Miracle-dz/UniFAR-FTUs huggingface dataset:
Development of Colorimetric Microtiter Plate Assay for Assessment of Antimicrobials against Acanthamoeba

ABSTRACT We have developed and optimized a 96-well microtiter plate assay, based on the reduction of alamarBlue, to assess the efficacies of much needed new antimicrobials against Acanthamoeba species. This assay has been optimized for determination of drug efficacy against two potentially pathogenic species, Acanthamoeba castellanii and Acanthamoeba polyphaga, and has been validated by comparison of their relative susceptibilities to chlorhexidine, a drug widely used to treat Acanthamoeba keratitis. The results demonstrate that the assay is comparable to a manual counting assay and that A. polyphaga is more resistant to chlorhexidine than A. castellanii. Thus, by use of the manual counting assay, 3.125 μM chlorohexidine was almost completely effective against A. castellanii, whereas this concentration was less than 20% effective against A. polyphaga. Similar results were obtained by the alamarBlue assay. The new assay was used to determine the relative susceptibilities of A. castellanii and A. polyphaga to the alkylphosphocholines (APCs) hexadecylphosphocholine (hexadecyl-PC; miltefosine) and octadecylphosphocholine (octadecyl-PC) as well as an alkylgycerolphosphocholine, edelfosine. Both APCs studied were equally effective against A. castellanii, but octadecyl-PC was less effective than hexadecyl-PC against A. polyphaga. Both APCs were more effective than edelfosine against both Acanthamoeba species. A. polyphaga was found to be significantly less susceptible to each of the phosphocholine analogues. The newly described assay offers a number of advantages over those described previously. It is less labor-intensive than previously described assays and is sensitive and rapid, and the results can be read in a nonsubjective manner. As it is based on a standard 96-well, microtiter plate, it is amenable to automation and high throughput.
Diagnostic criteria for inclusion body myositis

Inclusion body myositis (IBM) was first identified as a specific disorder about 40 years ago and is now recognized to be the most frequently presenting primary myopathy in middle age and beyond. Initial characterization was based on the observation of specific pathological features distinguishing it from polymyositis. It was soon appreciated that there were also distinguishing clinical features. The earliest diagnostic criteria were heavily biased towards pathological features, but over time revised criteria have given increasing importance to certain clinical features. Until the specific cause of IBM is determined, and the basic pathogenetic mechanisms are better understood, there can be no diagnostic gold‐standard against which to compare the sensitivity and specificity of any proposed diagnostic criteria, but such criteria are essential to ensure that patients entering clinical, epidemiological, genetic, pathological or therapeutic studies represent a homogeneous population. It is likely that any currently accepted diagnostic criteria will, once a gold‐standard is eventually established, be shown to have ‘missed’ patients with atypical features, but that has to be accepted to make certain that current studies are not contaminated by patients who do not have IBM. In other words, in everyday clinical practice there will be the occasional patient who an experienced myologist strongly suspects has IBM, but does not meet current criteria – the criteria lack sensitivity. But if the criteria are so broad as to include all such atypical cases, they would be likely to include patients who do not in fact have IBM – they would lack specificity. The sensitivity and specificity of existing criteria have been reviewed recently, in so far as it is possible to do so, and found to have high specificity but variable sensitivity.
Isolation and physicochemical characterisation of starch from cocoyam (Colocasia esculenta) grown in Malawi.

BACKGROUND The aim of this study was to determine the physicochemical properties of starches isolated from Malawian cocoyams and compare them with those of cassava and corn starches. RESULTS The purity of the isolated starches varied from 851 to 947 g kg(-1) and pH from 4.93 to 6.95. Moisture, ash, protein, fat and amylose contents ranged from 104 to 132, 0.3 to 1.5, 3.5 to 8.4, 0.9 to 1.6, and 111 to 237 g kg(-1), respectively. Cocoyam starches gave higher potassium and phosphorus but lower calcium levels than the other starches. The shape of starch granules varied from spherical to polygonal with cocoyam starches displaying smaller-sized granules than cassava and corn starches. Cocoyam starches gave a higher wavelength of maximum iodine absorption and blue value but lower reducing capacity values than cassava and corn starches. The extent of acid hydrolysis of the starches also differed. Cocoyam starches exhibited amylopectin molecules of higher molecular weights but amylose molecules of lower molecular weights than cassava and corn starches. Cocoyam starches exhibited lower water absorption capacity and swelling power, paste clarity and viscosity but higher solubility, gelatinisation temperatures and retrogradation tendencies than cassava and corn starches. CONCLUSIONS The physicochemical properties of native Malawian cocoyam starches vary among the different accessions and differ from those of cassava and corn starches.
Comparison of the quantitative performance of a Q-Exactive high-resolution mass spectrometer with that of a triple quadrupole tandem mass spectrometer for the analysis of illicit drugs in wastewater.

RATIONALE Analysis of drugs in wastewater is gaining more interest, as new approaches to estimate drug consumption from the amount of drug residues in wastewater have been proposed. The aim of this study was to compare the quantitative performance of high-resolution mass spectrometry with that of triple quadrupole mass spectrometry. METHODS A Q-Exactive mass spectrometer was operated in full scan (HRFS) (70 000 FWHM) and product scan (HRPS) (17 500 FWHM) modes. The first and third quadrupoles of the QqQ MS/MS instrument were operated at 0.7 FWHM. A mass-extracted window of 5 ppm around the theoretical m/z of each analyte was used to construct chromatograms. An HESI-II ion source was used for the ionization of target compounds. In-line-SPE-LC configuration was used for the extraction and separation of target analytes. RESULTS All three methods showed good linearity and repeatability. High-resolution detection of product ions exhibited better sensitivity and selectivity for some compounds. For most of the tested compounds, LOQs ranged from 0.46 to 20 ng L(-1) . Good agreement between measured and nominal concentrations was observed for most of the compounds at different levels of fortification. Both MS/MS methods showed good selectivity, while HRFS gave some false positive results. CONCLUSIONS The Q-Exactive mass spectrometer proved to be suitable for trace detection and quantification of most of the tested drugs in wastewater, with performance comparable to that of the commonly used MS/MS triple quadrupole, but with better selectivity.
Excess MCM proteins protect human cells from replicative stress by licensing backup origins of replication

The six main minichromosome maintenance proteins (Mcm2–7), which presumably constitute the core of the replicative DNA helicase, are present in chromatin in large excess relative to the number of active replication forks. To evaluate the relevance of this apparent surplus of Mcm2–7 complexes in human cells, their levels were down-regulated by using RNA interference. Interestingly, cells continued to proliferate for several days after the acute (>90%) reduction of Mcm2–7 concentration. However, they became hypersensitive to DNA replication stress, accumulated DNA lesions, and eventually activated a checkpoint response that prevented mitotic division. When this checkpoint was abrogated by the addition of caffeine, cells quickly lost viability, and their karyotypes revealed striking chromosomal aberrations. Single-molecule analyses revealed that cells with a reduced concentration of Mcm2–7 complexes display normal fork progression but have lost the potential to activate “dormant” origins that serve a backup function during DNA replication. Our data show that the chromatin-bound “excess” Mcm2–7 complexes play an important role in maintaining genomic integrity under conditions of replicative stress.
Endometrial Side Population Cells: Potential Adult Stem/Progenitor Cells in Endometrium1

ABSTRACT Uterine endometrium is one of the most important organs for species preservation. However, the physiology of human endometrium remains poorly understood, because the human endometrium undergoes rapid and large changes during each menstrual cycle and it is very difficult to investigate human endometrium as one organ. This remarkable regenerative capacity of human endometrium strongly suggests the existence of adult stem cells, and physiology of endometrium cannot be explained without adult stem cells. Therefore, investigating endometrial stem/progenitor cells should lead to a breakthrough in understanding the normal endometrial physiology and the pathophysiology of endometrial neoplastic disorders, such as endometriosis and endometrial cancer. Several cell populations have been discovered as putative endometrial stem/progenitor cells. Emerging evidence reveals that the endometrial side population (SP) is one of the potential endometrial stem/progenitor populations. Of all the endometrial stem/progenitor cell candidates, the endometrial SP (ESP) is best investigated in vitro and in vivo, and has the largest number of references. In this review, we provide an overview of the accumulating evidence for the ESP cells, both directly from human endometria and from cultured endometrial cells. Furthermore, SP cells are compared to other potential stem/progenitor cells, and we discuss their stem cell properties. We also discuss the difficulties and unsolved issues in endometrial stem cell biology.
Using a Support Vector Machine and a Land Surface Model to Estimate Large-Scale Passive Microwave Brightness Temperatures Over Snow-Covered Land in North America

A support vector machine (SVM), a machine learning technique developed from statistical learning theory, is employed for the purpose of estimating passive microwave (PMW) brightness temperatures over snow-covered land in North America as observed by the advanced microwave scanning radiometer (AMSR-E) satellite sensor. The capability of the trained SVM is compared relative to the artificial neural network (ANN) estimates originally presented in [16]. The results suggest that the SVM outperforms the ANN at 10.65, 18.7, and 36.5 GHz for both vertically and horizontally polarized PMW radiation. When compared against daily AMSR-E measurements not used during the training procedure and subsequently averaged across the North American domain over the 9-year study period, the root-mean-squared error (RMSE) in the SVM output is 8 K or less, while the anomaly correlation coefficient is 0.7 or greater. When compared relative to the results from the ANN at any of the six frequency and polarization combinations tested, the RMSE was reduced by more than 18%, while the anomaly correlation coefficient was increased by more than 52%. Furthermore, the temporal and spatial variability in the modeled brightness temperatures via the SVM more closely agrees with that found in the original AMSR-E measurements. These findings suggest that the SVM is a superior alternative to the ANN for eventual use as a measurement operator within a data assimilation framework.
Serum erythropoietin (EPO) levels correlate with survival and independently predict response to EPO treatment in patients with myelodysplastic syndromes

Abstract: Treatment with recombinant erythropoietin (EPO) can alleviate anaemia in patients with myelodysplastic syndromes (MDS). The present study, based on a long‐term follow‐up of 68 MDS patients (26RA, 16 RAS, 26 RAEB) treated with EPO alone, pinpoints pre‐treatment variables associated with response induction, response duration and overall survival. Response, defined as an increase in haemoglobin >15 g L−1 or eliminated erythrocyte transfusion requirements, was observed in 22 of 66 (33%) evaluable patients. The median response duration was 15 (range 3–64 +) months. Using univariate logistic regression models, responders displayed significantly lower baseline serum EPO levels (S‐EPO), more often normal bone marrow blast cell content (RA/RAS vs. RAEB), normal cytogenetics and no need for erythrocyte transfusion. In a multiple logistic regression model, S‐EPO (P = 0.009), marrow blast content (P = 0.031) and erythrocyte transfusion need (P = 0.024) remained associated with response induction. The probability of response for a patient with S‐EPO >50 U L−1, RA/RAS and no transfusion need was 0.79 (0.53–0.93, 95% CI). The median overall survival time from start of EPO treatment was 26 months, significantly longer for responders than for non‐responders (49 vs. 18 months, P = 0.018). Survival was also predicted by baseline S‐EPO; patients with S‐EPO > 50 U L−1 (n = 50) had a median survival of 17 months, as compared to 65 months for those with S‐EPO >50 U L−1 (n = 14, P = 0.024). The international prognostic scoring system (IPSS) for MDS predicted survival (P = 0.003) and progression to acute leukemia (P < 0.001) but not response to EPO treatment. Furthermore, in a logistic regression model with S‐EPO and IPSS, S‐EPO (but not IPSS) was again a significant predictor for response (P = 0.007). Our data facilitate the optimal selection of MDS patients suitable for EPO treatment and pinpoint S‐EPO as a powerful predictor of response and overall survival in MDS.
Effect of mixed hematopoietic chimerism on cardiac allograft survival in cynomolgus monkeys1

Background. We have previously reported the successful induction of mixed chimerism and long-term acceptance of renal allografts in MHC-mismatched nonhuman primates after nonmyeloablative conditioning and donor bone marrow transplantation. In this study, we extended our regimen to cardiac allotransplantation and compared the immunological responses of heart and kidney allograft recipients. Methods. Five cynomolgus monkeys were conditioned with low-dose total body irradiation (1.5 Gy on days −6 and −5), supplemental thymic irradiation (7 Gy on day −1), antithymocyte globulin (50 mg/kg on days −2, −1, and 0), splenectomy (day 0), donor bone marrow transplantation (day 0), and a 4-week posttransplant course of cyclosporine. Heart allografts from MHC-mismatched donors were transplanted heterotopically on day 0. Results. Two monkeys failed to develop multilineage chimerism and rejected their allografts soon after cyclosporine was stopped (postoperative days [PODs] 43 and 56). Three monkeys developed multilineage chimerism, which persisted 20 to 43 days posttransplant by flow cytometric analysis and to POD 124 by polymerase chain reaction analysis. Allograft survival in these recipients was prolonged to 138, 428, and 509 days, and in vitro mixed leukocyte reaction and cell-mediated lympholysis (CML) assays demonstrated donor-specific hyporesponsiveness. However, in contrast to kidney allograft recipients, long-term heart allograft recipients eventually developed humoral and cellular immunity against the donor and rejected the grafts. At the time of rejection, 1.3% to 9.5% of donor coronary arteries exhibited intimal proliferation. Conclusions. The induction of transient mixed hematopoietic chimerism leads to long-term heart allograft survival in MHC disparate monkeys without chronic immunosuppression. However, unlike kidney allografts, full tolerance to cardiac allografts was not achieved. Organ-specific modifications of the preparative regimen may be necessary to prevent the chronic cellular and humoral immune responses elicited by cardiac allografts.
Nestin Expression in Adult and Developing Human Kidney

Nestin is considered a marker of neurogenic and myogenic precursor cells. Its arrangement is regulated by cyclin-dependent kinase 5 (CDK5), which is expressed in murine podocytes. We investigated nestin expression in human adult and fetal kidney as well as CDK5 presence in adult human podocytes. Confocal microscopy demonstrated that adult glomeruli display nestin immunoreactivity in vimentin-expressing cells with the podocyte morphology and not in cells bearing the endothelial marker CD31. Glomerular nestin-positive cells were CDK5 immunoreactive as well. Western blotting of the intermediate filament-enriched cytoskeletal fraction and coimmunoprecipitation of nestin with anti-CDK5 antibodies confirmed these results. Nestin was also detected in developing glomeruli within immature podocytes and a few other cells. Confocal microscopy of experiments conducted with antibodies against nestin and endothelial markers demonstrated that endothelial cells belonging to capillaries invading the lower cleft of S-shaped bodies and the immature glomeruli were nestin immunoreactive. Similar experiments carried out with antibodies raised against nestin and α-smooth muscle actin showed that the first mesangial cells that populate the developing glomeruli expressed nestin. In conclusion, nestin is expressed in the human kidney from the first steps of glomerulogenesis within podocytes, mesangial, and endothelial cells. This expression, restricted to podocytes in mature glomeruli, appears associated with CDK5.
Antioxidant Treatment Reduces Expansion and Contraction of Antigen-Specific CD8+ T Cells during Primary but Not Secondary Viral Infection

ABSTRACT During many viral infections, antigen-specific CD8+ T cells undergo large-scale expansion. After viral clearance, the vast majority of effector CD8+ T cells undergo apoptosis. Previous studies have implicated reactive oxygen intermediates (ROI) in lymphocyte apoptosis. The purpose of the experiments presented here was to determine the role of ROI in the expansion and contraction of CD8+ T cells in vivo during a physiological response such as viral infection. Mice were infected with lymphocytic choriomeningitis virus (LCMV) and treated with Mn(III)tetrakis(4-benzoic acid)porphyrin chloride (MnTBAP), a metalloporphyrin-mimetic compound with superoxide dismutase activity, from days 0 to 8 postinfection. At the peak of CD8+-T-cell response, on day 8 postinfection, the numbers of antigen-specific cells were 10-fold lower in MnTBAP-treated mice than in control mice. From days 8 to 30, a contraction phase ensued where the numbers of antigen-specific CD8+ T cells declined 25-fold in vehicle-treated mice compared to a 3.5-fold decrease in MnTBAP-treated mice. Differences in contraction appeared to be due to greater proliferation in drug-treated mice. By day 38, the numbers of antigen-specific CD8+ memory T cells were equivalent for the two groups. The administration of MnTBAP during secondary viral infection had no effect on the expansion of antigen-specific CD8+ secondary effector T cells. These data suggest that ROI production is critical for the massive expansion and contraction of antigen-specific CD8+ T cells during primary, but not secondary, viral infection.
Imaging monitored loosening of dense fibrous tissues using high-intensity pulsed ultrasound

Pulsed high-intensity focused ultrasound (HIFU) is proposed as a new alternative treatment for contracture of dense fibrous tissue. It is hypothesized that the pulsed-HIFU can release the contracted tissues by attenuating tensile stiffness along the fiber axis, and that the stiffness reduction can be quantitatively monitored by change of B-mode images. Fresh porcine tendons and ligaments were adapted to an ex vivo model and insonated with pulsed-HIFU for durations ranging from 5 to 30 min. The pulse length was 91 µs with a repetition frequency of 500 Hz, and the peak rarefactional pressure was 6.36 MPa. The corresponding average intensities were kept around 1606 W cm−2 for ISPPA and 72.3 W cm−2 for ISPTA. B-mode images of the tissues were acquired before and after pulsed-HIFU exposure, and the changes in speckle intensity and organization were analyzed. The tensile stiffness of the HIFU-exposed tissues along the longitudinal axis was examined using a stretching machine. Histology examinations were performed by optical and transmission electron microscopy. Pulsed-HIFU exposure significantly decreased the tensile stiffness of the ligaments and tendons. The intensity and organization of tissue speckles in the exposed region were also decreased. The speckle changes correlated well with the degree of stiffness alteration. Histology examinations revealed that pulsed-HIFU exposure probably damages tissues via a cavitation-mediated mechanism. Our results suggest that pulsed-HIFU with a low duty factor is a promising tool for developing new treatment strategies for orthopedic disorders.
Clinical Outcome After Segmental Wire Fixation and Bone Grafting for Repair of the Defects in Multiple Level Lumbar Spondylolysis

The aim of this retrospective study was to assess clinical outcomes after segmental wire fixation and bone grafting for repair of pars defects in patients with multiple-level lumbar spondylolysis. Subjects were 7 patients (5 men and 2 women, mean age 26.7 y) with multiple-level lumbar spondylolysis treated by segmental wire fixation and bone grafting at one of our affiliated institutions between 1983 and 2004. Clinical outcomes were determined by comparing preoperative and postoperative Japanese Orthopaedic Association scores and Mancab criteria, and healing of pars defects was evaluated by radiographic and computed tomography study. The condition involved 2 levels in 5 cases and 3 levels in 2 cases. The mean postoperative follow-up period was 51.0 months. The mean Japanese Orthopaedic Association score improved significantly from 21.29 before surgery to 27.86 after surgery, and the recovery rate was 85.21%. An “excellent” result was achieved in 5 cases, a “good” result in 1 case and a “fair” result in 1 case according to the Macnab criteria. Postoperative radiographs revealed healing of all defects in 4 cases, healing of 3 out of 4 defects in 2 cases, and no healing of any defect in 1 case. Pseudoarthrosis was related to wire breakage, and patients who did not obtain complete healing were patients who did not fully comply with instructions to wear a lumbar corset or restrict activity postoperatively. Segmental wire fixation and bone grafting were shown to be effective for multiple-level lumbar spondylolysis.