval of 15 min using an indwelling jugular catheter, beginning 1 h prior to GRF injection up to 8 h post treatment. In all animals, a peak of GH was recorded within 5 to 20 min of GRF administration, which confirms the biological validation of the EIA. To confirm homogeneity of buffalo GH with bovine GH, a parallelism test was conducted between the buffer standard curve of bovine GH and GH measured from serial dilution of buffalo plasma containing a high level of endogenous growth hormone.     

Algorithm and performance of a clinical IMRT beam-angle optimization system

This paper describes the algorithm and examines the performance of an intensity-modulated radiation therapy (IMRT) beam-angle optimization (BAO) system. In this algorithm successive sets of beam angles are selected from a set of predefined directions using a fast simulated annealing (FSA) algorithm. An IMRT beam-profile optimization is performed on each generated set of beams. The IMRT optimization is accelerated by using a fast dose calculation method that utilizes a precomputed dose kernel. A compact kernel is constructed for each of the predefined beams prior to starting the FSA algorithm. The IMRT optimizations during the BAO are then performed using these kernels in a fast dose calculation engine. This technique allows the IMRT optimization to be performed more than two orders of magnitude faster than a similar optimization that uses a convolution dose calculation engine. Any type of optimization criterion present in the IMRT system can be used in this BAO system. An objective function based on clinically-relevant dose-volume (DV) criteria is used in this study. This facilitates the comparison between a BAO plan and the corresponding plan produced by a planner since the latter is usually optimized using a DV-based objective function. A simple prostate case and a complex head-and-neck (HN) case were used to evaluate the usefulness and performance of this BAO method. For the prostate case we compared the BAO results for three, five and seven coplanar beams with those of the same number of equispaced coplanar beams. For the HN case we compare the BAO results for seven and nine non-coplanar beams with that for nine equispaced coplanar beams. In each case the BAO algorithm was allowed to search up to 1000 different sets of beams. The BAO for the prostate cases were finished in about 1-2 h on a moderate 400 MHz workstation while that for the head-and-neck cases were completed in 13-17 h on a 750 MHz machine. No a priori beam-selection criteria have been used in achieving this performance. In both the prostate and the head-and-neck cases, BAO is shown to provide improvements in plan quality over that of the equispaced beams. The use of DV-based objective function also allows us to study the dependence of the improvement of plan quality offered by BAO on the DV criteria used in the optimization. We found that BAO is especially useful for cases that require strong DV criteria. The main advantages of this BAO system are its speed and its direct link to a clinical IMRT system.     

Muscle synergies during shifts of the center of pressure by standing persons

Movements by a standing person are commonly associated with adjustments in the activity of postural muscles to cause a desired shift of the center of pressure (COP) and keep balance. We hypothesize that such COP shifts are controlled (stabilized) using a small set of central variables (muscle modes, M-modes), while each M-mode induces changes in the activity of a subgroup of postural muscles. The main purpose of this study has been to explore the possibility of identification of muscle synergies in a postural task using the framework of the uncontrolled manifold (UCM) hypothesis employing the following three steps in data analysis: (i) Identification of M-modes: Subjects were asked to release a load from extended arms through a pulley system, resulting in a COP shift forward prior to load release. Electromyographic (EMG) activity of eleven postural muscles on one side of the body was integrated over a 100 ms interval corresponding to the early stage of the COP shift, and subjected to a principal component (PC) analysis across multiple repetitions of each task. Three PCs were identified and associated with a push-back M-mode, a push-forward M-mode and a mixed M-mode. (ii) Calculation of the Jacobian of the system, which relates changes in the magnitude of M-modes to COP shifts using regression techniques: Subjects performed three different tasks (releasing different loads at the back, voluntarily shifting body weight forward and backward, at different speeds) to verify if the relationship between magnitudes of M-modes and COP shifts is task or direction specific. (iii) UCM analysis: Three tasks were chosen (load release in the front, arm movement forward and backward) which were associated with an early shift in COP. A manifold was identified in the M-mode space corresponding to a certain average (across trials) shift of the COP and variance per degree of freedom within the UCM (V_UCM) and orthogonal (V_ORT) to the UCM was computed. Across subjects, V_UCM was significantly higher than V_ORT when analysis at the third step was performed using a Jacobian computed based on a set of tasks associated with a COP shift in the same direction but not in the opposite direction. This result confirms our hypothesis that the M-modes work together as a synergy to stabilize a desired shift of the COP. Forward and backward COP shifts are associated with different synergies based on the same three M-modes.An erratum to this article can be found at     

Abrogation of DTH response and mitogenic lectin- and alloantigen-induced activation of lymphocytes by calcium inhibitors TMB-8 and BAPTA-AM

In vitro treatment of mouse lymphocytes with an intracellular calcium chelator BAPTA-AM significantly decreased lectin Concanavalin-A (Con-A)-induced and mixed lymphocyte reaction (MLR) mediated, alloantigen-induced lymphocyte activation as indicated by decreased percentage of lymphoblasts among the BAPTA-treated lymphocytes. In vivo treatment of mice with intracellular Ca(2+) antagonist TMB-8 was found to substantially impair delayed type hypersensitivity (DTH; cell mediated immune) response, as indicated by decreased footpad swelling on tuberculin challenge of mice sensitized with BCG, after a single treatment with a low dose of 0.01mg of TMB-8 per mouse. Interestingly, a second injection of a higher dose of TMB-8 (0.1mg per mouse) resulted in very significant (p=0.001) abrogation of DTH as indicated by complete absence of swelling of foot pad after PPD challenge in BCG-primed treated mice. All mice in this group showed fully impaired DTH response. Lymphocytes of allosensitized mice gave a significantly higher (p<0.05) MLR response than the na?ve mice. However, a single treatment of allosensitized mice with 0.1mg TMB-8 resulted into a lower MLR response, comparable in magnitude to that of untreated na?ve mice.     

DNA fingerprinting of Ascochyta rabiei with synthetic oligodeoxynucleotides

Summary The ascomycete fungus Ascochyta rabiei, an important pathogen of the grain legume crop chickpea (Cicer arietinum L.) in the Mediterranean region, has not been adequately characterized in molecular terms. We therefore used DNA fingerprinting, with synthetic oligodeoxynucleotides complementary to simple repetitive sequences, to pathotype different isolates of the fungus. Six single-spored A. rabiei isolates were first categorized using a host differential set of nine chickpea genotypes. Seedlings were inoculated under controlled environmental conditions, and disease severity was recorded 9 days after inoculation. DNA was extracted from in vitro-grown mycelia of the six purified fungal isolates, restricted with EcoRI, HinfI, MboII and TaqI, and fingerprinted with radiolabeled (GATA)₄, (GTG)₅, (CA)₈, and (TCC)₅, respectively. High levels of polymorphism were detected with optimal enzyme/probe combinations that allow one to discriminate between the isolates. The potential of DNA fingerprinting with simple repetitive sequences can thus be expanded to the identification of fungal races and pathotypes. The characterization of the geographic distribution and genetic variability of pathotypes will facilitate the selection of suitable host cultivars to be grown in specific regions.